1. Results Table 1: Factors associated with advanced liver fibrosis in univariable analysis among 216 chronic HCV patients Background There is a clinical need for more accurate non-invasive biochemical markers of hepatic fibrosis in chronic hepatitis C (CHC) infection. The hepatitis C virus (HCV) is known to disrupt lipid metabolism, making serum lipids and lipoproteins good candidates to explore as markers of the disease states and pathogenesis of hepatitis C infection. Apolipoprotein (Apo) A-I is currently used as a predictor in fibrosis biomarker algorithms, but other lipoprotein markers have not been extensively explored. Aim To investigated the association between a panel of serum lipoproteins and hepatic fibrosis in a tertiary center cohort of CHC patients. Conclusions A low level of serum Apo-CIII, a lipoprotein constituent of very low density lipoprotein (VLDL), was associated with severe liver fibrosis in this cohort of CHC patients. This was independent of the well- documented association with Apo A-I. Apo C-III may be a marker of disease progression and should be considered for further prospective evaluation as a component of noninvasive biomarker panels for predicting hepatic fibrosis in patients with CHC infection. Further studies are also needed to elucidate the mechanistic relationship between low Apo C-III levels and hepatic fibrosis which may be mediated through transcriptional regulatory factors or viral effects on VLDL metabolism. Methods We identified 216 CHC patients from our repository who had serum and liver biopsies that had been obtained concurrently. Nonfasting serum lipids were measured with direct LDL-C and HDL-C measurements. Apolipoprotein A-I, -B, -C-III and -E were analyzed by standard immunoturbidimetric methods in a lipid/lipoprotein specialty lab at Children’s Hospital in Boston, Massachusetts. Liver biopsies were reviewed by an experienced histopathologist at Duke University and scored by METAVIR stage. We tested the association between each lipoprotein and advanced fibrosis using univariable and multivariable logistic regression analysis. Serum Apolipoprotein C-III levels are independently associated with hepatic fibrosis in patients with chronic hepatitis C infection J. Rowell, M.D. 1, A. Thompson, M.D., Ph.D. 2, J.R. Guyton, M.D. 1, X. Q. Lao, M.D., Ph.D. 3, K. Patel, M.D. 2, J. McHutchison, M.D. 2, and J.J. McCarthy, Ph.D. 3 The MURDOCK Study. Institutions: 1) Department of Medicine, Division of Endocrinology, Metabolism and Nutrition, Duke University, Durham, NC, USA 2) Department of GI/Hepatology Research Program, Duke Clinical Research Institute, Duke University, Durham, NC, USA 3) Institute for Genome Sciences and Policy, Duke University, Durham, NC, USA Results Mild Fibrosis (N = 144 ) Advanced Fibrosis (N= 72) Unadjusted OR (95% CI) Unadj. P -value Discrete traits N (%) Female Sex (vs. Male)61 (42%)22 (31%)0.60 (0.33, 1.10)0.094 African American Race (vs. Caucasian) 38 (26%)24 (33%)1.40 (0.76, 2.6)0.289 HCV Genotype0.786 1117 (81%)58 (80%)Reference 2 14 (10%) 5 (7%)0.72 (0.25, 2.10) 3 5 (3%) 4 (6%)1.61 (0.42, 6.24) Other 8 (6%) 5 (7%)1.26 (0.40, 4.02) Quantitative traits Age (years) 47.3 ± 8.9 50.5 ± 5.81.53 (1.12, 2.10)0.008 BMI (kg/m 2 ) 28.4 ± 5.3 30.5 ± 6.41.46 (1.08, 1.96)0.013 Total Cholesterol (mg/dL)181.0 ± 42.8158.6 ± 42.10.57 (0.42, 0.79)0.0005 LDL-C (mg/dL)105.4 ± 37.7 90.1 ± 34.20.65 (0.48, 0.88)0.005 Apo B (mg/dL) 80.0 ± 22.7 75.0 ± 24.70.81 (0.61, 1.08)0.143 HDL (mg/dL) 49.6 ± 17.0 39.9 ± 15.90.52 (0.38, 0.73)0.0002 Apo A-I (mg/dL)143.9 ± 27.8125.0 ± 30.10.49 (0.36, 0.69)<.0001 Apo C-III (mg/dL)* 11.0 ± 4.9 8.4 ± 3.20.46 (0.32, 0.65)<.0001 Apo E (mg/dL)* 4.6 ± 1.5 4.3 ± 1.30.80 (0.60,1.07)0.130 Triglyceride (mg/dL)*132.5 ± 70.3137.8 ± 53.41.20 (0.90,1.60)0.209 High levels of Apo C-III, Apo A-I, total cholesterol, LDL-C, and HDL-C were associated with a decreased odds of advanced fibrosis in univariate analysis, whereas age and BMI showed the opposite effect. Apo A-I and HDL-C were highly correlated (r 2 = 0.88), as were total and LDL cholesterol (r 2 = 0.91), so one of each pair was chosen for inclusion in the multivariate model with other covariates. Adjusted OR (95% CI) Adjusted P-value Age (years) 1.57 (1.10, 2.23)0.013 BMI (kg/m 2 ) 1.55 (1.10, 2.19)0.013 LDL-C (mg/dL) 0.72 (0.51, 1.03)0.072 Apo A-I (mg/dL) 0.65 (0.45, 0.94)0.022 Apo C-III (mg/dL)* 0.49 (0.33, 0.72)0.0004 Table 2: Factors associated with advanced liver fibrosis in multivariable analysis among 216 chronic HCV patients In a multivariable model including age, BMI, LDL-C, Apo A-I and Apo C-III, the effect of Apo A-I was greatly attenuated, and Apo C-III emerged as the most significant factor associated with fibrosis (p=0.0004). The area under the receiver operator curve (AUROC) for predicting advanced fibrosis using Apo A-I and Apo C-III, respectively, is 0.670 and 0.684. The AUROC increased to 0.721 when both are included in the model. The AUROC increased to 0.779 when Apo A-I, Apo C-III, age, LDL-C, and BMI were included in the model. Figure 1 illustrates how the mean Apo C-III level significantly decreased with increasing METVIR fibrosis score (<0.0001). Figure 1: Mean Apo C-III ± Standard Error in 216 Chronic HCV Subjects *ln transformed for analysis Odds ratios are presented as the odds per unit (standard deviation) Metavir F0-F2 = mild fibrosis Metavir F3-F4 = advanced fibrosis Adjusted for age, race, sex, HCV genotype, and BMI *ln transformed for analysis Odds ratios are presented as the odds per unit (standard deviation) P < 0.0001 Acknowledgements This study was funded in part by a generous grant from the David H. Murdock Institute for Business and Culture via the M.U.R.D.O.C.K. Study and NIH CTSA award 1 UL 1 RR024128-01 to Duke University (Califf, PI)