Martinez-Balibrea, E. | Search bioRxiv

bioRxiv2025

Loss-of-Function genetic Screen Unveils Synergistic Efficacy of PARG Inhibition with Combined 5-Fluorouracil and Irinotecan Treatment in Colorectal Cancer

Colorectal cancer (CRC) remains a major global health concern, partly due to resistance to therapy and the lack of new effective treatments for advanced disease. The combination of 5-Fluorouracil (5FU, a thymidylate synthase inhibitor) and irinotecan (a topoisomerase 1 inhibitor) is widely used in first-line and subsequent treatments. This study aimed to identify novel therapeutic targets to enhance combinatorial therapy, improving treatment efficacy and durability of response. We performed a loss-of-function screen using HT29 CRC cell line and a retroviral library containing 7296 shRNAs targeting 912 chromatin genes. Cells were then treated with 5FU and SN38 (the active metabolite of irinotecan) or left untreated for 4 weeks. Genes enriched in resistant clones were identified through next-generation sequencing. Among candidate genes, PARG was selected for functional validation. CRISPR/Cas9-mediated knockout (HT29 PARG-KO) resulted in increased global poly(ADP-ribosyl)ation after 5FU and SN38 treatment. PARG depletion led to reduced cell viability and increased apoptosis, particularly after 5FU exposure. Pharmacological PARG inhibition (PDD00017273) synergized with 5FU and SN38 across three CRC models (HT29, DLD1, HT115). In vivo, HT29 PARG-KO xenografts were more sensitive to 5FU. Immunohistochemical analysis of 170 CRC patient tumors revealed that positive PARG expression correlated with poor response to 5FU + Irinotecan, increased liver metastases, and worse long-term survival. Our findings highlight PARG as a promising therapeutic target for CRC, where its inhibition enhances the efficacy of standard chemotherapy.

cancer biology↗

bioRxiv2020

Taxane induces attenuation of the CXCR2/BCL-2 axis and sensitize prostate cancer to platinum-based treatments

BackgroundTaxanes are the most active chemotherapy agents in metastatic castration resistant prostate cancer (mCRPC) patients, yet resistance almost invariably occurs representing an important clinical challenge. Taxane-platinum combinations have shown clinical benefit in a subset of patients but the mechanistic basis and biomarkers remain elusive. ObjectiveTo identify mechanisms and response biomarkers for the antitumor efficacy of taxane-platinum combinations in mCRPC. Design, setting, and participantsTranscriptomic data from a publicly available mCRPC dataset of taxane-exposed and naive patients was analysed to identify response biomarkers and emerging vulnerabilities. Functional and preclinical validation was performed in taxane resistant mCRPC cell lines and genetically engineered mouse models (GEMM). InterventionmCRPC cells were treated with docetaxel, cisplatin, carboplatin and the CXCR2 inhibitor, SB265610. Gain and loss of function in culture of CXCR2 was achieved by overexpression or siRNA-silencing. Preclinical assays in GEMM mice tested the anti-tumor efficacy of taxane-platinum combinations. Outcome measurements and statistical analysisProliferation, apoptosis and colony assays measured drug activity in vitro. Preclinical endpoints in mice included growth, survival and histopathology. Changes in CXCR2, BCL-2 and chemokines were analysed by RT-qPCR and Western Blot. Human expression data was analyzed using GSEA, hierarchical clustering and correlation studies. GraphPad Prism software, R-studio, were used for statistical and data analyses. Results and limitationsTranscriptomic data from taxane-exposed human mCRPC tumors correlates with a marked negative enrichment of apoptosis and inflammatory response pathways accompanied by a marked downregulation of CXCR2 and BCL-2. Mechanistically, we show that docetaxel treatment inhibits CXCR2 and that BCL-2 downregulation occurs as a downstream effect. Further, we demonstrated that taxane resistance is directly associated to CXCR2 expression and that targeting of CXCR2 sensitizes prostate cancer (PC) cells to cisplatin. Finally, taxane-platinum combinations in vivo are highly synergistic and previous exposure to taxanes sensitizes mCRPC tumors to second line cisplatin treatment. ConclusionsTogether our data identifies an acquired vulnerability in taxane treated mCRPC patients with potential predictive activity for platinum-based treatments. Patient summaryA subset of patients with aggressive and therapy resistant PC benefits from taxane-platinum combination chemotherapy however, we lack biomarkers and mechanistic basis about how that synergistic effect occurs. Here, using patient data and preclinical models, we found that taxanes reduce cancer cell scape mechanisms to chemotherapy-induced cell death, hence turning these cells more vulnerable to additional platinum treatment.

cancer biology↗