Profiling of reversible and covalent HER2 kinase inhibitors for anti-tumor activity and broader biological effects to evaluate selectivity and functional activity

Hurman epidermal growth factor receptor 2 (HER2) is a member of the human epidermal growth factor receptor tyrosine kinase family that regulates cell proliferation, survival, and differentiation. Amplification or activating mutations of HER2 result in constitutive signaling through pathways such as PI3K/AKT and MAPK, driving oncogenic transformation and tumor progression in various cancers. This is particularly notable in breast and gastric cancers, making it an important therapeutic target for the treatment of tumors in these tissues. HER2-directed therapies, including monoclonal antibodies, antibody-drug conjugates, and small molecule tyrosine kinase inhibitors (TKIs), have markedly improved outcomes for HER2-positive malignancies. Among TKIs, lapatinib was the first dual HER2/EGFR inhibitor to be approved for clinical use. Its reversible inhibition of both kinases leads to broad pathway inhibition but also to off-target effects, limiting its use. Tucatinib, a third-generation reversible HER2 TKI, was designed for enhanced selectivity towards HER2 with minimal EGFR inhibition, reducing toxicity while preserving efficacy in HER2-driven tumors. Furthermore, tucatinib demonstrates clinical benefit in combination treatments, particularly for HER2-positive breast cancer with brain metastases, due to favorable CNS penetration. Zongertinib is a newly approved next-generation, highly selective covalent HER2 inhibitor with optimized pharmacologic and safety profiles. As such, it retains potent activity against HER2-amplified and HER2-mutant tumors while minimizing off-target kinase effects, which translates into improved tolerability and the potential for broader therapeutic use. We evaluated the activity and selectivity of lapatinib, tucatinib, and zongertinib in vitro across a panel of about 300 human tumor cell lines using the OncoPanel ® cellular phenotypic platform. Potencies were determined from ten-point dose-response curves to calculate IC 50 and EC 50 values, thus allowing the identification of similarities and differences in potency, efficacy, and genomic biomarkers of sensitivity and resistance relating to each drug. Activities are also being assessed in a human tumor xenograft model of the HER2-positive NCI-N87 human gastric cancer cell line, to further enable the comparison and contrasting of these drugs in vivo . To complement these in vitro and in vivo analyses, these HER2 inhibitors were further characterized in the BioMAP ® Diversity PLUS ® panel of primary human cell systems to assess functional activities across a diverse range of tissue and immune biology contexts. Together, these studies define distinct biological and mechanistic profiles among HER2-targeted TKIs and highlight the evolution of this drug class with increasing selectivity, tolerability, and efficacy.