Next Article in Journal
Potential Role of Phytochemicals as Glucagon-like Peptide 1 Receptor (GLP-1R) Agonists in the Treatment of Diabetes Mellitus
Previous Article in Journal
In Vitro Assessment of 177Lu-Labeled Trastuzumab-Targeted Mesoporous Carbon@Silica Nanostructure for the Treatment of HER2-Positive Breast Cancer
Previous Article in Special Issue
Synergistic Effect of Ginsenoside Rh2 Combines with Ionizing Radiation on CT26/luc Colon Carcinoma Cells and Tumor-Bearing Animal Model
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
Pharmaceuticals
Volume 17
Issue 6
10.3390/ph17060734
pharmaceuticals-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles thumb_up
...
Endorse textsms
...
Comment
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Review

Amicis Omnia Sunt Communia: NF-κB Inhibition as an Alternative to Overcome Osteosarcoma Heterogeneity

by
Mariana Medeiros
1,
Sophia Guenka
2,
David Bastos
2,
Karla Laissa Oliveira
3 and
María Sol Brassesco
2,*
1
Cell Biology Department, Ribeirão Preto Medical School, University of São Paulo, Avenida Bandeirantes, 3900-Vila Monte Alegre, Ribeirão Preto 14040-900, São Paulo, Brazil
2
Biology Department, Faculty of Philosophy, Sciences and Letters at Ribeirão Preto, University of São Paulo, Avenida Bandeirantes, 3900-Vila Monte Alegre, Ribeirão Preto 14040-900, São Paulo, Brazil
3
Regional Blood Center, University of São Paulo, Avenida Bandeirantes, 3900-Vila Monte Alegre, Ribeirão Preto 14051-140, São Paulo, Brazil
*
Author to whom correspondence should be addressed.
Pharmaceuticals 2024, 17(6), 734; https://doi.org/10.3390/ph17060734
Submission received: 30 April 2024 / Revised: 29 May 2024 / Accepted: 31 May 2024 / Published: 5 June 2024
(This article belongs to the Special Issue Recent Advances in Nuclear Factor Kappa-B (NF-kB) Inhibitors for Cancer Therapy)
Browse Figures
Versions Notes

Abstract

:
Tumor heterogeneity poses a significant challenge in osteosarcoma (OS) treatment. In this regard, the “omics” era has constantly expanded our understanding of biomarkers and altered signaling pathways (i.e., PI3K/AKT/mTOR, WNT/β-catenin, NOTCH, SHH/GLI, among others) involved in OS pathophysiology. Despite different players and complexities, many commonalities have been described, among which the nuclear factor kappa B (NF-κB) stands out. Its altered activation is pervasive in cancer, with pleiotropic action on many disease-relevant traits. Thus, in the scope of this article, we highlight the evidence of NF-κB dysregulation in OS and its integration with other cancer-related pathways while we summarize the repertoire of compounds that have been described to interfere with its action. In silico strategies were used to demonstrate that NF-κB is closely coordinated with other commonly dysregulated signaling pathways not only by functionally interacting with several of their members but also by actively participating in the regulation of their transcription. While existing inhibitors lack selectivity or act indirectly, the therapeutic potential of targeting NF-κB is indisputable, first for its multifunctionality on most cancer hallmarks, and secondly, because, as a common downstream effector of the many dysregulated pathways influencing OS aggressiveness, it turns complex regulatory networks into a simpler picture underneath molecular heterogeneity.

1. The NF-κB Pathway

The NF-κB transcription factor consists of five members of the Rel family generally found as hetero- or homodimers: RelA (p65), RelB, c-Rel, p50 (NF-κB1/p105), and p52 (NF-κB2/p100) [1]. Each of these proteins present a 300 amino acid N-terminal common domain called Rel Homology Domain (RHD), which is responsible for mediating DNA binding, dimerization, and interaction with IκB (IkappaB kinase or IKK) through the nuclear location signal (NLS) [2,3,4,5,6,7,8,9,10,11] (Figure 1A,B).
Under normal conditions, NF-κB activation occurs in response to many diverse stimuli and depends on a phosphorylation cascade initiating with the IκB proteins (IκBα, IκBβ, IκBε, IκBγ, Bcl-3, p100, and p105), which recognize the RHD domain and keep the nuclear localization signal (NLS) masked, resulting in extranuclear localization [12,13,14,15,16,17,18]. When phosphorylated, IκB proteins are degraded and expose the NLS of NF-κB dimers, resulting in nuclear translocation and gene transcription which is typically rapid and transient. This phosphorylation cascade can occur through two pathways: canonical and non-canonical (Figure 2).
The most common way to activate the canonical pathway is by the induction of tumor necrosis factor α (TNFα) [12,19]. The binding of TNFα to its toll-like receptor recruits adapter proteins responsible for activating the IκB inhibitor complex (IKK) via phosphorylation, which consists of two catalytic subunits IKKα and IKKβ and a regulatory subunit, IKKγ/NEMO kinase [20,21,22]. Once activated, IKK phosphorylates IκB, promoting its ubiquitination via a recombinant carboxyl-terminal ubiquitin hydrolase (β-TrCP) and subsequent degradation by the 26S proteasome [23,24,25]. In this way, NF-κB is translocated to the nucleus where it binds to the consensus sequence 5′-GGGRN W YYCC-3′ (R = purine base, N = any base, W = adenine or thymine, and Y = pyrimidine base) inducing gene transcription [26,27].
Alternatively, while the canonical pathway involves the RelA, c-Rel, RelB, and p50 subunits, the non-canonical pathway is responsible for processing the precursor p100 into the p52 subunit [28,29,30,31,32]. This precursor has a C-terminal processing-inhibitory domain (PID), which resides between the ankyrin repeat domain and acts as an IκB-like regulatory region. To be translocated to the nucleus in the form of a dimer with p50, p100 undergoes processing that removes the PID region. The protein responsible for initiating this process is NF-κB-inducing kinase (NIK), which promotes the activation of IKKα, which in turn phosphorylates p100, triggering its subsequent ubiquitination and degradation via the β-TrCP/26s proteasome. In this way, the newly formed dimer is translocated to the nucleus and binds to DNA at the consensus sequence [33,34,35] (Figure 2A).

2. NF-κB Dysregulation and Cancer

Inappropriate activation of NF-κB may mediate tumorigenesis [36]. Even though NF-κB dysregulation has been mostly ascribed to mutations in its subunits, resulting in the loss of interaction with cytoplasmatic inhibitors, other rearrangements such as amplification and gene fusions (e.g., ZFTA/RELA in supratentorial ependymomas), and crosstalk with other dysregulated signaling pathways have also been described [37,38,39,40,41,42,43,44,45,46,47,48] (Figure 2B).
Nevertheless, irrespective of the underlying mechanism, due to its pleiotropic nature, NF-κB represents an important point of convergence among different tumor hallmarks [49], affecting the expression of several survival factors, antiapoptotic genes, pro-angiogenic and pro-motility (including migration and invasion) genes, and can mediate radio- and chemoresistance mechanisms [50,51,52]. Accordingly, it greatly contributes to the appearance of more aggressive tumors and leads to worse prognoses and lower survival rates for treated patients.
Of note, this transcription factor is a known regulator in the differentiation of chondrocytes, osteoblasts, and osteocytes. Therefore, its dysregulation can initiate and/or promote sarcomagenesis [53,54,55].

Osteosarcoma

Primary bone tumors are rare neoplasms accounting for less than 0.2% of all cancers. Osteosarcoma (OS), chondrosarcoma (ChS), and Ewing sarcoma (EWS) are the most prevalent forms [56,57], occurring in a clear bimodal age distribution, with the first peak occurring in the 10–19 age group and the second peak in elderly (>60 years-old) [58]. Their incidence remains stable worldwide and is estimated around seven individuals per million per year [59,60,61]. In addition to their rarity, these tumors show high morphological heterogeneity and variable biologic behavior [62,63], making their treatment a challenge.
OS is the most common [64], constituting 56% of all existing bone sarcomas [65]. Most patients affected with this tumor comprise children and young adults (<30 years), with a peak incidence during the “puberty growth spurt” phase [66]. However, as mentioned earlier, a second peak incidence can be observed in people over 60 years of age, especially associated with pre-existing conditions, such as Paget’s disease, for example [66,67].
The tumor is characterized by the presence of malignant mesenchymal cells, which synthesize osteoids and/or immature bone [65], being typically found in the metaphysis of long bones, mainly at the distal femur, proximal tibia, and proximal humerus [68].
OS arises mainly on the bone surface (parosteal and periosteal) and less frequently outside the bone in other tissues of mesenchymal origin (extraskeletal). Nevertheless, it usually develops in the intramedullary space with the bone membrane (periosteum) potentially rupturing during periods of accelerated growth [55,69,70]. Also, despite developing in the bone, OS is understood to be a high-grade neoplasm that presents extreme metastasis to the lung [71,72].
Its treatment typically involves neoadjuvant chemotherapy (applied before surgery) with doxorubicin, cisplatin, and high doses of methotrexate, aiming to reduce the tumor volume before resection [73,74]. Subsequently, treatment response is evaluated by determining tumor necrosis through the method described by Huvos and collaborators in 1977 [75,76,77,78]. Surgical resection consists of total tumor ablation or even limb salvage. During such treatment, chemotherapy is stopped for about 2–3 weeks and can be resumed normally thereafter [64]. Then, the last stage of treatment consists of adjuvant or postoperative chemotherapy, used with the goal of exterminating remnant neoplastic cells, improving patients’ survival [79].
Nevertheless, chemoresistance remains a significant barrier to be overcome [80]. Despite the constantly increasing number of therapeutical strategies for neoadjuvant and adjuvant treatment that has allowed the field to successfully achieve a cure in 70% of patients with localized OS, for patients diagnosed with metastatic disease at presentation, survival outcomes have remained unchanged over the past four decades, with less than 30% of patients alive after 5 years [81,82]. Radiotherapy is administered only in palliative cases, when surgical resection is not an option [83].
From this perspective, many efforts have been made on the search for effective biomarkers that can be considered good therapeutic targets for the treatment of this tumor [84]. However, unlike other sarcomas, OS lacks recurrent genetic alterations; instead, it is highly heterogeneous with varied ploidy abnormalities, chromosomal losses and gains, and somatic DNA copy number alterations [85,86]. Inactivation of classical tumor suppressor genes such as TP53, RB1, and hyperactivation oncogenes, including MYC and MDM2, are also common [87,88]. Additionally, epigenomic, transcriptomic, proteomic, metabolomic, and functional genomic approaches have constantly expanded the number of altered signaling pathways in OS. Indeed, alterations in the major signaling pathways, such as PI3K/AKT/mTOR, JAK/STAT, WNT/β-catenin, NOTCH, Hedgehog/Gli, TGF-β, MAPK, and the receptor tyrosine kinases (RTKs) signaling pathways, have been identified in OS development and metastasis [89]. In all cases, the primary consequence of each signaling cascade is the activation of specific target genes by signal-regulated transcription factors. Of note, despite different players and complexities, many surprising and fundamental commonalities in the transcriptional mechanisms by which these pathways control the expression of their target genes have been described, among which NF-κB stands out (Figure 3). As a fact, coupling of signaling pathways enhances the functions of individual pathways and results in a more complex regulatory network [90].
The PI3K/AKT pathway is frequently hyperactivated in OS and contributes to increased proliferation and invasion, inhibition of apoptosis, angiogenesis, and chemoresistance [91,92]. Moreover, activation of this pathway has been associated with lung metastasis and poorer prognosis [93]. Constitutive Akt activity may lead to NF-κB activation. This kinase phosphorylates several substrates and downstream effectors including IkB proteins, which in turn are degraded and allow the exposure of the NLS of NF-κB dimers, their nuclear translocation, and gene transcription [94].
JAK/STAT activation has also been involved in OS development and metastasis. Even though its activation might be indirect (i.e., long non-coding RNAs), it has been demonstrated that while STAT3 signaling inhibits IKK activity in the context of a normal immune response, in tumors, STAT3 prolongs NF-κB nuclear retention through p300-mediated RelA acetylation, thereby interfering with NF-κB nuclear export [95].
Similarly, several reports showed aberrant constitutive activation of the WNT/β-catenin signaling pathway in OS tumor development and metastasis; however, current knowledge remains uncertain because of the high complexity of this pathway [96,97]. Higher β-catenin levels in OS have been associated with poor prognosis, lung metastatic dissemination [98], and stemness [99]. Spiegelman et al., showed that overexpression of β-catenin or Wnt proteins in 293T and HeLa cells increased the expression levels of βTrCP, an E3 ubiquitin ligase receptor that mediates the ubiquitination and subsequent degradation of both β-catenin and IκBα. Thus, increased βTrCP results in enhanced degradation of IκB-α and, consequently, NF-κB transactivation without affecting IKK activity [100]. Moreover, immunoprecipitation assays in murine osteoblasts have revealed that β-catenin can physically interact with the NF-κB subunits (p65 and p50) [101,102]. In line with this notion, the TCF/LEF transcription factors, which are downstream effectors of the WNT pathways and act together with β-catenin, bind to promoters of NF-κB target genes, leading to synergistic upregulation of gene expression [103]. Furthermore, the inhibition of GSK-3β (a key regulator of β-catenin) with SB216763 in OS cells eventually led to the inhibition of the NF-κB pathway and reduced the transcription of its targets [104]. Indeed, inhibition of GSK-3β with the same drug has been shown to repress IκBα phosphorylation, NF-κB (p65) nuclear translocation, and its DNA binding activity [101]. Of note, an inverse relationship was observed between (inactive) p-Ser9-GSK-3β and (active) nuclear p65 levels in OS samples, which denoted lower overall survival to OS patients. In that case, it was proposed that, when GSK-3β is impeded, IκBα is stabilized and retained in the cytoplasm enhancing apoptosis induced via chemotherapy [104].
Coactivation of NF-κB and NOTCH signaling has been previously demonstrated as well [105,106]. Notch activation can induce the expression of a large fraction of classical NF-κB gene targets in T-cell progenitors [107], and as a key player in osteogenic differentiation, bone healing, and in the development of the skeleton [108], its abnormal activation has been observed in most OS clinical specimens with a close relation with poorer prognosis [89,109]. In this regard, NOTCH3 knockdown has been shown to deeply impair proliferation, apoptosis, and invasion in OS cells, while it reduced the number of metastatic lesions in vivo. In addition, the expression of this receptor was considered a prognostic factor correlated with metastasis and poor patient outcome. In the same context, expression of HES1, a downstream effector of NOTCH signaling, was reduced after NOTCH3 silencing in the human OS cell line U2OS [110]. Activated NOTCH1 also induced HES1 and sustained NF-κB-signaling through NFKB1, NFKB2, RELA, and RELB. Moreover, simultaneous silencing of both receptors produced a greater drop in NF-κB activity, suggesting that NOTCH1 and NOTCH3 individually modulate NF-κB, and that both receptors are necessary for its maximal activity [111].
The role of SHH signaling in the pathogenesis of OS has also been extensively researched [81,112,113]. Evidence shows that dysregulation of this pathway occurred in both ligand-dependent and ligand-independent manners [114]. Higher expression levels of genes encoding SHH, DHH, PTCH1, GLI1, GLI2, and SMO were detected in OS cell lines [115] and were validated in the tumor samples. Moreover, associations with tumor volume and, consequently, with patient outcome were described [112]. Sustained aberrant SHH activity may result from autocrine and paracrine induction [114]. In this regard, integration between SHH signaling and NF-κB also occurs [116,117,118]. Indeed, evidence showed that NF-κB directly binds and transcriptionally activates the SHH and GLI1 promoters [119,120], supporting both ligand-dependent and ligand-independent tumor promotion.
Likewise, the TGF-β signaling pathway, which is activated in OS, affects the development of lung metastases [121] and mediates protumorigenic microenvironmental changes [122]. Its interaction with NF-κB may result from non-canonical cascades, where the activated receptor complex transmits the signal to the transcription factor [123].
Following this line, activation and autophosphorylation of RTKs results in the recruitment of a wide range of downstream signaling proteins to propagate critical cellular signaling pathways. Several mechanisms underlay the constitutive activation of RTK in human cancers including gain-of-function mutations, genomic amplification, chromosomal rearrangements, and/or autocrine activation [124]. In OS, dysregulation of many of these receptors has been associated with tumor development and metastasis. For instance, overexpression of VEGF, which leads to the autocrine activation of its receptor, is associated with a more aggressive phenotype [125,126] and was significantly associated with higher metastasis rates, higher clinical stage, and chemoresistance [127,128]. VEGF is a typical NF-κB target [129], which also represents an effector of VEGFR signaling [130]. Likewise, NF-κB-mediated HER2 overexpression is involved in radiation-induced repopulation in heterogeneous tumors [131]; this RTK is amplified in ~30% of patients with OS [132].
In a similar way, NF-κB is a downstream mediator of many other RTKs including the platelet-derived growth factor (PDGF) [133,134], the fibroblast growth factor receptor (FGFR) [135,136,137,138], AXL, RET, and EPHB2 [139], all of which are frequently activated in most OS tissues and cell lines, correlate with poor clinical outcomes, and have been suggested as promising therapeutic targets [140,141,142,143,144,145].
Moreover, RTK signaling generally converges to activate the RAS/RAF/MEK/ERK pathway [146]. It is well-known that activation of NF-κB represents a common outcome of the RAS/RAF/MEK/ERK signaling pathway exerting proliferative effects. ERK, for instance, can lead to its activation by phosphorylating IKK [48], and, in OS, the phosphorylation of ERK1/2 promotes invasion and metastasis. Moreover, an autocrine loop between RAF and NF-κB has also been described in some cell types [147]. Interaction with RAF has also been described to trigger NF-κB through the activation of MEKK1 and IKKβ [148]. Furthermore, RAS/RAF/MEK/ERK signaling in OS was significantly associated with immune infiltration and tumor microenvironment (TME) [149]. OS microenvironment is undeniably essential for growth and dissemination [150,151]. Beside immune evasion, stromal and other tumor-associated cells influence OS by secreting growth factors, cytokines, and exosomal and non-exosomal miRNAs that lead to metabolic reprogramming, extracellular matrix remodeling, neovascularization, drug resistance, and maintenance of the cancer stem cells phenotype [69,152].
Therefore, given all the above, within the complex circuit maze underlying the molecular basis of OS, NF-κB stands out not only as a key mediator of several hallmarks of cancer biology, but also as a common denominator of intricate crosstalk with other signaling pathways crucial to shaping specific responses. This feature is highlighted with protein–protein interaction analysis showing that between 25% and 50% of proteins involved in each of the pathways cited above are functionally connected with NF-κB subunits (Figure 4–Supplemental Table S1).
Even more, the NF-κB pathway itself is dysregulated in OS [55,153,154,155,156,157,158], and, even though the mechanisms by which it plays a causative role are not yet fully understood, it clearly contributes to tumor progression and chemoresistance [159,160]. Nuclear localization of NF-κB (p65) is frequently found in OS samples [104], and higher levels of this transcription factor detected with immunohistochemical analysis denote lower survival rates [161].
In this regard, an in silico analysis using gene expression of NF-κB subunits on three different OS datasets assessed at the R2 Genomics Analysis and Visualization Platform (http://r2.amc.nl accessed on 15 April 2024) (Mixed Osteosarcoma-Aqeilan-18-MAS5.0-u133p2, Mixed Osteosarcoma-Guenther-20-MAS5.0-u133a, and Tumor Osteosarcoma-Kobayashi-27-MAS5.0-u133p2) demonstrated higher levels of RELA, RELB, REL, and NFKB2 compared to normal osteoblasts (p < 0.05) (Figure 5A). The analysis of possible associations with clinical features using the Mixed Osteosarcoma (Mesenchymal)-Kuijjer-127-vst-ilmnhwg6v2 dataset showed significantly lower levels of NFKB1 expression in patients with metastasis at diagnosis and lower 5-year metastasis-free survival rates (Figure 5B and 5C, respectively). Also, despite a lack of differential expression between normal and OS samples, NFKB1 showed a positive correlation with RELA (Figure 5D), a nonsurprising result considering that these genes encode the most abundant NF-κB heterodimer, p65/p52 [162].
Correlation between mRNA expression levels of pathway-associated genes (with the functional interaction with NF-κB shown above) was also observed, with all the 68 genes present in the Kuijjer’s dataset correlated with the expression of one or more NF-κB subunits (Figure 6A). Even though the patterns are not similar for different NF-κB subunits, analysis through the TFLink gateway (available at https://tflink.net/ accessed on 17 April 2024), which provides comprehensive and highly accurate information on transcription factor–target gene interactions, showed that 61 out of 68 genes are indeed NF-κB predicted targets (Figure 6B). Of note, five of those genes (CDKN1A, BTRC, IL6, CTNNB1, and CCND1) are regulated by all NF-κB subunits and are among those with higher correlation scores. Differential expression analysis showed 12 downregulated genes (mostly associated with the negative regulation of transcription: NFKBIB, BTRC, CREBBP, HDAC1, NCOR2, and FBXW11) and 19 upregulated genes [mostly associated with protein ubiquitination (SOCS1, RBX1, RPS27A, UBB, UBA52, and UBE2N), positive regulation of cell proliferation (CTNNB1, NRAS, HRAS, CDKN1A, and STAT3), and cell surface receptor signaling pathways (IL6, IFNA5, IFNA7, IFNA16, and NGF) in OS samples compared to controls. Several of these genes have already been associated with OS pathophysiology [149,163,164,165,166,167,168,169,170,171,172,173,174,175,176,177]. Moreover, KEGG functional enrichment was assessed through STRING v12 (available at https://string-db.org/ accessed on 17 April 2024) and showed that under-expressed genes belonged mainly to MAPK and WNT signaling pathways, while upregulated genes belonged to the PI3K/AKT and JAK/STAT cascades (Figure 6C), reinforcing the concept of NF-κB as a common player underneath OS establishment and progression.
Consequently, the inhibition of this transcription factor not only could result in the suppression of the proliferative and invasive capacities and chemoresistance of OS but could also improve clinical outcomes [178,179,180,181,182].

