Search Results (300)

Glioblastoma (GBM) remains one of the most aggressive and treatment-resistant brain tumors, largely due to its profound intratumoral heterogeneity and immunosuppressive microenvironment. Various classifications of GBM subtypes were created based on transcriptional and methylation profiles. This effort, followed by the development of new technology such as single-nuclei sequencing (snRNAseq) and spatial transcriptomics, led to a better understanding of the glioma cells’ plasticity and their ability to transition between diverse cellular states. GBM cells can mimic neurodevelopmental programs to resemble oligodendrocyte or neural progenitor behavior and hitchhike the local neuronal network to support their growth. The tumor microenvironment, especially under hypoxic conditions, drives the tumor cell clonal selection, which then reshapes the immune cells’ functions. These adaptations contribute to immune evasion by progressively disabling T cell and myeloid cell functions, ultimately establishing a highly immunosuppressive tumor milieu. This complex and metabolically constrained environment poses a major barrier to effective antitumor immunity and limits the success of conventional therapies. Understanding the dynamic interactions between glioma cells and their microenvironment is essential for the development of more effective immunotherapies and rational combination strategies aimed at overcoming resistance and improving patient outcomes. Full article

Background/Objectives: Myelodysplastic syndrome (MDS) is a heterogeneous clonal hematopoietic disorder characterized by ineffective hematopoiesis and leukemic transformation risk. Current therapies show limited efficacy, with ~50% of patients failing hypomethylating agents. This review aims to synthesize recent discoveries through an integrated network model and examine translation into precision therapeutic approaches. Methods: We reviewed breakthrough discoveries from the past three years, analyzing single-cell multi-omics technologies, epitranscriptomics, stem cell architecture analysis, and precision medicine approaches. We examined cell-type-specific splicing aberrations, distinct stem cell architectures, epitranscriptomic modifications, and microenvironmental alterations in MDS pathogenesis. Results: Four interconnected mechanisms drive MDS: genetic alterations (splicing factor mutations), aberrant stem cell architecture (CMP-pattern vs. GMP-pattern), epitranscriptomic dysregulation involving pseudouridine-modified tRNA-derived fragments, and microenvironmental changes. Splicing aberrations show cell-type specificity, with SF3B1 mutations preferentially affecting erythroid lineages. Stem cell architectures predict therapeutic responses, with CMP-pattern MDS achieving superior venetoclax response rates (>70%) versus GMP-pattern MDS (<30%). Epitranscriptomic alterations provide independent prognostic information, while microenvironmental changes mediate treatment resistance. Conclusions: These advances represent a paradigm shift toward personalized MDS medicine, moving from single-biomarker to comprehensive molecular profiling guiding multi-target strategies. While challenges remain in standardizing molecular profiling and developing clinical decision algorithms, this systems-level understanding provides a foundation for precision oncology implementation and overcoming current therapeutic limitations. Full article

Conventional methods for generating knock-out or knock-in mammalian cell models using CRISPR-Cas9 genome editing often require tedious single-cell clone selection and expansion. In this study, we develop and optimise rapid and robust strategies to engineer homozygous fluorescent reporter knock-in cell pools with precise genome editing, circumventing clonal variability inherent to traditional approaches. To reduce false-positive cells associated with random integration, we optimise the design of donor DNA by removing the start codon of the fluorescent reporter and incorporating a self-cleaving T2A peptide system. Using fluorescence-assisted cell sorting (FACS), we efficiently identify and isolate the desired homozygous fluorescent knock-in clones, establishing stable cell pools that preserve parental cell line heterogeneity and faithfully reflect endogenous transcriptional regulation of the target gene. We evaluate the knock-in efficiency and rate of undesired random integration in the electroporation method with either a dual-plasmid system (sgRNA and donor DNA in two separate vectors) or a single-plasmid system (sgRNA and donor DNA combined in one vector). We further demonstrate that coupling our single-plasmid construct with an integrase-deficient lentivirus vector (IDLV) packaging system efficiently generates fluorescent knock-in reporter cell pools, offering flexibility between electroporation and lentivirus transduction methods. Notably, compared to the electroporation methods, the IDLV system significantly minimises random integration. Moreover, the resulting reporter cell lines are compatible with most of the available genome-wide sgRNA libraries, enabling unbiased CRISPR screens to identify key transcriptional regulators of a gene of interest. Overall, our methodologies provide a powerful genetic tool for rapid and robust generation of fluorescent reporter knock-in cell pools with precise genome editing by CRISPR-Cas9 for various research purposes. Full article

