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Anaplastic lymphoma kinase (ALK)-rearranged non-small cell lung cancer (NSCLC) is treated with ALK tyrosine kinase inhibitors (TKIs), but the lack of activity of immune checkpoint inhibitors (ICIs) is poorly understood. Here, we identified immunogenic ALK peptides to show that ICIs induced rejection of ALK + tumors in the flank but not in the lung. A single-peptide vaccination restored priming of ALK-specific CD8 + T cells, eradicated lung tumors in combination with ALK TKIs and prevented metastatic dissemination of tumors to the brain. The poor response of ALK + NSCLC to ICIs was due to ineffective CD8 + T cell priming against ALK antigens and is circumvented through specific vaccination. Finally, we identified human ALK peptides displayed by HLA-A*02:01 and HLA-B*07:02 molecules. These peptides were immunogenic in HLA-transgenic mice and were recognized by CD8 + T cells from individuals with NSCLC, paving the way for the development of a clinical vaccine to treat ALK + NSCLC., (© 2023. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Anaplastic lymphoma kinase (ALK) tyrosine kinase inhibitors (TKIs) show potent efficacy in several ALK-driven tumors, but the development of resistance limits their long-term clinical impact. Although resistance mechanisms have been studied extensively in ALK-driven non-small cell lung cancer, they are poorly understood in ALK-driven anaplastic large cell lymphoma (ALCL). Here, we identify a survival pathway supported by the tumor microenvironment that activates phosphatidylinositol 3-kinase γ (PI3K-γ) signaling through the C-C motif chemokine receptor 7 (CCR7). We found increased PI3K signaling in patients and ALCL cell lines resistant to ALK TKIs. PI3Kγ expression was predictive of a lack of response to ALK TKI in patients with ALCL. Expression of CCR7, PI3Kγ, and PI3Kδ were up-regulated during ALK or STAT3 inhibition or degradation and a constitutively active PI3Kγ isoform cooperated with oncogenic ALK to accelerate lymphomagenesis in mice. In a three-dimensional microfluidic chip, endothelial cells that produce the CCR7 ligands CCL19/CCL21 protected ALCL cells from apoptosis induced by crizotinib. The PI3Kγ/δ inhibitor duvelisib potentiated crizotinib activity against ALCL lines and patient-derived xenografts. Furthermore, genetic deletion of CCR7 blocked the central nervous system dissemination and perivascular growth of ALCL in mice treated with crizotinib. Thus, blockade of PI3Kγ or CCR7 signaling together with ALK TKI treatment reduces primary resistance and the survival of persister lymphoma cells in ALCL.

Immunotherapy has shown limited efficacy in patients with EGFR-mutated lung cancer. Efforts to enhance the immunogenicity of EGFR-mutated lung cancer have been unsuccessful to date. Here, we discover that MET amplification, the most common mechanism of resistance to third-generation EGFR tyrosine kinase inhibitors (TKI), activates tumor cell STING, an emerging determinant of cancer immunogenicity (1). However, STING activation was restrained by ectonucleosidase CD73, which is induced in MET-amplified, EGFR-TKI-resistant cells. Systematic genomic analyses and cell line studies confirmed upregulation of CD73 in MET-amplified and MET-activated lung cancer contexts, which depends on coinduction of FOSL1. Pemetrexed (PEM), which is commonly used following EGFR-TKI treatment failure, was identified as an effective potentiator of STING-dependent TBK1-IRF3-STAT1 signaling in MET-amplified, EGFR-TKI-resistant cells. However, PEM treatment also induced adenosine production, which inhibited T-cell responsiveness. In an allogenic humanized mouse model, CD73 deletion enhanced immunogenicity of MET-amplified, EGFR-TKI-resistant cells, and PEM treatment promoted robust responses regardless of CD73 status. Using a physiologic antigen recognition model, inactivation of CD73 significantly increased antigen-specific CD8+ T-cell immunogenicity following PEM treatment. These data reveal that combined PEM and CD73 inhibition can co-opt tumor cell STING induction in TKI-resistant EGFR-mutated lung cancers and promote immunogenicity., Significance: MET amplification upregulates CD73 to suppress tumor cell STING induction and T-cell responsiveness in TKI-resistant, EGFR-mutated lung cancer, identifying a strategy to enhance immunogenicity and improve treatment., (©2022 The Authors; Published by the American Association for Cancer Research.)