3. Experimental Evidence NF-κB Inhibition

As diagnosis and implementation of ideal treatment plans for a better quality of life require the integration of clinical, radiological, and histological characteristics of each malignancy, molecular markers have become key players for proper diagnosis, prognosis assessment, and prediction of treatment response. However, as the plethora of described point mutations, gene fusions, chromosomal aberrations, and epigenetic modifications increases, intra- and intertumoral heterogeneity still hampers tumor management and foment the design and subsequent development of drug candidates.
Certainly, drugs with genetically validated targets are more likely to obtain clinical approval than drugs whose targets are not supported with experimental evidence.
In this regard, when the phenotypic relevance of each NF-κB subunit knockdown/knockout in OS cell lines is analyzed through the DepMap portal (https://depmap.org/portal/ accessed on 20 April 2024), most of them are shown to be dependent (dependency score < 0) on RELA, NFKB1, and NFKB2 expression (Figure 7A).
Over the years, hundreds of molecules have been described as capable of interfering with the NF-κB signaling pathway. These compounds are basically divided into three types of general strategies: biomolecular inhibitors, natural products (and their derivatives), and synthetic compounds, all of which can act at different steps of the activation pathway. Within each category, these molecules can act as direct inhibitors, including IκB kinase inhibitors (IKKs), 26S proteasome inhibitors, ubiquitin-ligase complex inhibitors, and constituent NF-κB subunit inhibitors, or they can exert their activity indirectly as is the case for antioxidants [183,184]. Many NF-κB inhibitors have been extensively tested in cancer models (in vitro and in vivo) [185,186,187,188,189,190] and considered promising chemotherapeutics or chemo- or radiosensitizers. The literature shows several interesting findings regarding the use on NF-κB inhibitors for OS which are listed below and summarized in Table 1.
Amentoflavone—This is a natural biflavonoid formed from the oxidative process of two apigenin molecules commonly found in Ginkgo biloba [191]. Extensively researched for its wide range of activities, it notably affects the ERK/PI3K/Akt pathway and, more importantly, NF-κB signaling by inhibiting both IκBα degradation [192] and the nuclear translocation of dimers [193]. Its anticancer properties have been demonstrated in OS and various other cancer types, influencing cell proliferation, apoptosis, invasion, metastasis, autophagy, transcription, and drug-resistance. These effects are mediated through the regulation of NF-κB and related proteins such as FAS-L, TNF-alpha, and inflammatory cytokines [194]. Specifically, treatment of U2OS cells has shown impaired cell migration and invasion, evidenced by decreased expression of metastasis-associated proteins like uPA, MMP2, and MMP9 [195]. Furthermore, when combined with sorafenib, amentoflavone’s anticancer effects were enhanced, leading to cell death through extrinsic and intrinsic apoptosis signaling pathways [193]. Similar outcomes were observed in U2OS-derived xenografts treated with amentoflavone (100 mg/kg/day), where photon emission from the tumor area was three-times lower than in the control group after 15 days. Additionally, progression-associated proteins (phospho-p65, phospho-ERK, VEGF, MMP9, XIAP, and CyclinD1) were significantly reduced [196].
Andrographolide—Derived from Andrographis paniculata, this labdane diterpenoid exhibits notable therapeutic applications as an anti-inflammatory agent with possible antineoplastic properties [197]. Andrographolide weakens NF-κB activation by exhausting the expression levels of p65 and p50 [198]. Inhibition of p65 translocation and decreased expression levels of phosphorylated ERK and p38 have also been described after treatment [199]. In OS, this compound reduces cell viability, although the mechanism of cell death remains debated. Liu and colleagues [200] observed a dose-dependent induction of autophagy accompanied by suppression of PI3K/Akt/mTOR and enhancement of JNK signaling. Conversely, Wang and colleagues [201] demonstrated that treatment at lower concentrations induced apoptosis in U2OS cells, as evidenced by increased PARP [poly (ADP-ribose) polymerase], caspase-3, -8, and -9 cleavage. Furthermore, this treatment led to G2/M cell cycle arrest and inhibition of epithelial–mesenchymal transition, characterized by decreased levels of Snail, MMP-2, MMP-7, MMP-9, vimentin, and N-cadherin, along with increased E-cadherin expression. Additionally, andrographolide enhanced the action of pyrrolidinedithiocarbamate ammonium, another NF-κB inhibitor, and reduced the volume of orthotopic tumors and pulmonary micrometastases [202]. Consistently, andrographolide treatment showed antitumor effects on primary OS cells [201], decreasing both total and nuclear protein levels of p65 in 143B cells [202].
Bay 11-7082—Also known as Fenretinide (N-(4-hydroxyphenyl)retinamide), this is a synthetic compound that blocks IκB-α phosphorylation with an IC50 of 10 μM in HUVEC cells [203]. In OS, this vitamin A analogue induces apoptosis and impedes cell migration and invasion in vitro, while inhibiting intraosseous tumor growth in NOD/SCID mice [204]. Moreover, when combined with the PI3K inhibitor LY294002 or lithium chloride, the antiproliferative and pro-apoptotic effects were enhanced [104,204].
DHMEQ—DHMEQ (Dehydroxymethylepoxyquinomicin) is a synthetic and specific inhibitor of NF-κB, designed to block the DNA binding of p65, p50, RelB, and Rel subunits [205,206]. By doing so, it can inhibit both the canonical and non-canonical NF-κB pathway activation. DHMEQ treatment has been shown to reduce cell viability, colony formation, and the mitotic index while triggering apoptosis. Likewise, treatment reduced the migration and invasion capacities of HOS and MG-6 cell lines. When combined with standard chemotherapeutic drugs like cisplatin, doxorubicin, or methotrexate, DHMEQ showed synergistic effects, mainly in a sequential schedule [207].
Dihydroartemisinin—This semi-synthetic derivative of artemisinin, an herbal drug that has been used in traditional Chinese medicine for centuries and is commonly isolated from Artemisia annua, exerts its activity on NF-κB signaling by inhibiting IkBα phosphorylation and DNA binding [208,209]. This compound has demonstrated strong antitumor effects in OS cases, including decreased cell viability, proliferation, colony formation, and altered migrative capacity [210,211]. In part, these effects were attributed to caspase-dependent apoptosis and cell cycle arrest at the G2/M [212]. Similarly, treatment of orthotopic tumors can prevent OS formation and maintain intact bone structure in athymic mice treated with intragastric 20 mg/kg administration once a day for 37 days. However, the authors showed that the anticancer effects may result from inactivating Wnt/β-catenin signaling by increasing GSK3β activity and the consequent degradation of β-catenin [210]. When combined with Apatinib (VEGFR2 inhibitor), the effects were even greater, reducing cell viability, migration, and invasion in vitro, as well as reducing tumor volume in vivo [210,213]. Alternatively, it has also been demonstrated that dihydroartemisinin acts as a ROS (reactive oxygen species) generator causing mitochondrial damage, and it activated autophagy via stimulation of the ROS/Erk1/2 pathway [211,214]. Widespread changes in lipid metabolic programs in OS after treatment were also reported [213].
Dihydromyricetin—This is a natural compound extracted from Ampelopsis grossedentata representing a potent inhibitor of NF-κB, responsible for inhibiting IκBα phosphorylation and degradation [215]. Treatment of OS cells decreased cell viability and caused G2/M cell cycle arrest through the upregulation of p21 and induced DNA damage. Additionally, increased phosphorylation of cell cycle checkpoint proteins such as ATM, CHK2, and H2AX was also observed [216,217]. Dihydromyricetin also decreased the phosphorylation of IKKα/β, NF-κB, and IκBα in U2OS cells, while inhibited their migration and invasion ability by downregulating the expression NF-κB direct transcriptional target uPA [217]. However, there is also evidence that this flavonoid acts by interfering with p38 and the AMPKα–GSK3β–Sox2 signaling pathway [216]. Alternatively, it has also been reported that dihydromyricetin may prevent hydrogen peroxide-induced apoptosis in MG63 cells through downregulation of caspase activation and upregulation of Bcl-2 levels [218].
Ginsenoside Rh2—This bioactive compound found in Panax ginseng has a long-established use in Chinese traditional medicine and exhibits antiproliferative, anti-invasive, anti-metastatic, cell cycle arrest-inducing, and differentiation-promoting abilities by inhibiting NF-κB degradation [219,220]. In OS, treatment with this steroid glycoside led to reduced cell viability and increased the levels of cleaved caspase-3, caspase-8, and caspase-9, while decreasing Bcl-2 levels. Additionally, it promoted the MAPK signaling pathway while inhibiting cell migration and invasion thorough the negative regulation of MMP-2, MMP-7, MMP-9, and mesenchymal markers such as Snail, N-cadherin, and vimentin [221]. Moreover, when encapsulated in solid silica nanospheres, it reduced tumor volume and recruited immune cells in a murine OS model [222].
Isoalantolactone—This compound can be found in many medicinal plants, mainly those belonging to the Asteraceae family and other angiosperms known for producing sesquiterpene lactones (isoalantolactone, alantolactone, and 5-epoxyalantolactone) [223,224]. Isoalantolactone has shown antiproliferative effects against several cell types, including HeLa, B16F10, and MK-1 [225]. Treatment of OS cell lines (U2OS, MG63, and Saos-2) showed interesting results as well, with reduced viability and G2/M arrest. Increased apoptosis was associated with ROS generation and the dissipation of mitochondrial membrane potential. Furthermore, the study provided evidence of decreased levels of nuclear p65 in a dose-dependent manner (50% lower levels in cells treated with 40 µM compared to controls) [226].
Isoliquiritigenin—This chalcone-type flavonoid is extracted from the root or rhizome of the licorice plant Glycyrrhiza glabra [227]. With a broad range of pharmacological properties, it exhibits direct growth inhibitory effects in various types of cancers by blocking the nuclear translocation of NF-κB and IκBα degradation [228,229]. Also, as a broad metalloproteinase inhibitor, it caused a drastic reduction in the migration capacity of OS cells, while treatment with 0.9 mg kg−1 (injected intravenously into mice via the tail vein) once every 2 days for 6 days was able to reduce tumor size by about 80% in NOD/SCID mice, impeded distant organ metastasis, and prolonged the survival time (100% of the animal were alive at day 70) [230]. Other cellular processes are associated with the increased production of Bax and caspase-3 and the reduction in Bcl-2 [231,232].
β-lapachone—This naphthoquinone was originally isolated from the heartwood of the lapacho tree Handroanthus impetiginosus commonly found in South America [233]. With activity against several types of malignant tumors, treatment of U2-OS cells induced necrotic cell death, reductio n in mitochondrial transmembrane potential, and release of mitochondrial cytochrome c [234]. Additionally, Hori et al. (2011), demonstrated that this quinone was more efficient when combined with hyperthermia (42 °C) [235].
Licoricidin—This isoflavonoid is also extracted from the roots of the plant G. glabra [236,237]. Besides the known antimicrobial activities against Helicobacter pylori, it has shown anticancer potential [237,238]. In OS, treatment with this compound diminished viability in a dose-dependent manner. Additionally, reduced levels of p65 phosphorylation were observed both in vitro and in vivo, especially by enhancing gemcitabine-induced cytotoxicity [239].
Magnoflorine—This compound is described as an important alkaloid with a wide range of pharmacological applications that can be obtained from several members of the Ranunculaceae, Menispermaceae, and Magnoliaceae families [240]. Regarding bone tumors, magnoflorine reduced viability and invasion of MG-63 and U2OS cells in a dose-dependent manner, while it did not affect normal osteoblasts (hFOB1.19). Downregulation of p65 and IκBα phosphorylation was also correlated with increasing treatment concentrations. Additionally, enhanced sensibility of OS cells to cisplatin was also observed [241].
Matrine—This compound is the main monomer extracted from the medicinal plant Sophora flavescens Ait, with several pharmacological activities, including anti-inflammatory, antitumor, anti-viral, and others [242]. Initial testing in OS resulted in decreased p50 and p65 nuclear translocation and reduced levels of phosphorylated IκB-β. Cell proliferation and invasion were inhibited in a dose-dependent manner [243]. Similarly, the study by Zhou et al. (2019), showed that cotreatment with matrine significantly increased adriamycin cytotoxicity in a concentration-dependent manner. Decreased cell motility was also observed in vitro with reduced expression of MMP-9 and STAT3. In vivo, intragastric administration (50 mg/kg/day for 3 weeks) significantly reduced U2OS-derived xenografts volume [243]. Likewise, intratumoral matrine at 0.75 mg/mL for 5 weeks significantly inhibited growth of U2OS xenographic tumors [244].
Nimbolide—This chemical compound is extracted from the neem plant Azadirachta indica (Meliaceae family) and has shown anticancer activity through the modulation of various molecular pathways including p53, pTEN, PI3K/AKT, VEGF, Bcl-2, and NF-κB [245]. Treatment of OS cells (143B) with nimbolide reduced viability with an IC50 of around 4 µM [246]. Apoptosis was also observed after treatment of MG-63 and U2OS cells, as a result of endoplasmic reticulum (ER) stress, mitochondrial dysfunction, accumulation of ROS, and caspase activation. In these cells, nimbolide treatment decreased phosphorylation of IKKα/β, IκBα, and p65 as detected with luciferase activity assays [247].
Okadaic acid—This compound is considered a very potent toxin produced by dinoflagellates [248,249]. Treatment of OS cells reduced migration and induced apoptosis in a dose- and time-dependent manner, with an IC50 determined as 50 nM after 24 h [250].
Parthenolide—This sesquiterpene lactone of the germacranolide class, extracted from Tanacetum parthenium plants, acts as a covalently reactive compound that has shown selective toxicity against cancer cells at concentrations around 5–20 µM. With broad biological activity, it interferes with several pathways, albeit the most prominent and the first confirmed target was NF-κB, through alkylation of IKKβ [251] and inhibition of IκB phosphorylation [252,253]. Regarding OS, the study by D’Anneo et al. (2013) demonstrated drastic effects on viability with only 30% of cells alive after 5 h of treatment (25 µM); the DNA-binding activity of p65 also decreased rapidly (80% compared to control after 2 h of treatment), although cell death occurred in a caspase-independent manner [254]. Similar results were described by Kishida and Yoshikawa (2007) who also showed the ability of parthenolide to suppress metastasis to the lung when animals were treated soon after cell inoculation [255]. Furthermore, this compound sensitized LM7 (derived from SAOS-2) and LM8 cells to ionizing radiation [256,257].
Phloretin—This dihydrochalcone mainly found in the leaves of apple trees [258] has shown promise in cancer treatment by downregulating the expression of NF-κB, EGFR, and VEGF, and by blocking or decreasing the phosphorylation of MAP kinases among other mechanisms [259]. Huang et al. (2015) also described its effects on NF-κB signaling through the inhibition of IκB-α phosphorylation and p65 translocation to the nucleus [260]. In MG63, U2OS, and 143B cells, this inhibitor was able to reduce the viability and acted synergistically with daunomycin, 5-FU, etoposide, and methotrexate [261].
Punicalagin—This antioxidant extracted from pomegranate (Punica granatum) can regulate IκBα degradation and reduce p65 expression. Treatment of OS cells resulted in decreased viability and motility. Likewise, administration of punicalagin in xenograft mouse models inhibited tumor growth and diminished angiogenesis [262,263].
Raddeanin A—Extracted from the traditional Chinese herb Anemone raddeana Regel, this triterpenoid can inhibit p65, thereby promoting antitumor effects among other biological activities [264]. Tested as a single drug, it reduced cell viability in a panel composed of six human OS cell lines and was also responsible for reducing cell migration and invasion [265,266]. Raddeanin A also increased the cytotoxic potential of doxorubicin and downregulated MDR1 (a known target of NF-κB) in drug-resistant cells [267]. Moreover, intraperitoneal treatment with 5 mg/kg every 3 days was sufficient to reduce tumor growth in orthotopic OS models [266,267].
Sulphoraphene—Commonly found in cruciferous vegetables [268], this compound has an extensive anticancer effect by reducing p65 phosphorylation. In OS, treatment with this phytochemical reduced cell viability and colony formation in a dose-dependent manner, while impairing epithelial–mesenchymal transition (EMT). In vivo, a dose of 40 μmol/kg promoted a threefold reduction in tumor growth compared to controls after 42 days [269].
Tetramethylpyrazine—This alkaloid extracted from Chuanxiong (Ligusticum wallichii) exerts a variety of pharmacological effects [270] and has been shown to reduce nuclear p65 in OS. Indeed, treatment with this compound promoted G1/G0 cell cycle arrest and apoptosis by modulating cyclin D1 and BCL-2. Moreover, intraperitoneal injection (100 mg/kg doses every other day for 28 days) inhibited xenograft tumor growth with minimal effects on body weight [271].
Theabrownin—Extracted from pu-erh tea, this pigment modulates phosphorylation levels of p65 and IκBα. When tested in OS cultures, it diminished cell viability, reduced Ki67 expression, and increased cleavage of PARP and caspase-3 [272]. In addition, theabrownin inhibited the motility of U2OS cells, impairing microfilament and microtubule formation. Reinforcing this observation, treated cells expressed more E-cadherin than the mesenchymal markers vimentin, Snail-1, and Slug [273].
Thymoquinone—Naturally occurring in the seeds of Nigella Sativa, this compound presents well-known chemotherapeutic and chemopreventive effects, modulating NF-κB indirectly by inhibiting TNF-alpha activation [274]. Already described as a potent radiosensitizer [275], it has shown antitumor effects in OS models both as a single agent, and in combination with selenium [276], low doses of 5-FU, oxaliplatin [277], cisplatin, and methotrexate [278,279,280]. Mechanistically, in these models NF-κB inhibition increased pro-apoptotic proteins, blocked the cell cycle by promoting an increase in p21WAF1, and affected DNA metabolism proteins like g-H2AX and NBS1 [281,282]. Moreover, the administration of 6 mg/kg in tumor-bearing mice for 15 days arrested tumor growth (volume) without apparent side effects, downregulating Ki67, CD34, survivin, XIAP, and VEGF [282].
Ursolic Acid—This triterpenoid commonly used in traditional Asian medicine has been shown to inhibit the NF-κB pathway by interfering with IKKa activity, p65 phosphorylation, and DNA binding [283,284,285]. As a single agent, this compound exerted cytotoxic and antimigratory effects against several OS cell lines. The estimated half-maximum inhibitory concentration values for MG-63 were calculated to be around 11 μg/mL at 24 h and 8 μg/mL at 48 h, indicating dose- and time-dependent responses [286]. Oxidative stress and collapse of the mitochondrial membrane permeability were also observed [286,287]. These phenotypes were accompanied by the activation of ERK1/2, JNK, and MAPK signaling, and downregulation of MMP-2 and EGFR signaling. Inhibition of the JNK pathway was also reported in this tumor type [287]. In vivo, intraperitoneal administration of ursolic acid provoked a diminution of tumor growth, improving p53 expression and reducing the expression of β-catenin, NF-κB, and the phosphorylation of STAT3 [286,287]. Moreover, this antioxidant showed synergistic effects when combined with zoledronic acid [288] and cisplatin [289].
Mangostin—This compound represents a natural xanthonoid isolated from the bark and dried sap of Garcinia mangostana with the potential to inhibit IkBα and p65 phosphorylation [290]. Treating OS cultures promoted a reduction in cell viability and triggered apoptosis, increasing cleaved caspase-3 and PARP [291,292]. Regarding motility, mangostin inhibited invasion and migration of MG-63 cells, increasing E-cadherin and decreasing mesenchymal markers such as N-cadherin, Slug, and Snail [292]. In vivo, it promoted ER stress-mediated apoptosis caused by ROS accumulation while it restrained WNT/β-catenin signaling [291].
Genistein—This naturally occurring flavonoid acts as an antioxidant capable of downregulating NF-κB DNA binding [293]. Widely distributed in the Fabaceae family, it has been proven to exhibit good preclinical results against various types of human cancers, mainly from epithelial origin [294]. In OS models, treatment with genistein led to reduced cell viability and motility, while inducing morphological changes and differentiation denoted by increased osteocalcin [295,296,297]. Additionally, xenograft tumors derived from genistein-treated LM8 cells showed significant lower mass than controls and fewer metastases to lungs and liver [298]. When tested in combination, genistein reversed OS resistance to gemcitabine through the downregulation of NF-κB activity and the suppression of Akt [293]. Moreover, MNNG/HOS tumor-bearing mice treated daily with genistein, while receiving gemcitabine (80 mg/kg) once every other day, showed significant inhibition of tumor growth compared to controls and animals that received individual treatments [295].
Magnolol—As one of the main active components of Magnolia officinalis, this lignan has already shown antitumor effects through NF-κB pathway inactivation [299]. In the OS models, magnolol exposure promoted a reduction in cell viability and triggered apoptosis by inhibiting ERK/NF-κB signaling [300]. In addition, magnolol reduced colony formation, cell migration, and invasion of MG-63 cells, inducing G0/G1 cell cycle arrest and upregulation of pro-apoptosis proteins. However, a weak antiproliferative activity in normal human osteoblast cells (hFOB1.19) was also observed [301].
Bortezomib—This proteasome inhibitor, known for its ability to inhibit the NF-κB pathway, has been widely tested in OS models [302]. Its most relevant effect in this tumor type is its chemosensitizer ability. When combined with everolimus, an anti-angiogenic drug, bortezomib improved the inhibition of cell proliferation, induced cell cycle arrest, and enhanced apoptosis. Mechanistically, the combination induced higher levels of cleaved PARP, caspase-3, caspase-8, and caspase-9, while reducing the expression of c-MYC, survivin, and phospho-cyclin D1 [303]. Moreover, compared to monotherapy, the administration of everolimus and bortezomib significantly suppressed tumor growth in vivo [303,304].
Curcumin—This polyphenol derived from the turmeric rhizome of Curcuma longa L. is extensively described as an antitumor agent capable of constraining NF-κB pathway activation by interfering with IKK and blocking IκBα and p65 phosphorylation [305]. The literature reports numerous curcumin antiproliferative and antimigratory effects on OS with modulation of p21, Bax, Bcl-xl, Bcl2, caspase-3, PARP cleavage, cyclin D1, MMP-2, and MMP-9 expression [306,307,308,309,310,311].
However, the most interesting effects of curcumin and its analogues are their chemo- and radiosensitization properties [312,313]. For instance, combining curcumin and C6 ceramide (a type of sphingolipid that plays a role in cell differentiation, the cell cycle, cell growth, and cell death) optimized the anticancer effects of curcumin in vitro and in vivo [314]. Combinations with another plant derivative, JCTH-4 also presented synergistic effects provoking cells death by apoptosis and autophagy, in addition to an increase in ROS generation [315].
Metformin—This compound is a synthetic derivative of galegine and/or guanidine, natural products found in the herbal medicine Galega officinalis, and is widely used in the treatment of type 2 diabetes [316]. However, many studies discuss its anticancer capabilities, acting either directly or indirectly [317]. Accordingly, this compound has proved to be efficient against OS cell lines, including KHOS/NP, HOS, MG-63, and U2OS, and it has displayed potent in vivo antitumor effects in KHOS/NP xenografts [318]. Similarly, positive results were obtained treating 143B cells, with G2/M arrest and apoptosis attributed to ROS-dependent JNK/c-Jun activation [319].
Of note, metformin was included in a clinical trial involving children with relapsed or refractory solid and central nervous system tumors. The therapy protocol included vincristine, irinotecan, temozolomide for one cycle, and metformin given concurrently beginning in cycle 2. OS patients enrolled (n = 2) received 666 mg/m2/day. Grade 3 and 4 toxicities included anemia (16%), thrombocytopenia (9.6%), and neutropenia (29.8%). However, both patients presented stable diseases, reinforcing the prospects of its use in clinical setting [320].
Caffeine—This methylxanthine has long been widely used by the world population as a eugeroic or as a mild cognitive enhancer to increase alertness and attentional performance. Its antitumor effects, mainly when combined with chemotherapy, have been attributed to its potential to inhibit p65 phosphorylation [321]. In OS, simultaneous treatment with cisplatin markedly reduced cell proliferation, whereas exposure to either compound alone barely affected survival [322]. Likewise, when the effectiveness of combinations of caffeine and cisplatin were tested in cisplatin-resistant cells, the combination improved the cytotoxicity, a result not observed in cisplatin-sensitive cell lines [323]. The combination also showed positive results in vivo, both in xenografts and PDX [324,325]. In all cases, oral administration of caffeine improved the cytotoxic effects of cisplatin, reducing the tumor volume almost two times more than the administration of cisplatin alone [326]. Combined therapy led to a reduction in lung metastasis and an improvement in overall survival [327].
Table 1. NF-κB inhibitors tested in OS models.
Table 1. NF-κB inhibitors tested in OS models.
InhibitorTargetIC50In Vivo AntiproliferationAnti-MotilityChemosensitizer/SynergismRadiosensitizer Reference
AmentoflavoneIκBα degradation and p65 translocation50–100 μMyesyesyesyes-[193,196]
AndrographolideModification of p5050–70 μMyesyesyes--[201]
Bay 11-7085Inhibition of IκBα10 μMyesyesyesyes-[104,204]
BortezomibProteasome yesyesyesyes [303,304]
Caffeinep65 and antioxidant1–2.80 mMyesyesyesyes-[322,326]
CurcuminIKK activety, IkBα and p65 phosphorilation10–100 nMyesyesyesyes-[314,315]
DHMEQInhibition of p6512–48 μg/mL-yesyesyes -[206,207]
DihydroartemisininIκBα degradation and DNA binding4.6–16 yesyesyesyes-[212]
DihydromyricetinPhosphorylation and degradation of IκBα20–60 μmol/mLyesyesyesyes-[216]
Genisteinantioxidant20–80 μMyesyesyesyes-[295,296]
Ginsenoside Rh2NF-kB degradation2.52–7.85 μg/mLyesyesyesyes-[219,220,221,222]
Isoalantolactonep650–200 μM-yes---[226]
Isoliquiritigeninp65 translocation0–100 μMyesyesyes [230,231,232]
Lapachonep65 phosphorilation0–10 μM-yes---[234,235]
Licoricidinp650–32 μMyesyes-yes-[239]
Magnoflorinep65 phosphorilation, IkBα5–80 μM-yesyesyes-[241]
Magnololantioxidant25–41 μM-yesyes--[300,301]
MangostinIkBα and p65 phosphorilation30–40 μMyesyesyes--[291,292]
Matrinep50 and p65 translocation, IkB-β0–1.5 mg/mLyesyesyesyes-[243]
Metforminp65 phosphorylation0–50 mMyesyesyes--[316,317,318,319,320]
Nimbolidep-IKK-β/α, p-p650–250 µg/mL-yesyes--[246,247]
Okadaic acidp650–50 nM-yesyes--[248,249,250]
Parthenolidep-650–100 μMyesyesyes-yes[257]
PHLORETINIκB-α phosphorylation and p65 translocation100 μg-yes-yes-[261]
Punicalaginp6510–100 μMyesyesyes--[262,263]
Raddeanin Ap651512–10.05 μMyesyesyesyes-[266,267]
Sulphoraphenep6540 μMyesyesyes--[269]
Tetramethylpyrazinep6510.3, 24.7, 54.7 mg/mLyesyes---[271]
Theabrowninp6543.93 or 51.98 mg/mLyesyesyes--[272,273]
ThymoquinoneTNF-α17–40 μMyesyes-yes-[278,280,281,282]
Ursolic AcidIKK and p65 phosphorilation5–37 μMyesyesyesyes-[286,287]

