By blocking three survival pathways simultaneously, researchers caused sustained pancreatic tumor regression in mouse and human tumor-derived models, providing a cautious but promising route to overcoming drug resistance.
Research: Targeted combination therapy achieves effective regression of pancreatic cancer and prevents tumor resistance. Image credit: Crystal Light / Shutterstock
In a recent study published in PNASA group of researchers investigated whether simultaneous inhibition of Kirsten rat sarcoma virus oncogene homolog (KRAS), epidermal growth factor receptor (EGFR), and signal transducer and activator of transcription 3 (STAT3) could induce sustained pancreatic tumor regression and prevent treatment resistance.
Pancreatic cancer resistance pathway
Pancreatic ductal adenocarcinoma remains one of the most lethal cancers, and despite advances in cancer treatment, survival rates remain alarmingly low. Cancer cells become resistant to targeted therapies, limiting clinical efficacy, and tumors continue to grow after treatment.
Many pancreatic tumors are caused by KRAS mutations, so KRAS-targeted therapy would be a major advance. It is becoming clear that during treatment-resistant tumor growth, neoplastic cells can activate alternative signaling pathways to survive targeted therapies. Thus, understanding the mechanisms of escape should be central to achieving long-lasting responses in patients.
Preclinical pancreatic cancer model
Multiple preclinical models were used to study the combined effects of targeting KRAS, EGFR, and STAT3. Genetically engineered mouse models carrying mutant KRAS and tumor protein P53 (TP53) genes were used to replicate human pancreatic cancer, orthotopic pancreatic tumors in which tumors were transplanted directly into the pancreas of immunocompetent mice, and patient-derived xenografts prepared from human pancreatic ductal adenocarcinoma samples and transplanted into immunodeficient mice.
To determine the role of RAF1 (Raf-1 proto-oncogene), EGFR, and STAT3 in tumor survival, tumor cells of pancreatic tumors were isolated from mouse pancreas, cultured under controlled laboratory conditions, and genetic engineering techniques were used to selectively ablate or suppress RAF1, EGFR, and STAT3.
Various signaling molecules such as Janus kinase 1 (JAK1), Janus kinase 2 (JAK2), interleukin 6 receptor alpha (IL6RA), FYN proto-oncogene, and Src family tyrosine kinases (FYN) were also evaluated to determine their role in resistance mechanisms.
In pharmacological studies, mice were administered daraxone lasib, a RAS(ON) inhibitor that targets multiple active RAS proteins and KRAS-driven signaling, afatinib, an irreversible inhibitor of EGFR and human epidermal growth factor receptor 2 (HER2), and SD36, a proteolytic targeting chimera (PROTAC) designed to degrade STAT3.
Tumor growth was monitored by ultrasound imaging, and efficacy, toxicity, and long-term outcomes were evaluated across different experimental paradigms using tissue analysis, immunohistochemistry, blood tests, and survival assessments.
STAT3 promotes treatment resistance
This study revealed that inhibition of RAF1 and EGFR alone is insufficient to eliminate advanced pancreatic tumors. Although some small tumors responded, larger tumors survived by activating STAT3 signaling.
The researchers found that phosphorylation of STAT3 at tyrosine 705 was increased in resistant cancers, indicating an increase in active STAT3. When STAT3 is silenced, resistant cancer cells die, but when a permanently activated form of STAT3 is introduced into naturally sensitive cells, resistance develops.
These data indicate that STAT3 is an important factor in treatment resistance.
Further experiments demonstrated that STAT3 activation is not primarily controlled by conventional JAK1, JAK2, or IL6RA signaling pathways. Instead, increased FYN activity promoted STAT3 activation.
Inhibiting FYN reduced STAT3 signaling and eliminated resistant tumor cells, revealing a previously underappreciated resistance mechanism.
