Drug resistance is a major challenge in the treatment of advanced cholangiocarcinoma (CCA). Understanding the mechanisms of drug resistance can aid in identifying novel prognostic biomarkers and therapeutic targets to improve treatment efficacy. This study established 5-fluorouracil- (5-FU) and gemcitabine-resistant CCA cell lines, KKU-213FR and KKU-213GR, and utilized comparative proteomics to identify differentially expressed proteins in drug-resistant cells compared to parental cells. Additionally, bioinformatics analyses were conducted to explore the biological and clinical significance of key proteins. The drug-resistant phenotypes of KKU-213FR and KKU-213GR cell lines were confirmed. In addition, these cells demonstrated increased migration and invasion abilities. Proteomics analysis identified 81 differentially expressed proteins in drug-resistant cells, primarily related to binding functions, biological regulation, and metabolic processes. Protein–protein interaction analysis revealed a highly interconnected network involving MET, LAMB1, ITGA3, NOTCH2, CDH2, and NDRG1. siRNA-mediated knockdown of these genes in drug-resistant cell lines attenuated cell migration and cell invasion abilities and increased sensitivity to 5-FU and gemcitabine. The mRNA expression of these genes is upregulated in CCA patient samples and is associated with poor prognosis in gastrointestinal cancers. Furthermore, the functions of these proteins are closely related to the epithelial–mesenchymal transition (EMT) pathway. These findings elucidate the potential molecular mechanisms underlying drug resistance and tumor progression in CCA, providing insights into potential therapeutic targets.

Cervical cancer (CC) remains a major global health challenge due to chemotherapy resistance and recurrence. Mesenchymal stem cell-derived exosomes (MSC-exosomes) have dual roles, as they can act as therapeutic agents and contribute to chemoresistance. However, their role in response to chemotherapy in CC remains unclear. Therefore, our study investigated the effects of MSC-exosome pretreatment on chemotherapy sensitivity using three-dimensional spheroid models generated from HeLa and SiHa CC cell lines. Proteomic profiling of MSC-exosomes identified key proteins, including ANXA1, ANXA2, EEF2, LGALS1, and PKM2, associated with tumor regeneration and chemotherapy response. MSC-exosomes exhibited context-dependent effects in both chemoresistance and chemosensitization by modulating drug efflux, metabolic reprogramming, stress adaptation, apoptosis, DNA damage response, and integrin-mediated signaling. MSC-exosome pretreatment altered spheroid responses to paclitaxel in combination with cisplatin or carboplatin. MSC-exosomes significantly enhanced chemotherapy-induced cytotoxicity in HeLa spheroids, as evidenced by reduced cell viability, increased caspase activity, and upregulation of the pro-apoptotic marker Bax. In contrast, SiHa spheroids represented selective responses: MSC-exosome pretreatment did not enhance sensitivity to paclitaxel–cisplatin but improved responsiveness to paclitaxel–carboplatin, particularly within the spheroid core. Overall, MSC-exosome pretreatment exerts cell type and drug-specific effects in CC spheroids, supporting their potential to modulate chemotherapy response.