Abstract 6642: Inter-alpha-trypsin inhibitor heavy chain (ITIH) genes: expressional profile in colorectal cancer and potential targeting by using a ribosome inactivating protein
Authors: Aiman Shahzad, Asim Pervaiz, Afraz Numan, Sana Iqbal, Martin R. Berger
Journal: Cancer Research (2025)

Abstract Introduction: Expression deregulation of Inter-α-Trypsin Inhibitor Heavy (ITIH) genes at circulatory and tissue levels has been reported in various cancers. These altered levels are linked with worse pathogenesis and low treatment response. ITIH genes are being considered as novel prognostic/ therapeutic targets. Knowledge about ITIH expression profile in primary/metastatic cancers and regulation control via synthetic/natural anticancer compounds deemed crucial. Present study was designed to uncover these facets in colorectal cancer. Materials &amp; Methods: Total RNA was extracted from frozen colorectal cancer clinical isolates and normal mucosa specimens followed by cDNA synthesis and real-time PCR for expressional analysis of the genes. Labelled rat colorectal cancer cells (CC531) were transplanted into the rat liver via the hepatic portal vein and were re-isolated by FACS after discrete time intervals (3, 6, 9, 14 and 21 days) followed by RNA extraction and cDNA microarray analysis. A fraction of re-isolated cells was cultured in vitro for 14 and 22 days to compare the results with those from tumor cells grown in vivo. Primary (SW480 and HCT116) and metastatic (SW620) colorectal cancer cell lines were exposed to riproximin (10 and 20ng/ml, 48 hours) followed by RNA extraction, cDNA synthesis, expressional profiling of the genes (ITIH3 and ITIH4) and fold change identification by 2ΔΔCT method. The results were compared with those of untreated control cells. Results: Both members of the ITIH family were continuously down-regulated in the clinical isolates. However, ITIH3 gene was more substantially down-regulated (-26fold average, Stage II) in the tissue samples of colorectal cancer patients as compared to ITIH4 gene (-9fold average, Stage II). In vivo model mimicking the colorectal cancer liver metastasis depicted a marked increase in the expression of both genes throughout the experimental period (21 days). Precisely, a maximum induction was observed in ITIH3 gene at day 03 of implantation (295fold), while ITIH4 was maximally induced at day 21 (246fold). Exposure with the plant protein (riproximin) induced the expression of ITIH3 gene in the three colorectal cancer cell lines. In contrast, riproximin exposure inhibited the ITIH4 gene in the three colorectal cancer at all applied concentrations. Conclusion: ITIH3 and ITIH4 genes were consistently down-regulated in clinical isolates. In contrast, a substantial induction was observed in metastatic colorectal liver cancer animal model. Riproximin induced expressional alterations in gene specific format in the colorectal cancer cell lines. Further investigations are needed to understand substantial high levels of ITIH genes during colorectal cancer liver metastasis and potential targeting via riproximin. Citation Format: Aiman Shahzad, Asim Pervaiz, Afraz Numan, Sana Iqbal, Martin R. Berger. Inter-alpha-trypsin inhibitor heavy chain (ITIH) genes: expressional profile in colorectal cancer and potential targeting by using a ribosome inactivating protein [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 6642.