Novel EGFR-mutant mouse models of lung adenocarcinoma reveal adaptive immunity requirement for durable osimertinib response

Kleczko, Emily K.; Le, Anh‐Tuan; Hinz, Trista K.; Nguyen, Teresa T.; Navarro, Andre; Hu, Chengjun; Selman, Ana M.; Clambey, Eric T.; Merrick, Daniel T.; Lu, Sizhao; Weiser-Evans, Mary C.; Nemenoff, Raphael A.; Heasley, Lynn E.

doi:10.1016/j.canlet.2023.216062

Cited by 14 publications

(15 citation statements)

References 54 publications

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“…A testable hypothesis is that the extent of chemokine induction following KRAS G12C inhibition may contribute to the depth and durability of response. In addition to the previously noted studies demonstrating involvement of host immunity in therapeutic response, our own studies with murine EML4-ALK-driven ( 19 ) and EGFR-mutant ( 20 ) lung cancer cell lines demonstrate a contribution of adaptive immunity to durable responses to the ALK inhibitor, alectinib and the EGFR inhibitor, osimertinib, respectively. Current understanding surrounding the depth and duration of response to oncogene-targeted inhibitors suggests that the extent to which chemokine induction occurs with oncogene-specific therapies associates with the degree of response, as EGFR mutant lung cancer patients exhibiting greater interferon γ transcriptional responses to TKIs presented with longer progression-free survival ( 17 ).…”

Section: Discussionsupporting

confidence: 55%

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Evaluation of KRASG12C inhibitor responses in novel murine KRASG12C lung cancer cell line models

Sisler

Hinz²,

Le³

et al. 2023

Front. Oncol.

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IntroductionThe KRAS(G12C) mutation is the most common genetic mutation in North American lung adenocarcinoma patients. Recently, direct inhibitors of the KRASG12C protein have been developed and demonstrate clinical response rates of 37-43%. Importantly, these agents fail to generate durable therapeutic responses with median progression-free survival of ~6.5 months.MethodsTo provide models for further preclinical improvement of these inhibitors, we generated three novel murine KRASG12C-driven lung cancer cell lines. The co-occurring NRASQ61L mutation in KRASG12C-positive LLC cells was deleted and the KRASG12V allele in CMT167 cells was edited to KRASG12C with CRISPR/Cas9 methods. Also, a novel murine KRASG12C line, mKRC.1, was established from a tumor generated in a genetically-engineered mouse model.ResultsThe three lines exhibit similar in vitro sensitivities to KRASG12C inhibitors (MRTX-1257, MRTX-849, AMG-510), but distinct in vivo responses to MRTX-849 ranging from progressive growth with orthotopic LLC-NRAS KO tumors to modest shrinkage with mKRC.1 tumors. All three cell lines exhibited synergistic in vitro growth inhibition with combinations of MRTX-1257 and the SHP2/PTPN11 inhibitor, RMC-4550. Moreover, treatment with a MRTX-849/RMC-4550 combination yielded transient tumor shrinkage in orthotopic LLC-NRAS KO tumors propagated in syngeneic mice and durable shrinkage of mKRC.1 tumors. Notably, single-agent MRTX-849 activity in mKRC.1 tumors and the combination response in LLC-NRAS KO tumors was lost when the experiments were performed in athymic nu/nu mice, supporting a growing literature demonstrating a role for adaptive immunity in the response to this class of drugs.DiscussionThese new models of murine KRASG12C mutant lung cancer should prove valuable for identifying improved therapeutic combination strategies with KRASG12C inhibitors.

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Section: Discussionsupporting

confidence: 55%

“…Our lab and others have shown the importance of innate and adaptive immunity in driving pre-clinical and clinical responses to oncogene-targeted therapies (16)(17)(18)(19)(20)(21)(22)(23). Thus, comprehensive assessment of the therapeutic potential of oncogene-targeted agents requires immune competent murine models.…”

Section: Introductionmentioning

confidence: 99%

Evaluation of KRASG12C inhibitor responses in novel murine KRASG12C lung cancer cell line models

Sisler

Hinz²,

Le³

et al. 2023

Front. Oncol.

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“…Multiple groups including ours have recently demonstrated the involvement of anti-tumor immunity in the therapeutic response to KRAS G12C inhibitors 10,12,13,15 . We recently reported the involvement of adaptive immunity in the therapeutic response of murine lung cancer cells driven by oncogenic EGFR to the TKI, osimertinib 14 . Using scRNA analysis of biopsies from oncogene-defined lung cancers including EGFR mutant and ALK+ cancers, Maynard et al revealed a transient immunostimulatory effect in lung cancer patients bearing oncogenic RTKs after initial TKI therapy, followed by establishment of an immunosuppressive environment upon progression 33 .…”

Section: Quantification Of Lymphocytes and Pmns In Alk+ Patient Biopsiesmentioning

confidence: 99%

“…Our recent published studies demonstrate that EGFR-targeted inhibitors induce an interferon (IFN) response program that varies markedly between distinct EGFR mutant lung cancer cell lines and positively associates with the duration of therapeutic response in EGFR-mutant lung cancer patients 7 . In fact, there is a growing literature supporting the role of host immune cells in overall therapeutic response to precision oncology agents and cytotoxic drugs [8][9][10][11][12][13][14][15][16][17][18][19] . However, the mechanisms whereby TKIs induce factors mediating paracrine signaling to the immune microenvironment, and the contribution to the overall therapeutic response are not well understood.…”

