Alvaro Guimaraes Paula

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 trial3. 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. Alterations in the tumour suppressor genes STK11 and/or KEAP1 can identify patients with advanced non-small-cell lung cancer who are likely to benefit from combinations of PD-(L)1 and CTLA4 immune checkpoint inhibitors added to chemotherapy.

KRAS mutations frequently co-occur with alterations in STK11/LKB1 and/or KEAP1, defining an aggressive subset of lung cancers resistant to immuno- and chemotherapy. While LKB1 loss is associated with vulnerability to DNA damage response-based therapies, the impact of KEAP1 alterations remains unknown. We demonstrate that KEAP1-NRF2 pathway drives a compensatory modulation of ATR-CHK1 signaling, enhancing vulnerability to ATR inhibitors (ATRi), particularly in the setting of increased replication stress associated with LKB1 loss. ATRi shows enhanced anti-tumor activity in LKB1 and/or KEAP1-deficient non-small cell lung cancer (NSCLC) models and synergizes with gemcitabine. ATRi also enhances antitumor immunity and mitigates the immunosuppressed phenotype of LKB1/KEAP1-deficient tumors. In the HUDSON trial, LKB1/KEAP1-deficient NSCLC patients demonstrate enhanced benefits to the ATRi ceralasertib plus durvalumab. These findings suggest that alterations in the KEAP1-NRF2 pathway and/or LKB1 are associated with enhanced sensitivity to ATRi and could serve as biomarkers for predicting response to ATRi combination regimens.

e20694 Background: Selective RET tyrosine kinase inhibitors (RETi) revolutionized the treatment of patients (pts) with advanced NSCLC harboring RET fusions and is broadly considered first-line standard of care treatment. Chylous effusions have emerged as a RETi-associated adverse event (AE) and are challenging to distinguish from disease progression. Given the rarity of this AE, a greater appreciation of incidence is needed. Methods: The Genomic Marker-guided Therapy Initiative (GEMINI) database at MD Anderson Cancer Center was used to identify patients with RET fusion-positive NSCLC between 2011 and 2024. Pts were included in chylous effusion incidence analyses if they received at least one month of treatment and had a confirmed diagnosis of chylous effusion, based on both radiographic imaging and body fluids sampling, requiring medical intervention. We defined the time to chylous effusion diagnosis as the interval between the treatment start date and the chylous effusion collection date. This study protocol was approved by the MD Anderson Institutional Review Board. Results: We identified 103 pts with RET fusion-positive NSCLC, of whom 73 received a RETi. Among these, we identified 12% of pts (n=9) who developed a chylous effusion: 14% (n= 7 of 51) and 13% (n= 2 of 15) were treated with selpercatinib or pralsetinib, respectively. Additionally, 7 pts received both RETi, and none of them developed a chylous effusion. Among pts who developed a chylous effusion, the mean age was 64 years old, 88% were White, and all pts had either never-smoking or light-smoking (≤ 10 pack-years) tobacco histories. Eight of nine pts had adenocarcinoma histology, and of the 9 pts fully characterized fusions, KIF5B was the predominant RET fusion partner (n= 5). Of the nine pts with chylous effusions, two had chylopericardium, two had chylous ascites, one had chylothorax and four had both chylous ascites and chylothorax. Triglyceride levels across pts ranged from 478mg/dl to >5200mg/dl. The mean time from RETi initiation to chylous effusion diagnosis was 17 months (range: 1 to 60 months). All pts underwent fluid drainage for symptom relief, 2 of 10 successfully underwent lymphangiography and embolization of the chylous leak, and one patient was successfully treated with pleurodesis. Additionally, 3 of 9 pts required a RETi dose reduction, which one had improvement and two had no effect on the effusion. One patient switched from selpercatinib to pralsertinib which did not affect the effusion. No cases of drug discontinuation due to these AEs were reported. Conclusions: We identified chylous effusion as an AE following RETi exposure in 12% of pts in our cohort, including 2 cases of chylopericardium. These adverse events can occur with either selpercatinib or pralsetinib, and making a differential diagnosis, including ruling out disease progression, is crucial for effective management.

