Lung cancer is still the main cause of cancer death worldwide despite the availability of targeted therapies and immune-checkpoint inhibitors combined with chemotherapy. Cancer cell heterogeneity and primary or acquired resistance mechanisms cause the elusive behaviour of this cancer and new biomarkers and active drugs are urgently needed to overcome these limitations. p65BTK, a novel isoform of the Bruton Tyrosine Kinase may represent a new actionable target in non-small cell lung cancer (NSCLC). p65BTK expression was evaluated by immunohistochemistry in 382 NSCLC patients with complete clinico-pathological records including smoking habit, ALK and EGFR status, and in metastatic lymph nodes of 30 NSCLC patients. NSCLC cell lines mutated for p53 and/or a component of the RAS/MAPK pathway and primary lung cancer-derived cells from Kras/Trp53 null mice were used as a preclinical model. The effects of p65BTK inhibition by BTK Tyrosine Kinase Inhibitors (TKIs) (Ibrutinib, AVL-292, RN486) and first-generation EGFR-TKIs (Gefitinib, Erlotinib) on cell viability were evaluated by MTT. The effects of BTK-TKIs on cell growth and clonogenicity were assessed by crystal violet and colony assays, respectively. Cell toxicity assays were performed to study the effect of the combination of non-toxic concentrations of BTK-TKIs with EGFR-TKIs and standard-of-care (SOC) chemotherapy (Cisplatin, Gemcitabine, Pemetrexed). p65BTK was significantly over-expressed in EGFR-wild type (wt) adenocarcinomas (AdC) from non-smoker patients and its expression was also preserved at the metastatic site. p65BTK was also over-expressed in cell lines mutated for KRAS or for a component of the RAS/MAPK pathway and in tumors from Kras/Trp53 null mice. BTK-TKIs were more effective than EGFR-TKIs in decreasing cancer cell viability and significantly impaired cell proliferation and clonogenicity. Moreover, non-toxic doses of BTK-TKIs re-sensitized drug-resistant NSCLC cell lines to both target- and SOC therapy, independently from EGFR/KRAS status. p65BTK results as an emerging actionable target in non-smoking EGFR-wt AdC, also at advanced stages of disease. Notably, these patients are not eligible for EGFR-TKIs-based therapy due to a lack of EGFR mutation. The combination of BTK-TKIs with EGFR-TKIs is cytotoxic for EGFR-wt/KRAS-mutant/p53-null tumors and BTK-TKIs re-sensitizes drug-resistant NSCLC to SOC chemotherapy. Therefore, our data suggest that adding BTK-TKIs to SOC chemotherapy and EGFR-targeted therapy may open new avenues for clinical trials in currently untreatable NSCLC.

Gastric cancer (GC) continues to be a major cause of cancer-related deaths globally, primarily due to resistance to standard treatments like 5-fluorouracil (5FU). The transforming growth factor-β (TGF-β) signaling pathway is recognized as a key contributor to tumor progression and resistance to therapy. This work investigated the therapeutic potential of targeting TGF-β receptor I (TGFBR1) with the selective inhibitor SB431542 to enhance the effect of 5FU in GC. Analysis of public gene expression datasets revealed that increased levels of TGF-β and TGFBR1 are significantly connected with poor prognosis, particularly in high-grade GC. In vitro experiments using AGS and SNU-1 cell lines demonstrated that co-treatment with SB431542 and 5FU significantly reduced cell viability, making GC cells more sensitive to 5FU. This combination treatment led to a significant activation of caspase-dependent apoptosis, indicating an enhanced pro-apoptotic effect. These findings suggest that TGFBR1 inhibition could provide a strategic approach to reduce the dosage of 5FU, thereby minimizing its severe side effects in gastric cancer patients. Furthermore, these results underscore the potential of TGFBR1 as both a prognostic biomarker and a therapeutic target, warranting further investigation in aggressive forms of gastric cancer.

Background: High dissemination potential and resistance to standard therapy significantly contribute to high mortality associated with ovarian cancer. Cancer stem cells (CSCs) drive tumor progression, metastasis, and recurrence after treatments’ failure. Here, we provide the first evidence of TIMP1 overexpression in ovarian CSCs, suggesting its potential role as a prognostic biomarker. Methods: Different ovarian cancer cell models were used to explore the potential link between TIMP1 and stem-like phenotypes. Experiments included spheroid formation, drug treatments, gene expression, functional assays, and zebrafish xenograft models to assess cell behavior and molecular changes. Results: TIMP1 was overexpressed in CSCs, and its expression was also upregulated in chemoresistant and anoikis-resistant cells. Our database analysis revealed a correlation between TIMP1 expression levels and poor patient prognosis. Overexpression of TIMP1 in ovarian cancer cell lines was able to recapitulate several features of the ovarian cancer stem cell phenotype, including treatment resistance, expression of stem cell markers, and anoikis resistance. TIMP1-overexpressing cells also exhibited enhanced migration potential in vitro and increased metastatic potential in vivo. Moreover, TIMP1 overexpression significantly altered the transcriptome landscape of cells, highlighting its role in modulating critical pathways associated with cell migration and inflammation. Conclusions: This study identifies the pivotal role of TIMP1 in ovarian CSCs and its contribution to therapy resistance, recurrence, and metastasis.