Metastatik kolorektal kanserde hasta kökenli organoid biyobankasının oluşturulması ve pan-KRAS inhibitörüne direncin incelenmesi

Metastatic colorectal cancer (mCRC) displays marked inter- and intratumoral heterogeneity, leading to variable therapeutic responses and frequent treatment failure. To model this complexity, a living biobank of patient-derived organoids (PDOs) from twenty mCRC patients representing diverse genetic backgrounds was established. The findings showed the preservation of the histological, genomic, and transcriptional features of their tumors of origin. Treatment responses were assessed using a multiparametric three-dimensional drug screening platform that integrates measurements of organoid growth, nuclear content, and proliferation, providing greater sensitivity than conventional viability assays. Drug response patterns demonstrated KRAS wild-type organoids exhibited pronounced cytotoxic responses, whereas KRAS-mutant models showed reduced cytotoxicity and predominantly cytostatic behavior. To investigate resistance mechanisms to the novel pan-KRAS inhibitor BI-2865, resistance evolution was modeled through longitudinal drug exposure combined with DNA barcode based clonal tracing and single-cell RNA sequencing. This approach enabled analysis of pan-KRAS resistance from an evolutionary and clonal perspective. Resistance was primarily driven by transcriptional reprogramming rather than extensive genomic alterations. Distinct evolutionary trajectories emerged depending on KRAS genotype. In KRAS wildtype organoids, resistance arose through the emergence of new resistant clones, whereas in KRASG13D models it resulted from the selective expansion of pre-existing tolerant populations. Focusing on KRASG13D organoids, single-cell transcriptomic profiling revealed that resistant cells occupied multiple transcriptional states rather than converging on a single identity. Together, these findings demonstrate that resistance to pan-KRAS inhibition is a dynamic, genotype-dependent process shaped by transcriptional plasticity and clonal selection and establish an integrated PDObased framework for dissecting therapeutic resistance in metastatic colorectal cancer.