Abstract Background: STK-012 is a first-in-class α/β-IL-2R biased partial agonist that drives antitumor activity by selectively stimulating CD25+ antigen-activated T-cells and avoids hallmark IL-2 toxicities by sparing pleiotropic activation of lymphocytes including NK cells. In this phase 1a/b study, STK-012 is combined with standard of care pembrolizumab + chemotherapy (PCT) in 1L PD-L1 negative NSQ NSCLC where PCT alone has poor outcomes (ORR 32% and median PFS 6.2 months). Methods: 1L NSQ NSCLC subjects received STK-012 SC Q3W + PCT. Serial blood samples were analyzed for changes in cytokines in chemoluminescence assays, T cell proliferation and activation markers were analyzed in spectral flow cytometry, and T cell clonality was analyzed by TCR sequencing. Data are shown for a cohort of 22 efficacy-evaluable 1L NSCLC subjects (N=18 PD-L1<1%, N=4 PD-L1=1%) which was enriched for loss-of-function tumor suppressor gene (LoF TSG) mutations (n=11; STK11, KEAP1, SMARCA4) or mucinous histology (n=5) associated with a “cold” TME and primary immune resistance. Results: STK-012 + PCT demonstrated an ORR of 55% in all efficacy evaluable patients; 50% in patients with PD-L1<1% tumors; 55% in LoF TSG mutations; and 80% in mucinous histology. STK-012 + PCT induced sustained proliferation of CD8+ and CD4+ T cells (13X and 5x increase from baseline, respectively), proliferation of antigen activated T cells including PD-1+ CD8+ (14X) and 4-1BB+ CD8+ (14X) and re-invigoration and proliferation of previously exhausted CD39+ (11x) and TIM3+ (18x) CD8+ T cells. After the first cycle, 3.35% of the total T cell repertoire were T cell clones which were newly detected or expanded >10x (n=12). STK-012 + PCT induced key cytokines IFNγ and IL-18 characterizing an activated CD8+ T cell response and increased IP-10 which is associated with enhanced T cell trafficking into tumor tissues (13x,14x and 23x mean increase from baseline, respectively). Sustained elevation of cytokines across treatment cycles (IFNγ median peak of 71, 95, 96 pg/mL in C1, C2 and C6) was observed, supportive of the emerging durability of the combination. Induction of cytokines TNFα and IL-6 was limited (median peak 4, 6 pg/mL), consistent with limited activation of naïve T cells and NK cells and the lack of capillary leak syndrome with STK-012. Conclusions: STK-012 + PCT led to robust cytokine induction, proliferation and expansion of antigen activated T cells, proliferation of previously exhausted T cells, and remodeling of the T cell repertoire. The addition of STK-012 to PCT has the potential to overcome resistance in immune excluded populations including PD-L1<1% tumors, LoF TSG mutations and mucinous histology. A global, randomized Phase 2 study, SYNERGY-101, of STK-012 + PCT vs. PCT in 1L PD-L1<1% NSQ NSCLC is ongoing (NCT05098132). Citation Format: Salman Punekar, Adam Jacob Schoenfeld, Edward B. Garon, So Yeon Kim, Kai He, Jennifer Marks, Brian S. Henick, Stephen V. Liu, Nagashree Seetharamu, Alexander Spira, Justin Gainor, Tim Larson, Ticiana A. Leal, Cameron Oswalt, Benjamin Izar, Alize Marangoz-Stager, Vic Danh, Grace Lunardi, Krystle Bach, Naiyer A. Rizvi, Alex Azrilevich, Anita Mehta-Damani, Martin Oft. Selective immune activation of antigen activated T cells with STK-012, an a/b IL-2 receptor biased partial agonist, with pembrolizumab and chemotherapy in 1L PD-L1 negative non-squamous NSCLC [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 6740.

