BackgroundPleural malignant mesothelioma (MM) is a deadly tumour predominantly associated with asbestos exposure. A reliable diagnostic and prognostic marker for MM will significantly enhance clinical care and is an area of intense research. Soluble mesothelin is the most studied and an FDA-approved biomarker for MM. A recent report showed promising results using fibulin-3 as a new diagnostic marker. The aim of this study was to compare the utility of fibulin-3 versus mesothelin, singly or in combination.MethodsFibulin-3 and soluble mesothelin were determined by ELISA in the plasma and pleural fluid of 153 patients presenting with a pleural effusion including 82 with MM, 36 with non-MM malignant effusions and 35 with benign effusions. Biomarker concentrations were determined in the plasma of an additional 49 cases with benign asbestos-related disease.ResultsMesothelin provides better diagnostic accuracy than fibulin-3 for MM whether measured in plasma or pleural effusion: area under the curve (AUC) for plasma was 0.822 (95% CI 0.76 to 0.87) compared with 0.671 (0.61 to 0.73), respectively, and for pleural fluid AUC was 0.815 (0.74 to 0.87) compared with 0.588 (0.51 to 0.67), respectively. Effusion fibulin-3 was an independent significant prognostic factor for survival in MM patients; HR 2.08 (1.14 to 3.82), p=0.017. MM patients with effusion fibulin-3 levels below the median survived significantly longer than those with levels above the median (14.1 vs 7.9 months, p=0.012). Mesothelin and neutrophil to lymphocyte ratio were not significant prognostic markers.ConclusionsSoluble mesothelin is a superior diagnostic biomarker for MM compared with fibulin-3, whereas fibulin-3 provides superior prognostic information compared with mesothelin.

A key to improving cancer immunotherapy will be the identification of tumor-specific “neoantigens” that arise from mutations and augment the resultant host immune response. In this study we identified single nucleotide variants (SNVs) by RNA sequencing of asbestos-induced murine mesothelioma cell lines AB1 and AB1-HA. Using the NetMHCpan 2.8 algorithm, the theoretical binding affinity of predicted peptides arising from high-confidence, exonic, non-synonymous SNVs was determined for the BALB/c strain. The immunoreactivity to 20 candidate mutation-carrying peptides of increased affinity and the corresponding wild-type peptides was determined using interferon-γ ELISPOT assays and lymphoid organs of non-manipulated tumor-bearing mice. A strong endogenous immune response was demonstrated to one of the candidate neoantigens, Uqcrc2; this response was detected in the draining lymph node and spleen. Antigen reactive cells were not detected in non-tumor bearing mice. The magnitude of the response to the Uqcrc2 neoantigen was similar to that of the strong influenza hemagglutinin antigen, a model tumor neoantigen. This work confirms that the approach of RNAseq plus peptide prediction and ELISPOT testing is sufficient to identify natural tumor neoantigens.

Malignant mesothelioma (MM) is a uniformly fatal tumour caused predominantly by exposure to asbestos. It is not known why some exposed individuals get mesothelioma and others do not. There is some epidemiological evidence of host susceptibility. BAP1 gene somatic mutations and allelic loss are common in mesothelioma and recently a BAP1 cancer syndrome was described in which affected individuals and families had an increased risk of cancer of multiple types, including MM. To determine if BAP1 mutations could underlie any of the sporadic mesothelioma cases in our cohort of patients, we performed targeted deep sequencing of the BAP1 exome on the IonTorrent Proton sequencer in 115 unrelated MM cases. No exonic germline BAP1 mutations of known functional significance were observed, further supporting the notion that sporadic germline BAP1 mutations are not relevant to the genetic susceptibility of MM.

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