- Email: [email protected]
The Likelihood of Heterogeneity or Additional Mutation to Compromise Targeting of KRAS G12C
S1538
Investigating of Immunotherapy and Combination Treatments in Ex-Vivo Culture Model of NSCLC J. Bar, I. Kamer, L. Bab-Dinitz, I.D. Meshulam, O. Zadok, O. Amir Sheba Medical Center, IL, Tel HaShomer, Israel Background: NSCLC induces pro-tumorigenic immunosuppressive changes to evade the immune system. Response of tumors to therapy, including to immunotherapy, depends on interactions of epithelial tumor cells and the microenvironment. Ex vivo culture (EVOC) models directly established from fresh NSCLC tumor tissues can be used as a complete model that resembles real tumor and its microenvironment. Little is known about the impact of drugs such as chemotherapy, radiotherapy or targeted agents on the expression and activity of the PD1-PDL1 signaling in human cancers. Combining such treatments with immunotherapy seems to be a promising approach that is actively investigated. We are using an EVOC model to study the impact of anti-PDL1 agents in NSCLC, alone or in combination with other therapeutics (chemotherapy or radiotherapy). Results: Our results indicate our EVOC model applicability as modeling immunotherapy effects in cancer. Specifically, tumor and stromal cells are maintained with a stable structure over a time window of several days and cell death is seen in response to cytotoxic drugs. Importantly, PDL1 is dramatically induced in response to inflammation signals (e.g. IFNg), PDL1 is reduced in response to steroids and variability in PDL-1 expression is seen between EVOCs from different NSCLC patients. Conclusions: NSCLC EVOC is a promising model for elucidation of immunotherapy and other drugs impact on real human cancer with its native microenvironment and immune cells.
Unmasking the Role of Activated and Exhausted B Cells in Lung Cancer T.C. Bruno,1 J.A. Kern,2 D.A.A. Vignali,1 J.E. Slansky3 1University of Pittsburgh, Pittsburg, PA, US, 2National Jewish Health, Denver, CO, US, 3 University of Colorado, Denver, CO, US Background: Effective immunotherapy options for patients with none small cell lung cancer (NSCLC) are becoming increasingly available. The immunotherapy focus has been on tumor infiltrating T cells (TILs); however, tumor infiltrating B cells (TIL-Bs) have also been reported to correlate with NSCLC patient survival. The function of TIL-Bs in human cancer has been understudied with limited focus on their role as antigen presenting cells and their influence on CD4 TILs. Results: We observed an increase in the total number of intratumoral B cells compared to other immune subsets in fresh, primary tumors of NSCLC patients. Furthermore, we demonstrated that TIL-Bs can efficiently present antigen to CD4 TILs and alter CD4 TIL phenotype using an in vitro antigen presentation assay. Specifically, we identified three CD4 TIL responses to TIL-Bs, which we categorized as: activated, antigen-associated, and nonresponsive. Within the activated and antigen-associated CD4 TIL population, activated TIL-Bs (CD19+CD20+CD69+CD27+CD21+) were associated with an effector T cell response (IFN-g+ CD4 TIL). Alternatively, exhausted TIL-Bs (CD19+CD20+CD69+CD27-CD21-) were associated with a regulatory T cell phenotype (FoxP3+ CD4 TIL). Conclusions: Our results demonstrate a new role for TIL-Bs in NSCLC tumors in their interplay with CD4 TILs in the tumor microenvironment, establishing them as a potential therapeutic target in NSCLC immunotherapy.
Journal of Thoracic Oncology
Vol. 12 No. 8S
Background: Activating mutations in RAS genes are associated with approximately 20% of all human cancers. New targeted therapies show preclinical promise in inhibiting the KRAS G12C variant, however, concerns exist regarding the effectiveness of such therapies in vivo given the possibilities of existing intratumor heterogeneity or de novo mutation leading to treatment resistance. Results: We performed deep sequencing of 27 KRAS G12 positive lung tumors and found no evidence of other oncogenic mutations within KRAS or within commonly mutated downstream genes that could confer resistance at the time of treatment. Furthermore, we estimate the de novo mutation rate in KRAS position 12 and in genes downstream of KRAS. We find that mutations that confer resistance are about as likely to occur downstream of KRAS as within KRAS. Moreover, we present an approach for estimation of the selection intensity for these point mutations that explains their high prevalence in tumors. Our approach predicts that BRAF V600E would provide the highest fitness advantage for de novo resistant subclones. Conclusions: Overall, our findings suggest that resistance to targeted therapy of KRAS G12C positive tumors is unlikely to be present at the time of treatment and, among the de novo mutations likely to confer resistance, mutations in BRAF, a gene with targeted inhibitors presently available, result in subclones with the highest fitness advantage.