PURPOSE:Cergutuzumab amunaleukin (CA) is an immunocytokine comprising a variant form of interleukin 2 (IL2) [constructed to avoid CD25 binding and regulatory T-cell (Treg) stimulation] fused to a carcinoembryonic antigen (CEA)-targeted antibody. This phase Ib open-label, multicenter dose-escalation and -expansion study (NCT02350673) evaluated the safety, activity, pharmacokinetics, and pharmacodynamics of CA plus atezolizumab in patients with advanced/metastatic CEA-positive solid tumors. PATIENTS AND METHODS:Patients received escalating doses of CA (6-20/25 mg) with fixed dosages of atezolizumab (840 mg) every 2 weeks or escalating dosages of CA weekly (10-15/20 mg) with fixed dosages of atezolizumab (1,200 mg) every 3 weeks. Primary objectives include maximum tolerated dose (MTD), recommended dose for expansion (RDE), and safety. RESULTS:Twenty-four patients were randomized to receive CA plus atezolizumab every 2 weeks and 45 patients to CA weekly plus atezolizumab every 3 weeks. A subgroup of patients (n = 5) received obinutuzumab before treatment to study the prevention of antidrug antibodies. The MTD was not determined; 15 mg weekly or 20 mg every 2 weeks of CA plus atezolizumab was the RDE. The safety profile was consistent with CA monotherapy and atezolizumab-based therapies. The addition of atezolizumab did not affect the pharmacokinetic profile of CA, and treatment induced the proliferation of T and NK cells in the blood without Treg expansion. Increases in pharmacodynamic markers (C-reactive protein, lymphocytes, sCD25, and cytokines) suggested immune activation despite limited antitumor activity (overall response rate: 13.5% with weekly/every-3-week regimen). CONCLUSIONS:The safety profile of this combination was manageable. Prominent pharmacodynamic effects were elucidated; antitumor activity was limited.

Brain metastases frequently develop in patients with non-small cell lung cancer (NSCLC) and are a common cause of cancer-related deaths, yet our understanding of the underlying human biology is limited. Here we performed multimodal single-nucleus RNA and T cell receptor, single-cell spatial and whole-genome sequencing of brain metastases and primary tumors of patients with treatment-naive NSCLC. Chromosomal instability (CIN) is a distinguishing genomic feature of brain metastases compared with primary tumors, which we validated through integrated analysis of molecular profiling and clinical data in 4,869 independent patients, and a new cohort of 12,275 patients with NSCLC. Unbiased analyses revealed transcriptional neural-like programs that strongly enriched in cancer cells from brain metastases, including a recurring, CINhigh cell subpopulation that preexists in primary tumors but strongly enriched in brain metastases, which was also recovered in matched single-cell spatial transcriptomics. Using multiplexed immunofluorescence in an independent cohort of treatment-naive pairs of primary tumors and brain metastases from the same patients with NSCLC, we validated genomic and tumor-microenvironmental findings and identified a cancer cell population characterized by neural features strongly enriched in brain metastases. This comprehensive analysis provides insights into human NSCLC brain metastasis biology and serves as an important resource for additional discovery.

2656 Background: New immuno-oncology (IO) agents are commonly used in patients who previously received PD-(L)1 inhibitors. In order to facilitate clinical trial interpretation and better delineate therapeutic contributions of novel IO agents, consensus definitions of PD-(L)1 single-agent and combination immunotherapy resistance were published in 2020 (PMC7174063) and 2022 (PMC10016305) by the Society of Immunotherapy of Cancer (SITC); definitions outlined in Table. Validation of these expert-derived definitions is currently lacking. Herein we analyze two SWOG trials to evaluate the proposed definitions for the advanced and adjuvant settings. Methods: S1609/DART (NCT02834013) was a basket trial for patients with rare cancers treated with ipilimumab (1mg/kg intravenously [IV] every 6 weeks) plus nivolumab (240mg IV every 2 weeks). S1404 (NCT02506153) included an arm where patients with high-risk resectable Stage III melanoma received adjuvant pembrolizumab (200mg IV every 3 weeks for 1 year). In both trials, overall survival (OS) was measured from study registration to death from any cause with those last known to be alive censored. OS was evaluated with Kaplan-Meier and martingale residual plots and Cox regression models. Results: In S1609 (advanced setting), 733 participants were analyzed: 127 (17%) were not evaluable for primary resistance due to death or off treatment before 6 weeks. Among the 570 evaluable, 366 met the SITC primary resistance definition and 204 did not. Martingale residuals plots indicated a positive association between time to progression and OS with no evidence of a threshold. With a 6-month landmark, participants with primary resistance had significantly shorter OS compared to those who did not: hazard ratio (HR)=2.84, 95% confidence interval (CI) 2.28-3.55, p<0.001. In S1404 (adjuvant setting), 626 participants were analyzed with 12 (2%) meeting the definition of early recurrence/primary resistance and 138 (22%) meeting the definition of late recurrence/secondary resistance. Using a 12-month landmark, there was no significant difference in OS between early and late recurrences (HR=0.98, 95% CI:0.35-2.70, p=0.25), however late recurrences were associated with significantly shorter OS than no recurrence (HR=7.69, 95% CI:2.71-20.1,p<0.001). Conclusions: In the advanced cancer cohort, the SITC definitions were validated. In the adjuvant cohort, early recurrences were uncommon and there was no significant difference in OS between early and late recurrences suggesting a 12-month cutoff may be more appropriate than 12 weeks. Additional analyses in other patient cohorts are needed to further understand if the SITC definitions for advanced cancers validate more broadly and if data-drive refinements are needed for the adjuvant setting. Advanced Primary Treatment > 6 weeks; no response or < 6 months Secondary Treatment > 6 months; response/stable > 6 months Adjuvant Early/primary < 12 weeks last dose Late/secondary 12 weeks