Light-chain (AL) amyloidosis is a rare clonal plasma cell disorder that, if left untreated, carries a high risk of organ damage and mortality. Due to the rarity of the disease and the vulnerability of affected organ systems, treatment requires significant caution and nuance. As a plasma cell dyscrasia, AL amyloidosis treatment regimens are often adapted from those used for related disorders, particularly multiple myeloma. Despite substantial progress in research and drug development, optimal treatment strategies for relapsed/refractory (RR) AL amyloidosis remain unclear, and no FDA-approved therapies currently exist for this setting. B-cell maturation antigen (BCMA) has emerged as a promising immunotherapy target, with associated drug classes including antibody–drug conjugates, bispecific antibodies, and CAR-T cell therapies. These therapies have been extensively studied in relapsed/refractory multiple myeloma (RRMM) and are now being explored in the context of RR AL amyloidosis. This review summarizes the current literature on the efficacy and tolerability of BCMA-directed therapies in AL amyloidosis, with a particular emphasis on CAR-T cell therapy and offers comparisons to outcomes observed in RRMM. Full article

Lifileucel, a tumor-infiltrating lymphocyte (TIL) cell therapy approved for advanced melanoma, demonstrates promise for treating other solid tumors, including endometrial cancer (EC). The current study evaluates the feasibility of manufacturing TILs from EC tumors using Iovance’s proprietary 22-day Gen2 manufacturing process. Key parameters, including TIL yield, viability, immune phenotype, T-cell receptor clonality, and cytotoxic activity, were assessed. Of the 11 EC tumor samples processed at research scale, 10 (91%) successfully generated >1 × 10⁹ viable TIL cells, with a median yield of 1.1 × 10¹⁰ cells and a median viability of 82.8%. Of the four EC tumor samples processed at full scale, all achieved the pre-specified TVC and viability targets. Putative tumor-reactive T-cell clones were maintained throughout the manufacturing process. Functional reactivity was evidenced by the upregulation of 4-1BB in CD8+ T cells, OX40 in CD4+ T cells, and increased production of IFN-γ and TNF-α upon autologous tumor stimulation. Furthermore, antitumor activity was confirmed using an in vitro autologous tumor organoid killing assay. These findings demonstrate the feasibility of ex vivo TIL expansion from EC tumors. This study provides a rationale for the initiation of the phase II clinical trial IOV-END-201 (NCT06481592) to evaluate lifileucel in patients with advanced EC. Full article

Background: Mycosis fungoides (MF), the most prevalent cutaneous T cell lymphoma, features clonal CD4⁺ T cell proliferation within a Th2-dominant microenvironment. Interleukin-4 (IL-4) promotes disease progression while Brentuximab Vedotin (BV), an anti-CD30 antibody–drug conjugate, shows efficacy but faces resistance challenges. Methods: We conducted a narrative literature review (2010–2024) synthesizing evidence on IL-4 signaling and BV’s efficacy in MF to develop a theoretical framework for combination therapy. Results: IL-4 may modulate CD30 expression and compromise BV’s effectiveness through immunosuppressive microenvironment remodeling. Theoretical mechanisms suggest that IL-4 pathway inhibition could reprogram the microenvironment toward Th1 dominance and restore BV sensitivity. However, no direct experimental evidence validates this combination, and safety concerns including potential disease acceleration require careful evaluation. Conclusions: The proposed IL-4/BV combination represents a biologically compelling but unproven hypothesis requiring systematic preclinical validation and biomarker-driven clinical trials. This framework could guide future research toward transforming treatment approaches for CD30-positive MF by targeting both malignant cells and their immunologically permissive microenvironment. Full article

Mitochondria are critical for cellular energy, and while large deletions in their genome (mtDNA) are linked to primary mitochondrial diseases, their significance in cancer is less understood. Given cancer’s metabolic nature, investigating mtDNA deletions in tumors at various stages could provide insights into disease origins and treatment responses. In this study, we analyzed 148 bone marrow samples from 129 pediatric patients with B-cell (B-ALL) and T-cell (T-ALL) acute lymphoblastic leukemia at diagnosis, remission, and relapse using long-range PCR, next-generation sequencing, and the Splice-Break2 pipeline. Both T-ALL and B-ALL exhibited significantly more mtDNA deletions than did the controls, with T-ALL showing a ~100-fold increase and B-ALL a ~15-fold increase. The T-ALL samples also exhibited larger deletions (median size > 2000 bp) and greater heterogeneity, suggesting increased mitochondrial instability. Clustering analysis revealed distinct deletion profiles between ALL subtypes and across disease stages. Notably, large clonal deletions were detected in some B-ALL remission samples, including one affecting up to 88% of mtDNA molecules, which points toward treatment-driven selection or toxicity. A multivariate analysis confirmed that disease type, timepoint, and WHO subtype significantly influenced mtDNA deletion metrics, while age and gender did not. These findings suggest that mtDNA deletion profiling could serve as a biomarker for pediatric ALL and may indicate mitochondrial toxicity contributing to late effects in survivors. Full article

