Abstract 1128: MTORC1/2 and HDAC1/2 inhibition promote tumor response through inhibition of MYC
Authors: DeStefanis Rebecca A. , Olson Autumn M. , DeZeeuw Alyssa K. , Payne Susan N. , Pasch Cheri A. , Clipson Linda , Deming Dustin A. 
Published: June 2022

Background: The identification of treatment strategies targeting PIK3CA mutant colorectal cancer (CRC) are of great clinical interest. Previous work from our lab has identified MTORC1/2 and HDAC1/2 inhibition with copanlisib (cop; PI3K/MTOR) and romidepsin (romi; HDAC1/2) as a potential therapeutic strategy. We hypothesized that changes in c-MYC protein levels and c-MYC target gene (CTG) alterations might be a potential mechanism of action for this combination.

Methods: Known CTGs were identified and their expression levels were examined in PIK3CA mutant vs WT CRCs using the cBioPortal Colorectal Adenocarcinoma (TCGA, PanCancer Atlas) dataset. Murine derived cancer organoids (MDCO) were generated from adenocarcinomas of Apc and Pik3ca mutant transgenic mice (F1 (FVB x B6) Apcfl/+ Pik3caH1047R/+). MDCO results were corroborated using the human 2D isogenic cell lines SW48 and SW48PIK3CA-H1047R (SW48PK) and RAS/RAF WT patient derived cancer organoids (PDCO) generated from CRC patient samples under approved IRB protocols. Immunoblots (IB) were used to assess c-MYC levels after treatment with cop, romi, and the combination across all models. RNA sequencing was conducted on PDCOs and SW48PK cells and changes in 16 CTGs were examined. An aggregate score was created for each treatment group where a statistically significantly altered CTG with log fold change ≥1.5 added one point and ≤-1.5 subtracted one point. All others were scored 0.

Results: c-MYC target genes were assessed for differential expression in PIK3CA mutant CRC vs PIK3CA WT CRC with only 1/16 genes decreased in PIK3CA mutant CRC (GADD45A: log ratio -0.21, q=0.01). Next, MDCOs treated with the combination showed a decrease in total c-MYC levels in the combination therapy. These results were corroborated in two human 2D CRC cell lines, SW48 and SW48PK and a panel of PDCOs. Interestingly, c-MYC levels decreased in both romi alone and the combination in both the SW48 and SW48PK cell lines after 24 hours of treatment. However, the extent to which c-MYC levels were decreased was not as substantial in the panel of PDCOs. Across all in vitro models a decrease in PI3K signaling, as illustrated by decreased pRPS6 and p4EBP1 in response to cop treatment and an increase in H3K27 acetylation in response to romi, was observed in the single agent and combination therapies. RNA sequencing demonstrated that cop, romi, and combo had a CTG score of 0, -4, and -7 respectively in the PIK3CA mutant PDCO. Similar results were seen in an additional RAS/RAF WT PDCO (0, 1, and -4, respectively) and SW48PK cells (0, -3, and -7, respectively).

Conclusion: A potential mechanism by which cop and romi treatment promote tumor response is through a decrease in c-MYC protein levels and expression of downstream c-MYC target genes. This regimen deserves further mechanistic investigations in vivo.

Citation Format: Rebecca A. DeStefanis, Autumn M. Olson, Alyssa K. DeZeeuw, Susan N. Payne, Cheri A. Pasch, Linda Clipson, Dustin A. Deming. MTORC1/2 and HDAC1/2 inhibition promote tumor response through inhibition of MYC [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 1128.