Search Results (61)

Histopathological grading of oral squamous cell carcinoma is currently based on differentiation of cells, while additional histological parameters, such as the tumor–stroma ratio (TSR), tumor budding (TB), or the combined TSR/tumor budding model could better assess tumor biological behavior and monitoring of patients. Background/Objectives: To integrate risk factors associated with tumor progression: the TSR, TB and TSR/tumor budding model, whose prognostic significance in oral cancer has not yet been evaluated. Methods: An observational cohort retrospective study assembled according to STROBE guidelines on histological materials from 196 patients with invasive squamous cell carcinoma of the oral cavity. The goal of the analysis was to evaluate the association between the tumor stroma ratio, tumor budding, and the combined model of TSR/TB with the clinical and pathologic features of patients with squamous cell carcinoma of the oral cavity and to determine the prognostic value of this model in relation to disease-free survival (DFS) Results: The analysis did not show that the tumor stroma ratio (TSR), tumor budding, and the combined model of TSR/tumor budding were statistically significantly associated with the occurrence of metastatic disease at the start of treatment or during postoperative follow-up, but confirmed the value of depth-of-invasion (DOI) as a negative prognostic factor (HR 15.3, p < 0.001). Conclusions: The TSR, TB, and the combined TSR/TB model were not found to be statistically significant predictors for the disease progression in the Cox regression survival analysis but were found to have a significant correlation with known negative prognostic factors: DOI, neural invasion, and T category. Full article

Background/Purpose: Conventional three-dimensional in vitro tumor models often fail to fully capture the complexity of the tumor microenvironment, particularly the diverse populations of cancer-associated fibroblasts that contribute to poor prognosis and treatment resistance. The purpose of this study is to develop a patient-specific gastric cancer assembloid model that integrates tumor epithelial cells with matched stromal cell subtypes, each derived using tailored growth media to enhance cancer preclinical research and advance personalized therapeutic strategies. Methods: Tumor tissue was dissociated, and cells expanded in media for organoids, mesenchymal stem cells, fibroblasts, or endothelial cells. The resulting tumor-derived subpopulations were co-cultured in an optimized assembloid medium supporting each cell type’s growth. Biomarker expression was assessed by immunofluorescence staining, and transcriptomic profiles were analyzed by RNA sequencing. Drug responsiveness was evaluated using cell viability assays following treatment with various therapeutic agents. Results: The optimized co-culture conditions yielded assembloids that closely mimicked the cellular heterogeneity of primary tumors, confirmed by the expression of epithelial and stromal markers. Compared to monocultures, the assembloids showed higher expression of inflammatory cytokines, extracellular matrix remodeling factors, and tumor progression-related genes across different organoids and stromal ratios. Drug screening revealed patient- and drug-specific variability. While some drugs were effective in both organoid and assembloid models, others lost efficacy in the assembloids, highlighting the critical role of stromal components in modulating drug responses. Conclusions: This assembloid system offers a robust platform to study tumor–stroma interactions, identify resistance mechanisms, and accelerate drug discovery and personalized therapeutic strategies for gastric cancer. Full article

Background/Objectives: Three-dimensional (3D) cell culture models more accurately simulate the in vivo tissue environments as compared to conventional two-dimensional (2D) monolayer cultures. Among these, spheroid cultures are particularly valuable for pharmaceutical research, as they allow for the study of tumor growth, drug responses, and cell–cell interactions in a physiologically relevant manner. Superhydrophobic surfaces (SHSs) have shown a promise in enhancing spheroid formation by reducing cell–substrate adhesion and promoting cell–cell aggregation. This study aims to evaluate the effectiveness of two different SHS coatings (SHS1: fluorinated; SHS2: silicone-based) in generating co-culture spheroids composed of non-tumoral fibroblasts (3T3) and tumoral epidermoid carcinoma cells (A431), thereby mimicking aspects of the tumor microenvironment. Methods: Co-cultures of 3T3 and A431 cells were seeded at varying ratios onto SHS1 and SHS2 substrates to assess their ability to support 3D spheroid formation. Spheroids were characterized by measurements of circularity and size distribution, viability through live/dead staining, and surface topography using 3D profilometry. Results: Spheroid formation was significantly influenced by both the surface properties and the fibroblast-to-carcinoma cell ratio. The fluorinated SHS1 surface facilitated superior cell viability and promoted the formation of well-rounded, uniform spheroids. In contrast, the silicone-based SHS2 surface resulted in less defined spheroidal structures and lower overall viability. Profilometry confirmed more consistent and compact 3D architectures on SHS1. Conclusions: This study demonstrates that SHS1, a fluorinated superhydrophobic coating, is more effective than SHS2 in supporting the formation of viable and structurally coherent 3D co-culture spheroids. These findings underscore the potential of SHS1 as a low-cost, tunable platform for developing in vitro cancer models and advancing the study of tumor–stroma interactions. Full article

