Jennifer M. Lee’s research while affiliated with Concordia University Ann Arbor and other places

Lung cancer is the leading cause of cancer deaths in the United States. New targeted therapies against the once-deemed undruggable oncogenic KRAS are changing current therapeutic paradigms. However, resistance to targeted KRAS inhibitors almost inevitably occurs; resistance can be driven by tumor cell-intrinsic changes or by changes in the microenvironment. Here, we utilized a genetically engineered mouse model of KRASG12D-driven lung cancer that allows for inducible and reversible expression of the oncogene: activation of oncogenic KRASG12D induces tumor growth; conversely, inactivation of KRASG12D causes tumor regression. We showed that in addition to regulating cancer cell growth and survival, oncogenic KRAS regulated the transcriptional status of cancer-associated fibroblasts and macrophages in this model. Utilizing ex vivo approaches, we showed that secreted factors from cancer cells induced the expression of multiple cytokines in lung fibroblasts, and in turn drove expression of immunosuppressive factors, such as arginase 1, in macrophages. In summary, fibroblasts emerged as a key source of immune regulatory signals, and a potential therapeutic target for improving the efficacy of KRAS inhibitors in lung cancer.

Mechanisms of resistance to inhibitors against mutant KRAS are linked to the remodeling of the tumor microenvironment (TME). Understanding this remodeling process during intervention and tumor recurrence will likely guide the development of future therapeutic treatment paradigms with long-term responses. 56% of never-smokers with non-small cell lung cancer have KRASG12D mutant cancer and respond little to immune checkpoint therapy (ICI) compared to smokers, who often have mutations in KRASG12C. Our analysis of the TME in a unique KrasG12D inducible and reversible mouse model of NSCLC using single cell RNA sequencing data shows KrasG12D-dependent control of the TME, particularly of the expression of PDL1 in a subset of myeloid cells. PDL1 expression was found to be KrasG12D dependent as it was reduced significantly when KrasG12D was turned ON in mouse lungs. Interestingly, when KrasG12D was turned OFF, the expression of PDL1 increased, indicating mechanisms of resistance. Furthermore, the cytokine expression and secretion of IL6, a known regulator of PDL1 expression, increased in lung tissue and lung cancer cells derived from this model upon genetic and chemical (MRTX1133) Kras inhibition. Moreover, the mechanisms of resistance to oncogenic Kras inhibition appear to be mutant specific. Transcriptome and secretome analyses in human and murine KrasG12D or KrasG12C mutant lung cancer cells show distinct cytokine regulation upon KRAS inhibition. Identification of mechanisms of resistance like the induction of PDL1 expression upon KrasG12D inhibition may provide a strong rationale for co-treatment of KRAS inhibitors with ICI for KrasG12D never-smokers with NSCLC. Citation Format: Ivana Barravecchia, Emily Lasse-Opsahl, Sophia Cavanaugh, Lily Rober, Rachael Baliira, Marzia Robotti, Rachael Hinshaw, Jennifer M. Lee, Yaqing Zhang, Marina Pasca di Magliano, Stefanie Galbán. Mutation specific mechanisms of resistance to oncogenic KRAS inhibition [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6841.

Withdrawal Statement This manuscript has been withdrawn by the authors due to a dispute over co-first authorship that is currently being arbitrated by the medical school at our institution. Therefore, the authors do not wish this work to be cited as reference for the project. Upon completion of the arbitration process, we will take steps to revert the current withdrawn status. If you have any questions, please contact the corresponding author.

Idiopathic pulmonary fibrosis (IPF) is characterized by progressive, often fatal loss of lung function due to overactive collagen production and tissue scarring. Patients with IPF have a sevenfold-increased risk of developing lung cancer. The COVID-19 pandemic has increased the number of patients with lung diseases, and infection can worsen prognoses for those with chronic lung diseases and disease-associated cancer. Understanding the molecular pathogenesis of IPF-associated lung cancer is imperative for identifying diagnostic biomarkers and targeted therapies that will facilitate prevention of IPF and progression to lung cancer. To understand how IPF-associated fibroblast activation, matrix remodeling, epithelial-to-mesenchymal transition (EMT), and immune modulation influences lung cancer predisposition, we developed a mouse model to recapitulate the molecular pathogenesis of pulmonary fibrosis–associated lung cancer using the bleomycin and Lewis lung carcinoma models. We demonstrate that development of pulmonary fibrosis–associated lung cancer is likely linked to increased abundance of tumor-associated macrophages and a unique gene signature that supports an immune-suppressive microenvironment through secreted factors. Not surprisingly, preexisting fibrosis provides a pre-metastatic niche and results in augmented tumor growth, and tumors associated with bleomycin-induced fibrosis are characterized by a dramatic loss of cytokeratin expression, indicative of EMT. Implications This characterization of tumors associated with lung diseases provides new therapeutic targets that may aid in the development of treatment paradigms for lung cancer patients with preexisting pulmonary diseases.