4. Final Considerations

Despite the plethora of different NF-κB inhibitors, the majority lack selectivity or have small therapeutic indexes. For instance, an analysis on the druggability of NF-κB subunits using the CanSAR platform version 1.5.4 (an integrated knowledge base that brings together multidisciplinary data to provide useful predictions for drug discovery—https://cansarblack.icr.ac.uk/ accessed on 20 April 2024) showed that although many compounds have predicted potential for interaction, only about 45% demonstrate binding efficiency, and none of the compounds present on the platform show potential for clinical application (Figure 7B). Furthermore, interaction networks of NF-κB inhibitors and associated binding proteins according to STITCH (a search tool for known and predicted interactions between chemicals and proteins available at http://stitch.embl.de accessed on 20 April 2024) indicate that among all the compounds described above with alleged activity against NF-κB, only DHMEQ and BAY 11-7085 show direct action on this transcription factor (Figure 7C). Nevertheless, some of these substances show direct or indirect activity on many signaling pathways that ultimately prevent the activation of NF-κB. A clear example is genistein. While its cytotoxic effects are mainly attributed to the inhibition of the transcription factor, genistein also interferes with several signaling cascades that, as seen above, interplay with NF-κB: NOTCH, PI3K/Akt/mTOR, WNT/β-catenin, JAK-STAT, and RTKs pathways. Moreover, it modulates the expression of several micro-RNAs, expanding even more the range of molecular interactions to achieve the primary goal of killing cancer cells through NF-κB inhibition.
Thus, the potential to develop efficient treatments by interfering with NF-κB is undeniable, not only for its pleiotropic effects on most cancer hallmarks but also because it represents a common downstream effector of the many dysregulated pathways that influence OS aggressiveness. Deeper knowledge and understanding of how these cascades crosstalk would, over time, turn complex regulatory networks into simpler pictures underneath molecular heterogeneity, reflecting better options for controlling tumor growth and halting metastasis, which remains a major obstacle in OS treatment.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/ph17060734/s1, Supplemental Table S1: Functional connections between NF-κB subunits and members of signaling pathways commonly dysregulated in OS. Lists of proteins belonging to each signaling pathway were obtained though the GESEA—Gene Set Enrichment Analysis—available at https://www.gsea-msigdb.org. Then, PPI between each pathway and NF-κB subunits were assessed through STRING v12 (available at https://string-db.org/ accessed on 10 April 2024).

Author Contributions

Formal analysis, investigation, data curation, writing—original draft preparation M.M., S.G., D.B., K.L.O. and M.S.B. Conceptualization, project administration, funding acquisition, figures design, review, and editing, M.S.B. All authors have read and agreed to the published version of the manuscript.

Funding

This work was supported by the National Council for Scientific and Technological Development (CNPq) research productivity fellowships (nº 301464/2019-7 and 303593/2022-9) given to MSB.

Conflicts of Interest

The authors affirm that they do not have any conflicts of interest.