The immune landscape of tumor-draining lymph nodes (TDLNs) plays a critical role in shaping antitumor responses and influencing prognosis in oral squamous cell carcinoma (OSCC). Among patients with lymph node (LN) metastasis, clinical outcomes vary widely, yet reliable biomarkers for prognostic stratification remain limited. This study aimed to identify immune features in tumors and LNs that differentiate between favorable and poor outcomes in OSCC patients with nodal metastasis. We analyzed T cell receptor (TCR) CDR3 repertoires and the expression of immune-related genes in primary tumors and paired sentinel LNs from OSCC patients who underwent tumor resection and lymphadenectomy. Patients were divided into three groups: Group A (no nodal metastasis), Group B1 (metastasis without recurrence), and Group B2 (metastasis with recurrence). TCR diversity was assessed using the Shannon index. The expression of immune-related genes (e.g., CD3E, CD4, CD8B, FOXP3, CTLA4, IL2, IL4) was measured by quantitative PCR and normalized to GAPDH. TCR diversity was lower in tumors than in non-metastatic LNs, reflecting clonal expansion. Metastatic LNs exhibited tumor-like diversity, suggesting infiltration by tumor-reactive clones. Tumor gene expression did not differ across groups, but LNs from metastatic cases showed the reduced expression of several immune genes. Notably, CD3E, CD8B, CTLA4, IL2, and IL4 distinguished B1 from B2. The immune profiling of LNs offers superior prognostic value over tumor analysis in OSCC patients with LN metastasis. LN-based evaluation may aid in postoperative risk stratification and personalized postoperative management and could inform decisions regarding adjuvant therapy and follow-up strategies. Full article

In B-cell chronic lymphocytic leukemia (B-CLL), hypodiploidy is a rare but aggressive subtype of the disease with a very bad prognosis. Hypodiploidy, in contrast to normal B-CLL chromosomal aberrations, is marked by widespread genomic instability, which promotes treatment resistance and quick illness development. Its persistence after treatment implies that chromosomal loss gives cancerous clones a selection edge, which is made worse by telomere malfunction and epigenetic changes. Since thorough genetic profiling has a major impact on patient outcomes, advanced diagnostic methods are crucial for early detection. Treatment approaches must advance beyond accepted practices because of its resistance to traditional medicines. Hematopoietic stem cell transplantation (HSCT) and chimeric antigen receptor (CAR) T-cell therapy are two potential new therapeutic modalities. Relapse and treatment-related morbidity continue to be limiting concerns, despite the noteworthy improvements in outcomes in high-risk CLL patients receiving HSCT. Although more research is required, CAR T-cell treatment is effective in treating recurrent B-ALL and may also be used to treat B-CLL with hypodiploidy. Novel approaches are essential for enhancing patient outcomes and redefining therapeutic success when hypodiploidy challenges established treatment paradigms. Hypodiploidy is an uncommon yet aggressive form of B-CLL that has a very bad prognosis. Hypodiploidy represents significant chromosomal loss and structural imbalance, which contributes to a disordered genomic environment, in contrast to more prevalent cytogenetic changes. This instability promotes resistance to certain new drugs as well as chemoimmunotherapy and speeds up clonal evolution. Its persistence after treatment implies that hypodiploid clones have benefits in survival, which are probably strengthened by chromosomal segregation issues and damaged DNA repair pathways. Malignant progression and treatment failure are further exacerbated by telomere erosion and epigenetic dysregulation. The need for more sensitive molecular diagnostics is highlighted by the fact that standard karyotyping frequently overlooks hypodiploid clones, particularly those concealed by endoreduplication, despite the fact that these complications make early and correct diagnosis crucial. Hypodiploidy requires a move toward individualized treatment because of their link to high-risk genetic traits and resistance to conventional regimens. Although treatments like hematopoietic stem cell transplantation and CAR T-cells show promise, long-term management is still elusive. To improve long-term results and avoid early relapse, addressing this cytogenetic population necessitates combining high-resolution genomic technologies with changing therapy approaches. Full article

