Oncogenic KRAS mutations are a defining feature of pancreatic ductal adenocarcinoma (PDAC), one of the most lethal solid malignancies, characterized by poor responsiveness to chemotherapy. Recent clinical successes of direct KRAS inhibitors in other cancer types have renewed interest in KRAS-directed therapy for PDAC. However, early clinical experience has revealed often short-lived responses, highlighting the rapid emergence of resistance and the need to understand the mechanisms limiting durable benefit. This review summarizes current knowledge on molecular resistance to RAS-directed inhibitors in PDAC, organized into five categories: intrinsic resistance, KRAS-dependent mechanisms, KRAS-independent bypass signaling, downstream pathway reactivation, and tumor microenvironment (TME)-mediated resistance. Evidence from PDAC models is integrated with insights from other KRAS-driven malignancies, particularly non-small cell lung cancer, where direct KRAS inhibition has been studied extensively. Collectively, resistance appears to arise from layered adaptive processes rather than single alterations, including secondary KRAS mutations, receptor tyrosine kinase-driven bypass signaling, reactivation of mitogen-activated protein kinase (MAPK) and phosphoinositide 3-kinase (PI3K) pathways, stromal-mediated protection, and reduced drug exposure. Notably, until recently, most evidence in PDAC has been indirect, while direct investigation of resistance mechanisms in KRAS-targeted settings has expanded rapidly only in recent years. Building on these advances, ongoing clinical trials increasingly explore rational combination strategies targeting upstream regulators, downstream effectors, and the TME. However, critical challenges persist, including patient selection, treatment sequencing, toxicity, and the lack of pharmacodynamic frameworks. Overcoming resistance will require mechanistically guided combinations, improved disease-specific models, and biomarker-driven adaptive strategies to ultimately achieve durable clinical benefits in PDAC.
Tyrosine kinase inhibitors (TKIs) have significantly changed the treatment of non-small cell lung cancer (NSCLC) harbouring epidermal growth factor receptor (EGFR) mutations, and osimertinib is now established as first-line therapy for NSCLCs. Combination therapies (e.g., osimertininb plus chemotherapy; lazertinib plus amivantamab) have been shown to improve median progression-free survival (mPFS) and median overall survival (mOS) relative to monotherapy. However, due to several resistance mechanisms, patients experience disease progression following EGFR TKI treatment. On-target resistance mechanisms include additional mutations (e.g., C797S/G/N, L718Q, L844V, G724X). Within the heterogeneous group of off-target resistance mechanisms, human epidermal growth factor receptor 2 (HER2) amplifications, mesenchymal-epithelial transition factor (MET) alterations, oncogenic fusions (e.g., BRAF, FGFR, RET), histological changes, epithelial-mesenchymal transitions, and alterations of the RAS/MEK/ERK and the PI3K/AKT/mTOR signal transduction pathways are critical and can confer resistance to EGFR TKIs. Several drugs have been identified to inhibit these pathways, with some of them already approved for clinical use. Most fourth-generation EGFR TKIs are orally bioavailable and are mainly allosteric thiazole amide-based reversible inhibitors. Their activity results from selective binding to an allosteric site, which can alter the EGFR protein conformation, allowing them to bypass C797X. A recommendation for the optimal treatment strategy and sequence for NSCLC patients with acquired EGFR TKI-resistant tumours still cannot be given. An improved understanding of the underlying resistance mechanisms will help to pave the way for the development of innovative and highly specific drugs for the therapy of osimertinib-resistant NSCLCs. The putative clinical relevance of fourth-generation EGFR TKIs for NSCLC patients needs to be defined, and many development hurdles need to be cleared before victory can be declared.
