Background: Fam-trastuzumab deruxtecan-nxki (T-DXd) has shown clinical benefit in HER2-positive and HER2-low metastatic breast cancer (MBC), but resistance remains a barrier to durable response. To investigate resistance mechanisms, real-world data (RWD) from a large, clinically annotated cohort were analyzed to identify molecular correlates and potential therapeutic vulnerabilities.
Methods: A retrospective analysis of RWD from 300 MBC patients treated with T-DXd (Tempus database1) was conducted. Real-world clinical endpoints and tumor transcriptomics (Tempus xR2 RNA-seq) were integrated. Patients were stratified as responders (complete or partial response, CR/PR) or non-responders (stable or progressive disease, SD/PD) based on curated real-world best overall response (rwBOR). Criteria included: (1) SD ≥5 weeks post-treatment; (2) PR/CR sustained ≥ 4 weeks without reversion; and (3) manual adjudication of discordant cases with Eisai’s translational and clinical teams. Additional endpoints, real-world progression free survival (rwPFS), time to next treatment (rwTTNT), and overall survival (rwOS), were derived using validated Tempus algorithms. Differential gene expression (DGE) analysis was performed using Limma3, adjusting for key covariates (collection site, pre- T-DXd ER/PR status, age at treatment, time/careplan between sample collection and T-DXd start). Cox proportional hazards models were applied to survival endpoints. Gene set enrichment analysis4 was used to identify biologically relevant pathways. Analyses were stratified by HER2-status.
Results: After quality control, transcriptomic data were available for 231 patients with matched rwBOR (120 non-responders, 111 responders). Survival endpoints were evaluable in > 230 patients. DGE analysis revealed largely non-overlapping resistance-associated genes between HER2-positive and HER2-low subgroups, suggesting distinct resistance mechanisms. In HER2-low non-responders, significant enrichment (FDR q < 0.2) was observed in multiple oncogenic pathways including TNF/NF-κB, TGF-β, KRAS, Wnt/β-catenin signaling and inflammatory response – implicated in immune evasion, epithelial-mesenchymal transition (EMT), and resistance to antibody-drug conjugates. In contrast, HER2-positive non-responders showed negative enrichment of Wnt/β-catenin signaling and oxidative phosphorylation and, suggesting metabolic reprogramming. Survival analysis identified 45 genes in HER2-positive and 23 genes in the HER2-low subgroups, significantly associated with rwPFS, rwTTNT, and rwOS (q<0.2; HR > 2 or < 0.5). Notable genes included TMEM150A, DENND2D, and SLC6A14 (HER2-positive), and C1ORF61 and SERTAD1 (HER2-low), implicating roles in membrane transport, transcriptional regulation, and cellular stress responses.
Conclusions: This study represents one of the largest real-world transcriptomic analyses of resistance in T-DXd- -treated MBC. Distinct molecular signatures and pathway activations were identified in HER2-positive vs HER2-low tumors, highlighting biological heterogeneity of resistance. Enrichment of immune and EMT pathways in HER2-low non-responders suggests potential benefit from combination strategies, such as immune-checkpoint inhibitors or pathway-targeted agents. Gene-level correlates of real-world endpoints support biomarker-driven therapeutic optimization and warrant prospective validation to inform future resistance-mitigation strategies in T-DXd-treated MBC.
References:
1. www.tempus.com
2. Tempus-xR_Validation.pdf
3. Ritchie, Matthew E., et al., Nucleic acids research 43.7 e47-e47 (2015). 4. Subramanian, Aravind, et al., Proceedings of the National Academy of Sciences 102.43 15545-15550 (2005).