Low Exposures to Amphibole or Serpentine Asbestos in Germline Bap1-mutant Mice Induce Mesothelioma Characterized by an Immunosuppressive Tumor Microenvironment

Our plain-English summary of Low Exposures to Amphibole or Serpentine Asbestos in Germline Bap1-mutant Mice Induce Mesothelioma Characterized by an Immunosuppressive Tumor Microenvironment, by Y. Kadariya, E. Sementino, M. Ruan, et al (Cancer Research Communications, 2024).

The study

• Authors: Y. Kadariya, E. Sementino, M. Ruan, M. Cheung, P. Hadikhani, H.U. Osmanbeyoglu, A.J. Klein-Szanto, K. Cai, J.R. Testa
• Journal: Cancer Research Communications
• Published: 2024;4(4):1004-1015
• Cited by: 8 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Unlabelled: Asbestos and BAP1 germline mutations are risk factors for malignant mesothelioma (MM). While it is well accepted that amphibole asbestos is carcinogenic, the role of serpentine (chrysotile) asbestos in MM has been debated. To address this controversy, we assessed whether minimal exposure to chrysotile could significantly increase the incidence and rate of MM onset in germline Bap1-mutant mice. With either crocidolite or chrysotile, and at each dose tested, MMs occurred at a significantly higher rate and earlier onset time in Bap1-mutant mice than in wild-type littermates. To explore the role of gene-environment interactions in MMs from Bap1-mutant mice, we investigated proinflammatory and protumorigenic factors and the tumor immune microenvironment (TIME). IHC and immunofluorescence staining showed an increased number of macrophages in granulomatous lesions and MMs. The relative number of CD163-positive (CD163+) M2 macrophages in chrysotile-induced MMs was consistently greater than in crocidolite-induced MMs, suggesting that chrysotile induces a more profound immunosuppressive response that creates favorable conditions for evading immune surveillance. MMs from Bap1-mutant mice showed upregulation of CD39/CD73-adenosine and C-C motif chemokine ligand 2 (Ccl2)/C-C motif chemokine receptor 2 (Ccr2) pathways, which together with upregulation of IL6 and IL10, promoted an immunosuppressive TIME, partly by attracting M2 macrophages. Interrogation of published human MM RNA sequencing (RNA-seq) data implicated these same immunosuppressive pathways and connections with CD163+ M2 macrophages. These findings indicate that increased M2 macrophages, along with upregulated CD39/CD73-adenosine and Ccl2/Ccr2 pathways, contribute to an immunosuppressive TIME in chrysotile-induced MMs of Bap1-mutant mice, suggesting that immunotherapeutic strategies targeting protumorigenic immune pathways could be beneficial in human BAP1 mutation carriers who develop MM.

Significance: We show that germline Bap1-mutant mice have enhanced susceptibility to MM upon minimal exposure to chrysotile asbestos, not only amphibole fibers. Chrysotile induced a more profound immune tumor response than crocidolite in Bap1-mutant mice by upregulating CD39/CD73-adenosine and Ccl2/Ccr2 pathways and recruiting more M2 macrophages, which together contributed to an immunosuppressive tumor microenvironment. Interrogation of human MM RNA-seq data revealed interconnected immunosuppressive pathways consistent with our mouse findings.

© 2024 The Authors; Published by the American Association for Cancer Research.

In plain English

In this animal experiment, germline Bap1 heterozygous mutant mice (a knockout model, and a knock-in nonsense-mutation model for the lowest crocidolite dose) and wild-type (WT) littermates on an FVB background were injected intraperitoneally with crocidolite or chrysotile asbestos and followed for malignant mesothelioma (MM). With a total of 0.8 mg crocidolite, MM occurred in 19 of 21 (90%) Bap1-mutant mice versus 12 of 22 (55%) WT mice (median survival 56 versus 72 weeks); at 0.1 mg total, incidence was 47% (9/19) versus 13% (2/16). With chrysotile, incidence was 62% (16/26) versus 17% (4/23) at 3.2 mg total and 16 of 18 (89%) versus 5 of 15 (33%) at 0.4 mg total, with earlier death in mutant mice at every dose. Tumors were mostly biphasic. In tumors from Bap1-mutant mice, staining showed few T, B and natural killer cells but abundant macrophages, and CD163-positive M2 macrophages, CD39 and Ccr2 were higher in chrysotile-induced than crocidolite-induced tumors; among mice with chrysotile-induced MM, ascites IL6 and IL10 were higher in Bap1-mutant than WT mice. The authors concluded that chrysotile as well as crocidolite can be carcinogenic in this model and induces an immunosuppressive tumor microenvironment.

