Colorectal cancer (CRC) affects nearly 1.4 million individuals every year and is the third most common cause of deaths due to cancer. There is a need to identify novel prognostic biomarkers with accurate prognostic and therapeutic implications. In this study, we sought to identify prognostic gene expression signatures and explore their relationship with the tumor microenvironment. Under the IRB approved protocol, FFPE blocks of CRC patients were acquired from the Medical College of Georgia, Augusta, USA. Tumor regions and adjacent normal tissue regions were identified and marked by a board-certified pathologist. RNA isolation was performed from tumor sections (n = 88) and adjacent normal regions (n = 16). A panel of 17 genes with the highest perturbation in TCGA dataset was investigated on the FFPE tissues. The quantification of the mRNA molecules was performed using total RNA (300 ng) on the NanoString platform. It uses a specific molecular barcoding system to tag target nucleic acid molecules with very high sensitivity and specificity. The immunological dynamics of the prognostic genes were analyzed through TIMER (Tumor-Infiltrating Immune Cells) web portal using the TCGA-COAD dataset. In the clinical dataset, MCM4, YWHAB, LRRC59 and DPP7 were highly expressed in tumor tissue compared to adjacent control and were associated with worse prognosis (Log-rank test, p < 0.001*). However, PI4K2B, PCMT1 and PBX1P1 showed higher expression in adjacent normal tissue compared to tumor regions (Student's t-test, p < 0.001*). Further analysis using TIMER portal revealed that copy number alterations in YWHAB were significantly associated with decreased tumor infiltration of B cells, CD8+ T cells, neutrophils and dendritic cells (Wilcoxon test, p < 0.001*). The predominant alterations observed were arm-level gain and high amplification. Further, arm-level gain of MCM4 was associated with lower infiltration of B cells and CD8+ T cells (Wilcoxon test, p < 0.001*) and FBXO46 was associated with lower infiltration of B cells only (Wilcoxon test, p < 0.05*). Additional comparison between CRC tumor and adjacent normal tissue was explored using “DiffExp” module of TIMER web portal. The comparison showed that YWHAB, DPP7 and MCM4 were expressed at higher amounts in tumor tissues compared to adjacent normal tissue. These preliminary analyses, therefore reveal the significance of prognostic genes and their association with the infiltration of immune cells that in turn can be of therapeutic value. A comprehensive network of prognostic gene signatures and its perturbations in immune cell functioning can aid in efficient prognostic classification and design of new therapies.

Citation Format: Pankaj Kumar Ahluwalia, Ashis Mondal, Meenakshi Ahluwalia, Nikhil Sahajpal, Kimya Jones, Yasmeen Jilani, Allan Njau, Ravindra Kolhe. Exploring prognostic and immunological impact of a novel 4 gene signature in colorectal cancer patients [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4334.