Abstract 1017: Lipid metabolic reprogramming drives resistance to PD1 blockage
Authors: Cortez Maria A. , Niknam Sharareh , Cuko Efrosini , Schoenhals Jonathan E. , Barsoumian Hampartsoum , Younes Ahmed I. , Li Ailin , Vykoukal Jody V. , Ivan Cristina , Calin George A. , Hwu Patrick , Welsh James W. 
Published: July 2017

Abstract
The mechanisms underlying immunosuppression and resistance to PD1 inhibitors in cancer are not well understood. We attempted to fill this gap with an integrated analysis of mRNA, microRNA, and protein expression in an anti-PD1-resistant lung adenocarcinoma mouse model. The model was created by in vivo passage of 344SQ murine lung cancer cells (p53R172HΔg/+K-rasLA1/+) in a syngeneic host repeatedly dosed with anti-mouse PD1 antibodies. Anti-PD1-resistant 344SQ (344SQ_R) and 344SQ parental (344SQ_P) cells were then inoculated into syngeneic 129Sv/ev mice, which were then dosed twice with anti-PD1 or control IgG antibodies. Tumor tissues were collected and analyzed as follows: transcriptome with Affymetrix; protein levels by reverse phase protein array analysis; signature enrichment by gene set enrichment analysis; metabolome by mass spectrometry; and lipid content with fluorescent probes Oil O rad and BODIPY. We also isolated tumor-infiltrating immune cells for flow cytometry and gene expression analyses. We identified lipid-related metabolic pathways as being the most highly enriched in anti-PD1-resistant tumors (344SQ_R) vs. their 344SQ_P counterparts; the resistant cells also had more lipid droplets than the 344SQ_P cells. The anti-PD1-resistant tumors overexpressed several genes involved in lipogenesis and fatty acid pathways (e.g., fatty acid binding proteins [FABPs], fatty acid synthase, acetyl-coA-acyltransferase 2, fatty acid elongases). Specifically, FABP overexpression promoted fatty acid uptake and lipid-droplet accumulation in resistant tumors. Lipid-sensitive targets linked to inflammation and insulin signaling (e.g,. stress-activated kinases such as JNK and NFκB) were altered in 344SQ_R vs. 344SQ_P tumors. Mechanistically, JNK downregulation by NFκB-regulated microRNAs protected PD1-resistant tumors from lipotoxicity caused by FABPs upregulation and fatty acid uptake. FABP levels were higher in plasma from 344SQ_R than from 344SQ_P tumors. Tumor-infiltrating macrophages from 344SQ_R tumors had 4 times the amount of FABP mRNA than parental tumors and a correspondingly higher percentage of M2-like macrophages. 344SQ_R tumors promoted immune suppressive cells by upregulating FABPs expression in M2-like macrophages, marked by increased fatty acid intake and fatty acid oxidation. Conversely, percentages of CD4+ and CD8+ tumor-infiltrating lymphocytes were reduced in the resistant tumors. These results suggest that lipid metabolic rewiring drives resistance PD1 inhibitors supporting the accumulation of immunosuppressive cells, including M2-like macrophages, preventing type I immune responses elicited by T cells. Collectively, these findings reveal new potential lipid-related targets for drug development or new treatments combining inhibitors of these targets with anti-PD1 therapy.
Citation Format: Maria A. Cortez, Sharareh Niknam, Efrosini Cuko, Jonathan E. Schoenhals, Hampartsoum Barsoumian, Ahmed I. Younes, Ailin Li, Jody V. Vykoukal, Cristina Ivan, George A. Calin, Patrick Hwu, James W. Welsh. Lipid metabolic reprogramming drives resistance to PD1 blockage [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1017. doi:10.1158/1538-7445.AM2017-1017