Background: Colorectal cancer (CRC) heterogeneity is strongly influenced by molecular subtypes and tumor stroma interactions. The meprin/A5/PTPmu (MAM) domain, a conserved structural motif in transmembrane proteins, remains undercharacterized in CRC pathogenesis. Methods: We analyzed RNA-seq data from TCGA-COAD to evaluate MAM domain gene expression. Immunohistochemistry and Western blotting were conducted to validate the results of the database analysis. Results: Bioinformatics analysis revealed that MAM domain-containing protein 2 (MAMDC2) was enriched in mesenchymal subtype 4 (CMS4) colorectal cancer (p < 0.001). IHC confirmed MAMDC2 overexpression in MSS colorectal cancer with a high tumor stroma ratio (TSR) and peritoneal metastatic lesions (p < 0.01). WB and real-time PCR analyses confirmed that MAMDC2 has a role in regulating epithelial–mesenchymal transition (EMT) development in CRC. Importantly, we identified that cancer cell-derived MAMDC2 promotes MYLK expression in cancer-associated fibroblasts (CAFs) through paracrine signaling. Conclusions: Our findings suggest MAMDC2 may function as a stromal-associated regulator in MSS colorectal cancer with a high tumor stromal ratio (TSR). Full article

Ovarian cancer is a lethal malignancy, with most patients initially responding to chemotherapy but frequently experiencing recurrence. Previous studies primarily examined tumor characteristics using limited genetic markers or bulk RNA sequencing. Here, we used spatial transcriptomics via the GeoMx^® platform, alongside multispectral immune cell immunofluorescence (IF), to identify biomarkers associated with disease progression following first-line treatment of high-grade serous carcinoma (HGSC). We identified several spatial biomarkers linked to non-recurrence, including elevated NKG7 expression in CD45+ immune cell regions (p = 0.0011) and higher TFPI2 and PIGR expression in tumor areas (p = 2.09 × 10⁻⁶), both associated with improved progression-free survival. Multispectral IF revealed significantly higher regulatory T cell (Treg) to CD8+ T cell ratios in the tumor nests and stroma of recurrent patients (p = 0.016, 0.048). Tregs were also found closer to cancer cells or macrophages than CD8+ T cells in recurrent tumors (p = 0.048), correlating with poor survival. Integrated analysis showed that immune cell density and immune pathway scores in the recurrent group positively correlated with cancer pathway scores, except for NF-κB. This comprehensive analysis revealed clues to interactions between different immune cells and identified biomarkers that may be useful for predicting recurrence of HGSC. Full article

Pancreatic cancer (PC), with pancreatic ductal adenocarcinoma (PDAC) comprising about 90% of all cases, is one of the most aggressive and lethal solid tumors. PDAC remains one of the most significant challenges of oncology to this day due to its inadequate response to conventional treatment, gradual rise in incidence since 2004, and poor five-year survival rates. As cancer cells are the primary adversary in this uneven fight, they remain the primary research target. Nevertheless, increasing attention is being paid to the tumor microenvironment (TME). The most crucial TME constellation components are immune cells, especially macrophages, stellate cells and lymphocytes, fibroblasts, bacterial and fungal microflora, and neuronal cells. Depending on the particular phenotype of these cells, the composition of the microenvironment, and the cell ratio, patients can experience different disease outcomes and varying vulnerability to treatment approaches. This study aims to present the current knowledge and review the most up-to-date scientific findings regarding the microenvironment of PC. It contains detailed information on the structure and cellular composition of the stroma, including its impact on disease development, metastasis, and response to treatment, as well as the therapeutic opportunities that arise from targeting this tissue. Full article

