{"paper_id":"772dc209-41f9-4749-9d6d-256768568ed4","body_text":"Peritoneal mesothelioma (PM) is a rare malignant neoplasm that originates from the mesothelial cells lining the peritoneal serosa. PM was first reported in the early 1900s as a diffuse intraperitoneal neoplastic process associated with ascites in a young woman ( 1 ). The neoplasm represents 15-20% ( 2 ) of all mesotheliomas and shares some features with the most common pleural counterpart even if several substantial differences make it a separate and definite nosological entity. From an epidemiological point of view, PM more frequently affects females than males, with an earlier median age (52 years) ( 3 ) than pleural mesothelioma.\nPM usually arises as multiple serosal nodules with thickening of the peritoneum. Adipose tissue invasion and stromal invasion represent indicative features of malignancy ( 4 ). Histologically, it is classified into three histotypes: a) epithelioid, with round monomorphic cells arranged into different architectural patterns; b) sarcomatoid, composed of spindle elements; and c) biphasic, with at least 10% of both components. Well-differentiated papillary mesothelioma is included among the mesothelial tumours of the peritoneum, which represents a rare variant with an indolent behaviour that occurs mainly in women of reproductive age ( 2 ,  5 ). The diagnostic algorithm mirrors that of the pleural diagnostic algorithm, with a combination of positivity for mesothelial markers and negativity for BAP1 (BRCA1 associated protein 1) in 40-60% of cases. The histologic variants are clinically relevant, allowing a prognostic stratification of patients and guiding the treatment strategies ( 6 ).\nThe link between asbestos exposure is weaker than that in pleural tumours. Even if asbestos exposure represents the most important risk factor ( 7 ), it is found in approximately 33-50% of cases compared to the frequency of over 90% in pleural mesothelioma. Furthermore, the latency period between asbestos exposure and the development of mesothelioma is 20 years for PM, compared to 30-40 years for pleural mesothelioma ( 8 ,  9 ). Asbestos fibres have been identified in the omentum and mesentery of the gastrointestinal tract ( 10 ). Various hypotheses have been formulated to explain the mechanisms by which asbestos fibres can reach the peritoneal cavity. A study showed significant incidences of pulmonary asbestosis (17%) and pleural plaques (26%) in a cohort of patients with PM, suggesting a link between pleuropulmonary and peritoneal diseases probably secondary to the migration of asbestos fibres ( 11 ). Another hypothesis concerns the migration of asbestos fibres through the female genital system from the uterus to the fallopian tubes up to the peritoneal cavity. Such contamination could take place following sexual intercourse or following the application of talc contaminated with asbestos, as described by some papers that have focused on mesotheliomas that originate from the germinative epithelium of the ovary, which represents a specialization of mesothelial cells ( 12 ). Other rare causes related to the onset of PM are those attributable to chronic inflammatory states of the peritoneal serosa, such as chronic peritonitis, recurrent Mediterranean fever, germline mutations of  BRCA  (BReast CAncer), endometriosis, the period following radiation therapy for other peritoneal or pelvic neoplastic processes, and simian vacuolating virus infection ( 13 – 15 ).\nThe strict association between asbestos exposure and the development of malignant mesothelioma is indisputable. Our understanding of the mechanisms of action of asbestos fibres and their effects on mesothelial cells has deepened since the middle of the twentieth century. Knowledge about carcinogenesis related to asbestos has increased together with awareness about the molecular changes in this tumour. Mesothelioma appears to be characterized by chromosome rearrangements and gene mutations/deletions ( 16 ). More recently, the molecular landscape of mesothelioma has been enriched by the discovery of susceptibility familial factors that influence the impact of asbestos, target mutations in oncogenes and tumour suppressor genes, and epigenetic changes ( 17 ,  18 ).\nAs treatment options for PM, as well as pleural PM, are currently limited and target therapies are far away from being available, a better understanding of the molecular pathogenesis could suggest further therapeutic opportunities. In this review, we aimed to report recent research in the last 20 years focusing on the most promising molecular pathways and prognostic factors in PM in terms of diagnostic and therapeutic implications.