Because only a small fraction of asbestos-exposed individuals develop malignant mesothelioma, and because mesothelioma clustering is observed in some families, we searched for genetic predisposing ...factors. We discovered germline mutations in the gene encoding BRCA1 associated protein-1 (BAP1) in two families with a high incidence of mesothelioma, and we observed somatic alterations affecting BAP1 in familial mesotheliomas, indicating biallelic inactivation. In addition to mesothelioma, some BAP1 mutation carriers developed uveal melanoma. We also found germline BAP1 mutations in 2 of 26 sporadic mesotheliomas; both individuals with mutant BAP1 were previously diagnosed with uveal melanoma. We also observed somatic truncating BAP1 mutations and aberrant BAP1 expression in sporadic mesotheliomas without germline mutations. These results identify a BAP1-related cancer syndrome that is characterized by mesothelioma and uveal melanoma. We hypothesize that other cancers may also be involved and that mesothelioma predominates upon asbestos exposure. These findings will help to identify individuals at high risk of mesothelioma who could be targeted for early intervention.
Malignant mesotheliomas are highly aggressive tumors usually caused by exposure to asbestos. Germline-inactivating mutations of BAP1 predispose to mesothelioma and certain other cancers. However, why ...mesothelioma is the predominate malignancy in some BAP1 families and not others, and whether exposure to asbestos is required for development of mesothelioma in BAP1 mutation carriers are not known. To address these questions experimentally, we generated a Bap1(+/-) knockout mouse model to assess its susceptibility to mesothelioma upon chronic exposure to asbestos. Bap1(+/-) mice exhibited a significantly higher incidence of asbestos-induced mesothelioma than wild-type (WT) littermates (73% vs. 32%, respectively). Furthermore, mesotheliomas arose at an accelerated rate in Bap1(+/-) mice than in WT animals (median survival, 43 weeks vs. 55 weeks after initial exposure, respectively) and showed increased invasiveness and proliferation. No spontaneous mesotheliomas were seen in unexposed Bap1(+/-) mice followed for up to 87 weeks of age. Mesothelioma cells from Bap1(+/-) mice showed biallelic inactivation of Bap1, consistent with its proposed role as a recessive cancer susceptibility gene. Unlike in WT mice, mesotheliomas from Bap1(+/-) mice did not require homozygous loss of Cdkn2a. However, normal mesothelial cells and mesothelioma cells from Bap1(+/-) mice showed downregulation of Rb through a p16(Ink4a)-independent mechanism, suggesting that predisposition of Bap1(+/-) mice to mesothelioma may be facilitated, in part, by cooperation between Bap1 and Rb. Drawing parallels to human disease, these unbiased genetic findings indicate that BAP1 mutation carriers are predisposed to the tumorigenic effects of asbestos and suggest that high penetrance of mesothelioma requires such environmental exposure.
Malignant pleural mesothelioma (MPM), an aggressive cancer of the mesothelial cells lining the pleural cavity, lacks effective treatments. Multiple somatic mutations and copy number losses in tumor ...suppressor genes (TSGs)
,
, and
are frequently associated with MPM. The impact of single versus multiple genomic alterations of TSG on MPM biology, the immune tumor microenvironment, clinical outcomes, and treatment responses are unknown. Tumors with genomic alterations in
alone were associated with a longer overall patient survival rate compared to tumors with
and/or
alterations with or without
and formed a distinct immunogenic subtype with altered transcription factor and pathway activity patterns.
genomic alterations consistently contributed to an adverse clinical outcome. Since the genomic alterations of only
was associated with the PD-1 therapy response signature and higher
and
gene expression, it might be a candidate marker for immune checkpoint blockade therapy. Our results on the impact of TSG genotypes on MPM and the correlations between TSG alterations and molecular pathways provide a foundation for developing individualized MPM therapies.