Background/Objectives: While cytoskeleton-associated protein 4 (CKAP4) has been associated with prognosis in various malignancies, its prognostic value for bladder cancer (BCa) remains unclear. The aim of this study was to evaluate CKAP4 expression in tumor cells and cancer-associated fibroblasts (CAFs) following radical cystectomy (RC) in patients with BCa. Methods: In this study, CKAP4 in tumor cells was defined as CKAP4-1, while CKAP4 expressed in CAFs was defined as CKAP4-2. CKAP4-2 expression was evaluated to explore its potential association with tumor aggressiveness and patient outcomes. CKAP4 expression in 86 RC specimens was assessed using immunohistochemistry. CKAP4-1 positivity was considered when ≥5% cytoplasmic staining of cancer cells, with at least moderate staining intensity, was observed. CKAP4-2 positivity was evaluated using a point scale (0–3), with scores based on the number of CKAP4 positive CAFs in the tumor stroma. Scores of 2 (moderate number of CAFs) and 3 (significant number of CAFs) were considered to indicate positivity. Results: CKAP4-1 and CKAP4-2 were expressed in 53 (61.6%) and 34 (39.5%) patients, respectively. Kaplan–Meier analysis showed that patients with CKAP4-1 had significantly shorter cancer-specific survival and recurrence-free survival (RFS; p = 0.046 and p = 0.0173, respectively). Multivariate analysis showed that CKAP4-1 positivity was an independent predictor of RFS (p = 0.041, hazard ratio: 2.09, 95% confidence interval: 1.03–4.25). Conclusions: This study showed that CKAP4 expression in tumor cells may serve as a useful prognostic biomarker for patients with BCa who undergo RC. Full article

The management of head and neck squamous cell carcinoma (HNSCC) relies heavily on TNM staging and WHO histologic grading; however, in recent years, the analysis of prognostic markers expressed in the tumor stroma has gained attention. The tumor–stroma ratio (TSR) quantifies the proportion of tumor tissue relative to the surrounding stromal tissue; it is assessed with the percentage of stromal tissue within the tumor area, with a cutoff point of 50% being widely used to discriminate high-stroma cancer. In this systematic review and meta-analysis, we investigated the potential prognostic role of the TSR in HNSCC. After a literature screening, 24 studies dealing with the TSR and survival outcomes were included. The TSR showed a significant association with overall survival (OS) in both unadjusted and adjusted measures (RR 2.04, CI 1.57–2.65, p < 0.01; HR 2.36 CI 1.89–2.94, p < 0.00001), with an even stronger prognostic potential in oral cavity/oral tongue cancers (RR 2.44 CI 1.84–3.22, p < 0.00001). The TSR also showed prognostic value when dealing with cancer-specific survival and was associated with a reduction in disease-free survival (DFS). In particular, the TSR also retained its prognostic role in terms of DFS when specifically considering early-stage cancers in both unadjusted and adjusted analyses (RR 1.81 CI 1.57–2.10, p < 0.00001; HR 2.09 CI 1.58–2.76, p < 0.00001). Therefore, we conclude that the TSR is a reliable prognostic marker that is easy to assess in routine histological slides and can be effectively implemented in the routine evaluation of HNSCC. Full article

Triple-negative breast cancer (TNBC) is a subtype of breast cancer with a poor prognosis and limited treatment options. This study evaluates the prognostic value of stromal markers in TNBC, focusing on the tumor–stroma ratio (TSR) and overall stroma ratio (OSR) in whole slide images (WSI), as well as the expression of type-I collagen, type-III collagen, and fibrillin-1 on tissue microarrays (TMAs), using both visual assessment and digital image analysis (DIA). A total of 101 female TNBC patients, primarily treated with surgery between 2005 and 2016, were included. We found that high visual OSR correlates with worse overall survival (OS), advanced pN categories, lower stromal tumor-infiltrating lymphocyte count (sTIL), lower mitotic index, and patient age (p < 0.05). TSR showed significant connections to the pN category and mitotic index (p < 0.01). High expression levels of type-I collagen (>45%), type-III collagen (>30%), and fibrillin-1 (>20%) were linked to significantly worse OS (p = 0.004, p = 0.013, and p = 0.005, respectively) and progression-free survival (PFS) (p = 0.028, p = 0.025, and p = 0.002, respectively), validated at the mRNA level. Our results highlight the importance of stromal characteristics in promoting tumor progression and metastasis and that targeting extracellular matrix (ECM) components may offer novel therapeutic strategies. Furthermore, DIA can be more accurate and objective in evaluating TSR, OSR, and immunodetected stromal markers than traditional visual examination. Full article