Fatty Acid Synthase Is a Therapeutic Target in Mutant KRAS Lung Cancer C. Bartolacci,1 M. Padanad,1 C. Andreani,1 M. Melegari,1 S. Rindhe,1 K. George,2 A. Frankel,1 J. McDonald,1 P.P. Scaglioni1 1UT Southwestern Medical Center, Dallas, TX, US, 23-V Biosciences, Menlo Park, CA, US Background: Mutant KRAS is a high priority therapeutic target. Nevertheless, mutant KRAS remains non-targetable. Mutant KRAS, as other oncogenes, regulates the metabolism of cancer cells to promote the acquisition of fuel and building blocks. Cancer cells require large quantities of fatty acids that are conjugated to Acyl-CoA to participate to the synthesis of lipids used for cellular membranes, energy storage or metabolic reactions. Accordingly, cancer cells upregulate fatty acid synthetase (FASN), the rate limiting enzyme for the synthesis of fatty acids. Results: We found that FASN and Acyl-CoA synthetase long chain fatty acid 3 (ACSL3), the enzyme responsible for the conjugation of fatty acids to Acyl-CoA are required for the viability for mutant KRAS lung cancer cells. Furthermore, germline ablation of ACSL3 prevents the formation of lung cancer in vivo. Exogenous palmitate reverses the phenotype caused by FASN inhibition. Furthermore, with infusion lipidomics, we found that detrimental effects on cell viability correlate with depletion of intracellular long chain fatty acids and triglycerides. Conclusions: Mutant KRAS reprograms the metabolism of FA, establishing a dependency on FASN. The FASN inhibitor, TVB-2640, has been evaluated in a Phase 1 study in solid tumor patients (Clinicaltrials.gov NCT02223247); these results support the opportunity to test this hypothesis in the clinic.
Identification and Characterization of Oncogenic SOS1 Mutations in Lung Adenocarcinoma D. Cai,1 P. Choi,2 M. Meyerson2 1Harvard University, Boston, MA, US, 2 Dana Farber Cancer Institute, Boston, MA, US
The Likelihood of Heterogeneity or Additional Mutation to Compromise Targeting of KRAS G12C V. Cannataro,1 S. Gaffney,1 C. Stender,1 Z. Zhao,1 M. Philips,2 A. Greenstein,3 J. Townsend1 1Yale University, New Haven, CT, US, 2 New York University, New York, NY, US, 3Gilead Sciences, Foster City, CA, US
Background: Lung adenocarcinomas are characterized by genetic alterations along receptor tyrosine kinase pathways. Around 50% of lung adenocarcinomas contain alterations in KRAS and EGFR alone. However, genetic drivers in a large proportion of other cases remain to be determined. Recent exome sequencing analysis of lung adenocarcinomas in our lab has identified SOS1, a guanine nucleotide exchange factor, as being
Investigating of Immunotherapy and Combination Treatments in Ex-Vivo Culture Model of NSCLC J. Bar, I. Kamer, L. Bab-Dinitz, I.D. Meshulam, O. Zadok, O. Amir Sheba Medical Center, IL, Tel HaShomer, Israel Background: NSCLC induces pro-tumorigenic immunosuppressive changes to evade the immune system. Response of tumors to therapy, including to immunotherapy, depends on interactions of epithelial tumor cells and the microenvironment. Ex vivo culture (EVOC) models directly established from fresh NSCLC tumor tissues can be used as a complete model that resembles real tumor and its microenvironment. Little is known about the impact of drugs such as chemotherapy, radiotherapy or targeted agents on the expression and activity of the PD1-PDL1 signaling in human cancers. Combining such treatments with immunotherapy seems to be a promising approach that is actively investigated. We are using an EVOC model to study the impact of anti-PDL1 agents in NSCLC, alone or in combination with other therapeutics (chemotherapy or radiotherapy). Results: Our results indicate our EVOC model applicability as modeling immunotherapy effects in cancer. Specifically, tumor and stromal cells are maintained with a stable structure over a time window of several days and cell death is seen in response to cytotoxic drugs. Importantly, PDL1 is dramatically induced in response to inflammation signals (e.g. IFNg), PDL1 is reduced in response to steroids and variability in PDL-1 expression is seen between EVOCs from different NSCLC patients. Conclusions: NSCLC EVOC is a promising model for elucidation of immunotherapy and other drugs impact on real human cancer with its native microenvironment and immune cells.
