BAP1 Loss Is Associated with Higher ASS1 Expression in Epithelioid Mesothelioma: Implications for Therapeutic Stratification

Our plain-English summary of BAP1 Loss Is Associated with Higher ASS1 Expression in Epithelioid Mesothelioma: Implications for Therapeutic Stratification, by S.E. Barnett, J. Kenyani, M. Tripari, et al (Molecular Cancer Research, 2023).

The study

• Authors: S.E. Barnett, J. Kenyani, M. Tripari, Z. Butt, R. Grosman, F. Querques, L. Shaw, L.C. Silva, Z. Goate, S.J. Marciniak, D.M. Rassl, R. Jackson, L.Y. Lian, P.W. Szlosarek, J.J. Sacco, J.M. Coulson
• Journal: Molecular Cancer Research
• Published: 2023;21(5):411-427
• Cited by: 14 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Unlabelled: The nuclear deubiquitylase BRCA1-associated protein 1 (BAP1) is frequently inactivated in malignant pleural mesothelioma (MPM) and germline BAP1 mutation predisposes to cancers including MPM. To explore the influence on cell physiology and drug sensitivity, we sequentially edited a predisposition mutation (w-) and a promoter trap (KO) into human mesothelial cells. BAP1w-/KO MeT5A cells express less BAP1 protein and phenocopy key aspects of BAP1 loss in MPM. Stable isotope labeling with amino acids in cell culture-mass spectrometry revealed evidence of metabolic adaptation, with concomitant alteration of cellular metabolites. In MeT5A, BAP1 deficiency reduces glycolytic enzyme levels but increases enzymes involved in the tricarboxylic acid cycle and anaplerotic pathways. Notably both argininosuccinate synthase 1 (ASS1), essential for cellular synthesis of arginine, and its substrate aspartate, are elevated in BAP1w-/KO MeT5A cells. Likewise, ASS1 expression is higher in BAP1-altered MPM cell lines, and inversely correlates with BAP1 in The Cancer Genome Atlas MESO dataset. Elevated ASS1 is also evident by IHC staining in epithelioid MPM lacking nuclear BAP1 expression, with improved survival among patients with BAP1-negative/ASS1-expressing tumors. Alterations in arginine metabolism may sensitize cells to metabolic drugs and we find that BAP1-negative/ASS1-expressing MPM cell lines are more sensitive to ASS1 inhibition, although not to inhibition of purine synthesis by mizoribine. Importantly, BAP1w-/KO MeT5A become desensitized to arginine deprivation by pegylated arginine deiminase (ADI-PEG20), phenocopying BAP1-negative/ASS1-expressing MPM cell lines.

Implications: Our data reveal an interrelationship between BAP1 and arginine metabolism, providing a potential means of identifying patients with epithelioid MPM likely to benefit from ADI-PEG20.

©2023 The Authors; Published by the American Association for Cancer Research.