Viral evasion from effective human immunodeficiency virus type 1 (HIV-1)-specific CD8+ T-cell responses and from antiretroviral therapy through viral sequence variation is frequently accompanied by a loss in viral fitness. The impact of sequence variations on replication capacity in vitro was mostly studied by introducing single mutations into a specific clonal strain such as NL4-3. How the specific viral backbone itself impacts replicative fitness remains elusive. To test for a potential effect of the viral backbone, we constructed HIV-1 clade B clones with consensus sequences for gag and/or pol and evaluated the infectivity of viral variants harboring well-defined cytotoxic T-lymphocyte (CTL) escape mutations or drug resistance mutations within this backbone or the clonal NL4-3 strain. Viral variants with consensus sequences were replication-competent in vitro, although at lower rates than the NL4-3 virus. Introduction of the dominant CTL escape mutation R₂₆₄K into the newly constructed viruses or into NL4-3 led to a dramatic reduction in infection rates. In contrast to the NL4-3 backbone, the combination of R₂₆₄K with its compensatory mutation S₁₇₃A on the consensus backbone led to higher infection rates as compared to the same virus in the absence of R₂₆₄K and S₁₇₃A. Furthermore, 2 out of 10 drug resistance mutations in pol led to opposing effects, with an increase in infection rates on the consensus gag/pol backbone and a reduction on NL4-3. Therefore, the effect of the respective viral backbone on infectivity observed in vitro might constitute an additional factor to explain differential kinetics of mutational evasion from immune and pharmaceutical pressure. Full article

Background/Objectives: To reduce the early mortality of light-chain amyloidosis (AL), earlier diagnosis is needed. To pursue this goal, we conducted a multicenter study screening for AL λ-type (NCT04615572) in subjects > 60 years of age with λ smoldering myeloma (SMM) or monoclonal gammopathy of undetermined significance (MGUS), a light-chain differential (dFLC, λ minus κ) > 23 mg/L, and no prior amyloid diagnosis. Methods: Variables included AL-related IGVL gene usage and clonal plasma cell cytogenetic abnormalities, such as t(11;14) or gain 1q, which are present in 75% of AL cases. Here, 9 out of 33 λ IGVL genes, accounting for 90% of AL λ cases, were considered to be AL-related. Bone marrow was obtained, plasma cell cytogenetics and next generation sequencing for IGVL genes were performed, and subjects with AL-related IGVL genes were screened for AL using tissue studies. Results: From 2021 to 2023, we enrolled 30 subjects (19 M/11 F) with a median age of 68.5 years old (IQR 64.3–73), 17 SMM and 13 MGUS, with a median of 6% marrow plasma cells (range, 3.5–40). Here, 11 SMM and 4 MGUS cases had t(11;14) or gain 1q; 10/17 SMM and 12/13 MGUS had AL-related genes, and AL was ultimately confirmed by tissue biopsy in 3 with SMM. SMM, AL-related IGVL genes, and t(11;14) or gain 1q were found in 6 SMM subjects, including the 3 with AL (3/6 vs. 0/16; p < 0.05, Fisher’s exact, two-tailed). Conclusions: These results justify a larger study screening for AL in SMM to develop a likelihood algorithm for AL using dFLC, IGVL gene usage, and the presence of t(11;14) or gain 1q. Full article

Understanding the mechanisms of immune activation and deactivation is paramount. A host must initiate effective immunity against pathogenic infections while avoiding triggering immunity against self-antigens, which can lead to detrimental autoimmune disorders. Host immunological pathways can be categorized as Immunoglobulin (Ig)G-dominant eradicable immune reactions and IgA-dominant tolerable immune reactions. Eradicable immune reactions include Th1, Th2, Th22, and Thαβ immune responses against four different types of pathogens. Tolerable immune reactions include Th1-like, Th9, Th17, and Th3 immune responses against four different types of pathogens. Here, we try to determine the mechanisms of activation and deactivation of host immune reactions. The spleen and liver play contrasting roles in mediating immune responses: the spleen is primarily involved in immune activation, whereas the liver is responsible for immune deactivation. Similarly, the sympathetic and parasympathetic nervous systems have opposing functions in immune modulation, with the sympathetic system promoting pro-inflammatory responses and the parasympathetic system facilitating anti-inflammatory processes. Furthermore, adrenocorticotropic hormone (ACTH) and glucocorticosteroids exhibit contrasting effects on immune regulation: ACTH is involved in activating adaptive immunity while inhibiting innate immunity, whereas glucocorticosteroids activate natural IgM antibody associated with innate immunity while inhibiting adaptive immunity. Heat shock proteins, particularly molecular chaperones induced by fever, play pivotal roles in immune activation. Conversely, IgD B cells and gamma/delta T cells contribute to immune deactivation through mechanisms such as clonal anergy. Understanding these mechanisms provides insights into immunological pathways, aiding in the better management of infectious diseases and autoimmune disorders. Full article

