Expression of genes associated with poor prognosis from hepatocellular carcinoma are increased in peripheral blood from Hmong with active hepatitis B infection

BACKGORUND & AIMS: Hepatocellular carcinoma (HCC) disproportionally affects Asian Americans due to the high prevalence of chronic hepatitis B virus (HBV) infection and among Asian ethnic groups. The Hmong are diagnosed with HCC at a younger age, with more advanced stage disease, and have the highest cause specific mortality of any Asian ethnic group. Poor survival from HCC has also been associated with differential gene expression in HCC tumors leading us to hypothesize that quantitative traits loci lead to differences in the expression of genes in Hmong which reduce their HCC survival. METHODS: We performed bulk RNA-sequencing of whole blood total RNA from HBV-infected Hmong (n=30) and non-Hmong Asians (Chinese (n=46), and Vietnamese (n=19)). RESULTS: No significant differences between groups were identified in demographic, clinical, or viral factors. A total of 1,491 differentially-expressed transcripts (DETs) were identified across the 3 ethnic groups. When restricted to those with an HBV viral load >2,000 IU/L, 768 DETs were identified and accurately separated individuals by ethincity. Among 273 DETs identified by functional enrichment and pathway analysis and associated with annotated sets of HCC-related genes, 9 were found in 2 gene sets highly expressed in HCC with poor survival. After adjusting for age and sex, expression levels of 4 of these, including CD164, ECT2, HDAC2, and UBE2T, remained significantly elevated in Hmong compared to Chinese and Vietnamese. CONCLUSIONS: These results suggest expression quantitative trait loci predispose Hmong to a more agrressive form of HCC and may explain the poor survival among this Asian ethnic group. Overall design: The goal of these studies was to utilize comprehensive transcriptomic profiling in order to determine the differences in gene expression associated with hepatocellular carcinoma (HCC) survival in Laotian/Hmong relative to Chinese and Vietnamese. For this study, we analyzed a total of 97 whole blood RNA samples obtained from a patient cohort consisting of Laotian/Hmong (n=30), Chinese (n=46), and Vietnamese (n=19) patients with chronic hepatitis B. Of these, 42 patients were considered to have active disease (i.e., viral load >2,000 IU/ml). Blood was drawn into PAXgene Blood RNA vacutainer tubes (BD Biosciences) and total RNA isolated using the PAXgene Blood RNA Kit (Qiagen). Indexed, directional RNA-Seq libraries were then prepared from total RNA (1.0 ug) using the TruSeq Stranded Total RNA Library Prep Kit with Ribo-Zero Globin (Illumina, San Diego, CA). All libraries (n=97) were pooled and then multiplex sequenced on an Illumina HiSeq 4000 Sequencing System (100bp, paired-end, 95 samples/lane over 8 lanes).