KRAS-LKB1 (KL) mutant lung cancers silence STING owing to intrinsic mitochondrial dysfunction, resulting in T cell exclusion and resistance to programmed cell death (ligand) 1 (PD-[L]1) blockade. Here we discover that KL cells also minimize intracellular accumulation of 2'3'-cyclic GMP-AMP (2'3'-cGAMP) to further avoid downstream STING and STAT1 activation. An unbiased screen to co-opt this vulnerability reveals that transient MPS1 inhibition (MPS1i) potently re-engages this pathway in KL cells via micronuclei generation. This effect is markedly amplified by epigenetic de-repression of STING and only requires pulse MPS1i treatment, creating a therapeutic window compared with non-dividing cells. A single course of decitabine treatment followed by pulse MPS1i therapy restores T cell infiltration in vivo, enhances anti-PD-1 efficacy, and results in a durable response without evidence of significant toxicity., Competing Interests: Declaration of interests S.K., T.T., D.A.B., C.P.P, and P.G. are inventors on a pending patent for combination DAC and MPS1 inhibitor therapy to prime cancer immunogenicity. D.A.B. is a consultant for N of One/Qiagen and Tango Therapeutics; is a founder and shareholder in Xsphera Biosciences; has received honoraria from Merck, H3 Biomedicine/Esai, EMD Serono, Gilead Sciences, Abbvie, and Madalon Consulting; and has received research grants from BMS, Takeda, Novartis, Gilead, and Lilly. T.U.B is a consultant for N of One/Qiagen. P.C.G. has sponsored research agreements with Marengo Therapeutics, Epizyme, Daiichi Sankyo, and Foghorn Therapeutics. C.P.P is a consultant for DropWorks and Xsphera Biosciences; has stock and other ownership interests in Xsphera Biosciences; received honoraria from Bio-Rad; and has sponsored research agreements with Daiichi Sankyo, Bicycle Therapeutics, Transcenta, Bicara Therapeutics, AstraZeneca, Intellia Therapeutics, Janssen Pharmaceuticals, and Array Biopharma. S.K. has a sponsored research agreement with Boehringer-Ingelheim. E.H.K. is an employee and stockholder of Merck & Co. and previously had a Sponsored Research Agreement with Takeda Pharmaceuticals. H.K. has a Sponsored Research Agreement with Takeda Pharmaceuticals. P.A.J. has received consulting fees from AstraZeneca, Boehringer-Ingelheim, Pfizer, Roche/Genentech, Takeda Oncology, ACEA Biosciences, Eli Lilly and Company, Araxes Pharma, Ignyta, Mirati Therapeutics, Novartis, LOXO Oncology, Daiichi Sankyo, Sanofi Oncology, Voronoi, SFJ Pharmaceuticals, Takeda Oncology, Transcenta, Silicon Therapeutics, Syndax, Nuvalent, Bayer, Esai, Biocartis, Allorion Therapeutics, Accutar Biotech, and Abbvie; receives post-marketing royalties from DFCI; owned intellectual property on EGFR mutations licensed to Lab Corp; has sponsored research agreements with AstraZeneca, Daichi-Sankyo, PUMA, Boehringer Ingelheim, Eli Lilly and Company, Revolution Medicines; and Astellas Pharmaceuticals; and has stock ownership in LOXO Oncology and Gatekeeper Pharmaceuticals., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Small cell lung carcinoma (SCLC) is highly mutated, yet durable response to immune checkpoint blockade (ICB) is rare. SCLC also exhibits cellular plasticity, which could influence its immunobiology. Here we discover that a distinct subset of SCLC uniquely upregulates MHC I, enriching for durable ICB benefit. In vitro modeling confirms epigenetic recovery of MHC I in SCLC following loss of neuroendocrine differentiation, which tracks with derepression of STING. Transient EZH2 inhibition expands these nonneuroendocrine cells, which display intrinsic innate immune signaling and basally restored antigen presentation. Consistent with these findings, murine nonneuroendocrine SCLC tumors are rejected in a syngeneic model, with clonal expansion of immunodominant effector CD8 T cells. Therapeutically, EZH2 inhibition followed by STING agonism enhances T-cell recognition and rejection of SCLC in mice. Together, these data identify MHC I as a novel biomarker of SCLC immune responsiveness and suggest novel immunotherapeutic approaches to co-opt SCLC's intrinsic immunogenicity. SIGNIFICANCE: SCLC is poorly immunogenic, displaying modest ICB responsiveness with rare durable activity. In profiling its plasticity, we uncover intrinsically immunogenic MHC I hi subpopulations of nonneuroendocrine SCLC associated with durable ICB benefit. We also find that combined EZH2 inhibition and STING agonism uncovers this cell state, priming cells for immune rejection. This article is highlighted in the In This Issue feature, p. 1861 ., (©2021 American Association for Cancer Research.)