Aim: Despite the contribution of cancer stem-like cells (CSLCs) to acquired paclitaxel resistance in non-small cell lung cancer (NSCLC), the biomarkers and regulatory mechanisms sustaining their stemness under chemotherapy pressure remain poorly understood. This study aimed to identify the stemness-maintaining programs underlying CSLC-associated paclitaxel resistance. Methods: Paclitaxel-resistant NSCLC cell models were established. RNA-seq data from resistant spheres and adherent resistant cells were integrated with Gene Ontology/Kyoto Encyclopedia of Genes and Genomes/gene set enrichment analysis and patient transcriptomic datasets to identify CSLC maintenance-associated candidate biomarkers. Inhibitors, sphere-forming assays, CD104-CD166+CD49fhi flow cytometry, reverse transcription quantitative polymerase chain reaction, western blotting, and ST6GAL1 knockdown or overexpression were used for functional and mechanistic validation. Sambucus nigra agglutinin lectin blotting was performed to assess epidermal growth factor receptor (EGFR) α2,6-sialylation. Clinical relevance was assessed using ST6GAL1 immunohistochemistry on 46 clinical lung tumor tissues. Results: Paclitaxel-resistant NSCLC cells exhibited enhanced sphere formation and CD104-CD166+CD49fhi expansion. N-glycosylation was activated in resistant spheres. A seven-gene N-glycosylation signature was identified as a CSLC-associated candidate biomarker in acquired paclitaxel resistance. Inhibiting N-glycosylation suppressed the EGFR-mTOR-SOX2/BMI1 axis, decreased CSLCs, and restored paclitaxel sensitivity. ST6GAL1 regulated EGFR α2,6-sialylation. ST6GAL1 depletion also decreased EGFR abundance, suppressed mTOR-SOX2/BMI1 signaling, and sensitized paclitaxel-resistant spheres to paclitaxel rather than adherent cells. ST6GAL1 expression, which was higher in tumors from patients who underwent chemotherapy, showed a trend toward poorer survival among chemotherapy-treated patients. The seven-gene signature was associated with shorter disease-free survival but not overall survival in lung cancer patients. Conclusions: ST6GAL1-mediated α2,6-sialylation of EGFR contributes to the maintenance of CSLC-associated paclitaxel resistance through mTOR-SOX2/BMI1 signaling. These findings identify ST6GAL1-dependent EGFR sialylation as a potential therapeutic target in CSLC-associated chemoresistance.
Aim: Pancreatic ductal adenocarcinoma (PDAC) has a dismal prognosis, and innovative therapies are needed. We developed a phase I pilot, non-viral gene therapy (CYL-02) encoding SST2 and DCK::UMK genes complexed with a polyethyleneimine non-viral vector to sensitize tumours to gemcitabine chemotherapy, which showed excellent feasibility and safety. Methods: An open-label, multicentre, randomized Phase 2 trial (NCT02806687) compared CYL-02 [two intratumoral endoscopic ultrasound (EUS)-guided injections one month apart] + gemcitabine injected weekly vs. gemcitabine alone in 70 patients (n = 35 per arm) with locally advanced PDAC. The primary endpoint was progression-free survival (PFS). Secondary endpoints were overall survival (OS), tumour response, safety, and ancillary studies. Results: PFS did not differ between arms. Unexpectedly, OS was worse in the gene therapy + chemotherapy arm. Exploratory analyses revealed no differences in clinical status, baseline characteristics, or the number of patients receiving second-line therapy, with the exception of a more frequent use of FOLFOX post-progression in the control arm, which could explain this difference. Proteomic profiling identified clusterin (CLU) as a candidate treatment-induced resistance factor in patients: CYL-02 + gemcitabine significantly increased circulating CLU levels, while gemcitabine alone did not. In vitro and in vivo models confirmed CLU induction by DCK combined with gemcitabine and demonstrated that CLU silencing restored chemosensitivity, reducing experimental tumour volume in mice. Conclusion: CLU induction in patients receiving CYL-02 and gemcitabine may explain adaptive resistance and lack of efficacy of gene therapy. These results provide a rationale for combining CYL-02 and gemcitabine with CLU targeting to overcome resistance in pancreatic cancer.
Therapeutic resistance remains a major challenge in bladder cancer, encompassing resistance to intravesical Bacillus Calmette–Guérin (BCG) therapy, platinum- and gemcitabine-based chemotherapy and immune-checkpoint blockade. Although extracellular vesicles (EVs) have been implicated in cancer drug resistance, their roles in bladder cancer are often discussed within the broader context of EV biology, making it difficult to distinguish direct mechanisms of treatment resistance from treatment-associated remodelling or biomarker associations. In this review, we refocus EV-mediated therapeutic resistance in bladder cancer and propose an evidence-stratified framework, distinguishing EVs as causal mediators, permissive amplifiers or biomarkers of drug adaptation. Studies have provided substantial mechanistic evidence from chemotherapy-associated models, including EV-mediated regulation of gemcitabine resistance through H3C14-associated EV remodelling and chemotherapy-exposed bladder cancer EVs that support stem-like, treatment-tolerant tumour states. Evidence supporting EV involvement in platinum resistance, BCG failure and immunotherapy resistance remains biologically plausible but less directly established. Therefore, clearer drug-specific models, improved EV-subtype resolution, cargo perturbation and functional rescue experiments are needed. We also discuss urinary EVs for longitudinal monitoring of treatment response and emerging resistance in bladder cancer. By integrating drug-specific therapeutic contexts with EV-specific causality criteria, this review provides a focused framework for evaluating when EVs mediate bladder cancer drug resistance, amplify adaptive tumour–microenvironment states or primarily serve as biomarkers for therapeutic monitoring.
