American Association for Cancer Research
15357163mct130056-sup-fig_s1.pdf (87.18 kB)

Supplementary Figure S1 from Preclinical Evaluation of Transcriptional Targeting Strategy for Human Hepatocellular Carcinoma in an Orthotopic Xenograft Mouse Model

Download (87.18 kB)
journal contribution
posted on 2023-04-03, 14:07 authored by Kian Chuan Sia, Hung Huynh, Alexander Yaw Fui Chung, London Lucien Peng Jin Ooi, Kiat Hon Lim, Kam Man Hui, Paula Yeng Po Lam

Supplementary Figure S1 - PDF file 87K, Gene expression of AH-6CC-L2C in proliferating and G1-arrested PLC/PRF/5 cells. Cell cycle-regulated luciferase reporter activity in proliferating and G1-arrested PLC/PRF/5 cells after 48 h in A, post transfection with pAH-6CC-L2C or B, post infection with AH-6CC-L2C at MOI of 1. Total RLU was normalized with total protein contents and % of eGFP-positive cells representing the transfection efficiency. Data were presented as mean plus-minus SEM, n = 3. C, Corresponding cell cycle profiles of proliferating and G1-arrested PLC/PRF/5 cells as analyzed by FACS analysis



Gene regulation of many key cell-cycle players in S-, G2 phase, and mitosis results from transcriptional repression in their respective promoter regions during the G0 and G1 phases of cell cycle. Within these promoter regions are phylogenetically conserved sequences known as the cell-cycle–dependent element (CDE) and cell-cycle genes homology regions (CHR) sites. Thus, we hypothesize that transcriptional regulation of cell-cycle regulation via the CDE/CHR region together with liver-specific apolipoprotein E (apoE)-hAAT promoter could bring about a selective transgene expression in proliferating human hepatocellular carcinoma. We show that the newly generated vector AH-6CC-L2C could mediate hepatocyte-targeted luciferase gene expression in tumor cells and freshly isolated short-term hepatocellular carcinoma cultures from patient biopsy. In contrast, normal murine and human hepatocytes infected with AH-6CC-L2C expressed minimal or low luciferase activities. In the presence of prodrug 5-fluorocytosine (5-FC), AH-6CC-L2C effectively suppressed the growth of orthotopic hepatocellular carcinoma patient-derived xenograft mouse model via the expression of yeast cytosine deaminase (yCD) that converts 5-FC to anticancer metabolite 5-fluoruracil. More importantly, we show that combination treatment of AH-6CC-L2C with an EZH2 inhibitor, DZNep, that targets EpCAM-positive hepatocellular carcinoma, can bring about a greater therapeutic efficacy compared with a single treatment of virus or inhibitor. Our study showed that targeting proliferating human hepatocellular carcinoma cells through the transcriptional control of therapeutic gene could represent a feasible approach against hepatocellular carcinoma. Mol Cancer Ther; 12(8); 1651–64. ©2013 AACR.