The data presented in this article are companion materials to our manuscript titled “BAP1 maintains HIF-dependent interferon beta induction to suppress tumor growth in clear cell renal cell ...carcinoma” (Langbein et al., 2022), where we investigated the downstream effects of BAP1 (BRCA1-associated protein 1) expression in clear cell renal cell carcinoma (ccRCC) cell lines and mouse xenograft models. In the manuscript, we showed that BAP1 upregulates STING (stimulator of interferon genes) expression and activity in ccRCC cells, leading to IFN-β transcription and activation of interferon stimulated gene factor 3 (ISGF3), the transcription factor that mediates the effects of type I interferons (IFNs). Here, we suppressed additional components of the type I IFN pathway, including IRF9 (a component of ISGF3), IFNAR1 (the type I IFN receptor), and STING (a stimulator of IFN production) by shRNA to investigate their involvement in BAP1-mediated upregulation of ISGF3 activity. We also inhibited extracellular IFN-β via neutralizing antibody treatment in BAP1-expressing cells to ascertain the role of the secreted cytokine in this pathway. ISGF3 activity was assessed by western blot analysis and qPCR measurement of its transcriptional targets. To examine the relevance of our observations in another model system, we characterized primary kidney cells from WT and Bap1fl/fl mice by cytokeratin 8 immunohistochemistry and examined the effect of Bap1 knockout on Sting protein expression. Finally, we treated mice bearing BAP1 knockdown xenografted tumors with diABZI, a STING agonist, and measured immune cell recruitment via CD45 immunohistochemistry. These data can serve as a starting point for further investigation on the roles of BAP1 and other tumor suppressor genes in interferon pathway regulation.
Abstract
There is irrefutable evidence that germline BRCA1-associated protein 1 gene (BAP1) mutations contribute to malignant mesothelioma (MM) susceptibility. However, BAP1 mutations are not found ...in all cases with evidence of familial MM or in other high-risk cancer families affected by various cancers, including MM. The goal of this study was to use whole genome sequencing (WGS) to determine the frequency and types of germline gene variants occurring in 12 MM patients who were selected from a series of 141 asbestos-exposed MM patients with a family history of cancer but without a germline BAP1 mutation. WGS was also performed on two MM cases, a proband and sibling, from a previously reported family with multiple cases of MM without the inheritance of a predisposing BAP1 mutation. Altogether, germline DNA sequencing variants were identified in 21 cancer-related genes in 10 of the 13 probands. Germline indel, splice site and missense mutations and two large deletions were identified. Among the 13 MM index cases, 6 (46%) exhibited one or more predicted pathogenic mutations. Affected genes encode proteins involved in DNA repair (ATM, ATR, BRCA2, BRIP1, CHEK2, MLH3, MUTYH, POLE, POLE4, POLQ and XRCC1), chromatin modification (ARID1B, DNMT3A, JARID2 and SETD1B) or other cellular pathways: leucine-rich repeat kinase 2 gene (LRRK2) (two cases) and MSH4. Notably, somatic truncating mutation or deletions of LRRK2 were occasionally found in MMs in The Cancer Genome Atlas, and the expression of LRRK2 was undetectable or downregulated in a majority of primary MMs and MM cell lines we examined, implying that loss of LRRK2 expression is a newly recognized tumor suppressor alteration in MM.
Abstract
Constitutive activation of AKT is a frequent occurrence in the development of human T-cell acute lymphocytic leukemia/lymphoma (T-ALL), due largely to inactivation of PTEN. Up regulation of ...MYC is also commonly observed in T-ALL. We previously demonstrated that transgenic expression of a constitutively active form of Akt2 alone is sufficient to initiate T-cell lymphoma in mice, and that tumor formation typically requires up regulation of c-Myc or Dlx5 caused by specific chromosomal rearrangements. Here we report that transgenic overexpression of Dlx5 specifically in the mouse thymus can also drive T-cell lymphomagenesis. Tumors from these mice were characterized by loss of Pten expression and trisomy 15. Proliferation of these lymphoma cells was highly sensitive to the Akt pathway inhibitors BEZ235 and RAD001 as well as to the Myc inhibitor JQ1. Additionally, BEZ235 was found to cooperate with JQ1 to induce a more profound effect on cell cycle arrest and apoptosis. Moreover, BEZ235 potentiated vincristine-induced apoptosis when cells were treated with both drugs simultaneously, whereas pretreatment with BEZ235 antagonized the cell-killing effect of vincristine. These findings suggest that combination therapies with AKT and MYC inhibitors hold promise for the treatment of T-ALL in the clinic.