In plain English

This laboratory study examined how BAP1 loss, frequent in malignant pleural mesothelioma (MPM), alters mesothelial cell metabolism and drug response. The authors genome-edited human MeT5A pleural mesothelial cells, adding a familial predisposition splice mutation and then a promoter trap, giving clones with catalytically active BAP1 protein reduced to about 60% (C3.1) and about 40% (C5.1) of parental levels. Proteomic (SILAC-MS) and NMR metabolomic profiling showed lower glycolytic enzymes and higher tricarboxylic acid cycle and anaplerotic enzymes, with argininosuccinate synthase 1 (ASS1) and aspartate raised. Across 21 patient-derived MPM cell lines, ASS1 protein and mRNA were higher where BAP1 was altered. In tissue microarrays of 164 epithelioid MPM cases (66% lacking nuclear BAP1), ASS1 was higher in tumours without nuclear BAP1 (n = 108) than with it (n = 56; Mann-Whitney P = 0.001), and ASS1 mRNA was higher in BAP1-altered cases in the TCGA MESO dataset (P = 0.0016). The BAP1-mutated MeT5A cells were resistant to ADI-PEG20 and more resistant than parental cells to mizoribine, while across the MPM cell lines mizoribine sensitivity did not track BAP1 or ASS1 status.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory study: genome-edited isogenic human mesothelial cell lines with proteomic/metabolomic profiling and drug assays, plus patient-derived MPM cell lines, tissue microarray immunohistochemistry and analysis of the TCGA MESO dataset
• Who was studied: Isogenic MeT5A mesothelial cell lines; a panel of 21 patient-derived MPM cell lines; tissue microarrays from 164 epithelioid MPM patients (125 male, 33 female, 6 with no gender recorded; median age 71, range 53-88) from Mesobank; and 82 cases in the TCGA MESO dataset
• Exposure: BAP1 status: an engineered w-family predisposition splice mutation plus a promoter-trap knockout in mesothelial cells, and naturally BAP1-altered versus BAP1-normal MPM cells and tumours; drugs tested were alpha-methyl-DL-aspartic acid (ASS1 inhibition), mizoribine and ADI-PEG20
• Main results:
  • In the edited MeT5A cells, catalytically active BAP1 protein was reduced to about 60% (C3.1) and about 40% (C5.1) of parental levels
  • Proteomics: 22% of detected proteins changed by more than 1.5-fold in the BAP1w-/KO clones compared with parental or heterozygous cells
  • In BAP1-deficient MeT5A cells, enzymes of glycogen and glutamine metabolism, the TCA cycle and anaplerotic pathways including the urea cycle were increased, while glycolytic enzymes were reduced
  • Across the panel of 21 MPM cell lines, ASS1 protein and mRNA levels tracked with BAP1 status, being higher in BAP1-altered lines
  • In the 164 epithelioid tumour samples, ASS1 was higher in tumours lacking nuclear BAP1 (n = 108) than in those with nuclear BAP1 (n = 56), Mann-Whitney P = 0.001
  • Survival analysis at the 18-month landmark: among nuclear-BAP1-negative cases, moderate/high ASS1 (n = 26) versus low ASS1 (n = 15) P = 0.003; among nuclear-BAP1-positive cases, moderate/high ASS1 (n = 10) versus low ASS1 (n = 11) P = 0.383
  • In the TCGA MESO cohort (n = 82), ASS1 transcript was significantly higher in BAP1-altered cases (Mann-Whitney P = 0.0016)
  • In the TCGA MESO cohort, BAP1 alteration was more frequent among tumours with higher ASS1 expression (Fisher exact test P = 0.0002)
  • TCGA survival by group (69 uncensored patients): BAP1-altered ASS1-high 21.6-33.9 months (n = 19), BAP1-altered ASS1-low 2.7-10.2 months (n = 9), BAP1-normal ASS1-high 10.1-19.3 months (n = 17), BAP1-normal ASS1-low 11.8-25.8 months (n = 24); log-rank P = 0.001
  • Among the MPM cell lines tested with ADI-PEG20, the BAP1-normal lines were sensitive and the BAP1-altered lines more resistant
  • In the isogenic model, the BAP1w-/KO MeT5A cells were resistant to arginine deprivation with ADI-PEG20, matching the BAP1-altered MPM line MESO-23T
  • In the isogenic MeT5A model, the BAP1w-/KO cells with elevated ASS1 were more resistant to mizoribine than the BAP1-normal cells
  • In the MPM cell lines, mizoribine sensitivity was not related to ASS1 expression or BAP1 status
• The authors' conclusion: The authors concluded that BAP1 loss is linked to higher ASS1 expression and to resistance to arginine deprivation, and proposed that BAP1 immunohistochemistry could help select patients with epithelioid MPM for ADI-PEG20, a hypothesis they state still needs further preclinical work
• Limitations the authors state:
  • The survival advantage seen for nuclear-BAP1-negative, ASS1-moderate/high tumours involved a small subgroup with very late curve separation
  • ASS1 enzymatic activity was not formally measured
  • The initial proteomic comparison used only two MPM cell lines, from which the authors say it is hard to generalize
  • The authors stated their finding of reduced glycolytic enzymes appears at odds with the Warburg effect
  • Other contributory factors to the ASS1-inhibition results, such as extracellular arginine concentration or genetic background, remain to be dissected
  • The authors could not exclude a contribution of other genomic differences in the edited cells to survival of the BAP1-mutated MeT5A clones

Topics

The National Library of Medicine indexes this study under subjects our site covers: Mesothelioma.

Medical terms in this study, explained in our Lexicon: BAP1, Epithelioid, Mesothelioma, Pleural Mesothelioma, BAP1 Tumor Predisposition Syndrome, Mesothelial Cells, Pegargiminase (ADI-PEG20).

Related studies

• Loss of the deubiquitylase BAP1 alters class I histone deacetylase expression and sensitivity of mesothelioma cells to HDAC inhibitors — J.J. Sacco, J. Kenyani, et al. (2015)
• Loss of BAP1 expression is associated with genetic mutation and can predict outcomes in gallbladder cancer — T. Hirosawa, M. Ishida, et al. (2018)
• Loss of NF2 Induces TGFβ Receptor 1-mediated Noncanonical and Oncogenic TGFβ Signaling: Implication of the Therapeutic Effect of TGFβ Receptor 1 Inhibitor on NF2 Syndrome — J.H. Cho, A.Y. Oh, et al. (2018)
• Loss of expression of BAP1 predicts longer survival in mesothelioma — M. Farzin, C.W. Toon, et al. (2015)
• Loss of expression of BAP1 is a useful adjunct, which strongly supports the diagnosis of mesothelioma in effusion cytology — J. Andrici, A. Sheen, et al. (2015)
• BAP1 Is Altered by Copy Number Loss, Mutation, and/or Loss of Protein Expression in More Than 70% of Malignant Peritoneal Mesotheliomas — N. Leblay, F. Leprêtre, et al. (2017)

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