Gonzalo Sapisochín

The liver is the largest solid organ in the body and is critical for metabolic and immune functions. However, little is known about the cells that make up the human liver and its immune microenvironment. Here we report a map of the cellular landscape of the human liver using single-cell RNA sequencing. We provide the transcriptional profiles of 8444 parenchymal and non-parenchymal cells obtained from the fractionation of fresh hepatic tissue from five human livers. Using gene expression patterns, flow cytometry, and immunohistochemical examinations, we identify 20 discrete cell populations of hepatocytes, endothelial cells, cholangiocytes, hepatic stellate cells, B cells, conventional and non-conventional T cells, NK-like cells, and distinct intrahepatic monocyte/macrophage populations. Together, our study presents a comprehensive view of the human liver at single-cell resolution that outlines the characteristics of resident cells in the liver, and in particular provides a map of the human hepatic immune microenvironment.

IMPORTANCE: Several factors are associated with increased hepatocellular carcinoma (HCC) recurrence after liver transplantation (LT), but no reliable risk score has been established to determine the individual risk for HCC recurrence. OBJECTIVE: We aimed to develop and validate a Risk Estimation of Tumor Recurrence After Transplant (RETREAT) score for patients with HCC meeting Milan criteria by imaging. DESIGN, SETTING, AND PARTICIPANTS: Predictors of recurrence were tested in a development cohort of 721 patients who underwent LT between 2002 and 2012 at 3 academic transplant centers (University of California-San Francisco; Mayo Clinic, Rochester; and Mayo Clinic, Jacksonville) to create the RETREAT score. This was subsequently validated in a cohort of 341 patients also meeting Milan criteria by imaging who underwent LT at the University of Toronto transplant center using the C concordance statistic and net reclassification index. MAIN OUTCOMES AND MEASURES: Characteristics associated with post-LT HCC recurrence. RESULTS: A total of 1061 patients participated in the study; 77.8% (825) were men, and the median (IQR) age was 58.2 (53.3-63.9) years in the development cohort and 56.4 (51.7-61.0) years in the validation cohort (P < .001). In the development cohort of 721 patients (542 men), median α-fetoprotein (AFP) level at the time of LT was 8.3 ng/mL; 9.4% had microvascular invasion (n = 68), and 22.1% were beyond Milan criteria on explant (n = 159) owing to understaging by pretransplantation imaging. Cumulative probabilities of HCC recurrence at 1 and 5 years were 5.7% and 12.8%, respectively. On multivariable Cox proportional hazards regression, 3 variables were independently associated with HCC recurrence: microvascular invasion, AFP at time of LT, and the sum of the largest viable tumor diameter and number of viable tumors on explant. The RETREAT score was created using these 3 variables, with scores ranging from 0 to 5 or higher that were highly predictive of HCC recurrence (C statistic, 0.77). RETREAT was able to stratify 5-year post-LT recurrence risk ranging from less than 3% with a score of 0 to greater than 75% with a score of 5 or higher. The validation cohort (n = 340; 283 men) had significantly higher microvascular invasion (23.8% [n = 81], P < .001), explant beyond Milan criteria (37.3% [n = 159], P < .001), and HCC recurrence at 5 years (17.9% [n = 159], P = .03). RETREAT showed good model discrimination (C statistic, 0.82; 95% CI, 0.77-0.86) and superior recurrence risk classification compared with explant Milan criteria (net reclassification index, 0.40; P = .001) in the validation cohort. CONCLUSIONS AND RELEVANCE: We have developed and validated a simple and novel prognostic score that may improve post-LT HCC surveillance strategies and help identify patients who may benefit from future adjuvant therapies.

The selection of liver transplant candidates with hepatocellular carcinoma (HCC) relies mostly on tumor size and number. Instead of relying on these factors, we used poor tumor differentiation and cancer-related symptoms to exclude patients likely to have advanced HCC with aggressive biology. We initially reported similar 5-year survival for patients whose tumors exceeded (M+ group) and were within (M group) the Milan criteria. Herein, we validate our original data with a new prospective cohort and report the long-term follow-up (10-years) using an intention-to-treat analysis. The previously published study (cohort 1) included 362 listed (294 transplanted) patients from January 1996 to August 2008. The validation cohort (cohort 2) includes 243 listed (105 M+ group, 76 beyond University of California San Francisco criteria; 210 transplanted) patients from September 2008 to December 2012. Median follow-up from listing was 59.7 (26.8-103) months. For the validation cohort 2, the actuarial survival from transplant for the M+ group was similar to that of the M group at 1 year, 3 years, and 5 years: 94%, 76%, and 69% versus 95%, 82%, and 78% (P = 0.3). For the combined cohorts 1 and 2, there were no significant differences in the 10-year actuarial survival from transplant between groups. On an intention-to-treat basis, the dropout rate was higher in the M+ group and the 5-year and 10-year survival rates from listing were decreased in the M+ group. An alpha-fetoprotein level >500 ng/mL predicted poorer outcomes for both the M and M+ groups. CONCLUSION: Tumor differentiation and cancer-related symptoms of HCC can be used to select patients with advanced HCC who are appropriate candidates for liver transplantation; alpha-fetoprotein level limitations should be incorporated in the listing criteria for patients within or beyond the Milan criteria. (Hepatology 2016;64:2077-2088).