Breast cancer (BC) is the most common cancer in women, and therapeutic strategies for it are based on the molecular subtypes of luminal BC, HER2 BC, and triple-negative BC (TNBC) because each subtype harbors different unique genetic aberrations. Recently, features of the tumor microenvironment (TME), especially cancer-associated fibroblasts (CAFs), have been demonstrated to play a critical role in BC progression, and we would like to understand the molecular features of BC CAFs for novel therapeutic strategies. In a recent study, 115 CAF-associated genes (CAFGs) were identified in a public database of microdissection and microarray data (GSE35602) from 13 colorectal cancer (CRC) tumors. Using a public database (GSE10797) of 28 BC tumors, a similar analysis was performed. In BC, 59 genes from the 115 CAFGs identified in CRC (CRC CAFGs) were also closely associated with a CAFs marker, SPARC (R = 0.6 or beyond), and POSTN was of particular interest as one of the BC CAFGs with the highest expression levels and a close association with SPARC expression (R = 0.94) in the cancer stroma of BC tumors. In BC stroma, POSTN was followed in expression levels by DKK3, MMP2, PDPN, and ACTA2. Unexpectedly, FAP and VIM were not as highly associated with SPARC expression in the cancer stroma of BC tumors and exhibited low expression. These findings suggested that ACTA2 might be the most relevant conventional CAFs marker in BC, and ACTA2 was actually correlated in expression with many CRC CAFGs, such as SPARC. Surprisingly, the SE ratio values of the BC CAFGs were much lower (average SE = 3.8) than those of the CRC CAFGs (SE = 10 or beyond). We summarized the current understanding of BC CAFs from the literature. Finally, in triple-negative BC (TNBC) (n = 5), SPARC expression uniquely showed a close association with COL11A1 and TAGLN expression, representing a myofibroblast (myCAFs) marker in the cancer stroma of the BC tumors, suggesting that myCAFs may be molecularly characterized by TNBC in contrast to other BC phenotypes. In summary, CAFs could have unique molecular characteristics in BC, and such TME uniqueness could be therapeutically targeted in BC. Full article

About one-fourth of patients with pancreatic ductal adenocarcinoma (PDAC) are categorized as borderline resectable (BR) or locally advanced (LA). Chemotherapy and radiation therapy have not yielded the anticipated outcomes in curing patients with BR/LA PDAC. The surgical resection of these tumors presents challenges owing to the unpredictability of the resection margin, involvement of vasculature with the tumor, the likelihood of occult metastasis, a higher ratio of positive lymph nodes, and the relatively larger size of tumor nodules. Oncolytic virotherapy has shown promising activity in preclinical PDAC models. Unfortunately, the desmoplastic stroma within the PDAC tumor microenvironment establishes a barrier, hindering the infiltration of oncolytic viruses and various therapeutic drugs—such as antibodies, adoptive cell therapy agents, and chemotherapeutic agents—in reaching the tumor site. Recently, a growing emphasis has been placed on targeting major acellular components of tumor stroma, such as hyaluronic acid and collagen, to enhance drug penetration. Oncolytic viruses can be engineered to express proteolytic enzymes that cleave hyaluronic acid and collagen into smaller polypeptides, thereby softening the desmoplastic stroma, ultimately leading to increased viral distribution along with increased oncolysis and subsequent tumor size regression. This approach may offer new possibilities to improve the resectability of patients diagnosed with BR and LA PDAC. Full article

Fibrotic stroma and angiogenic tumor vessels play an important role in modulating tumor immunity. We previously reported a rationally designed protein (ProAgio) that targets integrin α_vβ₃ at a novel site. ProAgio induces the apoptosis of cells that express high levels of the integrin. Both activated cancer-associated fibroblasts (CAFs) and angiogenic endothelial cells (aECs) in tumors express high levels of integrin α_vβ₃. ProAgio simultaneously and specifically induces apoptosis in CAFs and aECs in tumors. We provide evidence here that the depletion of CAFs and the elimination of leaky tumor angiogenic vessels by ProAgio alter tumor immunity. ProAgio reduces CD₄⁺ Treg and Myeloid-derived suppressor cells (MDSCs), increases CD₈⁺ T-cells, and increases the M1/M2 macrophage ratio in the tumor. The depletion of dense fibrotic stroma (CAFs) by ProAgio decreases the Programmed Death Ligand 1 (PDL-1) levels in the stroma areas surrounding the tumors, and thus strongly increases the delivery of anti-PDL-1 antibody to the target cancer cells. The impact of ProAgio on tumor immunity provides strong synergistical effects of checkpoint inhibitors on lung cancer treatment. Full article