Abstract Background: KEAP1 is one of the most commonly mutated tumor suppressors in lung adenocarcinoma and is frequently co-altered with KRAS and STK11/LKB1 mutations. Mechanistically, KEAP1 mutations impair NRF2 degradation, resulting in the upregulation of antioxidant and ferroptosis response genes. Non-small cell lung cancer (NSCLC) tumors with KEAP1 mutations are resistant to various treatment modalities, including KRAS G12C inhibitors. KRAS inhibitors are changing the treatment landscape for several malignancies, including NSCLC, and understanding mechanisms of resistance to these agents is of utmost importance. Here, we examined the effects of the ferroptosis-resistant phenotype of KEAP1 mutation on sensitivity to KRAS inhibitors in KEAP1/KRAS-mutant lung adenocarcinoma. Methods: DepMap was used to assess the drug sensitivity profiles for sotorasib, MRTX1133, erastin, RSL3, ML162, and ML210 as well as RNAi and CRISPR screen data in human NSCLC cell lines. The Cancer Therapeutics Response Portal database was used to create a NSCLC ferroptosis resistance gene signature by identifying the genes with Pearson correlation z-score of >3 with resistance to both RSL3 and erastin. Kras MUT (K), Kras MUT/Keap1 KO (KK), Kras MUT/Lkb1 KO (KL), and Kras MUT/Lkb1 KO/Keap1 KO (KLK) tumor cells were created by knocking-out Keap1 and Stk11 from LKR13 KRAS G12C and KRAS G12D mutant murine cell lines. BODIPY™ 581/591 C11 was used to quantify lipid peroxidation by flow cytometry. Results: Using murine and human models, we found that KEAP1 deficiency promoted resistance to ferroptosis inducers (erastin, RSL3, ML162, ML210) as well as to KRAS G12C (sotorasib, adagrasib) and KRAS G12D (MRTX1133) inhibitors. MRTX1133 and adagrasib induced lipid peroxidation -a hallmark of ferroptosis- in LKR13 K G12D and G12C cell lines. RSL3 and erastin synergized with adagrasib and MRTX1133 in KEAP1-wt, but not KEAP1-mutant, LKR13 G12C and G12D models. Through RNAi and CRISPR screens conducted in a large panel of human KRAS-mutant NSCLC cell lines, we found that KEAP1 mutation and a higher NSCLC ferroptosis resistance signature also correlated with diminished sensitivity to genetic disruption of KRAS. Glutaminase 1 (GLS1) inhibition can deplete glutathione pools necessary for antioxidant response. While RSL3 alone could not effectively induce lipid peroxidation in the KEAP1-mutant LKR13 KLK G12D model, combining it with the GLS1 inhibitor CB-839 resulted in an increased lipid peroxidation in the KLK, but not KEAP1-wt, model and this approach increased the effectiveness of RSL3 treatment in KEAP1-mutant samples. Finally, CB-839 treatment enhanced the effectiveness of adagrasib and MRTX1133 in LKR13 KK and KLK G12C and G12D cell lines. Conclusions: KRAS inhibitors induce ferroptosis in KRAS-mutant lung adenocarcinoma cell lines. KEAP1 mutation and enhanced anti-ferroptosis response correlated with lack of response to KRAS inhibition. Our data indicates that GLS1 blockade could sensitize KEAP1-mutant lung adenocarcinoma to ferroptosis and KRAS inhibitors. Citation Format: Amirali Karimi, Yu Qian, David Molkentine, Ana Galan Cobo, Alvaro Guimaraes Paula, Büsra Ernhofer, David Peng, Monique Nilsson, John V. Heymach. Reversing the ferroptosis-resistant phenotype of KEAP1-mutant lung adenocarcinoma through glutaminase 1 inhibition enhances the efficacy of KRAS inhibitors [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 B013.