References

  1. Sen, R. David Baltimore Multiple Nuclear Factors Interact with the Immunoglobulin Enhancer Sequences. Cell 1986, 46, 705–716. [Google Scholar] [CrossRef]
  2. Bours, V.; Burd, P.R.; Brown, K.; Villalobos, J.; Park, S.; Ryseck, R.P.; Bravo, R.; Kelly, K.; Siebenlist, U. A Novel Mitogen-Inducible Gene Product Related to P50/P105-NF-Kappa B Participates in Transactivation through a Kappa B Site. Mol. Cell. Biol. 1992, 12, 685–695. [Google Scholar] [CrossRef]
  3. Bours, V.; Villalobos, J.; Burd, P.R.; Kelly, K.; Siebenlist, U. Cloning of a Mitogen-Inducible Gene Encoding a ΚB DNA-Binding Protein with Homology to the Rel Oncogene and to Cell-Cycle Motifs. Nature 1990, 348, 76–80. [Google Scholar] [CrossRef]
  4. Ghosh, S.; Gifford, A.M.; Riviere, L.R.; Tempst, P.; Nolan, G.P.; Baltimore, D. Cloning of the P50 DNA Binding Subunit of NF-ΚB: Homology to Rel and Dorsal. Cell 1990, 62, 1019–1029. [Google Scholar] [CrossRef]
  5. Kieran, M.; Blank, V.; Logeat, F.; Vandekerckhove, J.; Lottspeich, F.; Le Bail, O.; Urban, M.B.; Kourilsky, P.; Baeuerle, P.A.; Israël, A. The DNA Binding Subunit of NF-ΚB Is Identical to Factor KBF1 and Homologous to the Rel Oncogene Product. Cell 1990, 62, 1007–1018. [Google Scholar] [CrossRef]
  6. Mercurio, F.; Didonato, J.; Rosette, C.; Karin, M. Molecular Cloning and Characterization of a Novel Rel/NF-ΧB Family Member Displaying Structural and Functional Homology to NF-ΧB P50/P105. DNA Cell Biol. 1992, 11, 523–537. [Google Scholar] [CrossRef]
  7. Meyer, R.; Hatada, E.N.; Hohmann, H.P.; Haiker, M.; Bartsch, C.; Rothlisberger, U.; Lahm, H.W.; Schlaeger, E.J.; Van Loon, A.P.G.M.; Scheidereit, C. Cloning of the DNA-Binding Subunit of Human Nuclear Factor ΚB: The Level of Its MRNA Is Strongly Regulated by Phorbol Ester or Tumor Necrosis Factor α. Proc. Natl. Acad. Sci. USA 1991, 88, 966–970. [Google Scholar] [CrossRef]
  8. Miyamoto, S.; Verma, I.M. REL/NF-ΚB/IkB Story. Adv. Cancer Res. 1995, 66, 255–292. [Google Scholar]
  9. Ruben, S.M.; Dillon, P.J.; Schreck, R.; Henkel, T.; Chen, C.H.; Maher, M.; Baeuerle, P.A.; Rosen, C.A. Isolation of a Rel-Related Human CDNA That Potentially Encodes the 65-KD Subunit of NF-ΚB. Science 1991, 251, 1490–1493. [Google Scholar] [CrossRef]
  10. Ryseck, R.-P.; Bull, P.; Takamiya, M.; Bours, V.; Siebenlist, U.; Dobrzanski, P.; Bravo, R. New Rel Family Transcription Activator That Can Interact with P5O-NF-ΚB. Mol. Cell Biol. 1992, 12, 674–684. [Google Scholar]
  11. Steward, R. Dorsal, an Embryonic Polarity Gene in Drosophila, Is Homologous to the Vertebrate Proto-Oncogene, c-Rel. Science 1987, 238, 692–694. [Google Scholar] [CrossRef] [PubMed]
  12. Ghosh, S.; Hyden, M.S. Celebrating 25 Years of Ideas. Imunol. Rev. 2012, 49, 10–11. [Google Scholar]
  13. Karin, M. How NF-k B Is Activated: The Role of the I k B Kinase (IKK) Complex. Oncogene 1999, 18, 6867–6874. [Google Scholar] [CrossRef] [PubMed]
  14. Mercurio, F.; Didonato, J.A.; Rosette, C.; Karin, M. P105 and P98 Precursor Protein Play an Active Role in NF-ΚB-Mediated Signal Transduction. Genes Dev. 1993, 7, 705–718. [Google Scholar] [CrossRef] [PubMed]
  15. Naumann, M.; Wulczyn, F.G.; Scheidereit, C. The NF-Kappa B Precursor P105 and the Proto-Oncogene Product Bcl-3 Are I Kappa B Molecules and Control Nuclear Translocation of NF-Kappa B. EMBO J. 1993, 12, 213–222. [Google Scholar] [CrossRef] [PubMed]
  16. Rice, N.R.; MacKichan, M.L.; Israël, A. The Precursor of NF-ΚB P50 Has IκB-like Functions. Cell 1992, 71, 243–253. [Google Scholar] [CrossRef] [PubMed]
  17. Scheinman, R.I.; Beg, A.A.; Baldwin, A.S. NF-Kappa B P100 (Lyt-10) Is a Component of H2TF1 and Can Function as an I Kappa B-like Molecule. Mol. Cell. Biol. 1993, 13, 6089–6101. [Google Scholar] [CrossRef] [PubMed]
  18. Zabel, U.; Baeuerle, P.A. Purified Human IκB Can Rapidly Dissociate the Complex of the NF-ΚB Transcription Factor with Its Cognate DNA. Cell 1990, 61, 255–265. [Google Scholar] [CrossRef] [PubMed]
  19. Zhang, Q.; Lenardo, M.J.; Baltimore, D. 30 Years of NF-ΚB: A Blossoming of Relevance to Human Pathobiology. Cell 2017, 168, 37–57. [Google Scholar] [CrossRef]
  20. Li, Z.W.; Chu, W.; Hu, Y.; Delhase, M.; Deerinck, T.; Ellisman, M.; Johnson, R.; Karin, M. The IKKβ Subunit of IκB Kinase (IKK) Is Essential for Nuclear Factor ΚB Activation and Prevention of Apoptosis. J. Exp. Med. 1999, 189, 1839–1845. [Google Scholar] [CrossRef]
  21. Ling, L.; Cao, Z.; Goeddel, D.V. Nf-ΚB-Inducing Kinase Activates IKK-α by Phosphorylation of Ser-176. Proc. Natl. Acad. Sci. USA 1998, 95, 3792–3797. [Google Scholar] [CrossRef] [PubMed]
  22. Mercurio, F.; Zhu, H.; Murray, B.W.; Shevchenko, A.; Bennett, B.L.; Li, J.W.; Young, D.B.; Barbosa, M.; Mann, M.; Manning, A.; et al. IKK-1 and IKK-2: Cytokine-Activated IκB Kinases Essential for NF-ΚB Activation. Science 1997, 278, 860–866. [Google Scholar] [CrossRef] [PubMed]
  23. Alkalay, I.; Yaron, A.; Hatzubai, A.; Jung, S.; Avraham, A.; Gerlitz, O.; Pashut-Lavon, I.; Ben-Neriah, Y. In Vivo Stimulation of I Kappa B Phosphorylation Is Not Sufficient to Activate NF-Kappa B. Mol. Cell. Biol. 1995, 15, 1294–1301. [Google Scholar] [CrossRef] [PubMed]
  24. Roff, M.; Thompson, J.; Rodriguez, M.S.; Jacque, J.M.; Baleux, F.; Arenzana-Seisdedos, F.; Hay, R.T. Role of IκBα Ubiquitination in Signal-Induced Activation of NF-ΚB in Vivo. J. Biol. Chem. 1996, 271, 7844–7850. [Google Scholar] [CrossRef] [PubMed]
  25. Scherer, D.C.; Brockman, J.A.; Chen, Z.; Maniatis, T.; Ballard, D.W. Signal-Induced Degradation of IκBα Requires Site-Specific Ubiquitination. Proc. Natl. Acad. Sci. USA 1995, 92, 11259–11263. [Google Scholar] [CrossRef] [PubMed]
  26. Beg, A.A.; Ruben, S.M.; Scheinman, R.I.; Haskill, S.; Rosen, C.A.; Baldwin, A.S. IKB Interacts with the Nuclear Localization Sequences of the Subunits of NF-ΚB: A Mechanism for Cytoplasmic Retention. Genes Dev. 1992, 6, 2664–2665. [Google Scholar] [CrossRef] [PubMed]
  27. Sen, R.; Baltimore, D. Inducibility of κ Immunoglobulin Enhancer-Binding Protein NF-ΚB by a Posttranslational Mechanism. Cell 1986, 47, 921–928. [Google Scholar] [CrossRef] [PubMed]
  28. Gutierrez, H.; Hale, V.A.; Dolcet, X.; Davies, A. NF-ΚB Signalling Regulates the Growth of Neural Processes in the Developing PNS and CNS. Development 2005, 132, 1713–1726. [Google Scholar] [CrossRef] [PubMed]
  29. Meffert, M.K.; Chang, J.M.; Wiltgen, B.J.; Fanselow, M.S.; Baltimore, D. NF-ΚB Functions in Synaptic Signaling and Behavior. Nat. Neurosci. 2003, 6, 1072–1078. [Google Scholar] [CrossRef]
  30. Dejardin, E. The Alternative NF-ΚB Pathway from Biochemistry to Biology: Pitfalls and Promises for Future Drug Development. Biochem. Pharmacol. 2006, 72, 1161–1179. [Google Scholar] [CrossRef]
  31. Sun, S.C. The Non-Canonical NF-ΚB Pathway in Immunity and Inflammation. Nat. Rev. Immunol. 2017, 17, 545–558. [Google Scholar] [CrossRef] [PubMed]
  32. Xiao, G.; Harhaj, E.W.; Sun, S.-C. NF-ΚB-Inducing Kinase Regulates the Processing of NF-ΚB2 P100. Mol. Cell 2001, 7, 759–764. [Google Scholar] [CrossRef] [PubMed]
  33. Fong, A.; Sun, S.C. Genetic Evidence for the Essential Role of β-Transducin Repeat-Containing Protein in the Inducible Processing of NF-ΚB2/P100. J. Biol. Chem. 2002, 277, 22111–22114. [Google Scholar] [CrossRef] [PubMed]
  34. Liang, C.; Zhang, M.; Sun, S.C. β-TrCP Binding and Processing of NF-ΚB2/P100 Involve Its Phosphorylation at Serines 866 and 870. Cell Signal 2006, 18, 1309–1317. [Google Scholar] [CrossRef] [PubMed]
  35. Senftleben, U.; Cao, Y.; Xiao, G.; Greten, F.R.; Krähn, G.; Bonizzi, G.; Chen, Y.; Hu, Y.; Fong, A.; Sun, S.C.; et al. Activation by IKKα of a Second, Evolutionary Conserved, NF-ΚB Signaling Pathway. Science 2001, 293, 1495–1499. [Google Scholar] [CrossRef] [PubMed]
  36. Tanaka, K.; Babic, I.; Nathanson, D.; Akhavan, D.; Guo, D.; Gini, B.; Dang, J.; Zhu, S.; Yang, H.; de Jesus, J.; et al. Oncogenic EGFR Signaling Activates an MTORC2-NF-ΚB Pathway That Promotes Chemotherapy Resistance. Cancer Discov. 2011, 1, 524–538. [Google Scholar] [CrossRef] [PubMed]
  37. Ahmad, A.; Ahsan, H. Ras-Mediated Activation of NF-ΚB and DNA Damage Response in Carcinogenesis. Cancer Investig. 2020, 38, 185–208. [Google Scholar] [CrossRef] [PubMed]
  38. Bonavia, R.; Inda, M.M.; Vandenberg, S.; Cheng, S.Y.; Nagane, M.; Hadwiger, P.; Tan, P.; Sah, D.W.Y.; Cavenee, W.K.; Furnari, F.B. EGFRvIII Promotes Glioma Angiogenesis and Growth through the NF-ΚB, Interleukin-8 Pathway. Oncogene 2012, 31, 4054–4066. [Google Scholar] [CrossRef]
  39. Puliyappadamba, V.T.; Chakraborty, S.; Chauncey, S.S.; Li, L.; Hatanpaa, K.J.; Mickey, B.; Noorani, S.; Shu, H.K.G.; Burma, S.; Boothman, D.A.; et al. Opposing Effect of EGFRWT on EGFRvIII-Mediated NF-ΚB Activation with RIP1 as a Cell Death Switch. Cell Rep. 2013, 4, 764–775. [Google Scholar] [CrossRef]
  40. Spiller, S.E.; Logsdon, N.J.; Deckard, L.A.; Sontheimer, H. Inhibition of Nuclear Factor Kappa-B Signaling Reduces Growth in Medulloblastoma in Vivo. BMC Cancer 2011, 11, 136. [Google Scholar] [CrossRef]
  41. Parker, M.; Mohankumar, K.M.; Punchihewa, C.; Weinlich, R.; Dalton, J.D.; Li, Y.; Lee, R.; Tatevossian, R.G.; Phoenix, T.N.; Thiruvenkatam, R.; et al. C11orf95-RELA Fusions Drive Oncogenic NF-ΚB Signalling in Ependymoma. Nature 2014, 506, 451–455. [Google Scholar] [CrossRef] [PubMed]
  42. Pietsch, T.; Wohlers, I.; Goschzik, T.; Dreschmann, V.; Denkhaus, D.; Dörner, E.; Rahmann, S.; Klein-Hitpass, L. Supratentorial Ependymomas of Childhood Carry C11orf95-RELA Fusions Leading to Pathological Activation of the NF-ΚB Signaling Pathway. Acta Neuropathol. 2014, 127, 609–611. [Google Scholar] [CrossRef] [PubMed]
  43. Cherry, E.M.; Lee, D.W.; Jung, J.U.; Sitcheran, R. Tumor Necrosis Factor-like Weak Inducer of Apoptosis (TWEAK) Promotes Glioma Cell Invasion through Induction of NF-ΚB-Inducing Kinase (NIK) and Noncanonical NF-ΚB Signaling. Mol. Cancer 2015, 14, 1–13. [Google Scholar] [CrossRef] [PubMed]
  44. Friedmann-Morvinski, D.; Narasimamurthy, R.; Xia, Y.; Myskiw, C.; Soda, Y.; Verma, I.M. Targeting NF-ΚB in Glioblastoma: A Therapeutic Approach. Sci. Adv. 2016, 2, e1501292. [Google Scholar] [CrossRef] [PubMed]
  45. Ius, T.; Ciani, Y.; Ruaro, M.E.; Isola, M.; Sorrentino, M.; Bulfoni, M.; Candotti, V.; Correcig, C.; Bourkoula, E.; Manini, I.; et al. An NF-κ B Signature Predicts Low-Grade Glioma Prognosis: A Precision Medicine Approach Based on Patient-Derived Stem Cells. Neuro Oncol. 2018, 20, 776–787. [Google Scholar] [CrossRef] [PubMed]
  46. Luo, Y.; Li, M.; Pratap, U.P.; Viswanadhapalli, S.; Liu, J.; Venkata, P.P.; Altwegg, K.A.; Palacios, B.E.; Li, X.; Chen, Y.; et al. PELP1 Signaling Contributes to Medulloblastoma Progression by Regulating the NF-ΚB Pathway. Mol. Carcinog. 2020, 59, 281–292. [Google Scholar] [CrossRef] [PubMed]
  47. Arabzade, A.; Zhao, Y.; Varadharajan, S.; Chen, H.C.; Jessa, S.; Rivas, B.; Stuckert, A.J.; Solis, M.; Kardian, A.; Tlais, D.; et al. Zfta–Rela Dictates Oncogenic Transcriptional Programs to Drive Aggressive Supratentorial Ependymoma. Cancer Discov. 2021, 11, 2200–2215. [Google Scholar] [CrossRef] [PubMed]
  48. Oeckinghaus, A.; Hayden, M.S.; Ghosh, S. Crosstalk in NF-ΚB Signaling Pathways. Nat. Immunol. 2011, 12, 695–708. [Google Scholar] [CrossRef] [PubMed]
  49. Karin, M.; Cao, Y.; Greten, F.R.; Li, Z.W. NF-ΚB in Cancer: From Innocent Bystander to Major Culprit. Nat. Rev. Cancer 2002, 2, 301–310. [Google Scholar] [CrossRef]
  50. Galeaz, C.; Totis, C.; Bisio, A. Radiation Resistance: A Matter of Transcription Factors. Front. Oncol. 2021, 11, 662840. [Google Scholar] [CrossRef]
  51. Shiloh, Y. The ATM-Mediated DNA-Damage Response: Taking Shape. Trends Biochem. Sci. 2006, 31, 402–410. [Google Scholar] [CrossRef] [PubMed]
  52. Mokim Ahmed, K.; Li, J.J. NF-ΚB-Mediated Adaptive Resistance to Ionizing Radiation. Free Radic. Biol. Med. 2008, 44, 1–13. [Google Scholar] [CrossRef] [PubMed]
  53. Chen, Y.; Di Grappa, M.A.; Molyneux, S.D.; McKee, T.D.; Waterhouse, P.; Penninger, J.M.; Khokha, R. RANKL Blockade Prevents and Treats Aggressive Osteosarcomas. Sci. Transl. Med. 2015, 7, 317ra197. [Google Scholar] [CrossRef] [PubMed]
  54. Baud’huin, M.; Duplomb, L.; Ruiz Velasco, C.; Fortun, Y.; Heymann, D.; Padrines, M. Key Roles of the OPG–RANK–RANKL System in Bone Oncology. Expert. Rev. Anticancer Ther. 2007, 7, 221–232. [Google Scholar] [CrossRef] [PubMed]
  55. Kushlinskii, N.E.; Fridman, M.V.; Braga, E.A. Molecular Mechanisms and MicroRNAs in Osteosarcoma Pathogenesis. Biochemistry 2016, 81, 315–328. [Google Scholar] [CrossRef] [PubMed]
  56. Biermann, J.S.; Chow, W.; Reed, D.R.; Lucas, D.; Adkins, D.R.; Agulnik, M.; Benjamin, R.S.; Brigman, B.; Budd, G.T.; Curry, W.T.; et al. NCCN Guidelines Insights: Bone Cancer, Version 2.2017. J. Natl. Compr. Cancer Netw. 2017, 15, 155–167. [Google Scholar] [CrossRef] [PubMed]
  57. Reed, D.R.; Hayashi, M.; Wagner, L.; Binitie, O.; Steppan, D.A.; Brohl, A.S.; Shinohara, E.T.; Bridge, J.A.; Loeb, D.M.; Borinstein, S.C.; et al. Treatment Pathway of Bone Sarcoma in Children, Adolescents, and Young Adults. Cancer 2017, 123, 2206–2218. [Google Scholar] [CrossRef] [PubMed]
  58. Rosenberg, A.E. Bone Sarcoma Pathology: Diagnostic Approach for Optimal Therapy. Am. Soc. Clin. Oncol. Educ. Book 2017, 37, 794–798. [Google Scholar] [CrossRef]
  59. Alkazemi, B.; Ghazawi, F.M.; Lagacé, F.; Nechaev, V.; Zubarev, A.; Litvinov, I.V. Investigation of the Incidence and Geographic Distribution of Bone and Soft Tissue Sarcomas in Canada: A National Population-Based Study. Curr. Oncol. 2023, 30, 5631–5651. [Google Scholar] [CrossRef] [PubMed]
  60. Balmant, N.V.; de Reis, R.S.; de Santos, M.O.; Maschietto, M.; de Camargo, B. Incidence and Mortality of Bone Cancer among Children, Adolescents and Young Adults of Brazil. Clinics 2019, 74, e858. [Google Scholar] [CrossRef]
  61. Casali, P.G.; Blay, J.Y.; Bertuzzi, A.; Bielack, S.; Bjerkehagen, B.; Bonvalot, S.; Boukovinas, I.; Bruzzi, P.; Tos, A.P.D.; Dileo, P.; et al. Bone Sarcomas: ESMO Clinical Practice Guidelines for Diagnosis, Treatment and Follow-Up. Ann. Oncol. 2014, 25, iii113–iii123. [Google Scholar] [CrossRef]
  62. Zając, A.; Król, S.K.; Rutkowski, P.; Czarnecka, A.M. Biological Heterogeneity of Chondrosarcoma: From (Epi) Genetics through Stemness and Deregulated Signaling to Immunophenotype. Cancers 2021, 13, 1317. [Google Scholar] [CrossRef]
  63. Rajan, S.; Franz, E.M.; McAloney, C.A.; Vetter, T.A.; Cam, M.; Gross, A.C.; Taslim, C.; Wang, M.; Cannon, M.V.; Oles, A.; et al. Osteosarcoma Tumors Maintain Intra-Tumoral Transcriptional Heterogeneity during Bone and Lung Colonization. BMC Biol. 2023, 21, 98. [Google Scholar] [CrossRef]
  64. Belayneh, R.; Fourman, M.S.; Bhogal, S.; Weiss, K.R. Update on Osteosarcoma. Curr. Oncol. Rep. 2021, 23, 1–8. [Google Scholar] [CrossRef]
  65. Ritter, J.; Bielack, S.S. Osteosarcoma. Ann. Oncol. 2010, 21, vii320–vii325. [Google Scholar] [CrossRef]
  66. Meltzer, P.S.; Helman, L.J. New Horizons in the Treatment of Osteosarcoma. N. Engl. J. Med. 2021, 385, 2066–2076. [Google Scholar] [CrossRef]
  67. Mirabello, L.; Troisi, R.J.; Savage, S.A. International Osteosarcoma Incidence Patterns in Children and Adolescents, Middle Ages and Elderly Persons. Int. J. Cancer 2009, 125, 229–234. [Google Scholar] [CrossRef]
  68. Abate, M.E.; Longhi, A.; Galletti, S.; Ferrari, S.; Bacci, G. Non-Metastatic Osteosarcoma of the Extremities in Children Aged 5 Years or Younger. Pediatr. Blood Cancer 2010, 55, 652–654. [Google Scholar] [CrossRef]
  69. Yang, C.; Tian, Y.; Zhao, F.; Chen, Z.; Su, P.; Li, Y.; Qian, A. Bone Microenvironment and Osteosarcoma Metastasis. Int. J. Mol. Sci. 2020, 21, 6985. [Google Scholar] [CrossRef]
  70. Kempf-Bielack, B.; Bielack, S.S.; Jürgens, H.; Branscheid, D.; Berdel, W.E.; Exner, G.U.; Göbel, U.; Helmke, K.; Jundt, G.; Kabisch, H.; et al. Osteosarcoma Relapse after Combined Modality Therapy: An Analysis of Unselected Patients in the Cooperative Osteosarcoma Study Group (COSS). J. Clin. Oncol. 2005, 23, 559–568. [Google Scholar] [CrossRef]
  71. Jafari, F.; Javdansirat, S.; Sanaie, S.; Naseri, A.; Shamekh, A.; Rostamzadeh, D.; Dolati, S. Osteosarcoma: A Comprehensive Review of Management and Treatment Strategies. Ann. Diagn. Pathol. 2020, 49, 151654. [Google Scholar] [CrossRef]
  72. Yoshida, A. Osteosarcoma: Old and New Challenges. Surg. Pathol. Clin. 2021, 14, 567–583. [Google Scholar] [CrossRef]
  73. Chou, A.J.; Gorlick, R. Chemotherapy Resistance in Osteosarcoma: Current Challenges and Future Directions. Expert. Rev. Anticancer Ther. 2006, 6, 1075–1085. [Google Scholar] [CrossRef]
  74. Tsukamoto, S.; Errani, C.; Angelini, A.; Mavrogenis, A.F. Current Treatment Considerations for Osteosarcoma Metastatic at Presentation. Orthopedics 2020, 43, E345–E358. [Google Scholar] [CrossRef]
  75. Lewis, I.J.; Nooij, M.A.; Whelan, J.; Sydes, M.R.; Grimer, R.; Hogendoorn, P.C.; Memon, M.A.; Weeden, S.; Uscinska, B.M.; van Glabbeke, M.; et al. Improvement in Histologic Response but Not Survival in Osteosarcoma Patients Treated with Intensified Chemotherapy: A Randomized Phase III Trial of the European Osteosarcoma Intergroup. JNCI J. Natl. Cancer Inst. 2007, 99, 112–128. [Google Scholar] [CrossRef]
  76. Bacci, G.; Ferrari, S.; Bertoni, F.; Ruggieri, P.; Picci, P.; Longhi, A.; Casadei, R.; Fabbri, N.; Forni, C.; Versari, M.; et al. Long-Term Outcome for Patients with Nonmetastatic Osteosarcoma of the Extremity Treated at the Istituto Ortopedico Rizzoli according to the Istituto Ortopedico Rizzoli/Osteosarcoma-2 Protocol: An Updated Report. J. Clin. Oncol. 2016, 18, 4016–4027. [Google Scholar] [CrossRef]
  77. Bacci, G.; Ferrari, S.; Lari, S.; Mercuri, M.; Donati, D.; Longhi, A.; Forni, C.; Bertoni, F.; Versari, M.; Pignotti, E. Osteosarcoma of the Limb: Amputation Or Limb Salvage In Patients Treated by Neoadjuvant Chemotherapy. J. Bone Joint Surg. Br. 2002, 84, 88–92. [Google Scholar] [CrossRef]
  78. Isakoff, M.S.; Bielack, S.S.; Meltzer, P.; Gorlick, R. Osteosarcoma: Current Treatment and a Collaborative Pathway to Success. J. Clin. Oncol. 2015, 33, 3029–3035. [Google Scholar] [CrossRef]
  79. Ferrari, S.; Smeland, S.; Mercuri, M.; Bertoni, F.; Longhi, A.; Ruggieri, P.; Alvegard, T.A.; Picci, P.; Capanna, R.; Bernini, G.; et al. Neoadjuvant Chemotherapy with High-Dose Ifosfamide, High-Dose Methotrexate, Cisplatin, and Doxorubicin for Patients with Localized Osteosarcoma of the Extremity: A Joint Study by the Italian and Scandinavian Sarcoma Groups. J. Clin. Oncol. 2005, 23, 8845–8852. [Google Scholar] [CrossRef]
  80. Garcia-Ortega, D.Y.; Cabrera-Nieto, S.A.; Caro-Sánchez, H.S.; Cruz-Ramos, M. An Overview of Resistance to Chemotherapy in Osteosarcoma and Future Perspectives. Cancer Drug Resist. 2022, 5, 762. [Google Scholar] [CrossRef]
  81. Lézot, F.; Corre, I.; Morice, S.; Rédini, F.; Verrecchia, F. SHH Signaling Pathway Drives Pediatric Bone Sarcoma Progression. Cells 2020, 9, 536. [Google Scholar] [CrossRef] [PubMed]
  82. Simpson, E.; Brown, H.L. Understanding Osteosarcomas. J. Am. Acad. Physician Assist. 2018, 31, 15–19. [Google Scholar] [CrossRef]
  83. Kalavrezos, N.; Sinha, D. Head and Neck Sarcomas in Adulthood: Current Trends and Evolving Management Concepts. Br. J. Oral Maxillofac. Surg. 2020, 58, 890–897. [Google Scholar] [CrossRef] [PubMed]
  84. Dana, P.M.; Sadoughi, F.; Asemi, Z.; Yousefi, B. Molecular Signaling Pathways as Potential Therapeutic Targets in Osteosarcoma. Curr. Med. Chem. 2022, 29, 4436–4444. [Google Scholar] [CrossRef] [PubMed]
  85. Rajan, S.; Zaccaria, S.; Cannon, M.V.; Cam, M.; Gross, A.C.; Raphael, B.J.; Roberts, R.D. Structurally Complex Osteosarcoma Genomes Exhibit Limited Heterogeneity within Individual Tumors and across Evolutionary Time. Cancer Res. Commun. 2023, 3, 564–575. [Google Scholar] [CrossRef] [PubMed]
  86. Schiavone, K.; Garnier, D.; Heymann, M.F.; Heymann, D. The Heterogeneity of Osteosarcoma: The Role Played by Cancer Stem Cells. Adv. Exp. Med. Biol. 2019, 1139, 187–200. [Google Scholar] [CrossRef] [PubMed]
  87. Sun, Y.; Zhang, C.; Fang, Q.; Zhang, W.; Liu, W. Abnormal Signal Pathways and Tumor Heterogeneity in Osteosarcoma. J. Transl. Med. 2023, 21, 1–14. [Google Scholar] [CrossRef] [PubMed]
  88. Miller, C.W.; Aslo, A.; Won, A.; Tan, M.; Lampkin, B.; Koeffler, H.P. Alterations of the P53, Rb and MDM2 Genes in Osteosarcoma. J. Cancer Res. Clin. Oncol. 1996, 122, 559–565. [Google Scholar] [CrossRef] [PubMed]
  89. Nirala, B.K.; Yamamichi, T.; Yustein, J.T. Deciphering the Signaling Mechanisms of Osteosarcoma Tumorigenesis. Int. J. Mol. Sci. 2023, 24, 11367. [Google Scholar] [CrossRef]
  90. Poos, K.; Smida, J.; Nathrath, M.; Maugg, D.; Baumhoer, D.; Korsching, E. How MicroRNA and Transcription Factor Co-Regulatory Networks Affect Osteosarcoma Cell Proliferation. PLoS Comput. Biol. 2013, 9, e1003210. [Google Scholar] [CrossRef]
  91. Zhang, J.; Yu, X.H.; Yan, Y.G.; Wang, C.; Wang, W.J. PI3K/Akt Signaling in Osteosarcoma. Clin. Chim. Acta 2015, 444, 182–192. [Google Scholar] [CrossRef] [PubMed]
  92. Xu, X.; Yu, H. Ras-PI3K Pathway Promotes Osteosarcoma Progression via Regulating VRK1-Mediated H2A Phosphorylation at Threonine 120. Artif. Cells Nanomed. Biotechnol. 2019, 47, 4274–4283. [Google Scholar] [CrossRef] [PubMed]
  93. Jin, R.; Jin, Y.Y.; Tang, Y.L.; Yang, H.J.; Zhou, X.Q.; Lei, Z. GPNMB Silencing Suppresses the Proliferation and Metastasis of Osteosarcoma Cells by Blocking the PI3K/Akt/MTOR Signaling Pathway. Oncol. Rep. 2018, 39, 3034–3040. [Google Scholar] [CrossRef] [PubMed]
  94. Bai, D.; Ueno, L.; Vogt, P.K. Akt-Mediated Regulation of NFκB and the Essentialness of NFκB for the Oncogenicity of PI3K and Akt. Int. J. Cancer 2009, 125, 2863–2870. [Google Scholar] [CrossRef] [PubMed]
  95. Lee, H.; Herrmann, A.; Deng, J.H.; Kujawski, M.; Niu, G.; Li, Z.; Forman, S.; Jove, R.; Pardoll, D.M.; Yu, H. Persistently Activated Stat3 Maintains Constitutive NF-ΚB Activity in Tumors. Cancer Cell 2009, 15, 283–293. [Google Scholar] [CrossRef]
  96. Danieau, G.; Morice, S.; Rédini, F.; Verrecchia, F.; Royer, B.B. Le New Insights about the Wnt/β-Catenin Signaling Pathway in Primary Bone Tumors and Their Microenvironment: A Promising Target to Develop Therapeutic Strategies? Int. J. Mol. Sci. 2019, 20, 3751. [Google Scholar] [CrossRef] [PubMed]
  97. Ma, B.; Hottiger, M.O. Crosstalk between Wnt/β-Catenin and NF-ΚB Signaling Pathway during Inflammation. Front. Immunol. 2016, 7, 1–14. [Google Scholar] [CrossRef] [PubMed]
  98. Lu, Y.; Guan, G.F.; Chen, J.; Hu, B.; Sun, C.; Ma, Q.; Wen, Y.H.; Qiu, X.C.; Zhou, Y. Aberrant CXCR4 and β-Catenin Expression in Osteosarcoma Correlates with Patient Survival. Oncol. Lett. 2015, 10, 2123–2129. [Google Scholar] [CrossRef] [PubMed]
  99. Martins-Neves, S.R.; Corver, W.E.; Paiva-Oliveira, D.I.; van den Akker, B.E.W.M.; Briaire-de-Bruijn, I.H.; Bovée, J.V.M.G.; Gomes, C.M.F.; Cleton-Jansen, A.M. Osteosarcoma Stem Cells Have Active Wnt/β-Catenin and Overexpress SOX2 and KLF4. J. Cell. Physiol. 2016, 231, 876–886. [Google Scholar] [CrossRef]
  100. Spiegelman, V.S.; Slaga, T.J.; Pagano, M.; Minamoto, T.; Ronai, Z.; Fuchs, S.Y. Wnt/β-Catenin Signaling Induces the Expression and Activity of ΒTrCP Ubiquitin Ligase Receptor. Mol. Cell 2000, 5, 877–882. [Google Scholar] [CrossRef]
  101. Die, L.; Yan, P.; Jun Jiang, Z.; Min Hua, T.; Cai, W.; Xing, L. Glycogen Synthase Kinase-3 Beta Inhibitor Suppresses Porphyromonas Gingivalis Lipopolysaccharide-Induced CD40 Expression by Inhibiting Nuclear Factor-Kappa B Activation in Mouse Osteoblasts. Mol. Immunol. 2012, 52, 38–49. [Google Scholar] [CrossRef] [PubMed]
  102. Choi, Y.S.; Hur, J.; Jeong, S. Beta-Catenin Binds to the Downstream Region and Regulates the Expression C-Reactive Protein Gene. Nucleic Acids Res. 2007, 35, 5511–5519. [Google Scholar] [CrossRef] [PubMed]
  103. Yun, K.; So, J.-S.; Jash, A.; Im, S.-H. Lymphoid Enhancer Binding Factor 1 Regulates Transcription through Gene Looping. J. Immunol. 2009, 183, 5129–5137. [Google Scholar] [CrossRef] [PubMed]
  104. Tang, Q.L.; Xie, X.B.; Wang, J.; Chen, Q.; Han, A.J.; Zou, C.Y.; Yin, J.Q.; Liu, D.W.; Liang, Y.; Zhao, Z.Q.; et al. Glycogen Synthase Kinase-3β, NF-ΚB Signaling, and Tumorigenesis of Human Osteosarcoma. JNCI J. Natl. Cancer Inst. 2012, 104, 749–763. [Google Scholar] [CrossRef] [PubMed]
  105. Xiu, Y.; Dong, Q.; Fu, L.; Bossler, A.; Tang, X.; Boyce, B.; Borcherding, N.; Leidinger, M.; Sardina, J.L.; Xue, H.; et al. Coactivation of NF-ΚB and Notch Signaling Is Sufficient to Induce B-Cell Transformation and Enables B-Myeloid Conversion. Blood 2020, 135, 108–120. [Google Scholar] [CrossRef] [PubMed]
