Abstract 1729: Mutagenesis screens support potential best-in-class profile for neladalkib (NVL-655), a brain-penetrant and TRK-sparing ALK inhibitor

Abstract Introduction. ALK fusions drive ∼5% of non-small cell lung cancer (NSCLC). Three generations of ALK tyrosine kinase inhibitors (1G, 2G, and 3G TKIs) have been approved for the treatment of ALK-positive NSCLC. However, response to these TKIs can be limited by acquired drug resistance. Single ALK mutations have been reported in >50% of patients progressing on a 2G ALK TKI, whereas compound (i.e., ≥2 cis-allelic) ALK mutations have been observed after sequential treatment with 2G-3G ALK TKIs. NVL-655 is an investigational ALK TKI designed to be brain-penetrant, spare TRK, and maintain broad activity against single and compound ALK mutations that confer resistance to previous-generation therapies. Here, in vitro N-ethyl-N-nitrosourea (ENU) mutagenesis screens further support NVL-655 as more effective than previous-generation therapies at suppressing single and compound ALK mutations. Methods. Cells were treated with ENU for 1 day followed by alectinib (2G), lorlatinib (3G), or NVL-655 for 28 days. Wells with proliferating cells were counted on day 28, indicating TKI-resistant clones. Mutations in the ALK kinase domain were identified by Sanger sequencing. Some resistant clones harboring ALK kinase-domain mutations were isolated and used to profile the potency of ALK TKIs. Results. ENU screens were performed on Ba/F3 EML4-ALK, to simulate a first-line treatment setting, and on Ba/F3 EML4-ALK G1202R, the most common point mutation observed at progression on 2G ALK TKIs, to simulate a later-line treatment setting. NVL-655 unbound concentrations in the two screens were 8.0 and 3.2 nM, respectively, both below its 14 nM average clinical exposure at the recommended phase 2 dose (RP2D). Resistant clones emerged in only 9 (<2%) and 3 (<1%) of 480 wells, respectively, suggesting that resistance to NVL-655 would be unlikely in both treatment settings. By contrast, all 480 out of 480 wells became resistant to alectinib (in Ba/F3 EML4-ALK) or lorlatinib (in Ba/F3 EML4-ALK G1202R) at clinically relevant exposures, indicating lower effectiveness than NVL-655 at suppressing resistance in these screens. Conclusion. NVL-655 was highly effective at suppressing resistance in ENU mutagenesis screens simulating first-line (EML4-ALK) and later-line (EML4-ALK G1202R) treatment. At clinically relevant concentrations, <2% of all wells became resistant to NVL-655, compared to 100% for alectinib and lorlatinib. This preclinical study indicates the potential for NVL-655 to prevent acquisition of on-target resistance and thereby provide deeper and more durable target inhibition. The broad preclinical activity against ALK and diverse ALK mutations, together with its brain penetrance and TRK-sparing design, suggests NVL-655 as a potential best-in-class ALK therapy. Citation Format: Anupong Tangpeerachaikul, Henry E. Pelish. Mutagenesis screens support potential best-in-class profile for neladalkib (NVL-655), a brain-penetrant and TRK-sparing ALK inhibitor [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 1729.