The response of tumors to anti-cancer therapies is defined not only by cell-intrinsic therapy sensitivities but also by local interactions with the tumor microenvironment. Fibroblasts that make tumor stroma have been shown to produce paracrine factors that can strongly reduce the sensitivity of tumor cells to many types of targeted therapies. Moreover, a high stroma/tumor ratio is generally associated with poor survival and reduced therapy responses. However, in contrast to advanced knowledge of the molecular mechanisms responsible for stroma-mediated resistance, its effect on the ability of tumors to escape therapeutic eradication remains poorly understood. To a large extent, this gap of knowledge reflects the challenge of accounting for the spatial aspects of microenvironmental resistance, especially over longer time frames. To address this problem, we integrated spatial inferences of proliferation-death dynamics from an experimental animal model of targeted therapy responses with spatial mathematical modeling. With this approach, we dissected the impact of tumor/stroma distribution, magnitude and distance of stromal effects. While all of the tested parameters affected the ability of tumor cells to resist elimination, spatial patterns of stroma distribution within tumor tissue had a particularly strong impact. Full article

High intratumoral heterogeneity is thought to be a poor prognostic indicator. However, the source of heterogeneity may also be important, as genomic heterogeneity is not always reflected in histologic or ‘visual’ heterogeneity. We aimed to develop a predictor of histologic heterogeneity and evaluate its association with outcomes and molecular heterogeneity. We used VGG16 to train an image classifier to identify unique, patient-specific visual features in 1655 breast tumors (5907 core images) from the Carolina Breast Cancer Study (CBCS). Extracted features for images, as well as the epithelial and stromal image components, were hierarchically clustered, and visual heterogeneity was defined as a greater distance between images from the same patient. We assessed the association between visual heterogeneity, clinical features, and DNA-based molecular heterogeneity using generalized linear models, and we used Cox models to estimate the association between visual heterogeneity and tumor recurrence. Basal-like and ER-negative tumors were more likely to have low visual heterogeneity, as were the tumors from younger and Black women. Less heterogeneous tumors had a higher risk of recurrence (hazard ratio = 1.62, 95% confidence interval = 1.22–2.16), and were more likely to come from patients whose tumors were comprised of only one subclone or had a TP53 mutation. Associations were similar regardless of whether the image was based on stroma, epithelium, or both. Histologic heterogeneity adds complementary information to commonly used molecular indicators, with low heterogeneity predicting worse outcomes. Future work integrating multiple sources of heterogeneity may provide a more comprehensive understanding of tumor progression. Full article

Tumor-associated macrophages (TAMs) is a key element in the breast tumor microenvironment. CD163 and CD206 have been utilized for TAM identification, but the clinical implications of TAMs identified by these markers have not been thoroughly explored. This study conducted a comparative analysis of CD163 and CD206 TAMs using digital image analysis, focusing on their spatial distribution and prognostic significance in relation to tumor-infiltrating lymphocytes (TILs). Distinct clinico-pathological and prognostic characteristics were noted between the two types of TAMs. CD163 TAMs were linked to high-grade tumors (p = 0.006), whereas CD206 TAMs were associated with a higher incidence of nodal metastasis (p = 0.033). CD206 TAMs were predominantly found in the stroma, with more cases being stromal CD206-high (sCD206-high) than tumoral CD206-high (tCD206-high) (p = 0.024). Regarding prognostication, patients stratified according to stromal and tumoral densities of CD163 showed different disease-free survival (DFS) time. Specifically, those that were sCD163-low but tCD163-high exhibited the poorest DFS (chi-square = 10.853, p = 0.013). Furthermore, a high sCD163-to-stromal-TILs ratio was identified as an independent predictor of unfavorable survival outcomes (DFS: HR = 3.477, p = 0.018). The spatial distribution and interactions with TILs enhanced the prognostic value of CD163 TAMs, while CD206 TAMs appeared to have limited prognostic utility in breast cancer cases. Full article