\n\nA comprehensive review of the literature of the last decade was conducted in the Medline database, including research with the generic terms “peritoneal mesothelioma” AND “molecular”. All data were further confirmed by examining the list manually. The eligibility of studies was assessed by reading titles, abstracts, and full texts.\nIn  \n Tables 1 \n  and  \n 2 \n  are respectively listed in chronological order the research studies and the case reports in which molecular analyses of PM were performed. Below, the most important molecular alterations that have emerged are discussed and grouped into “Oncogenes”, “Tumour suppressor genes” and “Post-transcriptional alterations” sections. Two additional sections are dedicated to the description of the role of the tumour microenvironment and on therapeutic approaches in PM.\nPrevious molecular research studies.\nB, biphasic; E, epithelioid; FF, fresh-frozen; FFPE, formalin-fixed paraffin-embedded; FISH, Fluorescent In Situ Hybridization; IHC, immunohistochemistry; NGS, next-generation sequencing; NA, not available; qRT-PCR, quantitative real-time polymerase chain reaction; PCR, polymerase chain reaction; RNAseq, REN-sequencing; S, sarcomatoid; TMA, tissue micro-array; WDPM, well differentiated papillary mesothelioma; WES, whole-exome sequencing; WGS, whole-genome sequencing; WTS, whole-transcriptome sequencing.\nPrevious published case reports with molecular studies.\nArray CGH, Array comparative genomic hybridization; B, biphasic; E, epithelioid; FF, fresh-frozen; FFPE, formalin-fixed paraffin-embedded; FISH, Fluorescent In Situ Hybridization; NGS, next-generation sequencing; NA, not available; PCR, polymerase chain reaction; RNAseq, RNA-sequencing; S, sarcomatoid; WES, whole-exome sequencing; WGS, whole-genome sequencing.\nReceptor tyrosine kinases are surface receptors that link growth factors, cytokines, and hormones, functioning as key regulators of normal cellular processes. Mutations in receptor tyrosine kinases alter signalling cascades, leading to dysregulation of protein expression.\nEGFR  has been investigated in the pathogenesis of PM. Foster et al. ( 19 ) found  EGFR  (Epidermal Growth Factor Receptor) mutations, both in L858R and other catalytic domains, in a high percentage of PM patients. In different studies, ligand-dependent activation (e.g.,  HER2 - Human epidermal growth factor receptor 2,  HER3 , Axl, and  MET -mesenchymal epithelial transition factor) and coactivation of  EGFR  and  PDGFRB  (Platelet Derived Growth Factor Receptor Beta) were shown, together with cooperation of these receptors with the mTOR (mammalian target of rapamycin) pathway, suggesting the potential efficacy of the combined inhibition of these cascades in PM ( 20 ,  32 ). In this direction, a patient with multicystic PM ( 52 ) and two patients with papillary PM ( 53 ) were successfully treated with rapamycin, an mTOR inhibitor. The mTOR signalling pathway closely interacts with  PI3K  (Phosphoinositide 3-kinases). Varghese et al. ( 21 ) detected the overexpression of genes in these pathways in patients with the shortest survival in a group of 41 PM patients treated with surgical cytoreduction and regional intraoperative chemotherapy perfusion. In subsequent studies,  EGFR  alterations emerged as more complex than somatic mutations, with the detection of silent polymorphisms and cooperation with other receptors, such as the formation of heterodimers, showing that PM do not harbour somatic mutations in the  EGFR  tyrosine kinase domain that would make them sensitive to molecularly targeted therapy ( 22 ).\nIn 2016, Loharamtaweethong et al. ( 44 ) described an anecdotic case of  ALK  (Anaplastic lymphoma kinase)-rearranged PM in a child, thus opening a new perspective in the molecular dissection of this entity. The presence of  ALK  alteration was the topic of investigation for the group of Hung et al. ( 29 ), who detected this new promising pathogenetic mechanism in 3% of patients, mostly young women. This epidemiologic distribution of  ALK  rearrangement was confirmed in more recent studies ( 38 ,  48 ,  51 ), all representing by striatin ( STRN )- ALK  fusion, an extremely rare  ALK  rearrangement reported in only 56 cancers of different organs ( 51 ). This evidence suggests the need to explore the alteration in such groups, for the possible use of molecular target therapies.\nIn the complex scenario of hidden alterations, some studies focused on alterations that could also be present in mesothelioma, translating the experience of other better characterized neoplasms. This is the case of  EWSR1  (EWS RNA Binding Protein 1) rearrangements. In 2013, Panagopoulos et al. ( 54 ) detected a specific fusion gene in mesothelioma. These results were further confirmed by Desmeules et al. ( 26 ), who associated  EWSR1  alteration with a unique subset of mesothelioma arising in young, nonexposed,  BAP1 -retaining patients, resulting in an undirect activation of c- MET  gene.