Citation Format: Yinfei Tan, Eleonora Sementino, Joseph Testa. Akt and Myc cooperate to promote cell proliferation in Lck-Dlx5 T-cell lymphomas. abstract. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2543. doi:10.1158/1538-7445.AM2015-2543
The Dlx5 homeobox gene was first implicated as an oncogene in a T-ALL mouse model expressing myristoylated (Myr) Akt2. Furthermore, overexpression of Dlx5 was sufficient to drive T-ALL in mice by ...directly activating Akt and Notch signaling. These findings implied that Akt2 cooperates with Dlx5 in T-cell lymphomagenesis. To test this hypothesis, Lck-Dlx5;Lck-MyrAkt2 transgenic mice were generated. MyrAkt2 synergized with Dlx5 to greatly accelerate and enhance the dissemination of T-lymphomagenesis. RNA-seq analysis performed on lymphomas from Lck-Dlx5;Lck-MyrAkt mice revealed upregulation of genes involved in the Wnt and cholesterol biosynthesis pathways. Combined RNA-seq and ChIP-seq analysis of lymphomas from Lck-Dlx5;Lck-MyrAkt mice demonstrated that β-catenin directly regulates genes involved in sterol regulatory element binding transcription factor 2 (Srebf2)-cholesterol synthesis. These lymphoma cells had high Lef1 levels and were highly sensitive to β-catenin and Srebf2-cholesterol synthesis inhibitors. Similarly, human T-ALL cell lines with activated NOTCH and AKT and elevated LEF1 levels were sensitive to inhibition of β-catenin and cholesterol pathways. Furthermore, LEF1 expression positively correlated with expression of genes involved in the cholesterol synthesis pathway in primary human T-ALL specimens. Together, these data suggest that targeting β-catenin and/or cholesterol biosynthesis, together with AKT, could have therapeutic efficacy in a subset of T-ALL patients.
Pleural malignant mesothelioma is a therapy-resistant cancer affecting the serosal lining of the thoracic cavity. Mutations/deletions of
, and
are the most frequent genetic lesions in human malignant ...mesothelioma. We introduced various combinations of these deletions in the pleura of conditional knockout (CKO) mice, focusing on the contribution of
loss. While homozygous CKO of
, or
alone gave rise to few or no malignant mesotheliomas, inactivation of
cooperated with loss of either
or
to drive development of malignant mesothelioma in approximately 20% of double-CKO mice, and a high incidence (22/26, 85%) of malignant mesotheliomas was observed in
(triple)-CKO mice. Malignant mesothelioma onset was rapid in triple-CKO mice, with a median survival of only 12 weeks, and malignant mesotheliomas from these mice were consistently high-grade and invasive. Adenoviral-Cre treatment of normal mesothelial cells from
CKO mice, but not from mice with knockout of one or any two of these genes, resulted in robust spheroid formation
, suggesting that mesothelial cells from
mice have stem cell-like potential. RNA-seq analysis of malignant mesotheliomas from triple-CKO mice revealed enrichment of genes transcriptionally regulated by the polycomb repressive complex 2 (PRC2) and others previously implicated in known Bap1-related cellular processes. These data demonstrate that somatic inactivation of
, and
results in rapid, aggressive malignant mesotheliomas, and that deletion of
contributes to tumor development, in part, by loss of PRC2-mediated repression of tumorigenic target genes and by acquisition of stem cell potential, suggesting a potential avenue for therapeutic intervention. SIGNIFICANCE: Combinatorial deletions of
, and
result in aggressive mesotheliomas, with
loss contributing to tumorigenesis by circumventing PRC2-mediated repression of oncogenic target genes.
Malignant mesothelioma (MM) is an incurable cancer of the serosal lining that is often caused by exposure to asbestos. Therefore, novel agents for the prevention and treatment of this disease are ...urgently needed. Asbestos induces the release of pro-inflammatory cytokines such as IL-1β and IL-6, which play a role in MM development. IL-6 is a component of the JAK-STAT3 pathway that contributes to inflammation-associated tumorigenesis. Glycoprotein 130 (gp130), the signal transducer of this signaling axis, is an attractive drug target because of its role in promoting neoplasia via the activation of downstream STAT3 signaling. The anticancer drug, SC144, inhibits the interaction of gp130 with the IL-6 receptor (IL6R), effectively blunting signaling from this inflammatory axis. To test whether the inflammation-related release of IL-6 plays a role in the formation of MM, we evaluated the ability of SC144 to inhibit asbestos-induced carcinogenesis in a mouse model. The ability of sulindac and anakinra, an IL6R antagonist/positive control, to inhibit MM formation in this model was tested in parallel. Asbestos-exposed Nf2+/-;Cdkn2a+/- mice treated with SC144, sulindac or anakinra showed significantly prolonged survival compared to asbestos-exposed vehicle-treated mice. STAT3 activity was markedly decreased in MM specimens from SC144-treated mice. Furthermore, SC144 inhibited STAT3 activation by IL-6 in cultured normal mesothelial cells, and in vitro treatment of MM cells with SC144 markedly decreased the expression of STAT3 target genes. The emerging availability of newer, more potent SC144 analogs showing improved pharmacokinetic properties holds promise for future trials, benefitting individuals at high risk of this disease.
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