Abstract 3533: Effects of tyrosine kinase inhibitors imatinib, dasatinib, and nilotinib on cancer-associated fibroblast phenotypes in colorectal cancer
Authors: Johnson Katherine Anne , Lippert Anna L. , Kraus Sean G. , McGrath Grace E. , Emmerich Philip B. , Pasch Cheri A. , Clipson Linda , Matkowskyj Kristina A. , Zhang Wei , Deming Dustin A. 
Published: June 2022

Background: Cancer-associated fibroblasts (CAFs) are major regulators of the immune microenvironment and therapeutic response in colorectal cancer (CRC). Neutralizing their role in modulating the immune landscape could be the key to enhancing immunotherapy success. Imatinib, dasatinib, and nilotinib are tyrosine kinase inhibitors with several kinase targets.

Methods: Tissue microarrays spanning 153 patients were stained for αSMA, TAGLN, PDPN, ICAM1, and CD8. CD8 stains were quantified as number of tumor infiltrating lymphocytes per high powered field (TILs/HPF) in the epithelial compartment. All other stains were quantified by intensity on a 0-3+ scale. Scores for αSMA and TAGLN were combined into a myCAF score, and PDPN and ICAM1 into an iCAF score. myCAF gene expression signatures derived from a re-analysis of scRNA-seq data previously done by our lab were entered into the LINCS database to discover potential drugs to reverse the phenotype. Primary cancer associated fibroblasts were derived from patient tumor samples, then treated with clinically relevant concentrations of imatinib, dasatinib, or nilotinib for 96 hours. RNA was isolated and RT-qPCR was performed to quantify the myCAF genes ACTA2, COL11A1, and TAGLN, and the iCAF genes ICAM1, PDPN, IL1R1, CXCL1 and CXCL2. TGFB1 expression was also measured. Expression was normalized to untreated cells and GAPDH expression levels.

Results: Cancers with high expression of myCAF markers but low expression of iCAF markers had the most CD8+ TILs (average 10.2; median 1.5; range 0-73), while cancers with low myCAF scores and high iCAF scores had the least (average 1.5; median 0; range 0-19; p &lt; 0.01). Reversing the myCAF signature relative to iCAFs in the LINCs database revealed nilotinib as a top hit. Treatment with imatinib did not significantly alter the expression of myCAF genes (control vs. max dose: p = 0.06 for ACTA2, COL11A1 p =0.2, TAGLN p = 1), while treatment with dasatinib significantly increased these genes (ACTA2 1.4x higher, p &lt; 0.001; COL11A1 2.6x higher, p &lt; 0.01; TAGLN 1.5x higher, p &lt; 0.001). Only treatment with nilotinib significantly decreased myCAF genes (ACTA2 2.2x lower, p &lt;0.001; COL11A1 1.3x lower, p =0.05; TAGLN 1.9x lower, p &lt; 0.01). All three drugs decreased iCAF gene CXCL1, and all but dasatinib decreased CXCL2. All three drugs significantly decreased TGFB1, a potential functional marker for altering myCAF phenotype (dasatinib 1.1x lower, p = 0.1; imatinib 1.6x lower, p &lt; 0.001; nilotinib 1.5x lower, p &lt; 0.05).

Conclusions: myCAFs may be major actors in immune exclusion in the microenvironment, and the reversal of the myCAF phenotype may be a target for treatment with immunotherapy. Nilotinib, but not imatinib or dasatinib, is effective at decreasing expression of myCAF genes. Further research is warranted into the mechanisms of this drug on altering expression and whether these trends continue in vivo.

Citation Format: Katherine Anne Johnson, Anna L. Lippert, Sean G. Kraus, Grace E. McGrath, Philip B. Emmerich, Cheri A. Pasch, Linda Clipson, Kristina A. Matkowskyj, Wei Zhang, Dustin A. Deming. Effects of tyrosine kinase inhibitors imatinib, dasatinib, and nilotinib on cancer-associated fibroblast phenotypes in colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3533.