  106. Osipo, C.; Golde, T.E.; Osborne, B.A.; Miele, L.A. Off the Beaten Pathway: The Complex Cross Talk between Notch and NF-ΚB. Lab. Investig. 2007, 88, 11–17. [Google Scholar] [CrossRef]
  107. Espinosa, L.; Cathelin, S.; D’Altri, T.; Trimarchi, T.; Statnikov, A.; Guiu, J.; Rodilla, V.; Inglés-Esteve, J.; Nomdedeu, J.; Bellosillo, B.; et al. The Notch/Hes1 Pathway Sustains NF-ΚB Activation through CYLD Repression in T Cell Leukemia. Cancer Cell 2010, 18, 268–281. [Google Scholar] [CrossRef] [PubMed]
  108. Pakvasa, M.; Haravu, P.; Boachie-Mensah, M.; Jones, A.; Coalson, E.; Liao, J.; Zeng, Z.; Wu, D.; Qin, K.; Wu, X.; et al. Notch Signaling: Its Essential Roles in Bone and Craniofacial Development. Genes Dis. 2021, 8, 8–24. [Google Scholar] [CrossRef]
  109. Zhang, Z.; Wu, W.; Shao, Z. NOTCH Signaling in Osteosarcoma. Curr. Issues Mol. Biol. 2023, 45, 2266–2283. [Google Scholar] [CrossRef]
  110. Tang, X.F.; Cao, Y.; Peng, D.B.; Zhao, G.S.; Zeng, Y.; Gao, Z.R.; Lv, Y.F.; Guo, Q.N. Overexpression of Notch3 Is Associated with Metastasis and Poor Prognosis in Osteosarcoma Patients. Cancer Manag. Res. 2019, 11, 547–559. [Google Scholar] [CrossRef]
  111. López-López, S.; Monsalve, E.M.; de Ávila, M.J.R.; González-Gómez, J.; Hernández de León, N.; Ruiz-Marcos, F.; Baladrón, V.; Nueda, M.L.; García-León, M.J.; Screpanti, I.; et al. NOTCH3 Signaling Is Essential for NF-ΚB Activation in TLR-Activated Macrophages. Sci. Rep. 2020, 10, 1–16. [Google Scholar] [CrossRef] [PubMed]
  112. Lo, W.W.; Pinnaduwage, D.; Gokgoz, N.; Wunder, J.S.; Andrulis, I.L. Aberrant Hedgehog Signaling and Clinical Outcome in Osteosarcoma. Sarcoma 2014, 2014, 1–9. [Google Scholar] [CrossRef] [PubMed]
  113. Yao, Z.; Han, L.; Chen, Y.; He, F.; Sun, B.; Kamar, S.; Zhang, Y.; Yang, Y.; Wang, C.; Yang, Z. Hedgehog Signalling in the Tumourigenesis and Metastasis of Osteosarcoma, and Its Potential Value in the Clinical Therapy of Osteosarcoma. Cell Death Dis. 2018, 9, 1–12. [Google Scholar] [CrossRef] [PubMed]
  114. Lo, W.W.; Wunder, J.S.; Dickson, B.C.; Campbell, V.; McGovern, K.; Alman, B.A.; Andrulis, I.L. Involvement and Targeted Intervention of Dysregulated Hedgehog Signaling in Osteosarcoma. Cancer 2014, 120, 537–547. [Google Scholar] [CrossRef] [PubMed]
  115. Hirotsu, M.; Setoguchi, T.; Sasaki, H.; Matsunoshita, Y.; Gao, H.; Nagao, H.; Kunigou, O.; Komiya, S. Smoothened as a New Therapeutic Target for Human Osteosarcoma. Mol. Cancer 2010, 9, 1–14. [Google Scholar] [CrossRef] [PubMed]
  116. Wang, Y.; Wang, D.; Dai, Y.; Kong, X.; Zhu, X.; Fan, Y.; Wang, Y.; Wu, H.; Jin, J.; Yao, W.; et al. Positive Crosstalk between Hedgehog and NF-ΚB Pathways Is Dependent on KRAS Mutation in Pancreatic Ductal Adenocarcinoma. Front. Oncol. 2021, 11, 652283. [Google Scholar] [CrossRef] [PubMed]
  117. Vecchiotti, D.; Verzella, D.; Di Vito Nolfi, M.; D’andrea, D.; Flati, I.; Di Francesco, B.; Cornice, J.; Alesse, E.; Capece, D.; Zazzeroni, F. Elevated NF-ΚB/SHh/GLI1 Signature Denotes a Worse Prognosis and Represent a Novel Potential Therapeutic Target in Advanced Prostate Cancer. Cells 2022, 11, 2118. [Google Scholar] [CrossRef] [PubMed]
  118. Nakashima, H.; Nakamura, M.; Yamaguchi, H.; Yamanaka, N.; Akiyoshi, T.; Koga, K.; Yamaguchi, K.; Tsuneyoshi, M.; Tanaka, M.; Katano, M. Nuclear Factor-KappaB Contributes to Hedgehog Signaling Pathway Activation through Sonic Hedgehog Induction in Pancreatic Cancer. Cancer Res. 2006, 66, 7041–7049. [Google Scholar] [CrossRef] [PubMed]
  119. Kasperczyk, H.; Baumann, B.; Debatin, K.-M.; Fulda, S. Characterization of Sonic Hedgehog as a Novel NF-κB Target Gene That Promotes NF-κB-mediated Apoptosis Resistance and Tumor Growth in Vivo. FASEB J. 2009, 23, 21–33. [Google Scholar] [CrossRef]
  120. Colavito, S.A.; Zou, M.R.; Yan, Q.; Nguyen, D.X.; Stern, D.F. Significance of Glioma-Associated Oncogene Homolog 1 (GLI1) Expression in Claudin-Low Breast Cancer and Crosstalk with the Nuclear Factor Kappa-Light-Chain-Enhancer of Activated B Cells (NFκB) Pathway. Breast Cancer Res. 2014, 16, 444. [Google Scholar] [CrossRef]
  121. Lamora, A.; Talbot, J.; Bougras, G.; Amiaud, J.; Leduc, M.; Chesneau, J.; Taurelle, J.; Stresing, V.; Le Deley, M.C.; Heymann, M.F.; et al. Overexpression of Smad7 Blocks Primary Tumor Growth and Lung Metastasis Development in Osteosarcoma. Clin. Cancer Res. 2014, 20, 5097–5112. [Google Scholar] [CrossRef] [PubMed]
  122. Verrecchia, F.; Rédini, F. Transforming Growth Factor-β Signaling Plays a Pivotal Role in the Interplay between Osteosarcoma Cells and Their Microenvironment. Front. Oncol. 2018, 8, 133. [Google Scholar] [CrossRef] [PubMed]
  123. Akhurst, R.J.; Hata, A. Targeting the TGFβ Signalling Pathway in Disease. Nat. Rev. Drug Discov. 2012, 11, 790–811. [Google Scholar] [CrossRef]
  124. Paul, M.K.; Mukhopadhyay, A.K. Tyrosine Kinase—Role and Significance in Cancer. Int. J. Med. Sci. 2004, 1, 101–115. [Google Scholar] [CrossRef]
  125. Ohba, T.; Cates, J.M.M.; Cole, H.A.; Slosky, D.A.; Haro, H.; Ando, T.; Schwartz, H.S.; Schoenecker, J.G. Autocrine VEGF/VEGFR1 Signaling in a Subpopulation of Cells Associates with Aggressive Osteosarcoma. Mol. Cancer Res. 2014, 12, 1100–1111. [Google Scholar] [CrossRef]
  126. Daft, P.G.; Yang, Y.; Napierala, D.; Zayzafoon, M. The Growth and Aggressive Behavior of Human Osteosarcoma Is Regulated by a CaMKII-Controlled Autocrine VEGF Signaling Mechanism. PLoS ONE 2015, 10, e0121568. [Google Scholar] [CrossRef]
  127. Zhang, C.; Wang, L.; Xiong, C.; Zhao, R.; Liang, H.; Luo, X. The Role of Vascular Endothelial Growth Factor as a Prognostic and Clinicopathological Marker in Osteosarcoma: A Systematic Review and Meta-Analysis. J. Orthop. Surg. Res. 2021, 16, 738. [Google Scholar] [CrossRef]
  128. Oshiro, H.; Tome, Y.; Miyake, K.; Higuchi, T.; Sugisawa, N.; Kanaya, F.; Nishida, K.; Hoffman, R.M. An MTOR and VEGFR Inhibitor Combination Arrests a Doxorubicin Resistant Lung Metastatic Osteosarcoma in a PDOX Mouse Model. Sci. Rep. 2021, 11, 8583. [Google Scholar] [CrossRef] [PubMed]
  129. Fujioka, S.; Niu, J.; Schmidt, C.; Sclabas, G.M.; Peng, B.; Uwagawa, T.; Li, Z.; Evans, D.B.; Abbruzzese, J.L.; Chiao, P.J. NF-KappaB and AP-1 Connection: Mechanism of NF-KappaB-Dependent Regulation of AP-1 Activity. Mol. Cell. Biol. 2004, 24, 7806–7819. [Google Scholar] [CrossRef]
  130. Taniguchi, K.; Karin, M. NF-ΚB, Inflammation, Immunity and Cancer: Coming of Age. Nat. Rev. Immunol. 2018, 18, 309–324. [Google Scholar] [CrossRef]
  131. Cao, N.; Li, S.; Wang, Z.; Ahmed, K.M.; Degnan, M.E.; Fan, M.; Dynlach, J.R.; Li, J.J. NF-KappaB-Mediated HER2 Overexpression in Radiation-Adaptive Resistance. Radiat. Res. 2009, 171, 9–21. [Google Scholar] [CrossRef] [PubMed]
  132. Nakano, K. The Future of HER2-Targeted Treatment for Osteosarcoma: Lessons from the Negative Trastuzumab Deruxtecan Results. Int. J. Mol. Sci. 2023, 24, 16823. [Google Scholar] [CrossRef] [PubMed]
  133. Sulzbacher, I.; Träxler, M.; Mosberger, I.; Lang, S.; Chott, A. Platelet-Derived Growth Factor-AA and -Alpha Receptor Expression Suggests an Autocrine and/or Paracrine Loop in Osteosarcoma. Mod. Pathol. 2000, 13, 632–637. [Google Scholar] [CrossRef] [PubMed]
  134. Xu, J.; Xie, L.; Guo, W. PDGF/PDGFR Effects in Osteosarcoma and the “Add-on” Strategy. Clin. Sarcoma Res. 2018, 8, 15. [Google Scholar] [CrossRef] [PubMed]
  135. Brown, L.M.; Ekert, P.G.; Fleuren, E.D.G. Biological and Clinical Implications of FGFR Aberrations in Paediatric and Young Adult Cancers. Oncogene 2023, 42, 1875–1888. [Google Scholar] [CrossRef] [PubMed]
  136. Weekes, D.; Kashima, T.; Grigoriadis, A. P36. The Role of FGF-Signalling in Osteosarcoma: A Potential Therapeutic Target? Cancer Treat. Rev. 2008, 34, 26. [Google Scholar] [CrossRef]
  137. Zhou, W.Y.; Zheng, H.; Du, X.L.; Yang, J.L. Characterization of FGFR Signaling Pathway as Therapeutic Targets for Sarcoma Patients. Cancer Biol. Med. 2016, 13, 260–268. [Google Scholar] [CrossRef] [PubMed]
  138. Fernanda Amary, M.; Ye, H.; Berisha, F.; Khatri, B.; Forbes, G.; Lehovsky, K.; Frezza, A.M.; Behjati, S.; Tarpey, P.; Pillay, N.; et al. Fibroblastic Growth Factor Receptor 1 Amplification in Osteosarcoma Is Associated with Poor Response to Neo-Adjuvant Chemotherapy. Cancer Med. 2014, 3, 980–987. [Google Scholar] [CrossRef] [PubMed]
  139. Rettew, A.N.; Getty, P.J.; Greenfield, E.M. Receptor Tyrosine Kinases in Osteosarcoma: Not Just the Usual Suspects. Adv. Exp. Med. Biol. 2014, 804, 47–66. [Google Scholar] [CrossRef]
  140. Greenfield, E.M.; Collier, C.D.; Getty, P.J. Receptor Tyrosine Kinases in Osteosarcoma: 2019 Update. Adv. Exp. Med. Biol. 2020, 1258, 141–155. [Google Scholar] [CrossRef]
  141. Fleuren, E.D.G.; Versleijen-Jonkers, Y.M.H.; Boerman, O.C.; van der Graaf, W.T.A. Targeting Receptor Tyrosine Kinases in Osteosarcoma and Ewing Sarcoma: Current Hurdles and Future Perspectives. Biochim. Et Biophys. Acta (BBA)—Rev. Cancer 2014, 1845, 266–276. [Google Scholar] [CrossRef] [PubMed]
  142. Rettew, A.N.; Young, E.D.; Lev, D.C.; Kleinerman, E.S.; Abdul-Karim, F.W.; Getty, P.J.; Greenfield, E.M. Multiple Receptor Tyrosine Kinases Promote the in Vitro Phenotype of Metastatic Human Osteosarcoma Cell Lines. Oncogenesis 2012, 1, e34. [Google Scholar] [CrossRef]
  143. Abdeen, A.; Chou, A.J.; Healey, J.H.; Khanna, C.; Osborne, T.S.; Hewitt, S.M.; Kim, M.; Wang, D.; Moody, K.; Gorlick, R. Correlation between Clinical Outcome and Growth Factor Pathway Expression in Osteogenic Sarcoma. Cancer 2009, 115, 5243–5250. [Google Scholar] [CrossRef]
  144. Chaiyawat, P.; Klangjorhor, J.; Settakorn, J.; Champattanachai, V.; Phanphaisarn, A.; Teeyakasem, P.; Svasti, J.; Pruksakorn, D. Activation Status of Receptor Tyrosine Kinases as an Early Predictive Marker of Response to Chemotherapy in Osteosarcoma. Transl. Oncol. 2017, 10, 846–853. [Google Scholar] [CrossRef] [PubMed]
  145. Tian, Z.; Niu, X.; Yao, W. Receptor Tyrosine Kinases in Osteosarcoma Treatment: Which Is the Key Target? Front. Oncol. 2020, 10, 1642. [Google Scholar] [CrossRef] [PubMed]
  146. Chandhanayingyong, C.; Kim, Y.; Staples, J.R.; Hahn, C.; Lee, F.Y. MAPK/ERK Signaling in Osteosarcomas, Ewing Sarcomas and Chondrosarcomas: Therapeutic Implications and Future Directions. Sarcoma 2012, 2012, 404810. [Google Scholar] [CrossRef] [PubMed]
  147. Norris, J.L.; Baldwin, A.S. Oncogenic Ras Enhances NF-KappaB Transcriptional Activity through Raf-Dependent and Raf-Independent Mitogen-Activated Protein Kinase Signaling Pathways. J. Biol. Chem. 1999, 274, 13841–13846. [Google Scholar] [CrossRef] [PubMed]
  148. Baumann, B.; Weberjakob Troppmair, C.K.; Whiteside, S.; Israel, A.; Rapp, U.R.; Wirth, T. Raf Induces NF-KappaB by Membrane Shuttle Kinase MEKK1, a Signaling Pathway Critical for Transformation. Proc. Natl. Acad. Sci. USA 2000, 97, 4615–4620. [Google Scholar] [CrossRef] [PubMed]
  149. Chen, Q.; Zhou, X.; Jin, J.; Feng, J.; Xu, Z.; Chen, Y.; Zhao, H.; Li, Z. A Novel Defined RAS-Related Gene Signature for Predicting the Prognosis and Characterization of Biological Function in Osteosarcoma. J. Oncol. 2022, 2022, 5939158. [Google Scholar] [CrossRef]
  150. Corre, I.; Verrecchia, F.; Crenn, V.; Redini, F.; Trichet, V. The Osteosarcoma Microenvironment: A Complex But Targetable Ecosystem. Cells 2020, 9, 976. [Google Scholar] [CrossRef]
  151. Jin, J.; Cong, J.; Lei, S.; Zhang, Q.; Zhong, X.; Su, Y.; Lu, M.; Ma, Y.; Li, Z.; Wang, L.; et al. Cracking the Code: Deciphering the Role of the Tumor Microenvironment in Osteosarcoma Metastasis. Int. Immunopharmacol. 2023, 121, 110422. [Google Scholar] [CrossRef] [PubMed]
  152. Malla, R.R.; Kiran, P. Tumor Microenvironment Pathways: Cross Regulation in Breast Cancer Metastasis. Genes Dis. 2022, 9, 310–324. [Google Scholar] [CrossRef] [PubMed]
  153. Zhang, C.; Chen, B.; Jiang, K.; Lao, L.; Shen, H.; Chen, Z. Activation of TNF-α/NF-ΚB Axis Enhances CRL4BDCAF11 E3 Ligase Activity and Regulates Cell Cycle Progression in Human Osteosarcoma Cells. Mol. Oncol. 2018, 12, 476–494. [Google Scholar] [CrossRef] [PubMed]
  154. Zhou, J.L.; Deng, S.; Fang, H.S.; Yu, G.; Peng, H. Hsa-Let-7g Promotes Osteosarcoma by Reducing HOXB1 to Activate NF-ΚB Pathway. Biomed. Pharmacother. 2019, 109, 2335–2341. [Google Scholar] [CrossRef] [PubMed]
  155. Lu, S.; Liao, Q.S.; Tang, L. MiR-155 Affects Osteosarcoma Cell Proliferation and Invasion through Regulating NF-ΚB Signaling Pathway. Eur. Rev. Med. Pharmacol. Sci. 2018, 22, 7633–7639. [Google Scholar] [CrossRef] [PubMed]
  156. Gao, W.; Gao, J.; Chen, L.; Ren, Y.; Ma, J. Targeting XIST Induced Apoptosis of Human Osteosarcoma Cells by Activation of NF-ΚB/PUMA Signal. Bioengineered 2019, 10, 261–270. [Google Scholar] [CrossRef] [PubMed]
  157. Han, J.; Shen, X. Long Noncoding RNAs in Osteosarcoma via Various Signaling Pathways. J. Clin. Lab. Anal. 2020, 34, e23317. [Google Scholar] [CrossRef] [PubMed]
  158. Iyer, S.V.; Ranjan, A.; Elias, H.K.; Parrales, A.; Sasaki, H.; Roy, B.C.; Umar, S.; Tawfik, O.W.; Iwakuma, T. Genome-Wide RNAi Screening Identifies TMIGD3 Isoform1 as a Suppressor of NF-ΚB and Osteosarcoma Progression. Nat. Commun. 2016, 7, 13561. [Google Scholar] [CrossRef] [PubMed]
  159. Liu, J.F.; Chen, P.C.; Chang, T.M.; Hou, C.H. Thrombospondin-2 Stimulates MMP-9 Production and Promotes Osteosarcoma Metastasis via the PLC, PKC, c-Src and NF-ΚB Activation. J. Cell. Mol. Med. 2020, 24, 12826–12839. [Google Scholar] [CrossRef]
  160. Feng, Z.M.; Guo, S.M. Tim-3 Facilitates Osteosarcoma Proliferation and Metastasis through the NF-ΚB Pathway and Epithelial-Mesenchymal Transition. Genet. Mol. Res. 2016, 1–9. [Google Scholar] [CrossRef]
  161. Gong, T.; Su, X.; Xia, Q.; Wang, J.; Kan, S. Expression of NF-ΚB and PTEN in Osteosarcoma and Its Clinical Significance. Oncol. Lett. 2017, 14, 6744–6748. [Google Scholar] [CrossRef] [PubMed]
  162. Oeckinghaus, A.; Ghosh, S. The NF-KappaB Family of Transcription Factors and Its Regulation. Cold Spring Harb. Perspect. Biol. 2009, 1, a000034. [Google Scholar] [CrossRef] [PubMed]
  163. Martin, J.W.; Chilton-MacNeill, S.; Koti, M.; Van Wijnen, A.J.; Squire, J.A.; Zielenska, M. Digital Expression Profiling Identifies RUNX2, CDC5L, MDM2, RECQL4, and CDK4 as Potential Predictive Biomarkers for Neo-Adjuvant Chemotherapy Response in Paediatric Osteosarcoma. PLoS ONE 2014, 9, e95843. [Google Scholar] [CrossRef] [PubMed]
  164. Xie, X.; Li, Y.; Zhu, H.; Kuang, Z.; Chen, D.; Fan, T. Prognostic Significance of β-Catenin Expression in Osteosarcoma: A Meta-Analysis. Front. Oncol. 2020, 10, 402. [Google Scholar] [CrossRef] [PubMed]
  165. Gross, A.C.; Cam, H.; Phelps, D.A.; Saraf, A.J.; Bid, H.K.; Cam, M.; London, C.A.; Winget, S.A.; Arnold, M.A.; Brandolini, L.; et al. IL-6 and CXCL8 Mediate Osteosarcoma-Lung Interactions Critical to Metastasis. JCI Insight 2018, 3, 1–17. [Google Scholar] [CrossRef] [PubMed]
  166. Zhang, C.; Hong, C.S.; Hu, X.; Yang, C.; Wang, H.; Zhu, D.; Moon, S.; Dmitriev, P.; Lu, J.; Chiang, J.; et al. Inhibition of Protein Phosphatase 2A with the Small Molecule LB100 Overcomes Cell Cycle Arrest in Osteosarcoma after Cisplatin Treatment. Cell Cycle 2015, 14, 2100–2108. [Google Scholar] [CrossRef] [PubMed]
  167. Zhou, Y.; Li, G.; Li, H.; Lai, F.; Duan, P.; Cheng, M. Epithelial to Mesenchymal Transition Relevant Subtypes with Distinct Prognosis and Responses to Chemo- or Immunotherapies in Osteosarcoma. J. Immunol. Res. 2022, 2022, 1377565. [Google Scholar] [CrossRef] [PubMed]
  168. Bakkalci, D.; Al-Badri, G.; Yang, W.; Nam, A.; Liang, Y.; Fisher, J.; Cheema, U. Engineering a Metastatic Stroma Directs the Osteosarcoma Tumour Transcriptome in a Spatially Specific Manner. Appl. Mater. Today 2023, 35, 101994. [Google Scholar] [CrossRef]
  169. Zu, D.; Dong, Q.; Chen, S.; Chen, Y.; Yao, J.; Zou, Y.; Lin, J.; Fang, B.; Wu, B. MiRNA-331-3p Affects the Proliferation, Metastasis, and Invasion of Osteosarcoma through SOCS1/JAK2/STAT3. J. Oncol. 2022, 2022, 6459029. [Google Scholar] [CrossRef]
  170. Li, T.; Tang, Z.; Li, S.; Lu, M. Development of a Novel Six DNA Damage Response-Related Prognostic Signature in Osteosarcoma. Cell Mol. Biol. 2024, 70, 110–115. [Google Scholar] [CrossRef]
  171. Tu, B.; Du, L.; Fan, Q.M.; Tang, Z.; Tang, T.T. STAT3 Activation by IL-6 from Mesenchymal Stem Cells Promotes the Proliferation and Metastasis of Osteosarcoma. Cancer Lett. 2012, 325, 80–88. [Google Scholar] [CrossRef] [PubMed]
  172. Squire, J.A.; Pei, J.; Marrano, P.; Beheshti, B.; Bayani, J.; Lim, G.; Moldovan, L.; Zielenska, M. High-Resolution Mapping of Amplifications and Deletions in Pediatric Osteosarcoma by Use of CGH Analysis of CDNA Microarrays. Genes Chromosomes Cancer 2003, 38, 215–225. [Google Scholar] [CrossRef] [PubMed]
  173. Yang, Y.; Zhang, Y.; Qu, X.; Xia, J.; Li, D.; Li, X.; Wang, Y.; He, Z.; Li, S.; Zhou, Y.; et al. Identification of Differentially Expressed Genes in the Development of Osteosarcoma Using RNA-Seq. Oncotarget 2016, 7, 87194–87205. [Google Scholar] [CrossRef] [PubMed]
  174. Gaeta, R.; Morelli, M.; Lessi, F.; Mazzanti, C.M.; Menicagli, M.; Capanna, R.; Andreani, L.; Coccoli, L.; Aretini, P.; Franchi, A. Identification of New Potential Prognostic and Predictive Markers in High-Grade Osteosarcoma Using Whole Exome Sequencing. Int. J. Mol. Sci. 2023, 24, 86. [Google Scholar] [CrossRef] [PubMed]
  175. Sonaglio, V.; de Carvalho, A.C.; Toledo, S.R.C.; Salinas-Souza, C.; Carvalho, A.L.; Petrilli, A.S.; De Camargo, B.; Vettore, A.L. Aberrant DNA Methylation of ESR1 and P14ARF Genes Could Be Useful as Prognostic Indicators in Osteosarcoma. Onco Targets Ther. 2013, 6, 713–723. [Google Scholar] [CrossRef] [PubMed]
  176. Zhang, Q.; Yin, X.; Zhang, Y. MicroRNA-221 Promotes Cell Proliferation and Inhibits Apoptosis in Osteosarcoma Cells by Directly Targeting FBXW11 and Regulating Wnt Signaling. Arch. Med. Res. 2021, 52, 191–199. [Google Scholar] [CrossRef] [PubMed]
  177. Dalle Carbonare, L.; Gomez Lira, M.; Minoia, A.; Bertacco, J.; Orsi, S.; Lauriola, A.; Li Vigni, V.; Gandini, A.; Antoniazzi, F.; Zipeto, D.; et al. Expression of FBXW11 in Normal and Disease-Associated Osteogenic Cells. J. CellMol. Med. 2023, 27, 1580–1591. [Google Scholar] [CrossRef] [PubMed]
  178. Li, F.; Sethi, G. Targeting Transcription Factor NF-KappaB to Overcome Chemoresistance and Radioresistance in Cancer Therapy. Biochim. Biophys. Acta 2010, 1805, 167–180. [Google Scholar] [CrossRef] [PubMed]
  179. Labbozzetta, M.; Notarbartolo, M.; Poma, P. Can NF-ΚB Be Considered a Valid Drug Target in Neoplastic Diseases? Our Point of View. Int. J. Mol. Sci. 2020, 21, 70. [Google Scholar] [CrossRef]
  180. Yu, H.; Lin, L.; Zhang, Z.; Zhang, H.; Hu, H. Targeting NF-ΚB Pathway for the Therapy of Diseases: Mechanism and Clinical Study. Signal Transduct. Target. Ther. 2020, 5, 209. [Google Scholar] [CrossRef]
  181. Chauhan, A.; Islam, A.U.; Prakash, H.; Singh, S. Phytochemicals Targeting NF-ΚB Signaling: Potential Anti-Cancer Interventions. J. Pharm. Anal. 2022, 12, 394–405. [Google Scholar] [CrossRef] [PubMed]
  182. Lin, Y.; Bai, L.; Chen, W.; Xu, S. The NF-KappaB Activation Pathways, Emerging Molecular Targets for Cancer Prevention and Therapy. Expert Opin. Ther. Targets 2010, 14, 45–55. [Google Scholar] [CrossRef] [PubMed]
  183. Letícia de Castro Barbosa, M.; Alves da Conceicao, R.; Guerra Manssour Fraga, A.; Dias Camarinha, B.; Cristina de Carvalho Silva, G.; Gilcler Ferreira Lima, A.; Azevedo Cardoso, E.; de Oliveira Freitas Lione, V. NF-ΚB Signaling Pathway Inhibitors as Anticancer Drug Candidates. Anticancer Agents Med. Chem. 2017, 17, 483–490. [Google Scholar] [CrossRef] [PubMed]
  184. Gilmore, T.D.; Herscovitch, M. Inhibitors of NF-ΚB Signaling: 785 and Counting. Oncogene 2006, 25, 6887–6899. [Google Scholar] [CrossRef] [PubMed]
  185. Ramadass, V.; Vaiyapuri, T.; Tergaonkar, V. Small Molecule NF-ΚB Pathway Inhibitors in Clinic. Int. J. Mol. Sci. 2020, 21, 5164. [Google Scholar] [CrossRef] [PubMed]
  186. Baud, V.; Karin, M. Is NF-KappaB a Good Target for Cancer Therapy? Hopes and Pitfalls. Nat. Rev. Drug Discov. 2009, 8, 33–40. [Google Scholar] [CrossRef] [PubMed]
  187. Braun, T.; Carvalho, G.; Fabre, C.; Grosjean, J.; Fenaux, P.; Kroemer, G. Targeting NF-KappaB in Hematologic Malignancies. Cell Death Differ. 2006, 13, 748–758. [Google Scholar] [CrossRef] [PubMed]
  188. Di Francesco, B.; Verzella, D.; Capece, D.; Vecchiotti, D.; Di Vito Nolfi, M.; Flati, I.; Cornice, J.; Di Padova, M.; Angelucci, A.; Alesse, E.; et al. NF-ΚB: A Druggable Target in Acute Myeloid Leukemia. Cancers 2022, 14, 3557. [Google Scholar] [CrossRef] [PubMed]
  189. Rasmi, R.R.; Sakthivel, K.M.; Guruvayoorappan, C. NF-ΚB Inhibitors in Treatment and Prevention of Lung Cancer. Biomed. Pharmacother. 2020, 130, 110569. [Google Scholar] [CrossRef]
  190. Zakaria, N.; Yusoff, N.M.; Zakaria, Z.; Widera, D.; Yahaya, B.H. Inhibition of NF-ΚB Signaling Reduces the Stemness Characteristics of Lung Cancer Stem Cells. Front. Oncol. 2018, 8, 366363. [Google Scholar] [CrossRef]
  191. Amentoflavone|C30H18O10|CID 5281600—PubChem. Available online: https://pubchem.ncbi.nlm.nih.gov/compound/Amentoflavone (accessed on 27 April 2024).
  192. Banerjee, T.; Valacchi, G.; Ziboh, V.A.; Van Der Vliet, A. Inhibition of TNFalpha-Induced Cyclooxygenase-2 Expression by Amentoflavone through Suppression of NF-KappaB Activation in A549 Cells. Mol. Cell. Biochem. 2002, 238, 105–110. [Google Scholar] [CrossRef]
  193. Su, C.M.; Li, C.H.; Huang, M.C.; Yueh, P.F.; Hsu, F.T.; Lin, R.F.; Hsu, L.C. Reinforcement of Sorafenib Anti-Osteosarcoma Effect by Amentoflavone Is Associated with the Induction of Apoptosis and Inactivation of ERK/NF-ΚB. In Vivo 2022, 36, 1136–1143. [Google Scholar] [CrossRef]
  194. Xiong, X.; Tang, N.; Lai, X.; Zhang, J.; Wen, W.; Li, X.; Li, A.; Wu, Y.; Liu, Z. Insights Into Amentoflavone: A Natural Multifunctional Biflavonoid. Front. Pharmacol. 2021, 12, 768708. [Google Scholar] [CrossRef]
  195. Pan, P.J.; Tsai, J.J.; Liu, Y.C. Amentoflavone Inhibits Metastatic Potential through Suppression of ERK/NF-ΚB Activation in Osteosarcoma U2OS Cells. Anticancer Res. 2017, 37, 4911–4918. [Google Scholar] [CrossRef] [PubMed]
  196. Lee, Y.J.; Chung, J.G.; Chien, Y.T.; Lin, S.S.; Hsu, F.T. Suppression of ERK/NF-ΚB Activation Is Associated with Amentoflavone-Inhibited Osteosarcoma Progression In Vivo. Anticancer Res. 2019, 39, 3669–3675. [Google Scholar] [CrossRef] [PubMed]
  197. Andrographolide|C20H30O5|CID 5318517—PubChem. Available online: https://pubchem.ncbi.nlm.nih.gov/compound/Andrographolide (accessed on 27 April 2024).
  198. Xia, Y.-F.; Ye, B.-Q.; Li, Y.-D.; Wang, J.-G.; He, X.-J.; Lin, X.; Yao, X.; Ma, D.; Slungaard, A.; Hebbel, R.P.; et al. Andrographolide Attenuates Inflammation by Inhibition of NF-Kappa B Activation through Covalent Modification of Reduced Cysteine 62 of P50. J. Immunol. 2004, 173, 4207–4217. [Google Scholar] [CrossRef]
  199. Tao, L.; Zhang, L.; Gao, R.; Jiang, F.; Cao, J.; Liu, H. Andrographolide Alleviates Acute Brain Injury in a Rat Model of Traumatic Brain Injury: Possible Involvement of Inflammatory Signaling. Front. Neurosci. 2018, 12, 657. [Google Scholar] [CrossRef] [PubMed]
  200. Liu, Y.; Zhang, Y.; Zou, J.; Yan, L.; Yu, X.; Lu, P.; Wu, X.; Li, Q.; Gu, R.; Zhu, D. Andrographolide Induces Autophagic Cell Death and Inhibits Invasion and Metastasis of Human Osteosarcoma Cells in An Autophagy-Dependent Manner. Cell. Physiol. Biochem. 2017, 44, 1396–1410. [Google Scholar] [CrossRef]
  201. Wang, S.; Li, H.; Chen, S.; Wang, Z.; Yao, Y.; Chen, T.; Ye, Z.; Lin, P. Andrographolide Induces Apoptosis in Human Osteosarcoma Cells via the ROS/JNK Pathway. Int. J. Oncol. 2020, 56, 1417–1428. [Google Scholar] [CrossRef]
  202. Huang, H.; Lu, Q.; Yuan, X.; Zhang, P.; Ye, C.; Wei, M.; Yang, C.; Zhang, L.; Huang, Y.; Luo, X.; et al. Andrographolide Inhibits the Growth of Human Osteosarcoma Cells by Suppressing Wnt/β-Catenin, PI3K/AKT and NF-ΚB Signaling Pathways. Chem. Biol. Interact. 2022, 365, 110068. [Google Scholar] [CrossRef]
  203. Pierce, J.W.; Schoenleber, R.; Jesmok, G.; Best, J.; Moore, S.A.; Collins, T.; Gerritsen, M.E. Novel Inhibitors of Cytokine-Induced IκBα Phosphorylation and Endothelial Cell Adhesion Molecule Expression Show Anti-Inflammatory Effects in Vivo. J. Biol. Chem. 1997, 272, 21096–21103. [Google Scholar] [CrossRef]
  204. Tan, B.; Yuan, Z.; Zhang, Q.; Xiqiang, X.; Dong, J. The NF-ΚB Pathway Is Critically Implicated in the Oncogenic Phenotype of Human Osteosarcoma Cells. J. Appl. Biomed. 2021, 19, 190–201. [Google Scholar] [CrossRef] [PubMed]
  205. Fukushima, T.; Kawaguchi, M.; Yorita, K.; Tanaka, H. Antitumor Effect of Dehydroxymethylepoxyquinomicin, a Small Molecule Inhibitor of Nuclear Factor-KB, on Glioblastoma. Neuro Oncol. 2012, 14, 19–28. [Google Scholar] [CrossRef] [PubMed]