Intratumoral recruitment of immune cells following innate immune activation is critical for anti-tumor immunity and involves cytosolic dsDNA sensing by the cGAS/STING pathway. We have previously shown that KRAS-LKB1 (KL) mutant lung cancer, which is resistant to PD-1 blockade, exhibits silencing of STING, impaired tumor cell production of immune chemoattractants, and T cell exclusion. Since the vasculature is also a critical gatekeeper of immune cell infiltration into tumors, we developed a novel microfluidic model to study KL tumor-vascular interactions. Notably, dsDNA priming of LKB1-reconstituted tumor cells activates the microvasculature, even when tumor cell STING is deleted. cGAS-driven extracellular export of 2'3' cGAMP by cancer cells activates STING signaling in endothelial cells and cooperates with type 1 interferon to increase vascular permeability and expression of E selectin, VCAM-1, and ICAM-1 and T cell adhesion to the endothelium. Thus, tumor cell cGAS-STING signaling not only produces T cell chemoattractants, but also primes tumor vasculature for immune cell escape., (Copyright © 2020 Campisi, Sundararaman, Shelton, Knelson, Mahadevan, Yoshida, Tani, Ivanova, Cañadas, Osaki, Lee, Thai, Han, Piel, Gilhooley, Paweletz, Chiono, Kamm, Kitajima and Barbie.)

KRAS -driven lung cancers frequently inactivate TP53 and/or STK11/LKB1 , defining tumor subclasses with emerging clinical relevance. Specifically, KRAS - LKB1 (KL)-mutant lung cancers are particularly aggressive, lack PD-L1, and respond poorly to immune checkpoint blockade (ICB). The mechanistic basis for this impaired immunogenicity, despite the overall high mutational load of KRAS -mutant lung cancers, remains obscure. Here, we report that LKB1 loss results in marked silencing of stimulator of interferon genes (STING) expression and insensitivity to cytoplasmic double-strand DNA (dsDNA) sensing. This effect is mediated at least in part by hyperactivation of DNMT1 and EZH2 activity related to elevated S-adenylmethionine levels and reinforced by DNMT1 upregulation. Ectopic expression of STING in KL cells engages IRF3 and STAT1 signaling downstream of TBK1 and impairs cellular fitness, due to the pathologic accumulation of cytoplasmic mitochondrial dsDNA associated with mitochondrial dysfunction. Thus, silencing of STING avoids these negative consequences of LKB1 inactivation, while facilitating immune escape. SIGNIFICANCE: Oncogenic KRAS -mutant lung cancers remain treatment-refractory and are resistant to ICB in the setting of LKB1 loss. These results begin to uncover the key underlying mechanism and identify strategies to restore STING expression, with important therapeutic implications because mitochondrial dysfunction is an obligate component of this tumor subtype. See related commentary by Corte and Byers, p. 16 . This article is highlighted in the In This Issue feature, p. 1 ., (©2018 American Association for Cancer Research.)