Abstract CT041: Identification of gene signatures associated with response in a Phase II trial of entinostat (ENT) plus pembrolizumab (PEMBRO) in non-small cell lung cancer (NSCLC) patients whose disease has progressed on or after anti-PD-(L)1 therapy
Authors: Ordentlich Peter , Wang Lei , Tamang David , Sullivan Amy H. , Church Sally E. , Rozelle Dan , Meyers Michael L. , Ramalingam Suresh S. , Hellmann Matthew D. 
Published: August 2019

Abstract
Background: We previously reported that ENT + PEMBRO demonstrated promising clinical activity in anti-PD-(L)1 pre-treated NSCLC patients. Identification of circulating and/or tumor-resident biomarkers are essential for optimizing the utility of this combination. Here we report clinical data and the identification of gene expression signatures in tumors associated with response to PEMBRO + ENT in patients previously treated with PD-(L)1.
Methods: Gene expression analysis of pre-treatment tumor samples by RNAseq and NanoString’s PanCancer IO360™ assay was conducted in a subset of patients with sufficient RNA yield (n=43 of 76). Comparisons of differential expression were made between responders (R) vs non-responders (NR) and monocyte high (MHi) vs low (MLo) subset; significance was defined by BH adjusted p &lt;= 0.05 and &gt;1.5 fold difference. Pre-ranked gene set enrichment analysis was performed against all human MSigDB collections and gene sets with positive normalized enrichment score and p-value &lt;=0.05 were considered significantly enriched. IO360 signature analysis was performed as previously described and contrasts performed as for RNAseq.
Results: 76 patients were enrolled, of which 72 were evaluable for efficacy. The confirmed ORR with ENT + PEMBRO was 10% (7 of 72, 95% CI: 4-19%), with a median duration of response of 5.3 mos (4 patients ongoing) and median PFS of 2.8 mos. Results from gene set analysis revealed targets of Myc regulation (HALLMARK_MYC_TARGETS_V1) as the most significant enriched Hallmark collections in R (n=4) vs NR (n=39). Patients were also evaluated based on levels of baseline circulating classical monocytes (CD14+CD16-HLA-DRhi), which associated with response (MHi ORR 21.1%; PFS 5.3 mos vs MLo ORR 6.5%; PFS 2.7 mos). Myc target gene sets were also significantly enriched in MHi (n=11) vs MLo (n= 32) subjects. Oxidative phosphorylation, and E2F signaling were also among the top five for both R and MHi groups. 49.2% of statistically significant differentially expressed pathways were up-regulated in both R and MHi groups suggesting a strong overlap in differential gene expression in tumors from R and MHi patient subsets. Tumor adaptive immune resistance, as measured by the Tumor Inflammation Signature was also associated with both clinical response and longer PFS. Additionally, PD-L1, CTLA4, IFNγ-signaling and Th1 cell signatures correlated with response.
Conclusions: Gene expression analysis identified up-regulated Myc signaling in R vs NR as well as in MHi vs MLo. Recent reports have implicated Myc signaling as an anti-PD-(L)1 resistance factor and we and others have shown Myc can be down-regulated by entinostat. Our data may provide a mechanistic basis for the responses seen with ENT + PEMBRO in this anti-PD-(L)1 pre-treated population.
Citation Format: Peter Ordentlich, Lei Wang, David Tamang, Amy H. Sullivan, Sally E. Church, Dan Rozelle, Michael L. Meyers, Suresh S. Ramalingam, Matthew D. Hellmann. Identification of gene signatures associated with response in a Phase II trial of entinostat (ENT) plus pembrolizumab (PEMBRO) in non-small cell lung cancer (NSCLC) patients whose disease has progressed on or after anti-PD-(L)1 therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr CT041.