  206. Katsman, A.; Umezawa, K.; Bonavida, B. Reversal of Resistance to Cytotoxic Cancer Therapies: DHMEQ as a Chemo-Sensitizing and Immuno-Sensitizing Agent. Drug Resist. Updates 2007, 10, 1–12. [Google Scholar] [CrossRef] [PubMed]
  207. Castro-Gamero, A.M.; Borges, K.S.; Silveira, V.d.S.; Lira, R.C.P.; Queiroz, R.d.P.G.; Valera, F.C.P.; Scrideli, C.A.; Umezawa, K.; Tone, L.G. Inhibition of Nuclear Factor-ΚB by Dehydroxymethylepoxyquinomicin Induces Schedule-Dependent Chemosensitivity to Anticancer Drugs and Enhances Chemoinduced Apoptosis in Osteosarcoma Cells. Anticancer Drugs 2012, 23, 638–650. [Google Scholar] [CrossRef] [PubMed]
  208. Dai, X.; Zhang, X.; Chen, W.; Chen, Y.; Zhang, Q.; Mo, S.; Lu, J. Dihydroartemisinin: A Potential Natural Anticancer Drug. Int. J. Biol. Sci. 2021, 17, 603–622. [Google Scholar] [CrossRef] [PubMed]
  209. Hu, W.; Chen, S.S.; Zhang, J.L.; Lou, X.E.; Zhou, H.J. Dihydroartemisinin Induces Autophagy by Suppressing NF-ΚB Activation. Cancer Lett. 2014, 343, 239–248. [Google Scholar] [CrossRef] [PubMed]
  210. Liu, Y.; Wang, W.; Xu, J.; Li, L.; Dong, Q.; Shi, Q.; Zuo, G.; Zhou, L.; Weng, Y.; Tang, M.; et al. Dihydroartemisinin Inhibits Tumor Growth of Human Osteosarcoma Cells by Suppressing Wnt/β-Catenin Signaling. Oncol. Rep. 2013, 30, 1723–1730. [Google Scholar] [CrossRef] [PubMed]
  211. Shen, Y.; Zhang, B.; Su, Y.; Badshah, S.A.; Wang, X.; Li, X.; Xue, Y.; Xie, L.; Wang, Z.; Yang, Z.; et al. Iron Promotes Dihydroartemisinin Cytotoxicity via ROS Production and Blockade of Autophagic Flux via Lysosomal Damage in Osteosarcoma. Front. Pharmacol. 2020, 11, 493314. [Google Scholar] [CrossRef]
  212. Ji, Y.; Zhang, Y.C.; Pei, L.B.; Shi, L.L.; Yan, J.L.; Ma, X.H. Anti-Tumor Effects of Dihydroartemisinin on Human Osteosarcoma. Mol. Cell Biochem. 2011, 351, 99–108. [Google Scholar] [CrossRef]
  213. Ding, X.; Zhang, Y.; Liang, J.; Li, Q.; Hu, H.; Zhou, Y.; Zhang, B. Dihydroartemisinin Potentiates VEGFR-TKIs Antitumorigenic Effect on Osteosarcoma by Regulating Loxl2/VEGFA Expression and Lipid Metabolism Pathway. J. Cancer 2023, 14, 809–820. [Google Scholar] [CrossRef] [PubMed]
  214. Wang, J.; Xu, P.; Zhang, Y.; Han, S.; Wang, G.; Wang, H.; Song, H.; Li, S. Dynamic Nanoassemblies Derived from Small-Molecule Homodimeric Prodrugs for in Situ Drug Activation and Safe Osteosarcoma Treatment. iScience 2023, 26, 107409. [Google Scholar] [CrossRef] [PubMed]
  215. Tang, N.; Ma, J.; Wang, K.S.; Mi, C.; Lv, Y.; Piao, L.X.; Xu, G.H.; Li, X.; Lee, J.J.; Jin, X. Dihydromyricetin Suppresses TNF-α-Induced NF-ΚB Activation and Target Gene Expression. Mol. Cell Biochem. 2016, 422, 11–20. [Google Scholar] [CrossRef] [PubMed]
  216. Zhao, Z.; Yin, J.Q.; Wu, M.S.; Song, G.; Xie, X.B.; Zou, C.; Tang, Q.; Wu, Y.; Lu, J.; Wang, Y.; et al. Dihydromyricetin Activates AMP-Activated Protein Kinase and P38(MAPK) Exerting Antitumor Potential in Osteosarcoma. Cancer Prev. Res. 2014, 7, 927–938. [Google Scholar] [CrossRef] [PubMed]
  217. Chou, C.H.; Lu, K.H.; Yang, J.S.; Hsieh, Y.H.; Lin, C.W.; Yang, S.F. Dihydromyricetin Suppresses Cell Metastasis in Human Osteosarcoma through SP-1- and NF-ΚB-Modulated Urokinase Plasminogen Activator Inhibition. Phytomedicine 2021, 90, 153642. [Google Scholar] [CrossRef] [PubMed]
  218. Wang, Y.; Wang, W.; Qiu, E. Protection of Oxidative Stress Induced Apoptosis in Osteosarcoma Cells by Dihydromyricetin through Down-Regulation of Caspase Activation and up-Regulation of BcL-2. Saudi J. Biol. Sci. 2017, 24, 837–842. [Google Scholar] [CrossRef] [PubMed]
  219. Kim, S.Y.; Kim, D.H.; Han, S.J.; Hyun, J.W.; Kim, H.S. Repression of Matrix Metalloproteinase Gene Expression by Ginsenoside Rh2 in Human Astroglioma Cells. Biochem. Pharmacol. 2007, 74, 1642–1651. [Google Scholar] [CrossRef] [PubMed]
  220. Jang, W.Y.; Hwang, J.Y.; Cho, J.Y. Ginsenosides from Panax Ginseng as Key Modulators of NF-ΚB Signaling Are Powerful Anti-Inflammatory and Anticancer Agents. Int. J. Mol. Sci. 2023, 24, 6119. [Google Scholar] [CrossRef] [PubMed]
  221. Liu, D.; Wang, H.; Zhou, Z.; Mao, X.; Ye, Z.; Zhang, Z.; Tu, S.; Zhang, Y.; Cai, X.; Lan, X.; et al. Integrated Bioinformatic Analysis and Experiment Confirmation of the Antagonistic Effect and Molecular Mechanism of Ginsenoside Rh2 in Metastatic Osteosarcoma. J. Pharm. Biomed. Anal. 2021, 201, 114088. [Google Scholar] [CrossRef]
  222. Fu, L.; Zhang, W.; Zhou, X.; Fu, J.; He, C. Tumor Cell Membrane-Camouflaged Responsive Nanoparticles Enable MRI-Guided Immuno-Chemodynamic Therapy of Orthotopic Osteosarcoma. Bioact. Mater. 2022, 17, 221–233. [Google Scholar] [CrossRef]
  223. Rasul, A.; Khan, M.; Ali, M.; Li, J.; Li, X. Targeting Apoptosis Pathways in Cancer with Alantolactone and Isoalantolactone. ScientificWorldJournal 2013, 2013, 248532. [Google Scholar] [CrossRef] [PubMed]
  224. Sun, S.I.; Ju, R.K.; Hyun, A.L.; Chan, H.J.; Young, K.K.; Konishi, T.; Eun, J.K.; Park, J.H.Y.; Kim, J.S. Induction of Detoxifying Enzyme by Sesquiterpenes Present in Inula Helenium. J. Med. Food 2007, 10, 503–510. [Google Scholar] [CrossRef]
  225. Konishi, T.; Shimada, Y.; Nagao, T.; Okabe, H.; Konoshima, T. Antiproliferative Sesquiterpene Lactones from the Roots of Inula Helenium. Biol. Pharm. Bull. 2002, 25, 1370–1372. [Google Scholar] [CrossRef] [PubMed]
  226. Di, W.; Khan, M.; Rasul, A.; Sun, M.; Sui, Y.; Zhong, L.; Yang, L.; Zhu, Q.; Feng, L.; Ma, T. Isoalantolactone Inhibits Constitutive NF-ΚB Activation and Induces Reactive Oxygen Species-Mediated Apoptosis in Osteosarcoma U2OS Cells through Mitochondrial Dysfunction. Oncol. Rep. 2014, 32, 1585–1593. [Google Scholar] [CrossRef]
  227. Wang, K.L.; Yu, Y.C.; Hsia, S.M. Perspectives on the Role of Isoliquiritigenin in Cancer. Cancers 2021, 13, 115. [Google Scholar] [CrossRef]
  228. Zhang, Z.; Yung, K.K.L.; Ko, J.K.S. Therapeutic Intervention in Cancer by Isoliquiritigenin from Licorice: A Natural Antioxidant and Redox Regulator. Antioxidants 2022, 11, 1349. [Google Scholar] [CrossRef]
  229. Blockade of Cytokine-Induced Endothelial Cell Adhesion Molecule Expression by Licorice Isoliquiritigenin through NF-KappaB Signal Disruption—PubMed. Available online: https://pubmed.ncbi.nlm.nih.gov/17259331/ (accessed on 27 April 2024).
  230. Jian, M.; Sun, X.; Cheng, G.; Zhang, H.; Li, X.; Song, F.; Liu, Z.; Wang, Z. Discovery of Phenolic Matrix Metalloproteinase Inhibitors by Peptide Microarray for Osteosarcoma Treatment. J. Nat. Prod. 2022, 85, 2424–2432. [Google Scholar] [CrossRef]
  231. Chen, J.; Liu, C.; Yang, Q.Q.; Ma, R.B.; Ke, Y.; Dong, F.; Wu, X.E. Isoliquiritigenin Suppresses Osteosarcoma U2OS Cell Proliferation and Invasion by Regulating the PI3K/Akt Signalling Pathway. Chemotherapy 2018, 63, 155–161. [Google Scholar] [CrossRef] [PubMed]
  232. Li, C.; Zhou, X.; Sun, C.; Liu, X.; Shi, X.; Wu, S. Isoliquiritigenin Inhibits the Proliferation, Apoptosis and Migration of Osteosarcoma Cells. Oncol. Rep. 2019, 41, 2502–2510. [Google Scholar] [CrossRef]
  233. Ferraz da Costa, D.C.; Rangel, L.P.; Duarte da Cunha Martins-Dinis, M.M.; da Silva Ferretti, G.D.; Ferreira, V.F.; Silva, J.L. Anticancer Potential of Resveratrol, β-Lapachone and Their Analogues. Molecules 2020, 25, 893. [Google Scholar] [CrossRef]
  234. Liu, T.J.; Lin, S.Y.; Chau, Y.P. Inhibition of Poly(ADP-Ribose) Polymerase Activation Attenuates β-Lapachone-Induced Necrotic Cell Death in Human Osteosarcoma Cells. Toxicol. Appl. Pharmacol. 2002, 182, 116–125. [Google Scholar] [CrossRef] [PubMed]
  235. Hori, T.; Kondo, T.; Lee, H.; Song, C.W.; Park, H.J. Hyperthermia Enhances the Effect of β-Lapachone to Cause ΓH2AX Formations and Cell Death in Human Osteosarcoma Cells. Int. J. Hyperth. 2011, 27, 53–62. [Google Scholar] [CrossRef] [PubMed]
  236. Shibata, S.; Saitoh, T. The Chemical Studies on the Oriental Plant Drugs. XIX. Some New Constituents of Licorice Root. 1. The Structure of Licoricidin. Chem. Pharm. Bull. 1968, 16, 1932–1936. [Google Scholar] [CrossRef] [PubMed]
  237. Ma, H.; Wu, F.; Bai, Y.; Wang, T.; Ma, S.; Guo, L.; Liu, G.; Leng, G.; Kong, Y.; Zhang, Y. Licoricidin Combats Gastric Cancer by Targeting the ICMT/Ras Pathway in Vitro and in Vivo. Front. Pharmacol. 2022, 13, 972825. [Google Scholar] [CrossRef] [PubMed]
  238. Ji, S.; Tang, S.; Li, K.; Li, Z.; Liang, W.; Qiao, X.; Wang, Q.; Yu, S.; Ye, M. Licoricidin Inhibits the Growth of SW480 Human Colorectal Adenocarcinoma Cells in Vitro and in Vivo by Inducing Cycle Arrest, Apoptosis and Autophagy. Toxicol. Appl. Pharmacol. 2017, 326, 25–33. [Google Scholar] [CrossRef] [PubMed]
  239. Wang, Y.; Wang, S.; Liu, J.; Lu, Y.; Li, D. Licoricidin Enhances Gemcitabine-Induced Cytotoxicity in Osteosarcoma Cells by Suppressing the Akt and NF-ΚB Signal Pathways. Chem. Biol. Interact. 2018, 290, 44–51. [Google Scholar] [CrossRef] [PubMed]
  240. Xu, T.; Kuang, T.; Du, H.; Li, Q.; Feng, T.; Zhang, Y.; Fan, G. Magnoflorine: A Review of Its Pharmacology, Pharmacokinetics and Toxicity. Pharmacol. Res. 2020, 152, 104632. [Google Scholar] [CrossRef]
  241. Wang, Y.; Shang, G.; Wang, W.; Qiu, E.; Pei, Y.; Zhang, X. Magnoflorine Inhibits the Malignant Phenotypes and Increases Cisplatin Sensitivity of Osteosarcoma Cells via Regulating MiR-410-3p/HMGB1/NF-ΚB Pathway. Life Sci. 2020, 256, 117967. [Google Scholar] [CrossRef] [PubMed]
  242. Lin, Y.; He, F.; Wu, L.; Xu, Y.; Du, Q. Matrine Exerts Pharmacological Effects through Multiple Signaling Pathways: A Comprehensive Review. Drug Des. Devel. Ther. 2022, 16, 533–569. [Google Scholar] [CrossRef]
  243. Li, Y.; Zhang, Z.N.; Zhao, H.M.; Tong, Z.C.; Yang, J.; Wang, H.; Liang, X.J. Matrine Inhibits the Invasive Properties of Human Osteosarcoma Cells by Downregulating the ERK-NF-ΚB Pathway. Anticancer Drugs 2014, 25, 1035–1043. [Google Scholar] [CrossRef]
  244. Zhou, H.; Chen, S.; Yang, Y.; Yang, C.; Chen, D.; Yao, Z.; Sun, B. Matrine Enhances the Efficacy of Adriamycin Chemotherapy in Osteosarcoma Cells by the STAT3 Pathway. Anticancer Drugs 2019, 30, 1006–1012. [Google Scholar] [CrossRef] [PubMed]
  245. Alzohairy, M.A. Therapeutics Role of Azadirachta Indica (Neem) and Their Active Constituents in Diseases Prevention and Treatment. Evid. Based Complement. Alternat Med. 2016, 2016, 7382506. [Google Scholar] [CrossRef] [PubMed]
  246. Cohen, E.; Quistad, G.B.; Casida, J.E. Cytotoxicity of Nimbolide, Epoxyazadiradione and Other Limonoids from Neem Insecticide. Life Sci. 1996, 58, 1075–1081. [Google Scholar] [CrossRef] [PubMed]
  247. Liu, J.F.; Hou, C.H.; Lin, F.L.; Tsao, Y.T.; Hou, S.M. Nimbolide Induces ROS-Regulated Apoptosis and Inhibits Cell Migration in Osteosarcoma. Int. J. Mol. Sci. 2015, 16, 23405–23424. [Google Scholar] [CrossRef] [PubMed]
  248. Blanco, J.; Martín, H.; Mariño, C.; Rossignoli, A.E. Okadaic Acid Depuration from the Cockle Cerastoderma Edule. Toxins 2022, 14, 216. [Google Scholar] [CrossRef] [PubMed]
  249. Fu, L.; Zhao, X.Y.; Ji, L.D.; Xu, J. Okadaic Acid (OA): Toxicity, Detection and Detoxification. Toxicon 2019, 160, 1–7. [Google Scholar] [CrossRef] [PubMed]
  250. Yang, D.; Okamura, H.; Morimoto, H.; Teramachi, J.; Haneji, T. Protein Phosphatase 2A Cα Regulates Proliferation, Migration, and Metastasis of Osteosarcoma Cells. Lab. Investig. 2016, 96, 1050–1062. [Google Scholar] [CrossRef] [PubMed]
  251. Freund, R.R.A.; Gobrecht, P.; Fischer, D.; Arndt, H.D. Advances in Chemistry and Bioactivity of Parthenolide. Nat. Prod. Rep. 2020, 37, 541–565. [Google Scholar] [CrossRef] [PubMed]
  252. Pareek, A.; Suthar, M.; Rathore, G.S.; Bansal, V. Feverfew (Tanacetum Parthenium L.): A Systematic Review. Pharmacogn. Rev. 2011, 5, 103–110. [Google Scholar] [CrossRef]
  253. Kwok, B.H.B.; Koh, B.; Ndubuisi, M.I.; Elofsson, M.; Crews, C.M. The Anti-Inflammatory Natural Product Parthenolide from the Medicinal Herb Feverfew Directly Binds to and Inhibits IκB Kinase. Chem. Biol. 2001, 8, 759–766. [Google Scholar] [CrossRef]
  254. D’Anneo, A.; Carlisi, D.; Lauricella, M.; Emanuele, S.; Di Fiore, R.; Vento, R.; Tesoriere, G. Parthenolide Induces Caspase-Independent and AIF-Mediated Cell Death in Human Osteosarcoma and Melanoma Cells. J. Cell Physiol. 2013, 228, 952–967. [Google Scholar] [CrossRef]
  255. Kishida, Y.; Yoshikawa, H.; Myoui, A. Parthenolide, a Natural Inhibitor of Nuclear Factor-KappaB, Inhibits Lung Colonization of Murine Osteosarcoma Cells. Clin. Cancer Res. 2007, 13, 59–67. [Google Scholar] [CrossRef]
  256. Zuch, D.; Giang, A.H.; Shapovalov, Y.; Schwarz, E.; Rosier, R.; O’Keefe, R.; Eliseev, R.A. Targeting Radioresistant Osteosarcoma Cells with Parthenolide. J. Cell Biochem. 2012, 113, 1282–1291. [Google Scholar] [CrossRef]
  257. Sugiyasu, K.; Nanno, K.; Tamai, N.; Hashimoto, N.; Kishida, Y.; Yoshikawa, H.; Myoui, A. Radio-Sensitization of the Murine Osteosarcoma Cell Line LM8 with Parthenolide, a Natural Inhibitor of NF-ΚB. Oncol. Lett. 2011, 2, 407–412. [Google Scholar] [CrossRef] [PubMed]
  258. Duan, H.; Wang, R.; Yan, X.; Liu, H.; Zhang, Y.; Mu, D.; Han, J.; Li, X. Phloretin Induces Apoptosis of Human Esophageal Cancer via a Mitochondria-Dependent Pathway. Oncol. Lett. 2017, 14, 6763–6768. [Google Scholar] [CrossRef]
  259. Tuli, H.S.; Rath, P.; Chauhan, A.; Ramniwas, S.; Vashishth, K.; Varol, M.; Jaswal, V.S.; Haque, S.; Sak, K. Phloretin, as a Potent Anticancer Compound: From Chemistry to Cellular Interactions. Molecules 2022, 27, 8819. [Google Scholar] [CrossRef]
  260. Huang, W.C.; Dai, Y.W.; Peng, H.L.; Kang, C.W.; Kuo, C.Y.; Liou, C.J. Phloretin Ameliorates Chemokines and ICAM-1 Expression via Blocking of the NF-ΚB Pathway in the TNF-α-Induced HaCaT Human Keratinocytes. Int. Immunopharmacol. 2015, 27, 32–37. [Google Scholar] [CrossRef] [PubMed]
  261. Hsieh, I.S.; Yang, R.S.; Fu, W.M. Osteopontin Upregulates the Expression of Glucose Transporters in Osteosarcoma Cells. PLoS ONE 2014, 9, e109550. [Google Scholar] [CrossRef] [PubMed]
  262. Huang, T.; Zhang, X.; Wang, H. Punicalagin Inhibited Proliferation, Invasion and Angiogenesis of Osteosarcoma through Suppression of NF-κB Signaling. Mol. Med. Rep. 2020, 22, 2386–2394. [Google Scholar] [CrossRef]
  263. Wang, X.Z.; Zhang, S.F.; Yang, Z.H.; Ye, Z.W.; Liu, J. Punicalagin Suppresses Osteosarcoma Growth and Metastasis by Regulating NF-ΚB Signaling. J. Biol. Regul. Homeost. Agents 2020, 5, 1699–1708. [Google Scholar] [CrossRef]
  264. Naz, I.; Ramchandani, S.; Khan, M.R.; Yang, M.H.; Ahn, K.S. Anticancer Potential of Raddeanin A, a Natural Triterpenoid Isolated from Anemone Raddeana Regel. Molecules 2020, 25, 1035. [Google Scholar] [CrossRef] [PubMed]
  265. Ma, B.; Zhu, J.; Zhao, A.; Zhang, J.; Wang, Y.; Zhang, H.; Zhang, L.; Zhang, Q. Raddeanin A, a Natural Triterpenoid Saponin Compound, Exerts Anticancer Effect on Human Osteosarcoma via the ROS/JNK and NF-ΚB Signal Pathway. Toxicol. Appl. Pharmacol. 2018, 353, 87–101. [Google Scholar] [CrossRef] [PubMed]
  266. Wang, Z.; Shen, J.; Sun, W.; Zhang, T.; Zuo, D.; Wang, H.; Wang, G.; Xu, J.; Yin, F.; Mao, M.; et al. Antitumor Activity of Raddeanin A Is Mediated by Jun Amino-Terminal Kinase Activation and Signal Transducer and Activator of Transcription 3 Inhibition in Human Osteosarcoma. Cancer Sci. 2019, 110, 1746–1759. [Google Scholar] [CrossRef] [PubMed]
  267. Wang, Z.; Wang, C.; Zuo, D.; Zhang, T.; Yin, F.; Zhou, Z.; Wang, H.; Xu, J.; Mao, M.; Wang, G.; et al. Attenuation of STAT3 Phosphorylation Promotes Apoptosis and Chemosensitivity in Human Osteosarcoma Induced by Raddeanin A. Int. J. Biol. Sci. 2019, 15, 668–679. [Google Scholar] [CrossRef] [PubMed]
  268. Yagishita, Y.; Fahey, J.W.; Dinkova-Kostova, A.T.; Kensler, T.W. Broccoli or Sulforaphane: Is It the Source or Dose That Matters? Molecules 2019, 24, 3593. [Google Scholar] [CrossRef]
  269. Zhang, G.; Jin, C.; Zhu, Y.; Fu, F.; Wang, G.; Li, S. Sulforaphene Inhibits the Progression of Osteosarcoma via Regulating FSTL1/NF-ΚB Pathway. Life Sci. 2020, 263, 118485. [Google Scholar] [CrossRef] [PubMed]
  270. Lin, J.; Wang, Q.; Zhou, S.; Xu, S.; Yao, K. Tetramethylpyrazine: A Review on Its Mechanisms and Functions. Biomed. Pharmacother. 2022, 150, 113005. [Google Scholar] [CrossRef] [PubMed]
  271. Wang, Y.; Fu, Q.; Zhao, W. Tetramethylpyrazine Inhibits Osteosarcoma Cell Proliferation via Downregulation of NF-ΚB in Vitro and in Vivo. Mol. Med. Rep. 2013, 8, 984–988. [Google Scholar] [CrossRef] [PubMed]
  272. Jin, W.; Zhou, L.; Yan, B.; Yan, L.; Liu, F.; Tong, P.; Yu, W.; Dong, X.; Xie, L.; Zhang, J.; et al. Theabrownin Triggers DNA Damage to Suppress Human Osteosarcoma U2OS Cells by Activating P53 Signalling Pathway. J. Cell Mol. Med. 2018, 22, 4423–4436. [Google Scholar] [CrossRef]
  273. Jin, W.; Gu, C.; Zhou, L.; Yang, X.; Gui, M.; Zhang, J.; Chen, J.; Dong, X.; Yuan, Q.; Shan, L. Theabrownin Inhibits the Cytoskeleton-dependent Cell Cycle, Migration and Invasion of Human Osteosarcoma Cells through NF-κB Pathway-related Mechanisms. Oncol. Rep. 2020, 44, 2621–2633. [Google Scholar] [CrossRef]
  274. Alhmied, F.; Alammar, A.; Alsultan, B.; Alshehri, M.; Pottoo, F.H. Molecular Mechanisms of Thymoquinone as Anticancer Agent. Comb. Chem. High. Throughput Screen. 2021, 24, 1644–1653. [Google Scholar] [CrossRef]
  275. Homayoonfal, M.; Asemi, Z.; Yousefi, B. Potential Anticancer Properties and Mechanisms of Thymoquinone in Osteosarcoma and Bone Metastasis. Cell. Mol. Biol. Lett. 2022, 27, 1–28. [Google Scholar] [CrossRef] [PubMed]
  276. Effects of Thymoquinone and Selenium on the Proliferation of Mg 63 Cells in Tissue Culture—PubMed. Available online: https://pubmed.ncbi.nlm.nih.gov/19141954/ (accessed on 27 April 2024).
  277. Sarman, H.; Bayram, R.; Benek, S.B. Anticancer Drugs with Chemotherapeutic Interactions with Thymoquinone in Osteosarcoma Cells. Eur. Rev. Med. Pharmacol. Sci. 2016, 20, 1263–1270. [Google Scholar] [PubMed]
  278. Ahmadzadeh, H.; Ahmadi, M.; Golchin, A.; Malakoti, F.; Maleki, M.; Alemi, F.; Bazavar, M.; Yousefi, B. The Effect of TQ and Cis in OS. Drug Res. 2022, 72, 171–176. [Google Scholar] [CrossRef] [PubMed]
  279. Sanapour, N.; Malakoti, F.; Shanebandi, D.; Targhazeh, N.; Yousefi, B.; Soleimanpour, J.; Majidinia, M. Thymoquinone Augments Methotrexate-Induced Apoptosis on Osteosarcoma Cells. Drug Res. 2022, 72, 220–225. [Google Scholar] [CrossRef] [PubMed]
  280. Khyavi, P.A.; Valizadeh, A.; Shanehbandi, D.; Yousefi, B.; Soleimanpour, J. Thymoquinone Potentiates Methotrexate Mediated-Apoptosis in Saos-2 Osteosarcoma Cell Line. Drug Res. 2022, 72, 390–395. [Google Scholar] [CrossRef]
  281. Roepke, M.; Diestel, A.; Bajbouj, K.; Walluscheck, D.; Schonfeld, P.; Roessner, A.; Schneider-Stock, R.; Gali-Muhtasib, H. Lack of P53 Augments Thymoquinone-Induced Apoptosis and Caspase Activation in Human Osteosarcoma Cells. Cancer Biol. Ther. 2007, 6, 160–169. [Google Scholar] [CrossRef] [PubMed]
  282. Peng, L.; Liu, A.; Shen, Y.; Xu, H.Z.; Yang, S.Z.; Ying, X.Z.; Liao, W.; Liu, H.X.; Lin, Z.Q.; Chen, Q.Y.; et al. Antitumor and Anti-Angiogenesis Effects of Thymoquinone on Osteosarcoma through the NF-ΚB Pathway. Oncol. Rep. 2013, 29, 571–578. [Google Scholar] [CrossRef] [PubMed]
  283. Manu, K.A.; Kuttan, G. Ursolic Acid Induces Apoptosis by Activating P53 and Caspase-3 Gene Expressions and Suppressing NF-KappaB Mediated Activation of Bcl-2 in B16F-10 Melanoma Cells. Int. Immunopharmacol. 2008, 8, 974–981. [Google Scholar] [CrossRef]
  284. Hsu, Y.L.; Kuo, P.L.; Lin, C.C. Proliferative Inhibition, Cell-Cycle Dysregulation, and Induction of Apoptosis by Ursolic Acid in Human Non-Small Cell Lung Cancer A549 Cells. Life Sci. 2004, 75, 2303–2316. [Google Scholar] [CrossRef]
  285. Zhang, R.X.; Li, Y.; Tian, D.D.; Liu, Y.; Nian, W.; Zou, X.; Chen, Q.Z.; Zhou, L.Y.; Deng, Z.L.; He, B.C. Ursolic Acid Inhibits Proliferation and Induces Apoptosis by Inactivating Wnt/β-Catenin Signaling in Human Osteosarcoma Cells. Int. J. Oncol. 2016, 49, 1973–1982. [Google Scholar] [CrossRef] [PubMed]
  286. Wu, C.C.; Cheng, C.H.; Lee, Y.H.; Chang, I.L.; Chen, H.Y.; Hsieh, C.P.; Chueh, P.J. Ursolic Acid Triggers Apoptosis in Human Osteosarcoma Cells via Caspase Activation and the ERK1/2 MAPK Pathway. J. Agric. Food Chem. 2016, 64, 4220–4226. [Google Scholar] [CrossRef] [PubMed]
  287. Chen, J.; Fu, H.; Wang, Z.; Yin, F.; Li, J.; Hua, Y.; Cai, Z. A New Synthetic Ursolic Acid Derivative IUA with Anti-Tumor Efficacy Against Osteosarcoma Cells via Inhibition of JNK Signaling Pathway. Cell. Physiol. Biochem. 2014, 34, 724–733. [Google Scholar] [CrossRef] [PubMed]
  288. Wu, C.C.; Huang, Y.F.; Hsieh, C.P.; Chueh, P.J.; Chen, Y.L. Combined Use of Zoledronic Acid Augments Ursolic Acid-Induced Apoptosis in Human Osteosarcoma Cells through Enhanced Oxidative Stress and Autophagy. Molecules 2016, 21, 1640. [Google Scholar] [CrossRef] [PubMed]
  289. Fu, D.; Ni, Z.; Wu, K.; Cheng, P.; Ji, X.; Li, G.; Shang, X. A Novel Redox-Responsive Ursolic Acid Polymeric Prodrug Delivery System for Osteosarcoma Therapy. Drug Deliv. 2021, 28, 195–205. [Google Scholar] [CrossRef] [PubMed]
  290. Zhang, W.; Jiang, G.; Zhou, X.; Huang, L.; Meng, J.; He, B.; Qi, Y. α-Mangostin Inhibits LPS-Induced Bone Resorption by Restricting Osteoclastogenesis via NF-ΚB and MAPK Signaling. Chin. Med. 2022, 17, 34. [Google Scholar] [CrossRef]
  291. Yang, S.; Zhou, F.; Dong, Y.; Ren, F. α-Mangostin Induces Apoptosis in Human Osteosarcoma Cells Through ROS-Mediated Endoplasmic Reticulum Stress via the WNT Pathway. Cell Transplant. 2021, 30, 09636897211035080. [Google Scholar] [CrossRef] [PubMed]
  292. Park, S.J.; Park, B.S.; Yu, S.B.; Kang, H.M.; Kim, H.J.; Kim, I.R. Induction of Apoptosis and Inhibition of Epithelial Mesenchymal Transition by α-Mangostin in MG-63 Cell Lines. Evid.-Based Complement. Altern. Med. 2018, 2018, 3985082. [Google Scholar] [CrossRef]
  293. Zhang, B.; Shi, Z.L.; Liu, B.; Yan, X.B.; Feng, J.; Tao, H.M. Enhanced Anticancer Effect of Gemcitabine by Genistein in Osteosarcoma: The Role of Akt and Nuclear Factor-KappaB. Anticancer Drugs 2010, 21, 288–296. [Google Scholar] [CrossRef]
  294. Sharifi-Rad, J.; Quispe, C.; Imran, M.; Rauf, A.; Nadeem, M.; Gondal, T.A.; Ahmad, B.; Atif, M.; Mubarak, M.S.; Sytar, O.; et al. Genistein: An Integrative Overview of Its Mode of Action, Pharmacological Properties, and Health Benefits. Oxid. Med. Cell Longev. 2021, 2021, 3268136. [Google Scholar] [CrossRef]
  295. Liang, C.; Li, H.; Shen, C.; Lai, J.; Shi, Z.; Liu, B.; Tao, H. Genistein Potentiates the Anti-Cancer Effects of Gemcitabine in Human Osteosarcoma via the Downregulation of Akt and Nuclear Factor-ΚB Pathway. Anticancer Agents Med. Chem. 2012, 12, 554–563. [Google Scholar] [CrossRef] [PubMed]
  296. Nakamura, A.; Aizawa, J.; Sakayama, K.; Kidani, T.; Takata, T.; Norimatsu, Y.; Miura, H.; Masuno, H. Genistein Inhibits Cell Invasion and Motility by Inducing Cell Differentiation in Murine Osteosarcoma Cell Line LM8. BMC Cell Biol. 2012, 13, 24. [Google Scholar] [CrossRef] [PubMed]
  297. Hagiwara, H.; Wako, H.; Nakata, K.; Aida, R. Genistein Induces Antiproliferative Activity and Apoptosis in Human Osteosarcoma Saos-2 Cells. Anticancer Res. 2023, 43, 5387–5392. [Google Scholar] [CrossRef] [PubMed]
  298. Kidani, T.; Nakamura, A.; Kamei, S.; Norimatsu, Y.; Miura, H.; Masuno, H. Overexpression of Cytoplasmic β-Catenin Inhibits the Metastasis of the Murine Osteosarcoma Cell Line LM8. Cancer Cell Int. 2014, 14, 31. [Google Scholar] [CrossRef] [PubMed]
  299. Lee, J.; Jung, E.; Park, J.; Jung, K.; Lee, S.; Hong, S.; Park, J.; Park, E.; Kim, J.; Park, S.; et al. Anti-Inflammatory Effects of Magnolol and Honokiol Are Mediated through Inhibition of the Downstream Pathway of MEKK-1 in NF-KappaB Activation Signaling. Planta Med. 2005, 71, 338–343. [Google Scholar] [CrossRef] [PubMed]
  300. Li, C.H.; Ku, M.C.; Lee, K.C.; Yueh, P.F.; Hsu, F.T.; Lin, R.F.; Yang, C.C.; Wang, W.C.; Chen, J.H.; Hsu, L.C.; et al. Magnolol Suppresses ERK/NF-ΚB Signaling and Triggers Apoptosis Through Extrinsic/Intrinsic Pathways in Osteosarcoma. Anticancer Res. 2022, 42, 4403–4410. [Google Scholar] [CrossRef] [PubMed]
  301. Zhou, S.; Wen, H.; Li, H. Magnolol Induces Apoptosis in Osteosarcoma Cells via G0/G1 Phase Arrest and P53-Mediated Mitochondrial Pathway. J. Cell Biochem. 2019, 120, 17067–17079. [Google Scholar] [CrossRef]
  302. Van Stiphout, C.M.; Luu, A.K.; Viloria-Petit, A.M. Proteasome Inhibitors and Their Potential Applicability in Osteosarcoma Treatment. Cancers 2022, 14, 4544. [Google Scholar] [CrossRef] [PubMed]
  303. Asanuma, K.; Nakamura, T.; Nakamura, K.; Hagi, T.; Okamoto, T.; Kita, K.; Matsuyama, Y.; Yoshida, K.; Asanuma, Y.; Sudo, A. Compound Library Screening for Synergistic Drug Combinations: MTOR Inhibitor and Proteasome Inhibitor Effective against Osteosarcoma Cells. Anticancer Res. 2022, 42, 4319–4328. [Google Scholar] [CrossRef]