\nWT1  (Wilm’s tumor 1) gene mutations have been rarely described in PM, especially in those non-asbestos related ( 43 ). The immunohistochemical expression of WT1, which represents a useful diagnostic tool to assess the mesothelial origin of neoplastic cells, could also have a prognostic role associated with a better prognosis, as described by Pezzuto et al. ( 37 ).\nTumour suppressor genes regulate cell division and replication, thus leading to growth abnormalities when mutated, and their function is lost or reduced. This event seems to occur frequently in the development of PM.  BAP1  plays a key role in PM susceptibility and oncogenesis. Alakus et al. ( 23 ) described for the first time that the loss of  BAP1  occurred in PM in the absence of any other oncogenic drivers, such as  NF2  (neurofibromatosis type 2) and  CDKN2A  (Cyclin Dependent Kinase Inhibitor 2A). Although these genes have also been detected in the pleural form, some differences have been found between the two entities ( 25 ). This is mainly true for the prevalence of these alterations. In PM, a higher frequency of  BAP1  and a lower frequency of  CDKN2A  and  NF2  have been described ( 27 ,  55 ,  56 ). This could be related to the lower frequency in the peritoneum of biphasic and sarcomatoid mesothelioma that typically harbour these mutations ( 38 ). While  BAP1  was independent of the clinical outcome, the latter two have a negative prognostic significance, thus becoming interesting targets for therapeutic approaches. In contrast, Leblay et al. found that BAP1 protein nuclear expression mirrored molecular status, and its detection was a good and reliable prognostic marker for the complete loss of  BAP1  activity in PM ( 28 ). Particularly, the authors found a better overall survival for patients with  BAP1  mutations, protein expression loss, or at least one of these alterations independently of tumour histological subtype, age, and sex. In terms of biomarker discovery, Lai et al. ( 42 ) identified a tumour-specific neoantigen for  BAP1  following insertion of a frameshift mutation translated into a truncated protein which was predicted to be presented by the patient’s HLA-B molecule as a tumour-specific neo-antigen. A separate issue is represented by  BAP1  germline mutations. Together with somatic mutations, a  BAP1 -related cancer syndrome characterized by mesothelioma, uveal melanoma, and possibly other cancer types has been identified. This possibility should be taken into account when identifying patients at high risk. One group described a significant proportion of patients with mesothelioma carry germline mutations in cancer susceptibility genes, especially those with PM, absent asbestos exposure, second cancer diagnosis, and young age ( 31 ). As for the germline mutations of  NF2 , only very rare reports described the onset of a PM in the context of a type 2 neurofibromatosis ( 50 ). Two cases of PM have been described respectively associated with Cowden ( 46 ) and Li-Fraumeni syndrome ( 40 ). Von Hippel-Lindau (VHL) disease tumour suppressor gene  VHL  was found mutated in a unique case of clear cell epithelioid PM in a non-exposed women ( 49 ). Two tumour suppressor genes,  TRAF7  and  CDC42 , respectively involved in activation of mitogen‐ activated protein kinases (MAPKs) and in Rho GTPase signalling, were found mutually exclusively mutated in a series of well-differentiated papillary mesothelioma of the peritoneum ( 35 ), in absence of the typical mutations of the malignant counterpart, as those involving  BAP1 ,  NF2 ,  CDKN2A ,  ALK , contributing to a clear-cut separation between the two entities. These last exceptional cases show how a specific molecular signature could correspond an unusual morphological variant and this should be kept in mind especially for diagnostic purposes to avoid misinterpretations.\nAlthough the genomics of PM has deepened, much less is known about the epigenomic landscape of this tumour. Starting from the evidence of the pleural form, epigenetic alterations in PM have also been suggested to contribute to carcinogenesis. Hama et al. ( 39 ) quantitatively analysed the methylation of  KAZALD1  (Kazal Type Serine Peptidase Inhibitor Domain 1),  TMEM308  (Tumor Microenvironment of Metastasis 308), and  MAPK13  (Mitogen-Activated Protein Kinase 13) and reported hypermethylation of these genes in PM. The authors found a correlation between  KAZALD1  and the sarcomatoid variant, showing a certain histotype distribution of molecular alterations. Bozzi et al. focused on epithelioid PM, where tumour cells were characterized by stemness and plasticity supported by epigenetic reprogramming in the context of mesenchymal epithelial reverse transcription. Thus, the authors suggesting that the PM is likely to be