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ObjectiveWe previously reported a characterisation of the hepatocellular carcinoma (HCC) immune contexture and described an immune-specific class. We now aim to further delineate the immunogenomic classification of HCC to incorporate features that explain responses/resistance to immunotherapy.DesignWe performed RNA and whole-exome sequencing, T-cell receptor (TCR)-sequencing, multiplex immunofluorescence and immunohistochemistry in a novel cohort of 240 HCC patients and validated our results in other cohorts comprising 660 patients.ResultsOur integrative analysis led to define: (1) the inflamed class of HCC (37%), which includes the previously reported immune subclass (22%) and a new immune-like subclass (15%) with high interferon signalling, cytolytic activity, expression of immune-effector cytokines and a more diverse T-cell repertoire. A 20-gene signature was able to capture ~90% of these tumours and is associated with response to immunotherapy. Proteins identified in liquid biopsies recapitulated the inflamed class with an area under the ROC curve (AUC) of 0.91; (2) The intermediate class, enriched in TP53 mutations (49% vs 29%, p=0.035), and chromosomal losses involving immune-related genes and; (3) the excluded class, enriched in CTNNB1 mutations (93% vs 27%, p<0.001) and PTK2 overexpression due to gene amplification and promoter hypomethylation. CTNNB1 mutations outside the excluded class led to weak activation of the Wnt-βcatenin pathway or occurred in HCCs dominated by high interferon signalling and type I antigen presenting genes.ConclusionWe have characterised the immunogenomic contexture of HCC and defined inflamed and non-inflamed tumours. Two distinct CTNNB1 patterns associated with a differential role in immune evasion are described. These features may help predict immune response in HCC.

Mucopolysaccharidoses (MPS) are caused by deficiency of lysosomal enzyme activities needed to degrade glycosaminoglycans (GAGs), which are long unbranched polysaccharides consisting of repeating disaccharides. GAGs include: chondroitin sulfate (CS), dermatan sulfate (DS), heparan sulfate (HS), keratan sulfate (KS), and hyaluronan. Their catabolism may be blocked singly or in combination depending on the specific enzyme deficiency. There are 11 known enzyme deficiencies, resulting in seven distinct forms of MPS with a collective incidence of higher than 1 in 25,000 live births. Accumulation of undegraded metabolites in lysosomes gives rise to distinct clinical syndromes. Generally, the clinical conditions progress if untreated, leading to developmental delay, systemic skeletal deformities, and early death. MPS disorders are potentially treatable with enzyme replacement therapy or hematopoietic stem cell transplantation. For maximum benefit of available therapies, early detection and intervention are critical. We recently developed a novel high-throughput multiplex method to assay DS, HS, and KS simultaneously in blood samples by using high performance liquid chromatography/tandem mass spectrometry for MPS. The overall performance metrics of HS and DS values on MPS I, II, and VII patients vs. healthy controls at newborns were as follows using a given set of cut-off values: sensitivity, 100%; specificity, 98.5–99.4%; positive predictive value, 54.5–75%; false positive rate, 0.62–1.54%; and false negative rate, 0%. These findings show that the combined measurements of these three GAGs are sensitive and specific for detecting all types of MPS with acceptable false negative/positive rates. In addition, this method will also be used for monitoring therapeutic efficacy. We review the history of GAG assay and application to diagnosis for MPS.

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