Section: Introductionmentioning

confidence: 99%

Durable responses to alectinib in murine models of EML4-ALK lung cancer requires adaptive immunity

et al. 2023

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Lung cancers bearing oncogenic EML4-ALK fusions respond to targeted tyrosine kinase inhibitors (TKIs; e.g., alectinib), with variation in the degree of shrinkage and duration of treatment (DOT). However, factors that control this response are not well understood. While the contribution of the immune system in mediating the response to immunotherapy has been extensively investigated, less is known regarding the contribution of immunity to TKI therapeutic responses. We previously demonstrated a positive association of a TKI-induced interferon gamma (IFNγ) transcriptional response with DOT in EGFR-mutant lung cancers. Herein, we used three murine models of EML4-ALK lung cancer to test the role for host immunity in the alectinib therapeutic response. The cell lines (EA1, EA2, EA3) were propagated orthotopically in the lungs of immunocompetent and immunodeficient mice and treated with alectinib. Tumor volumes were serially measured by μCT and immune cell content was measured by flow cytometry and multispectral immunofluorescence. Transcriptional responses to alectinib were assessed by RNAseq and secreted chemokines were measured by ELISA. All cell lines were similarly sensitive to alectinib in vitro and as orthotopic tumors in immunocompetent mice, exhibited durable shrinkage. However, in immunodeficient mice, all tumor models rapidly progressed on TKI therapy. In immunocompetent mice, EA2 tumors exhibited a complete response, whereas EA1 and EA3 tumors retained residual disease that rapidly progressed upon termination of TKI treatment. Prior to treatment, EA2 tumors had greater numbers of CD8+ T cells and fewer neutrophils compared to EA1 tumors. Also, RNAseq of cancer cells recovered from untreated tumors revealed elevated levels of CXCL9 and 10 in EA2 tumors, and higher levels of CXCL1 and 2 in EA1 tumors. Analysis of pre-treatment patient biopsies from ALK+ tumors revealed an association of neutrophil content with shorter time to progression. Combined, these data support a role for adaptive immunity in durability of TKI responses and demonstrate that the immune cell composition of the tumor microenvironment is predictive of response to alectinib therapy.

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“…The pace of discovery of the oncogenic drivers in histologically-defined cancers such as lung adenocarcinoma is not always matched with access to human cell lines or PDXs as preclinical models. Moreover, a growing literature reveals that the tumor microenvironment (TME) including host immune cells significantly contribute to the therapeutic responses achieved with oncogene-targeted agents like tyrosine kinase inhibitors (TKIs) and KRAS G12C -targeted agents [11][12][13][14][15][16][17]. Thus, rigorous preclinical modeling of oncogene-targeted agents relevant to that observed in patients requires input from the host adaptive immune system.…”

Section: Introductionmentioning

confidence: 99%

A Rapid, Functional sgRNA Screening Method for Generating Murine RET and NTRK1 Fusion Oncogenes

Heasley

Schubert²,

Le³

et al. 2023

Preprint

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CRISPR/Cas9 gene editing technology is an indispensable and powerful tool in the field of cancer biology. To conduct successful CRISPR-based experiments, it is crucial that sgRNAs generate their designed alterations. Here, we describe a simple and efficient sgRNA screening method for validating sgRNAs that generate oncogenic gene rearrangements. We used IL3-independence in Ba/F3 cells as an assay to identify sgRNA pairs that generate fusion oncogenes involving the Ret and Ntrk1 tyrosine kinases. We confirmed these rearrangements with PCR or RT-PCR as well as sequencing. Ba/F3 cells harboring Ret or Ntrk1 rearrangements acquired sensitivity to RET and TRK inhibitors, respectively. Adenoviruses encoding Cas9 and sgRNAs that catalyze the Kif5b-Ret and Trim24-Ret rearrangements were intratracheally instilled into mice and yielded lung adenocarcinomas. A cell line (TR.1) was established from a Trim24-Ret positive tumor that exhibited high in vitro sensitivity to RET-specific TKIs. Moreover, orthotopic transplantation of TR.1 cells into the left lung yielded well-defined tumors that shrank in response to LOXO-292 treatment. The method offers an efficient means to validate sgRNAs that successfully target their intended loci for the generation of novel murine oncogene-driven tumor models.

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Novel EGFR-mutant mouse models of lung adenocarcinoma reveal adaptive immunity requirement for durable osimertinib response

Cited by 14 publications

References 54 publications

Evaluation of KRASG12C inhibitor responses in novel murine KRASG12C lung cancer cell line models

Evaluation of KRASG12C inhibitor responses in novel murine KRASG12C lung cancer cell line models

Durable responses to alectinib in murine models of EML4-ALK lung cancer requires adaptive immunity

A Rapid, Functional sgRNA Screening Method for Generating Murine RET and NTRK1 Fusion Oncogenes

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