In orthotopic tumors derived from genetically engineered mouse pancreatic cancer cells, simultaneous deletion of RAF1, EGFR, and STAT3 induced apoptosis, resulting in complete tumor disappearance within 3 to 4 weeks and no evidence of recurrence during up to 300 days of follow-up.
Histopathological analysis of the specimen showed complete removal of all tumor tissue and associated stroma, indicating tumor regression.
Three-drug combination therapy prevents tumor recurrence
The researchers then developed a drug-based approach based on their results. Dalaxone lasib alone inhibited the extracellular signal-regulated kinase (ERK) signaling pathway and slowed tumor growth, but the tumors later developed resistance and resumed growth. The combination of dalaxonelasib and afatinib produced a better response but was unable to completely destroy the tumor.
Combination therapy of daraxone lasib, afatinib, and SD36 induced strong apoptosis and complete regression in orthotopic murine tumors, with no detectable tumors in the mice for more than 200 days after treatment and no evidence of tumor recurrence or resistance. In genetically engineered mouse tumors, triple therapy resulted in regression in all treated tumors, with half showing complete regression within 30 to 60 days.
Importantly, this three-drug combination was well tolerated in mice, with no weight loss, major organ damage, hematologic abnormalities, or metabolic changes observed in the assay.
The therapeutic effect was not limited to mouse tumor cells. In the genetically engineered mice, some tumors were significantly reduced in size and disappeared completely.
Patient-derived xenografts derived from human tumors transplanted into immunodeficient mice harboring various mutations including KRAS, TP53, SMAD family member 4 (SMAD4), and cyclin-dependent kinase inhibitor 2A (CDKN2A) all showed a significant reduction in tumor size after initiation of triple therapy.
Similar tumor responses were observed with MRTX-1133, a selective inhibitor of KRAS G12D, compared to dalaxonelasib, demonstrating that this approach is applicable to other KRAS-directed therapies. Across the models tested, dual-drug therapy was less effective than triple-drug therapy.
However, the authors highlighted several translational limitations. The dose of afatinib used in mice was much higher than doses previously tested in clinical trials for pancreatic cancer, raising tolerability concerns, but SD36 does not yet have optimal pharmacological properties for clinical trials.
Multi-target pancreatic cancer strategy
Researchers showed that pancreatic tumors rely on an interconnected KRAS, EGFR, and STAT3 signaling network for survival. By blocking just one or two pathways, cancer cells can adapt and develop resistance. In contrast, simultaneous inhibition of all three pathways resulted in durable tumor regression and prevented recurrence across multiple experimental models, including human tumor-derived xenografts. The combination of daraxonelasib, afatinib, and SD36 achieved long-term tumor control with an acceptable safety profile in preclinical mouse models.
These results provide strong evidence that a multitargeted therapeutic approach may provide a promising new strategy to help improve outcomes for patients with pancreatic ductal adenocarcinoma, if a clinically relevant and tolerable version of the regimen can be developed.
This discovery remains in the preclinical stage and requires further drug development and clinical validation. Further research is needed to identify combinations of treatments that can overcome resistance while remaining safe and effective for patients.
The authors note that an earlier version of this study has been retracted and the current article is a revised version of that study.
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Reference magazines:
- Liacchia, V., Balambana, S., Kostopoulou, M., Lechuga, C.G., Zamorano-Dominguez, E., Acosta-Gallego, D., Morales-Cacho, L., Alvarez, R., San, P., Rosas-Perez, B., Valero, R., Jiménez-Parrado, S., López-Garcia, A., San. Roman, M., López-Gil, J.C., Drosten, M., Sainz, B., Jr., Musteanu, M., Callas, E., Doucetti, N., Poli, V., Sánchez-Bueno, F., Guerra, C., Barbacid, M. (2026). Targeted combination therapy achieves effective regression of pancreatic cancer and prevents tumor resistance. Annual Report of the National Academy of Sciences of the United States of America. 123(23). Doi: 10.1073/pnas.2610708123, https://www.pnas.org/doi/10.1073/pnas.2610708123