  304. Nakamura, K.; Asanuma, K.; Okamoto, T.; Iino, T.; Hagi, T.; Nakamura, T.; Sudo, A. Combination of Everolimus and Bortezomib Inhibits the Growth and Metastasis of Bone and Soft Tissue Sarcomas via JNK/P38/ERK MAPK and AKT Pathways. Cancers 2023, 15, 2468. [Google Scholar] [CrossRef]
  305. Shishodia, S.; Amin, H.M.; Lai, R.; Aggarwal, B.B. Curcumin (Diferuloylmethane) Inhibits Constitutive NF-ΚB Activation, Induces G1/S Arrest, Suppresses Proliferation, and Induces Apoptosis in Mantle Cell Lymphoma. Biochem. Pharmacol. 2005, 70, 700–713. [Google Scholar] [CrossRef] [PubMed]
  306. Yu, D.; An, F.; He, X.; Cao, X. Curcumin Inhibits the Proliferation and Invasion of Human Osteosarcoma Cell Line MG-63 by Regulating MiR-138. Int. J. Clin. Exp. Pathol. 2015, 8, 14946. [Google Scholar] [PubMed]
  307. Aziz, M.N.M.; Rahim, N.F.C.; Hussin, Y.; Yeap, S.K.; Masarudin, M.J.; Mohamad, N.E.; Akhtar, M.N.; Osman, M.A.; Cheah, Y.K.; Alitheen, N.B. Anti-Metastatic and Anti-Angiogenic Effects of Curcumin Analog DK1 on Human Osteosarcoma Cells In Vitro. Pharmaceuticals 2021, 14, 532. [Google Scholar] [CrossRef] [PubMed]
  308. Lu, K.H.; Lu, P.W.A.; Lin, C.W.; Yang, S.F. Curcumin in Human Osteosarcoma: From Analogs to Carriers. Drug Discov. Today 2023, 28, 103437. [Google Scholar] [CrossRef] [PubMed]
  309. Maran, A.; Yaszemski, M.J.; Kohut, A.; Voronov, A. Curcumin and Osteosarcoma: Can Invertible Polymeric Micelles Help? Materials 2016, 9, 520. [Google Scholar] [CrossRef] [PubMed]
  310. Xu, C.; Wang, M.; Zandieh Doulabi, B.; Sun, Y.; Liu, Y. Paradox: Curcumin, a Natural Antioxidant, Suppresses Osteosarcoma Cells via Excessive Reactive Oxygen Species. Int. J. Mol. Sci. 2023, 24, 11975. [Google Scholar] [CrossRef] [PubMed]
  311. Yuan, C.; Fan, R.; Zhu, K.; Wang, Y.; Xie, W.; Liang, Y. Curcumin Induces Ferroptosis and Apoptosis in Osteosarcoma Cells by Regulating Nrf2/GPX4 Signaling Pathway. Exp. Biol. Med. 2023, 248, 2183. [Google Scholar] [CrossRef]
  312. Zahedipour, F.; Bolourinezhad, M.; Teng, Y.; Sahebkar, A. The Multifaceted Therapeutic Mechanisms of Curcumin in Osteosarcoma: State-of-the-Art. J. Oncol. 2021, 2021, 3006853. [Google Scholar] [CrossRef] [PubMed]
  313. Xu, C.; Wang, M.; Guo, W.; Sun, W.; Liu, Y. Curcumin in Osteosarcoma Therapy: Combining with Immunotherapy, Chemotherapeutics, Bone Tissue Engineering Materials and Potential Synergism with Photodynamic Therapy. Front. Oncol. 2021, 11, 672490. [Google Scholar] [CrossRef]
  314. Dhule, S.S.; Penfornis, P.; He, J.; Harris, M.R.; Terry, T.; John, V.; Pochampally, R. The Combined Effect of Encapsulating Curcumin and C6 Ceramide in Liposomal Nanoparticles against Osteosarcoma. Mol. Pharm. 2014, 11, 417–427. [Google Scholar] [CrossRef]
  315. Ma, D.; Tremblay, P.; Mahngar, K.; Collins, J.; Hudlicky, T.; Pandey, S. Selective Cytotoxicity against Human Osteosarcoma Cells by a Novel Synthetic C-1 Analogue of 7-Deoxypancratistatin Is Potentiated by Curcumin. PLoS ONE 2011, 6, e28780. [Google Scholar] [CrossRef] [PubMed]
  316. Lamoia, T.E.; Shulman, G.I. Cellular and Molecular Mechanisms of Metformin Action. Endocr. Rev. 2021, 42, 77–96. [Google Scholar] [CrossRef] [PubMed]
  317. Sui, X.; Xu, Y.; Wang, X.; Han, W.; Pan, H.; Xiao, M. Metformin: A Novel but Controversial Drug in Cancer Prevention and Treatment. Mol. Pharm. 2015, 12, 3783–3791. [Google Scholar] [CrossRef] [PubMed]
  318. Ko, Y.; Choi, A.; Lee, M.; Lee, J.A. Metformin Displays in Vitro and in Vivo Antitumor Effect against Osteosarcoma. Korean J. Pediatr. 2016, 59, 374. [Google Scholar] [CrossRef] [PubMed]
  319. Li, B.; Zhou, P.; Xu, K.; Chen, T.; Jiao, J.; Wei, H.; Yang, X.; Xu, W.; Wan, W.; Xiao, J. Metformin Induces Cell Cycle Arrest, Apoptosis and Autophagy through ROS/JNK Signaling Pathway in Human Osteosarcoma. Int. J. Biol. Sci. 2020, 16, 74–84. [Google Scholar] [CrossRef]
  320. Metts, J.L.; Trucco, M.; Weiser, D.A.; Thompson, P.; Sandler, E.; Smith, T.; Crimella, J.; Sansil, S.; Thapa, R.; Fridley, B.L.; et al. A Phase I Trial of Metformin in Combination with Vincristine, Irinotecan, and Temozolomide in Children with Relapsed or Refractory Solid and Central Nervous System Tumors: A Report from the National Pediatric Cancer Foundation. Cancer Med. 2023, 12, 4270–4281. [Google Scholar] [CrossRef]
  321. Miwa, S.; Sugimoto, N.; Yamamoto, N.; Shirai, T.; Nishida, H.; Hayashi, K.; Kimura, H.; Takeuchi, A.; Igarashi, K.; Yachie, A.; et al. Caffeine Induces Apoptosis of Osteosarcoma Cells by Inhibiting AKT/MTOR/S6K, NF-ΚB and MAPK Pathways. Anticancer Res. 2012, 32, 3643–3649. [Google Scholar] [PubMed]
  322. Miwa, S.; Sugimoto, N.; Shirai, T.; Hayashi, K.; Nishida, H.; Ohnari, I.; Takeuchi, A.; Yachie, A.; Tsuchiya, H. Caffeine Activates Tumor Suppressor PTEN in Sarcoma Cells. Int. J. Oncol. 2011, 39, 465–472. [Google Scholar] [CrossRef] [PubMed]
  323. Ii, S.; Ueda, Y.; Shimazaki, M.; Katsuta, S.; Takazawa, K.; Kanazawa, Y.; Tomita, K.; Tsuchiya, H. Identification of Novel Genes Involved in the Synergistic Antitumor Effect of Caffeine in Osteosarcoma Cells Using CDNA Macroarray. Anticancer Res. 2008, 28, 645–653. [Google Scholar]
  324. Kawahara, M.; Takahashi, Y.; Takazawa, K.; Tsuchiya, H.; Tomita, K.; Yokogawa, K.; Miyamoto, K.I. Caffeine Dose-Dependently Potentiates the Antitumor Effect of Cisplatin on Osteosarcomas. Anticancer Res. 2008, 28, 1681–1685. [Google Scholar]
  325. Abe, K.; Yamamoto, N.; Hayashi, K.; Takeuchi, A.; Tsuchiya, H. Caffeine Citrate Enhanced Cisplatin Antitumor Effects in Osteosarcoma and Fibrosarcoma in Vitro and in Vivo. BMC Cancer 2019, 19, 689. [Google Scholar] [CrossRef] [PubMed]
  326. Higuchi, T.; Oshiro, H.; Miyake, K.; Sugisawa, N.; Han, Q.; Tan, Y.; Park, J.; Zhang, Z.; Razmjooei, S.; Yamamoto, N.; et al. Oral Recombinant Methioninase, Combined with Oral Caffeine and Injected Cisplatinum, Overcome Cisplatinum-Resistance and Regresses Patient-Derived Orthotopic Xenograft Model of Osteosarcoma. Anticancer Res. 2019, 39, 4653–4657. [Google Scholar] [CrossRef] [PubMed]
  327. Igarashi, K.; Kawaguchi, K.; Kiyuna, T.; Murakami, T.; Yamamoto, N.; Hayashi, K.; Kimura, H.; Miwa, S.; Tsuchiya, H.; Hoffman, R.M. Antimetastatic Efficacy of the Combination of Caffeine and Valproic Acid on an Orthotopic Human Osteosarcoma Cell Line Model in Nude Mice. Anticancer Res. 2017, 37, 1005–1011. [Google Scholar] [CrossRef] [PubMed]
Pharmaceuticals 17 00734 g001
Figure 1. (A) The NF-κB family consists of 5 protein members: RELA (p65), RELB, c-REL, NF-κB1 (p105/p50), and NF-κB2 (p100/p52). Each of these proteins contains a stretch of 300 amino acids called the Rel Homology Domain (RHD), which is responsible for mediating DNA binding, dimerization, and interaction with IκB (IkappaB kinase or IKK) through the nuclear localization signal (NLS). All members also contain a transactivation domain (TAD), which mediates transcriptional induction. RELB also contains a leucine-zipper region that cooperates with the TAD. NF-κB1 and NF-κB2, which require proteolytic activation, present a glycine rich region (GRR) and multiple copies of ankyrin repeats (ANK) (within which resides the processing-inhibitory domain—PID) that are characteristic for the IκB protein family. (B) These proteins are found as hetero- or homodimers in the cytoplasm. Fifteen different dimers can be formed, among which RELA/p52, RELB/p52, and p52/p52 are the most commonly found in vivo. Once activated, the dimers are translocated to the cell nucleus where they exert their transcriptional function. This figure was created using Servier Medical Art templates, which are licensed under a Creative Commons Attribution 3.0 Unported License; https://smart.servier.com accessed on 10 April 2024.
Figure 1. (A) The NF-κB family consists of 5 protein members: RELA (p65), RELB, c-REL, NF-κB1 (p105/p50), and NF-κB2 (p100/p52). Each of these proteins contains a stretch of 300 amino acids called the Rel Homology Domain (RHD), which is responsible for mediating DNA binding, dimerization, and interaction with IκB (IkappaB kinase or IKK) through the nuclear localization signal (NLS). All members also contain a transactivation domain (TAD), which mediates transcriptional induction. RELB also contains a leucine-zipper region that cooperates with the TAD. NF-κB1 and NF-κB2, which require proteolytic activation, present a glycine rich region (GRR) and multiple copies of ankyrin repeats (ANK) (within which resides the processing-inhibitory domain—PID) that are characteristic for the IκB protein family. (B) These proteins are found as hetero- or homodimers in the cytoplasm. Fifteen different dimers can be formed, among which RELA/p52, RELB/p52, and p52/p52 are the most commonly found in vivo. Once activated, the dimers are translocated to the cell nucleus where they exert their transcriptional function. This figure was created using Servier Medical Art templates, which are licensed under a Creative Commons Attribution 3.0 Unported License; https://smart.servier.com accessed on 10 April 2024.
Pharmaceuticals 17 00734 g001
Pharmaceuticals 17 00734 g002
Figure 2. (A) The canonical activation of the NF-κB pathway begins with the activation of receptors located on the cell surface (toll-like receptors, TLR; receptor tyrosine kinases, RTK; and tumor necrosis factor receptors, TNFR, for instance), which then emit signals through adaptor proteins to activate the IkB kinase (IKK) complex (composed of IKK-α, the catalytic subunit IKK-β, and the regulatory subunit NEMO). Once activated, the IKK complex catalyzes the polyubiquitination and the phosphorylation of the IkBs leading to their degradation by proteasome 26S. After this, the NF-κB dimers that are released from their inhibitors and translocate to the nucleus, where they activate the transcription of many genes through binding to the consensus sequence 5′-GGGRN W YYCC-3′ (R = purine base, N = any base, W = adenine or thymine, and Y = pyrimidine base). Alternatively, the non-canonical signaling cascade begins with the activation of LT-β or BAFF receptors, which emit signals through the cytoplasm leading to the phosphorylation of the NIK protein, which, in turn, phosphorylates the IKK complex (formed by only two IKK-α subunits). Once activated, this complex phosphorylates NF-κB2/p100, in two C-terminal sites, leading to its proteolytic cleavage. This process partially degrades p100 into p52. Subsequently, the NF-κB dimer is released and translocates to the nucleus, activating gene transcription. (B) In cancer cells, mutations, gene amplification and gene fusions involving its subunits, or interplay with other dysregulated signaling pathways may lead to the constitutive activation of NF-κB which contributes to cancer development and progression by increasing the expression of genes associated with antiapoptosis, survival, chemo- and radioresistance, adherence, invasion, and metastasis. This figure was created using Servier Medical Art templates, which are licensed under a Creative Commons Attribution 3.0 Unported License; https://smart.servier.com accessed on 10 April 2024.
Figure 2. (A) The canonical activation of the NF-κB pathway begins with the activation of receptors located on the cell surface (toll-like receptors, TLR; receptor tyrosine kinases, RTK; and tumor necrosis factor receptors, TNFR, for instance), which then emit signals through adaptor proteins to activate the IkB kinase (IKK) complex (composed of IKK-α, the catalytic subunit IKK-β, and the regulatory subunit NEMO). Once activated, the IKK complex catalyzes the polyubiquitination and the phosphorylation of the IkBs leading to their degradation by proteasome 26S. After this, the NF-κB dimers that are released from their inhibitors and translocate to the nucleus, where they activate the transcription of many genes through binding to the consensus sequence 5′-GGGRN W YYCC-3′ (R = purine base, N = any base, W = adenine or thymine, and Y = pyrimidine base). Alternatively, the non-canonical signaling cascade begins with the activation of LT-β or BAFF receptors, which emit signals through the cytoplasm leading to the phosphorylation of the NIK protein, which, in turn, phosphorylates the IKK complex (formed by only two IKK-α subunits). Once activated, this complex phosphorylates NF-κB2/p100, in two C-terminal sites, leading to its proteolytic cleavage. This process partially degrades p100 into p52. Subsequently, the NF-κB dimer is released and translocates to the nucleus, activating gene transcription. (B) In cancer cells, mutations, gene amplification and gene fusions involving its subunits, or interplay with other dysregulated signaling pathways may lead to the constitutive activation of NF-κB which contributes to cancer development and progression by increasing the expression of genes associated with antiapoptosis, survival, chemo- and radioresistance, adherence, invasion, and metastasis. This figure was created using Servier Medical Art templates, which are licensed under a Creative Commons Attribution 3.0 Unported License; https://smart.servier.com accessed on 10 April 2024.
Pharmaceuticals 17 00734 g002
Pharmaceuticals 17 00734 g003
Figure 3. Alterations in major signaling pathways that have been identified in OS development and metastasis: PI3K/AKT/mTOR, JAK/STAT, Wnt/β-catenin, NOTCH, Hedgehog/Gli, TGF-β, MAPK, and receptor tyrosine kinases (RTKs). Despite different players and complexities, NF-κB stands out as a common downstream effector, coupling the variety of molecular cascades underneath the characteristic molecular heterogeneity of this tumor. This figure was created using Servier Medical Art templates, which are licensed under a Creative Commons Attribution 3.0 Unported License; https://smart.servier.com accessed on 11 April 2024.
Figure 3. Alterations in major signaling pathways that have been identified in OS development and metastasis: PI3K/AKT/mTOR, JAK/STAT, Wnt/β-catenin, NOTCH, Hedgehog/Gli, TGF-β, MAPK, and receptor tyrosine kinases (RTKs). Despite different players and complexities, NF-κB stands out as a common downstream effector, coupling the variety of molecular cascades underneath the characteristic molecular heterogeneity of this tumor. This figure was created using Servier Medical Art templates, which are licensed under a Creative Commons Attribution 3.0 Unported License; https://smart.servier.com accessed on 11 April 2024.
Pharmaceuticals 17 00734 g003
Pharmaceuticals 17 00734 g004
Figure 4. Pathways crosstalk illustration through in silico protein–protein interactions (PPI). Lists of proteins belonging to each signaling pathway were obtained though the GESEA—Gene Set Enrichment Analysis—available at https://www.gsea-msigdb.org accessed on 13 April 2024. Then, PPI between each pathway and NF-κB subunits were assessed through STRING v12 (available at https://string-db.org/ accessed on 14 April 2024). The parameters evaluated were co-occurrence, experiments, and databases, and the minimum required interaction score was 0.700, considered high. Chord plots to show individual interactions with RELA/p65, RELB, REL/c-REL, NFKB1/p52, and NFKB2/p50 were generated through the SRplot online platform for data analysis and visualization (available at http://www.bioinformatics.com.cn/srplot accessed on 14 April 2024), and the percentage of proteins involved in PPI are expressed in pie charts. As an example, there are 233 proteins involved in the WNT pathway; from these, 82 (35.2%) interact with NF-κB subunits distributed as follows: 30 proteins with RELA/p65, 8 with RELB, 10 with REL/c-REL, and 28 with NFKB2/p50. Similar patterns are seen for other pathways which, as expected, show more interactions with RELA/p65 and NFKB1/p52.
Figure 4. Pathways crosstalk illustration through in silico protein–protein interactions (PPI). Lists of proteins belonging to each signaling pathway were obtained though the GESEA—Gene Set Enrichment Analysis—available at https://www.gsea-msigdb.org accessed on 13 April 2024. Then, PPI between each pathway and NF-κB subunits were assessed through STRING v12 (available at https://string-db.org/ accessed on 14 April 2024). The parameters evaluated were co-occurrence, experiments, and databases, and the minimum required interaction score was 0.700, considered high. Chord plots to show individual interactions with RELA/p65, RELB, REL/c-REL, NFKB1/p52, and NFKB2/p50 were generated through the SRplot online platform for data analysis and visualization (available at http://www.bioinformatics.com.cn/srplot accessed on 14 April 2024), and the percentage of proteins involved in PPI are expressed in pie charts. As an example, there are 233 proteins involved in the WNT pathway; from these, 82 (35.2%) interact with NF-κB subunits distributed as follows: 30 proteins with RELA/p65, 8 with RELB, 10 with REL/c-REL, and 28 with NFKB2/p50. Similar patterns are seen for other pathways which, as expected, show more interactions with RELA/p65 and NFKB1/p52.
Pharmaceuticals 17 00734 g004
Pharmaceuticals 17 00734 g005
Figure 5. (A) Gene expression of RELA, RELB, REL, NFKB1, and NFKB2 was assessed in R2 Genomics Analysis and Visualization Platform (http://r2.amc.nl accessed on 16 April 2024). Three databases were selected containing 4 control osteoblasts samples—white dots—and 61 OS samples—gray dots—(Mixed Osteosarcoma-Aqeilan-18-MAS5.0-u133p2, Mixed Osteosarcoma-Guenther-20-MAS5.0-u133a, and Tumor Osteosarcoma-Kobayashi-27-MAS5.0-u133p2; Probes: RELA 209878_s_at; RELB 205205_at; REL 235242_at; NFKB1 209239_at; NFKB2 209636_at) (ns = not significant). (B) Gene expression of NF-κB subunits according to the presence (gray violin) or absence (white violin) of metastasis at diagnosis using data from Mixed Osteosarcoma (Mesenchymal)-Kuijjer-127-vst-ilmnhwg6v2 also present at R2; (C) Kaplan–Meier curve of distant metastasis-free survival in months according to NFKB1 expression (low levels in blue, high levels in red)(generated at the R2 platform from Kuijjer’s dataset); and (D) Spearman correlation between RELA and NFKB1 expression in OS samples (data obtained from Guenther’s dataset). All data are expressed as log2. * = p < 0.05.
Figure 5. (A) Gene expression of RELA, RELB, REL, NFKB1, and NFKB2 was assessed in R2 Genomics Analysis and Visualization Platform (http://r2.amc.nl accessed on 16 April 2024). Three databases were selected containing 4 control osteoblasts samples—white dots—and 61 OS samples—gray dots—(Mixed Osteosarcoma-Aqeilan-18-MAS5.0-u133p2, Mixed Osteosarcoma-Guenther-20-MAS5.0-u133a, and Tumor Osteosarcoma-Kobayashi-27-MAS5.0-u133p2; Probes: RELA 209878_s_at; RELB 205205_at; REL 235242_at; NFKB1 209239_at; NFKB2 209636_at) (ns = not significant). (B) Gene expression of NF-κB subunits according to the presence (gray violin) or absence (white violin) of metastasis at diagnosis using data from Mixed Osteosarcoma (Mesenchymal)-Kuijjer-127-vst-ilmnhwg6v2 also present at R2; (C) Kaplan–Meier curve of distant metastasis-free survival in months according to NFKB1 expression (low levels in blue, high levels in red)(generated at the R2 platform from Kuijjer’s dataset); and (D) Spearman correlation between RELA and NFKB1 expression in OS samples (data obtained from Guenther’s dataset). All data are expressed as log2. * = p < 0.05.
Pharmaceuticals 17 00734 g005
Pharmaceuticals 17 00734 g006
Figure 6. (A) Spearman correlation values of mRNA levels between NF-κB and pathway-associated genes depicted as a circular heatmap generated through the SRplot online platform for data analysis and visualization (available at http://www.bioinformatics.com.cn/srplot accessed on 18 April 2024). (B) TFLink gateway (available at https://tflink.net/ accessed on 18 April 2024) was used to investigate transcription factor–target gene interactions. A total of 61 out of 68 genes are indeed NF-κB predicted targets, of which, 5 are regulated by all NF-κB subunits and are among those with higher correlations. (C) Differential gene expression analysis showed 12 downregulated genes and 19 upregulated genes in OS samples and controls (Vulcano Plot generated using the Mixed Osteosarcoma (Mesenchymal)-Kuijjer-127-vst-ilmnhwg6v2 dataset). KEGG enrichment analysis through the STRING platform showed that downregulated genes belong mainly to MAPK and WNT signaling pathways, while upregulated genes are from the PI3K/AKT and JAK/STAT cascades.
Figure 6. (A) Spearman correlation values of mRNA levels between NF-κB and pathway-associated genes depicted as a circular heatmap generated through the SRplot online platform for data analysis and visualization (available at http://www.bioinformatics.com.cn/srplot accessed on 18 April 2024). (B) TFLink gateway (available at https://tflink.net/ accessed on 18 April 2024) was used to investigate transcription factor–target gene interactions. A total of 61 out of 68 genes are indeed NF-κB predicted targets, of which, 5 are regulated by all NF-κB subunits and are among those with higher correlations. (C) Differential gene expression analysis showed 12 downregulated genes and 19 upregulated genes in OS samples and controls (Vulcano Plot generated using the Mixed Osteosarcoma (Mesenchymal)-Kuijjer-127-vst-ilmnhwg6v2 dataset). KEGG enrichment analysis through the STRING platform showed that downregulated genes belong mainly to MAPK and WNT signaling pathways, while upregulated genes are from the PI3K/AKT and JAK/STAT cascades.
Pharmaceuticals 17 00734 g006
Pharmaceuticals 17 00734 g007
Figure 7. (A) Gene dependency on NF-κB subunits was assessed through the DepMap platform (https://depmap.org/portal/ accessed on 21 April 2024), based on CRISPR and RNAi knockout experiments on pediatric OS cell lines. Score greater than zero (>0) indicates that the cell line is not dependent, less than zero (<0) indicates that the cell line is dependent, and the closer to −1 indicates that the gene is essential for the survival of the cell line. (B) Schematic illustrations of NF-κB druggability identified with the CanSAR database, including the total number of compounds with predicted interaction capacity with each NF-κB subunit, binding efficiency, and the lack of proven clinical application. (C) Interaction networks of NF-κB inhibitors and associated binding proteins according to STITCH (available at http://stitch.embl.de accessed on 21 April 2024). From all the cited compounds with alleged activity against NF-κB, only DHMEQ and BAY 11-7085 show direct action on this transcription factor. Compounds are represented as pill-shaped nodes, while proteins are shown as spheres (smaller nodes represent proteins of unknown 3D structures). Nodes that are associated to each other are linked by an edge: thicker lines represent stronger binding affinities. Networks were constructed considering a minimum required interaction score of 0.700 and were based on associations reported in curated databases (gray lines) or on both databases and experimental/biochemical data (green lines).
Figure 7. (A) Gene dependency on NF-κB subunits was assessed through the DepMap platform (https://depmap.org/portal/ accessed on 21 April 2024), based on CRISPR and RNAi knockout experiments on pediatric OS cell lines. Score greater than zero (>0) indicates that the cell line is not dependent, less than zero (<0) indicates that the cell line is dependent, and the closer to −1 indicates that the gene is essential for the survival of the cell line. (B) Schematic illustrations of NF-κB druggability identified with the CanSAR database, including the total number of compounds with predicted interaction capacity with each NF-κB subunit, binding efficiency, and the lack of proven clinical application. (C) Interaction networks of NF-κB inhibitors and associated binding proteins according to STITCH (available at http://stitch.embl.de accessed on 21 April 2024). From all the cited compounds with alleged activity against NF-κB, only DHMEQ and BAY 11-7085 show direct action on this transcription factor. Compounds are represented as pill-shaped nodes, while proteins are shown as spheres (smaller nodes represent proteins of unknown 3D structures). Nodes that are associated to each other are linked by an edge: thicker lines represent stronger binding affinities. Networks were constructed considering a minimum required interaction score of 0.700 and were based on associations reported in curated databases (gray lines) or on both databases and experimental/biochemical data (green lines).
Pharmaceuticals 17 00734 g007
Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content.