Unmasking the Role of Activated and Exhausted B Cells in Lung Cancer T.C. Bruno,1 J.A. Kern,2 D.A.A. Vignali,1 J.E. Slansky3 1University of Pittsburgh, Pittsburg, PA, US, 2National Jewish Health, Denver, CO, US, 3 University of Colorado, Denver, CO, US Background: Effective immunotherapy options for patients with none small cell lung cancer (NSCLC) are becoming increasingly available. The immunotherapy focus has been on tumor infiltrating T cells (TILs); however, tumor infiltrating B cells (TIL-Bs) have also been reported to correlate with NSCLC patient survival. The function of TIL-Bs in human cancer has been understudied with limited focus on their role as antigen presenting cells and their influence on CD4 TILs. Results: We observed an increase in the total number of intratumoral B cells compared to other immune subsets in fresh, primary tumors of NSCLC patients. Furthermore, we demonstrated that TIL-Bs can efficiently present antigen to CD4 TILs and alter CD4 TIL phenotype using an in vitro antigen presentation assay. Specifically, we identified three CD4 TIL responses to TIL-Bs, which we categorized as: activated, antigen-associated, and nonresponsive. Within the activated and antigen-associated CD4 TIL population, activated TIL-Bs (CD19+CD20+CD69+CD27+CD21+) were associated with an effector T cell response (IFN-g+ CD4 TIL). Alternatively, exhausted TIL-Bs (CD19+CD20+CD69+CD27-CD21-) were associated with a regulatory T cell phenotype (FoxP3+ CD4 TIL). Conclusions: Our results demonstrate a new role for TIL-Bs in NSCLC tumors in their interplay with CD4 TILs in the tumor microenvironment, establishing them as a potential therapeutic target in NSCLC immunotherapy.
Journal of Thoracic Oncology
Vol. 12 No. 8S
Background: Activating mutations in RAS genes are associated with approximately 20% of all human cancers. New targeted therapies show preclinical promise in inhibiting the KRAS G12C variant, however, concerns exist regarding the effectiveness of such therapies in vivo given the possibilities of existing intratumor heterogeneity or de novo mutation leading to treatment resistance. Results: We performed deep sequencing of 27 KRAS G12 positive lung tumors and found no evidence of other oncogenic mutations within KRAS or within commonly mutated downstream genes that could confer resistance at the time of treatment. Furthermore, we estimate the de novo mutation rate in KRAS position 12 and in genes downstream of KRAS. We find that mutations that confer resistance are about as likely to occur downstream of KRAS as within KRAS. Moreover, we present an approach for estimation of the selection intensity for these point mutations that explains their high prevalence in tumors. Our approach predicts that BRAF V600E would provide the highest fitness advantage for de novo resistant subclones. Conclusions: Overall, our findings suggest that resistance to targeted therapy of KRAS G12C positive tumors is unlikely to be present at the time of treatment and, among the de novo mutations likely to confer resistance, mutations in BRAF, a gene with targeted inhibitors presently available, result in subclones with the highest fitness advantage.
Fatty Acid Synthase Is a Therapeutic Target in Mutant KRAS Lung Cancer C. Bartolacci,1 M. Padanad,1 C. Andreani,1 M. Melegari,1 S. Rindhe,1 K. George,2 A. Frankel,1 J. McDonald,1 P.P. Scaglioni1 1UT Southwestern Medical Center, Dallas, TX, US, 23-V Biosciences, Menlo Park, CA, US Background: Mutant KRAS is a high priority therapeutic target. Nevertheless, mutant KRAS remains non-targetable. Mutant KRAS, as other oncogenes, regulates the metabolism of cancer cells to promote the acquisition of fuel and building blocks. Cancer cells require large quantities of fatty acids that are conjugated to Acyl-CoA to participate to the synthesis of lipids used for cellular membranes, energy storage or metabolic reactions. Accordingly, cancer cells upregulate fatty acid synthetase (FASN), the rate limiting enzyme for the synthesis of fatty acids. Results: We found that FASN and Acyl-CoA synthetase long chain fatty acid 3 (ACSL3), the enzyme responsible for the conjugation of fatty acids to Acyl-CoA are required for the viability for mutant KRAS lung cancer cells. Furthermore, germline ablation of ACSL3 prevents the formation of lung cancer in vivo. Exogenous palmitate reverses the phenotype caused by FASN inhibition. Furthermore, with infusion lipidomics, we found that detrimental effects on cell viability correlate with depletion of intracellular long chain fatty acids and triglycerides. Conclusions: Mutant KRAS reprograms the metabolism of FA, establishing a dependency on FASN. The FASN inhibitor, TVB-2640, has been evaluated in a Phase 1 study in solid tumor patients (Clinicaltrials.gov NCT02223247); these results support the opportunity to test this hypothesis in the clinic.
Identification and Characterization of Oncogenic SOS1 Mutations in Lung Adenocarcinoma D. Cai,1 P. Choi,2 M. Meyerson2 1Harvard University, Boston, MA, US, 2 Dana Farber Cancer Institute, Boston, MA, US
The Likelihood of Heterogeneity or Additional Mutation to Compromise Targeting of KRAS G12C V. Cannataro,1 S. Gaffney,1 C. Stender,1 Z. Zhao,1 M. Philips,2 A. Greenstein,3 J. Townsend1 1Yale University, New Haven, CT, US, 2 New York University, New York, NY, US, 3Gilead Sciences, Foster City, CA, US
Background: Lung adenocarcinomas are characterized by genetic alterations along receptor tyrosine kinase pathways. Around 50% of lung adenocarcinomas contain alterations in KRAS and EGFR alone. However, genetic drivers in a large proportion of other cases remain to be determined. Recent exome sequencing analysis of lung adenocarcinomas in our lab has identified SOS1, a guanine nucleotide exchange factor, as being