Abstract Background: NRG1-fusion positive (NRG1+) cancers define a new molecular subtype of solid tumors. NRG1 chimeric proteins signal through ERRB3/ERRB2 dimerization, activating PI3K/AKT and MAPK signalling pathways, promoting pro-tumoral properties. NRG1+ cancers targeting relies on bispecific anti-ERBB2/ERBB3 antibodies such as zenocutuzumab (zeno) or ERBB2 inhibitors such as afatinib (afa). These therapies have limited clinical efficacy, suggesting that the biology of the NRG1/ERBB3 pathway is not totally understood. Our objective is to understand the mechanisms of resistance to NRG1-directed therapies in NRG1+ cancers. Material and Methods: We have generated afa-resistant models by continuous exposure to increasing concentrations of afa, using MDA-MB-175 cells. They grew in presence of 256 nM afatinib. Afa-sensitivity was assessed by Prestoblue test. MDA-MB-175 cells exhibit an endogenous PPP6R3-TENM4-NRG1 fusion and are sensitive to afa, zeno and other NRG1-directed therapy. Bulk RNA sequencing data from afa-naive and afa-resistant MDA-MB-175 were subjected to unsupervised and comparative analysis. Characterization of these models was completed by quantitative RT-PCR and western blotting. Results: Five afa-resistant MDA-MB-175 models were generated, showing 10-to-100-fold increase in IC50 as compared to sensitive cells. RNA-seq data unsupervised analysis showed that afa-resistant (n=5) and afa-naive (n=5) MDA-MB-175 formed two distinct clusters. ALPP was one of the ten most differentially expressed genes in afa-resistant models (log2FoldChange = 8.1, p = 2.46.10-20) and the only one with a confirmed lack of expression in afa-naive and expression in afa-resistant MDA-MB-175 at the RNA and protein levels. In afa-resistant models, ALPP expression disappeared after 2-8 weeks when cultured without afa. In afa-naive and afa-resistant MDA-MB-175, an increase of ALPP expression was observed after one or two days of afa exposure. Investigation of NRG1-ERBB3 pathway in afa-resistant models showed a decrease of phospho-ERBB3 and phospho-ERK with no variation of total ERBB3 and ERK. Conclusion: Afa induced early ALPP expression in NRG1+ MDA-MB-175 cells, and afa-resistance was associated with high ALPP expression and a decrease of ERBB3 and ERK phosphorylation. Validation of ALPP as a relevant target is ongoing using ALPP KO and overexpressing NRG1+ models, including NRG1+ engineered preclinical models and NRG1+ patient-derived models from patients exposed to NRG1-directed therapies. Zeno and zongertinib impact on ALPP expression in preclinical models as well as ALPP expression in a cohort of NRG1+ non-small cell lung cancer and pancreatic ductal adenocarcinoma is ongoing. Overall, ALPP may represent an attractive target in NRG1+ cancers that may prevent resistance to NRG1-directed therapies. Citation Format: Manon Barre, Clarisse Thiollier-Schmitt, Marie Issenmann, Célia Cortay, Emeline Cros-Perrial, Sandra Ortiz-Cuaran, Nicolas Gadot, Elodie Voilin, Sylvie Lantuejoul, Anthony Ferrari, Eric Cumunel, Charles Dumontet, Lars Petter Jordheim, Michael Duruisseaux. Neuregulin-1(NRG1)-directed therapies induce alkaline phosphatase placental (ALPP) expression in NRG1-fusion positive cancers: Preclinical data and therapeutic perspective abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 7048.
Barré et al. (Fri,) studied this question.