Share and Cite

MDPI and ACS Style

Medeiros, M.; Guenka, S.; Bastos, D.; Oliveira, K.L.; Brassesco, M.S. Amicis Omnia Sunt Communia: NF-κB Inhibition as an Alternative to Overcome Osteosarcoma Heterogeneity. Pharmaceuticals 2024, 17, 734. https://doi.org/10.3390/ph17060734

AMA Style

Medeiros M, Guenka S, Bastos D, Oliveira KL, Brassesco MS. Amicis Omnia Sunt Communia: NF-κB Inhibition as an Alternative to Overcome Osteosarcoma Heterogeneity. Pharmaceuticals. 2024; 17(6):734. https://doi.org/10.3390/ph17060734

Chicago/Turabian Style

Medeiros, Mariana, Sophia Guenka, David Bastos, Karla Laissa Oliveira, and María Sol Brassesco. 2024. "Amicis Omnia Sunt Communia: NF-κB Inhibition as an Alternative to Overcome Osteosarcoma Heterogeneity" Pharmaceuticals 17, no. 6: 734. https://doi.org/10.3390/ph17060734

APA Style

Medeiros, M., Guenka, S., Bastos, D., Oliveira, K. L., & Brassesco, M. S. (2024). Amicis Omnia Sunt Communia: NF-κB Inhibition as an Alternative to Overcome Osteosarcoma Heterogeneity. Pharmaceuticals, 17(6), 734. https://doi.org/10.3390/ph17060734

Note that from the first issue of 2016, this journal uses article numbers instead of page numbers. See further details here.

Article Metrics

Back to TopTop