For patients with advanced non-small-cell lung cancer (NSCLC), dual immune checkpoint blockade (ICB) with CTLA4 inhibitors and PD-1 or PD-L1 inhibitors (hereafter, PD-(L)1 inhibitors) is associated with higher rates of anti-tumour activity and immune-related toxicities, when compared with treatment with PD-(L)1 inhibitors alone. However, there are currently no validated biomarkers to identify which patients will benefit from dual ICB1,2. Here we show that patients with NSCLC who have mutations in the STK11 and/or KEAP1 tumour suppressor genes derived clinical benefit from dual ICB with the PD-L1 inhibitor durvalumab and the CTLA4 inhibitor tremelimumab, but not from durvalumab alone, when added to chemotherapy in the randomized phase III POSEIDON trial(3). Unbiased genetic screens identified loss of both of these tumour suppressor genes as independent drivers of resistance to PD-(L)1 inhibition, and showed that loss of Keap1 was the strongest genomic predictor of dual ICB efficacy-a finding that was confirmed in several mouse models of Kras-driven NSCLC. In both mouse models and patients, KEAP1 and STK11 alterations were associated with an adverse tumour microenvironment, which was characterized by a preponderance of suppressive myeloid cells and the depletion of CD8(+) cytotoxic T cells, but relative sparing of CD4(+) effector subsets. Dual ICB potently engaged CD4(+) effector cells and reprogrammed the tumour myeloid cell compartment towards inducible nitric oxide synthase (iNOS)-expressing tumoricidal phenotypes that-together with CD4(+) and CD8(+) T cells-contributed to anti-tumour efficacy. These data support the use of chemo-immunotherapy with dual ICB to mitigate resistance to PD-(L)1 inhibition in patients with NSCLC who have STK11 and/or KEAP1 alterations.

Abstract Background: STK-012 is a first-in-class α/β-IL-2R biased partial agonist designed to drive antitumor activity by selectively stimulating CD25+ antigen activated T cells, while avoiding hallmark IL-2 toxicities by sparing pleotropic activation of lymphocytes including NK cells. Methods: STK-012-101 is a Phase 1a/b study of STK-012 administered subcutaneously in subjects with advanced, relapsed/refractory (r/r) solid tumors. During Phase 1a, subjects are enrolled in a 3+3 dose escalation to STK-012 as monotherapy (QW or Q3W) or STK-012 + pembrolizumab (Q3W) followed by Phase 1b expansions. Results: Phase 1a and preliminary Phase 1b data are being presented for STK-012 monotherapy. As of Oct 23rd, 2023, 45 subjects were treated at 7 dose levels (DL) across 2 schedules (QW at 0.375 mg and 0.75mg; Q3W at 0.75mg-3mg). The most common tumors were NSCLC (35.6%) and RCC (20%). The most common treatment related AEs (TRAEs) were maculo-papular rash (38%), injection site reactions (28.9%), fatigue (28.9%), and nausea (24.4%). Grade 3 TRAEs occurred in 12 subjects (26.6%) and included maculo-papular rash (4), vomiting (2), nausea (1), diarrhea (1), enterocolitis (1), arthralgia (1), urticaria (1), periorbital edema (1), blood creatinine increase (1) and leukocytosis (1). One subject had a Grade 4 TRAE of anaphylaxis in Cycle 9 after the data extract date. TRAEs were reversible, and no subjects had Grade 5 TRAEs. No subjects had capillary leak syndrome and <5% experienced other IL-2 hallmark TRAEs (hypotension [4.4%], AST/ALT increase [0%], pyrexia [4.4%], peripheral edema [4.4%]). No DLTs were observed. The maximum administered dose of STK-012 was 3mg Q3W. A DL of 2.25mg Q3W was advanced into Phase 1b based on safety, pharmacokinetic (PK) and pharmacodynamic (PD) data. Peripheral PD data showed STAT5 phosphorylation in CD25+ T cells and a dose dependent increase in activated T cells (Ki-67+CD38+CD8+) and serum IFNγ. Expansion of NK cells and Tregs was limited. STK-012 has a half-life of 4 days. Of 38 efficacy evaluable subjects, 3 had a partial response (PR) and 17 had stable disease as best overall response by RECIST V1.1. The PRs included 2 confirmed (anti-PD-1 r/r NSCLC and RCC) and 1 unconfirmed (anti-PD-1 r/r SCCHN) with the RCC subject in response for over 9 months with treatment ongoing. After the data extract date, a fourth subject achieved a PR (confirmed) with 80% reduction in target lesions (anti-PD-1/CTLA-4 and TKI r/r RCC) with treatment ongoing. Monotherapy Phase 1b dose expansions in r/r NSCLC and r/r RCC are ongoing. Conclusions: In our ongoing Phase 1a/b trial, STK-012 demonstrated a favorable safety, PK, and PD profile which is distinct from that of aldesleukin and non-α IL-2 analogues. Peripheral PD and PK data support selectivity for IL-2R α/β. Preliminary monotherapy efficacy in subjects who are r/r to prior immunotherapy warrants further development. Citation Format: Benjamin Izar, Dmitriy Zamarin, David R. Spigel, Christopher J. Hoimes, David F. McDermott, Kartik Sehgal, Yana G. Najjar, Adam J. Schoenfeld, Edward B. Garon, Ryan J. Sullivan, Brian S. Henick, Ticiana A. Leal, Michael E. Hurwitz, Rana R. McKay, Natalie Busby, Anita Mehta-Damani, Alex Azrilevich, Tony Tran, Naiyer Rizvi, Martin Oft, Alexander I. Spira. Initial results from a phase 1a/1b study of STK-012, a first-in-class α/β IL-2 receptor biased partial agonist in advanced solid tumors (NCT05098132) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr CT183.