Idiopathic Pulmonary Fibrosis (IPF) is characterized by progressive, often fatal loss of lung function due to overactive collagen production and tissue scarring. IPF patients have a sevenfold-increased risk of developing lung cancer. The COVID-19 pandemic has increased the number of patients with lung diseases, and infection can worsen prognoses for those with chronic lung diseases and disease-associated cancer. Understanding the molecular pathogenesis of IPF associated lung cancer is imperative for identifying diagnostic biomarkers and targeted therapies that will facilitate prevention of IPF and progression to lung cancer. To understand how IPF-associated fibroblast activation, matrix remodeling, epithelial-mesenchymal transition, and immune modulation influences lung cancer predisposition, we developed a mouse model to recapitulate the molecular pathogenesis of pulmonary fibrosis-associated lung cancer using the bleomycin and the Lewis Lung Carcinoma models. Models of pulmonary fibrosis, particularly bleomycin-induced fibrosis, do not recapitulate all aspects of human disease; however, to simplify nomenclature, we refer to our bleomycin-induced fibrosis model as IPF. We demonstrate that development of pulmonary fibrosis-associated lung cancer is linked to increased recruitment or reprogramming of tumor-associated macrophages and a unique gene signature that supports an immune-suppressive microenvironment through secreted factors. Not surprisingly, pre-existing fibrosis provides a pre-metastatic niche and results in augmented tumor growth. Tumors associated with bleomycin-induced fibrosis are characterized by an epithelial-to-mesenchymal transition characterized by dramatic loss of cytokeratin expression. Implications: We provide new therapeutic targets that may aid the characterization of tumors associated with lung diseases and development of treatment paradigms for lung cancer patients with pre-existing pulmonary diseases.

Lung cancer remains the leading cause of cancer-related deaths worldwide, with an estimated 1.6 million deaths each year. Non-small cell lung cancer (NSCLC) is with 85% by far the most common subtype of lung cancer, comprising adenocarcinomas and lung squamous cell carcinoma. Mutations in Kirsten rat sarcoma viral oncogene homolog (KRAS), epidermal growth factor receptor (EGFR) and anaplastic lymphoma receptor tyrosine kinase (ALK) genes are common with the worst overall survival for KRAS mutant adenocarcinoma patients. We have established a murine model of lung cancer, wherein expression of oncogenic can be controlled genetically, allowing activation of oncogenic G12D (*) to initiate tumor growth, tumor eradication upon * depletion and re-activation as a means to model relapse. Oncogenic depletion (deactivation) has previously been reported to result in tumor cell apoptosis even in the presence of tumor suppressor loss. However, the mechanisms of apoptosis, the role of the immune system on these changes, and the mechanisms allowing some tumor cells to escape apoptosis, which typically results in tumor relapse, are unknown. Here, we interrogated the immune response in mediating tumor regression and relapse using this genetically engineered models. Multiplex immunohistochemistry as well as CyTOF provided insight into the changes in immune contexture upon * depletion in mice haploinsufficient for tumor suppressor p53 or mutant for p53 (R172H). Interestingly, total number of T cells including cytotoxic T cells (CTLs) was elevated in lung tumors from p53 mutant mice supporting findings of heightened immune activation and overall response to immune therapy with an increased mutational burden. * inactivation and thus inhibition of MAPK signaling resulted in an overall decrease in abundance of CTLs and antigen presenting cells (APC) as well as engagement of CTL with tumor cells and APCs indicating a decrease in immune presence likely due to proceeding tumor cell kill and immune recruitment. However, intracellular distance of CTL with tumor cells indicated active tumor cell kill of the CTLs to eradicate remaining tumor cells. In summary, these findings support recent observation of increased immune activation in tumors with higher mutational load as well as changes mediated by inhibition of MAPK signaling which both maybe harnessed for enhancing future immunotherapies. Citation Format: Nina Steele, Kristena Y. Abdelmalak, Sarah F. Ferris, Jennifer M. Lee, Carlos Espinoza, Yaqing Zhang, Sundaresh Ram, Craig Galban, Nithya Ramnath, Timothy L. Frankel, Marina Pasca di Magliano, Stefanie Galbán. Oncogenic -mediated regulation of the tumor microenvironment in lung cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3855.