Background: SARS-CoV-2 was the causing agent of the COVID-19 pandemic, which resulted in millions of deaths worldwide and massive economic losses. Although there are already several vaccines licensed, as novel variants develop, understanding the immune response induced by vaccination and natural infection is key for the development of future vaccines. Methods: In this study, we have used flow cytometry and next-generation sequencing to assess the longitudinal CD8⁺ T-cell response against natural infection and vaccination in convalescent and vaccinated individuals, from early activation to immune memory establishment. Moreover, we have characterized the T-cell receptor clonality and diversity at different stages post-infection and post-vaccination. Results: We have found no significant differences in CD8⁺ T-cell activation during the first three weeks post-infection compared to the first three weeks after first vaccination. Conversely, natural infection resulted in sustained high levels of T-cell activation at four weeks post-infection, a point in which we observed a decline in T-cell activation post-vaccination despite boosting with a second vaccination shot. Moreover, additional vaccination did not result in enhanced T-cell activation. Of note, we have observed variations in the memory subset structure at every stage of disease and vaccination. Overall, both infection and immunization induced a highly diverse T-cell receptor repertoire, which was observed both between study groups and between patients inside a given group. Conclusions: These data contribute to expand our knowledge about the immune response to SARS-CoV-2 infection and vaccination and call for additional strategies to enhance T-cell responses by booster immunization. Full article

It is unknown whether intestinal ultrasonographic measurements differ between lymphoplasmacytic enteritis (LPE) and low-grade intestinal T-cell lymphoma (LGITL) in cats if the diagnosis is based either on histology/immunohistochemistry (IHC) or on clonality assay results. The effects of treatment on intestinal ultrasonographic measurements are also unknown. Therefore, we prospectively compared small intestinal wall layering between cats with LPE and LGITL and investigated whether there were differences between the groups when the diagnostic gold standard was either histology/IHC or clonality testing. We evaluated the effects of standardized treatment in a subset of cats. The thicknesses of the total wall, mucosa, muscularis, and submucosa were measured in the duodenum, jejunum, and ileum, and ratios (muscularis to submucosa, muscularis to total wall thickness) were calculated. The thickness of the largest mesenteric lymph nodes was also determined. Ultrasonographic measurements from duodenal and jejunal segments were grouped together, and ileal segments were assessed separately. Sixteen cats with standardized full-thickness biopsies from the stomach, duodenum, jejunum, and ileum were included. Samples for clonality testing were fresh-frozen and analyzed later, and the standardized treatment was based on histologic/IHC diagnoses. Ultrasonographic measurements were compared between LPE and LGITL when diagnoses were either based on histology/IHC or clonality testing using a linear mixed model. Repeated ultrasonographic measurements of segments were available for seven cats after 12 weeks (five LPE, two LGITL) and five cats after 24 weeks (three LPE, two LGITL) of standardized treatment. We found that none of the ultrasonographic measurements differed between LPE and LGITL regardless of the diagnostic gold standard used. During treatment, only the ratio of lamina muscularis thickness to total wall thickness decreased significantly in LPE cats after 12 and 24 weeks compared to baseline. In conclusion, the herein evaluated ultrasonographic variables did not differ between LPE and LGITL and the diagnostic gold standard used had no influence on the results. The detected change over time during treatment in LPE cats requires further study. Full article

Non-Hodgkin lymphomas (NHLs) encompass a diverse group of neoplasms arising from the clonal proliferation of B-cell progenitors, T-cell progenitors, mature B-cells, mature T-cells, and natural killer (NK) cells. These malignancies account for over 90% of lymphoid neoplasms. The link between the gut microbiome and neoplasms has been extensively studied in recent years. Growing evidence suggests that the gut microbiome may be involved not only in the development of the disease, but also in modulating the efficacy of implemented therapies. In this review, we summarize the current knowledge on the potential involvement of the gut microbiome in the development of diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL), mucosa-associated lymphoid tissue (MALT) lymphoma, and NK/T-cell lymphoma, including cutaneous T-cell lymphoma (CTCL). Moreover, we discuss the relationship between gut microbiome changes before and after treatment and their association with treatment outcomes, focusing on chemotherapy and CAR T-cell therapy. Full article