Neoantigen immunoediting drives immune checkpoint blockade efficacy, yet the molecular features of neoantigens and how neoantigen immunogenicity shapes treatment response remain poorly understood. To address these questions, 80 patients with non-small cell lung cancer were enrolled in the biomarker cohort of CheckMate 153 (CA209-153), which collected radiographic guided biopsy samples before treatment and during treatment with nivolumab. Early loss of mutations and neoantigens during therapy are both associated with clinical benefit. We examined 1,453 candidate neoantigens, including many of which that had reduced cancer cell fraction after treatment with nivolumab, and identified 196 neopeptides that were recognized by T cells. Mapping these neoantigens to clonal dynamics, evolutionary trajectories and clinical response revealed a strong selection against immunogenic neoantigen-harboring clones. We identified position-specific amino acid and physiochemical features related to immunogenicity and developed an immunogenicity score. Nivolumab-induced microenvironmental evolution in non-small cell lung cancer shared some similarities with melanoma, yet critical differences were apparent. This study provides unprecedented molecular portraits of neoantigen landscapes underlying nivolumab's mechanism of action. In this biomarker cohort analysis of CheckMate 153, analyses of genomic alterations and neoepitope immunogenicity in tumor tissue samples from patients with non-small cell lung cancer treated with nivolumab show the evolution of neoantigens during nivolumab-induced selection pressure and how it associates with clinical response.

Introduction Neoadjuvant chemoimmunotherapy has achieved overall survival (OS) benefit for patients with resectable non-small cell lung cancer (NSCLC). Here, we present outcomes after 3 years of follow-up from the first reported study of neoadjuvant atezolizumab+chemotherapy.Methods This open-label, multicenter single-arm investigator-initiated phase II study conducted at three US hospitals tested up to four cycles of atezolizumab, carboplatin, and nab-paclitaxel prior to surgery. Major pathological response (MPR, primary endpoint) was previously reported; here, we report 3-year disease-free survival (DFS), OS, and clinical characteristics of patients developing brain metastases (BM) with integrated data from tumor genomics, gene expression, and quantitative immunofluorescent measurement of immune markers.Results Of 30 enrolled patients, 29 were taken to the operating room. 26 underwent R0 resection, with 17 experiencing MPR (10 pCR). With a median follow-up of 39.5 months, the median OS was 55.8 months, and the median DFS was 34.5 months. Landmark OS at 36 months was 77%. Among 14 patients with recurrent disease, 6 patients had BM. Patients whose tumors had mutations in STK11 and KEAP1 did not have a significantly higher incidence of BM. Reduced copy number of STK11 and KEAP1, both residing on chromosome 19p, was observed in ~1/3 of tumors. Reduced CN of STK11 was significantly associated with worse pathological response and incidence of BM.Conclusions Consistent with recent phase III studies, 3-year OS data with neoadjuvant atezolizumab+chemotherapy was associated with prolonged PFS and OS. Establishing associations between STK11 and KEAP1 genomic alterations and key clinical outcomes in early-stage NSCLC requires further study.