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: animal (mouse) carcinogenicity experiment using two germline Bap1 heterozygous models (knockout and knock-in), with tumor immune microenvironment analyses, plus re-analysis of published human mesothelioma RNA-seq data
• Who was studied: Germline Bap1 heterozygous mutant mice and wild-type littermates on an FVB background: a knockout model (deletion of exons 6 and 7) in three asbestos experiments and a knock-in model (nonsense mutation in exon 16) for the 0.1 mg crocidolite experiment; mice injected at 8-10 weeks of age, cohorts of 15-26 per group, experiments run over a 4-year period at Fox Chase Cancer Center. Also 100 human mesothelioma RNA-seq samples from a published NCI dataset, 11 with germline BAP1 mutations
• Exposure: Intraperitoneal injections of crocidolite (amphibole) at 0.1 or 0.0125 mg per injection, 8 injections every 21 days (totals 0.8 and 0.1 mg/mouse), or chrysotile (serpentine) at 0.05 mg or 0.4 mg per injection, 8 injections (totals 0.4 and 3.2 mg/mouse)
• Main results:
  • At a total crocidolite dose of 0.8 mg/mouse, peritoneal MM occurred in 19 of 21 (90%) Bap1+/- mice versus 12 of 22 (55%) WT mice, with median survival 56 weeks versus 72 weeks
  • At a total crocidolite dose of 0.1 mg/mouse (knock-in model), MM incidence was 47% (9/19) in Bap1+/- mice versus 13% (2/16) in WT mice; median survival 73 versus 83 weeks
  • At a total chrysotile dose of 3.2 mg/mouse, MM incidence was 62% (16/26) in Bap1+/- mice versus 17% (4/23) in WT mice, with median survival 25 versus 31 weeks
  • At a total chrysotile dose of 0.4 mg/mouse, MM occurred in 16 of 18 (89%) Bap1+/- mice versus 5 of 15 (33%) WT mice; median survival 65 versus 77 weeks
  • Tumor histology in both crocidolite- and chrysotile-induced tumors was mostly biphasic with predominant sarcomatoid morphology
  • Few T, B or natural killer cells were seen in crocidolite- or chrysotile-induced tumors from Bap1-mutant mice, while macrophages (F4/80) were abundant
  • In tumors from Bap1-mutant mice, CD163-positive M2 macrophages were a significantly higher percentage in chrysotile-induced than crocidolite-induced tumors (P less than 0.005), while CD206 staining did not differ significantly (P greater than 0.05)
  • CD39 and Ccr2 staining was markedly increased in chrysotile-induced compared with crocidolite-induced tumors of Bap1-mutant mice
  • In ascites from mice with chrysotile-induced MM (4 Bap1+/- and 5 Bap1+/+), IL6 and IL10 were significantly elevated in Bap1-mutant compared with WT mice
  • Immunoblotting of six MMs from chrysotile-exposed Bap1+/- mice showed loss or greatly reduced Bap1 with retained p16Ink4a; the single MM from a WT mouse expressed Bap1 but lost detectable p16Ink4a
  • In the published human dataset of 100 mesotheliomas, tumors from patients with germline BAP1 mutations had higher CCL2 expression than sporadic tumors (P = 0.043); gene symbols are OCR-damaged in the text
• The authors' conclusion: The authors concluded that chrysotile, like crocidolite, can be carcinogenic in this mouse model, inducing a profound inflammatory and immunosuppressive response in germline Bap1 heterozygous mice even at low doses, and they drew parallels suggesting BAP1 mutation carriers may be highly susceptible to minimal amphibole or serpentine asbestos exposure
• Limitations the authors state:
  • The study was not designed to compare the carcinogenic potency of crocidolite and chrysotile directly, and the low doses used for the two fiber types were not the same
  • Competing deaths from adhesion-related intestinal obstruction and liver fibrosis, especially at the higher chrysotile dose, may have prevented mesothelioma from developing in some animals
  • Fibers were given intraperitoneally, which is not the usual route of human exposure, though the authors state it is standard for mouse asbestos studies
  • The human data analysis is presented by the authors as preliminary

Topics

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

Medical terms in this study, explained in our Lexicon: Amphibole Group, Serpentine Group, BAP1, Mesothelioma, Tumor Microenvironment, Germline Mutation, Malignant Mesothelioma, Amphibole Asbestos.

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