Abstract PR011: KRAS amplification creates a targetable pMHC antigen for T cell engager therapy to overcome KRAS inhibitor resistance

Abstract KRAS amplification (amp) has emerged as a shared mechanism of resistance to KRAS-targeted therapies, including KRASG12C and KRAS-multi small molecule inhibitors. Wild-type KRAS allele amplification also defines a subset of aggressive gastroesophageal cancers with poor outcomes. To exploit this alteration, we sought a mutation-agnostic, HLA-A*02:01–restricted KRAS pMHC antigen selectively presented in KRAS-amplified tumors. Using peptide prediction and targeted immunopeptidomics, we identified the amplified antigen pMHC pKA-A2, enabling a strategy to selectively target KRAS-amplified tumors with a bispecific pMHC×CD3 T cell engager (TCE). Quantitative immunopeptidomics across multiple models estimated pKA-A2 at ∼102–103 copies-per-cell (CPC) in KRAS-amplified cell lines (MKN1*, CPC = 119; COR-L23*, 555; HSKTC*, 1432) and undetectable (ND) or below 10 CPC in non-amplified cell lines (A375, NCI-H661, ND; NCI-H520*, 9). A*02:01-negative lines (*) were HLA-A*02:01-engineered. KRAS pathway inhibition by small molecules further increased antigen levels: In COR-L23 cells (KRASG12V; KRAS CN=16), partial resistance to 10 nM RMC-7977 produced a 2.5× rise in surface HLA and an 8-fold increase in pKA-A2, reaching ∼4,000 CPC at 48 h. KRAS amplification thereby establishes an exploitable pMHC therapeutic window—analogous to gp100 or PRAME—that is further broadened by KRAS inhibition. Importantly, NRAS and HRAS paralogs of pKA-A2 were not detected in any model. Consistent with this, stability profiling showed pKA-A2 to be markedly more stable at 37 °C (t½ ≈ 45 min) than NRAS/HRAS peptides (t½ < 5 min), supporting pKA-A2 as a selective TCE target with minimal paralog cross-reactivity. Screening our ultra-large human naïve antibody library with phage/yeast display and stringent counter-selection yielded the TCR-mimicking antibody PK313, exhibiting high affinity (KD = 5.3 nM) and strict specificity confirmed by SPR as well as by X-scan and healthy-tissue pMHC libraries (>102 and >104 peptides, respectively) with no high-affinity off-targets. A 2.8-Å cryo-EM structure revealed extensive pKA-A2-specific contacts. Reformatted as a TCE, PK313 induced potent cytotoxicity in KRAS-amplified cell line models (MKN1*, EC50 = 1.1 nM; COR-L23*, 150 pM; HSKTC*, 160 pM) and showed enhanced activity with KRAS inhibitors. Structure-guided maturation is underway to advance PK313 toward development candidate nomination. Our data demonstrate that KRAS amplification—whether intrinsic or emerging under therapeutic pressure—creates a robust, tumor-amplified pMHC antigen with exceptional potential for therapeutic exploitation. PK313 enables selective targeting of KRAS-amplified tumors as a monotherapy or in combination with KRAS small molecule inhibitors to enhance treatment durability and provides a therapeutic strategy for A*02:01 patients (∼42% in US population), who are currently excluded from A*03/A*11 KRAS-mutant neoantigen TCR-T/CAR-T/TCE programs. Citation Format: Lorenzo Maso, Diamond N. Mensah, Angelina Pizzo, Sergio A. Rodriguez-Aponte, Samantha Sze, Sean T. Toenjes, Paul Da Silva. Da Silva Jardine, Christoph Rader, Lauren E. Stopfer. KRAS amplification creates a targetable pMHC antigen for T cell engager therapy to overcome KRAS inhibitor resistance [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: RAS Oncogenesis and Therapeutics; 2026 Mar 5-8; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(5_Suppl_1):Abstract nr PR011.