Background: STK-012 is a first-in-class α/β-IL-2R biased partial agonist designed to drive antitumor activity by selectively stimulating CD25+ antigen activated T cells, while avoiding hallmark IL-2 toxicities by sparing pleotropic activation of lymphocytes including NK cells. Methods: STK-012-101 is a Phase 1a/b study of STK-012 administered subcutaneously in subjects with advanced, relapsed/refractory (r/r) solid tumors. During Phase 1a, subjects are enrolled in a 3+3 dose escalation to STK-012 as monotherapy (QW or Q3W) or STK-012 + pembrolizumab (Q3W) followed by Phase 1b expansions. Results: Phase 1a and preliminary Phase 1b data are being presented for STK-012 monotherapy. As of Oct 23rd, 2023, 45 subjects were treated at 7 dose levels (DL) across 2 schedules (QW at 0.375 mg and 0.75mg; Q3W at 0.75mg-3mg). The most common tumors were NSCLC (35.6%) and RCC (20%). The most common treatment related AEs (TRAEs) were maculo-papular rash (38%), injection site reactions (28.9%), fatigue (28.9%), and nausea (24.4%). Grade 3 TRAEs occurred in 12 subjects (26.6%) and included maculo-papular rash (4), vomiting (2), nausea (1), diarrhea (1), enterocolitis (1), arthralgia (1), urticaria (1), periorbital edema (1), blood creatinine increase (1) and leukocytosis (1). One subject had a Grade 4 TRAE of anaphylaxis in Cycle 9 after the data extract date. TRAEs were reversible, and no subjects had Grade 5 TRAEs. No subjects had capillary leak syndrome and <5% experienced other IL-2 hallmark TRAEs (hypotension [4.4%], AST/ALT increase [0%], pyrexia [4.4%], peripheral edema [4.4%]). No DLTs were observed. The maximum administered dose of STK-012 was 3mg Q3W. A DL of 2.25mg Q3W was advanced into Phase 1b based on safety, pharmacokinetic (PK) and pharmacodynamic (PD) data. Peripheral PD data showed STAT5 phosphorylation in CD25+ T cells and a dose dependent increase in activated T cells (Ki-67+CD38+CD8+) and serum IFNγ. Expansion of NK cells and Tregs was limited. STK-012 has a half-life of 4 days. Of 38 efficacy evaluable subjects, 3 had a partial response (PR) and 17 had stable disease as best overall response by RECIST V1.1. The PRs included 2 confirmed (anti-PD-1 r/r NSCLC and RCC) and 1 unconfirmed (anti-PD-1 r/r SCCHN) with the RCC subject in response for over 9 months with treatment ongoing. After the data extract date, a fourth subject achieved a PR (confirmed) with 80% reduction in target lesions (anti-PD-1/CTLA-4 and TKI r/r RCC) with treatment ongoing. Monotherapy Phase 1b dose expansions in r/r NSCLC and r/r RCC are ongoing. Conclusions: In our ongoing Phase 1a/b trial, STK-012 demonstrated a favorable safety, PK, and PD profile which is distinct from that of aldesleukin and non-α IL-2 analogues. Peripheral PD and PK data support selectivity for IL-2R α/β. Preliminary monotherapy efficacy in subjects who are r/r to prior immunotherapy warrants further development. Citation Format: Benjamin Izar, Dmitriy Zamarin, David R. Spigel, Christopher J. Hoimes, David F. McDermott, Kartik Sehgal, Yana G. Najjar, Adam J. Schoenfeld, Edward B. Garon, Ryan J. Sullivan, Brian S. Henick, Ticiana A. Leal, Michael E. Hurwitz, Rana R. McKay, Natalie Busby, Anita Mehta-Damani, Alex Azrilevich, Tony Tran, Naiyer Rizvi, Martin Oft, Alexander I. Spira. Initial results from a phase 1a/1b study of STK-012, a first-in-class α/β IL-2 receptor biased partial agonist in advanced solid tumors (NCT05098132) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr CT183.

Supplementary Table 1. Mutation hotspots interrogated by Sequenom.

PDF file - 308K, Complete list of target sequences for NanoString probes

PDF file - 76K, Examples of negative results with the NanoString assay for kinase fusions.

Effects of hybridization on HLA baiting efficiency