responsive to epigenetic regulator inhibitors, basing also on the inverse correlation between strong  EZH2  expression and the loss of the  BAP1  ( 24 ). One group studied splicing alterations in PM reporting an upregulation of spliceosomal genes with a high expression of  SF3B1  (Splicing factor 3B subunit 1) which correlated with a worse prognosis ( 34 ). An important posttranslational modification to target for the development of new therapeutic approaches for cancer treatment is ubiquitination. Recent studies have observed that ubiquitination is involved in the metabolic reprogramming of cancer cells. In a recent study, Serio et al. found several losses among which loss of function of ubiquitination and defensins in PM ( 30 ,  45 ,  57 ), suggesting an important role in the initial development and progression of neoplasia or in combination with other mechanisms.\nThe interactions between tumour cells and immune cells are complex. This strict association is well recognized in several thoracic malignancies, such as lung cancer ( 58 ,  59 ). An increasing emphasis has been attributed to the immune milieu and thus to the employment of immunotherapy against mesothelioma. Most research has been conducted on the pleural form ( 60 ,  61 ), while much less is known about the characterization of the TME in the peritoneal form. It seems that the role of the TME is not detached from MM carcinogenesis. Shrestha et al. ( 33 ) found high checkpoint receptor activation in  BAP1  haploinsufficient PM, thus suggesting predictive value for tumour response to this marker. Similarly, White et al. ( 62 ) demonstrated a significant increase in PD-L1 (Programmed death-ligand 1) expression in PM patients with a high mutational burden and germline mutations.\n\nStrictly connected to the topic of molecular alterations in PM is the development of new treatment strategies. To date, MM remains a rare cancer with only a few promising changes in treatment. This is mainly true for PM, as most of the knowledge is extrapolated from the pleural form. Together with the deeper awareness of the genomic alterations, therapeutic strategies targeting new pathways have been explored in the pleura with multiple trials available. As previously described,  BAP1  inactivating mutations are frequently detected, with itself or its multiple downstream pathways considered fascinating targets. In this scenario,  PARP  ( 63 ) and  EZH2  (Enhancer of zeste homolog 2) ( 64 ) inhibitors have been investigated in  BAP1 -negative tumours, with disappointing results. Similarly,  CDKN2A , commonly deleted in mesothelioma with the loss of p16 protein expression, has been targeted in a clinical trial ( 65 ) with only limited success. Another well-known inactivation concerns  NF2 , leading to the loss of merlin protein and the dysregulation of several streams, among which the Hippo-YAP/TAZ pathways ( 66 ), potentially blocked by mTOR inhibitors ( 52 ,  53 ), warrant further analysis. An attractive target has also been found in mesothelin, a membrane glycoprotein that has been blocked in epithelioid pleural mesothelioma ( 67 – 69 ). Regarding rare alterations, anecdotal reports about  ALK  fusions in PM need to be reported in terms of the response to ALK inhibitors, such as in lung cancers ( 48 ). Immunotherapy deserves a separate discussion and is widely studied in MM, for which the best results have been obtained ( 70 ) with different administration strategies, alone or in combination, which represents a considerable goal.\n\nThe molecular events that cause PM have not been clearly defined, and comprehensive genetic characterization remains challenging. Important steps have been made towards the definition of the molecular signature of neoplasia to identify therapeutic targets, along with the goals of other tumours. The recent published studies, particularly those based on next-generation or other high-throughput sequencing methodologies, show heterogenous molecular alterations, mostly involving  BAP1  and other DNA repair, chromatin, and cell cycle regulators. Awareness of the molecular and physiological changes leading to the neoplastic degeneration of mesothelial cells and the identification of gene mutations, epigenetic alterations, gene expression profiles and altered pathways could be helpful for selecting targetable mechanisms and molecules.\n\nConceptualization, FF, FP, GS, and LV. Methodology, AM and FP. Data curation DC, DR, and Ad’A. Writing—original draft preparation FF and FP. Writing—review and editing GS. Supervision, GS and LV. All authors have read and agreed to the published version of the manuscript.\n\nThe authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.\n\nAll claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.","source_license":"CC-BY-4.0","license_restricted":false}