Co-Targeting FASN and mTOR Suppresses Uveal Melanoma Growth.

Uveal melanoma (UM) displays a high frequency of metastasis; however, effective therapies for metastatic UM are limited. Identifying unique metabolic features of UM may provide a potential targeting strategy. A lipid metabolism protein expression signature was induced in a normal choroidal melanocyte (NCM) line transduced with GNAQ (Q209L), a driver in UM growth and development. Consistently, UM cells expressed elevated levels of fatty acid synthase (FASN) compared to NCMs. FASN upregulation was associated with increased mammalian target of rapamycin (mTOR) activation and sterol regulatory element-binding protein 1 (SREBP1) levels. FASN and mTOR inhibitors alone significantly reduced UM cell growth. Concurrent inhibition of FASN and mTOR further reduced UM cell growth by promoting cell cycle arrest and inhibiting glucose utilization, TCA cycle metabolism, and de novo fatty acid biosynthesis. Our findings indicate that FASN is important for UM cell growth and co-inhibition of FASN and mTOR signaling may be considered for treatment of UM.

Multi-omics Profiling Shows BAP1 Loss Is Associated with Upregulated Cell Adhesion Molecules in Uveal Melanoma.

BRCA1-associated protein 1 (BAP1) is a tumor suppressor gene that is mutated in cancer, including uveal melanoma. Loss-of-function BAP1 mutations are associated with uveal melanoma metastasis and poor prognosis, but the mechanisms underlying these effects remain unclear. Upregulation of cell-cell adhesion proteins is involved with collective migration and metastatic seeding of cancer cells. Here, we show that BAP1 loss in uveal melanoma patient samples is associated with upregulated gene expression of multiple cell adhesion molecules (CAM), including E-cadherin (CDH1), cell adhesion molecule 1 (CADM1), and syndecan-2 (SDC2). Similar findings were observed in uveal melanoma cell lines and single-cell RNA-sequencing data from uveal melanoma patient samples. BAP1 reexpression in uveal melanoma cells reduced E-cadherin and CADM1 levels. Functionally, knockdown of E-cadherin decreased spheroid cluster formation and knockdown of CADM1 decreased growth of BAP1-mutant uveal melanoma cells. Together, our findings demonstrate that BAP1 regulates the expression of CAMs which may regulate metastatic traits. IMPLICATIONS: BAP1 mutations and increased metastasis may be due to upregulation of CAMs.

The AMP-dependent kinase pathway is upregulated in BAP1 mutant uveal melanoma.

Metastatic uveal melanoma (UM) responds poorly to targeted therapies and immune checkpoint inhibitors. Loss of BRCA1-associated protein 1 (BAP1) via inactivating mutations in the BAP1 gene is associated with UM progression. Thus, molecular alterations caused by BAP1 dysfunction may be novel therapeutic targets for metastatic UM. Here, we found that phosphorylation of AMP-dependent kinase (AMPK) was elevated in BAP1-altered (or mutant) compared to BAP1-unaltered (or wild-type [WT]) UM tumors. As a readout of AMPK pathway activation, phosphorylation of an AMPK downstream effector, acetyl-CoA-carboxylase (ACC), was also elevated. BAP1 re-expression in BAP1-null UM cell lines decreased phospho-AMPK (pAMPK) and phospho-ACC (pACC) levels. AMPK phosphorylation is mediated by calcium/calmodulin dependent protein kinase kinase 2 (CaMKK2) and potentially liver kinase B1 (LKB1) in BAP1 mutant UM cells. Knockdown of AMPKα1/2 reduced the viability of BAP1 mutant UM cells, indicating a survival function of AMPK in BAP1 mutant UM. Our data suggest that the AMPK pathway is an important mechanism mediating the survival of BAP1 mutant UM. Targeting the AMPK pathway may be a novel therapeutic strategy for metastatic UM.

Pyruvate dehydrogenase inactivation causes glycolytic phenotype in BAP1 mutant uveal melanoma.

Effective therapeutic options are still lacking for uveal melanoma (UM) patients who develop metastasis. Metastatic traits of UM are linked to BRCA1-associated protein 1 (BAP1) mutations. Cell metabolism is re-programmed in UM with BAP1 mutant UM, but the underlying mechanisms and opportunities for therapeutic intervention remain unclear. BAP1 mutant UM tumors have an elevated glycolytic gene expression signature, with increased expression of pyruvate dehydrogenase (PDH) complex and PDH kinase (PDHK1). Furthermore, BAP1 mutant UM cells showed higher levels of phosphorylated PDHK1 and PDH that was associated with an upregulated glycolytic profile compared to BAP1 wild-type UM cells. Suppressing PDHK1-PDH phosphorylation decreased glycolytic capacity and cell growth, and induced cell cycle arrest of BAP1 mutant UM cells. Our results suggest that PDHK1-PDH phosphorylation is a causative factor of glycolytic phenotypes found in BAP1 mutant UM and propose a therapeutic opportunity for BAP1 mutant UM patients.

BAP1 mutant uveal melanoma is stratified by metabolic phenotypes with distinct vulnerability to metabolic inhibitors.

Cancer cell metabolism is a targetable vulnerability; however, a precise understanding of metabolic heterogeneity is required. Inactivating mutations in BRCA1-associated protein 1 (BAP1) are associated with metastasis in uveal melanoma (UM), the deadliest adult eye cancer. BAP1 functions in UM remain unclear. UM patient sample analysis divided BAP1 mutant UM tumors into two subgroups based on oxidative phosphorylation (OXPHOS) gene expression suggesting metabolic heterogeneity. Consistent with patient data, transcriptomic analysis of BAP1 mutant UM cell lines also showed OXPHOShigh or OXPHOSlow subgroups. Integrated RNA sequencing, metabolomics, and molecular analyses showed that OXPHOShigh BAP1 mutant UM cells utilize glycolytic and nucleotide biosynthesis pathways, whereas OXPHOSlow BAP1 mutant UM cells employ fatty acid oxidation. Furthermore, the two subgroups responded to different classes of metabolic suppressors. Our findings indicate that targeting cancer metabolism is a promising therapeutic option for BAP1 mutant UM; however, tailored approaches may be required due to metabolic heterogeneities.

Metabolic Adaptations to MEK and CDK4/6 Cotargeting in Uveal Melanoma.

Frequent GNAQ and GNA11 mutations in uveal melanoma hyperactivate the MEK-ERK signaling pathway, leading to aberrant regulation of cyclin-dependent kinases (CDK) and cell-cycle progression. MEK inhibitors (MEKi) alone show poor efficacy in uveal melanoma, raising the question of whether downstream targets can be vertically inhibited to provide long-term benefit. CDK4/6 selective inhibitors are FDA-approved in patients with estrogen receptor (ER)-positive breast cancer in combination with ER antagonists/aromatase inhibitors. We determined the effects of MEKi plus CDK4/6 inhibitors (CDK4/6i) in uveal melanoma. In vitro, palbociclib, a CDK4/6i, enhanced the effects of MEKi via downregulation of cell-cycle proteins. In contrast, in vivo CDK4/6 inhibition alone led to cytostasis and was as effective as MEKi plus CDK4/6i treatment at delaying tumor growth. RNA sequencing revealed upregulation of the oxidative phosphorylation (OxPhos) pathway in both MEKi-resistant tumors and CDK4/6i-tolerant tumors. Furthermore, oxygen consumption rate was increased following MEKi + CDK4/6i treatment. IACS-010759, an OxPhos inhibitor, decreased uveal melanoma cell survival in combination with MEKi + CDK4/6i. These data highlight adaptive upregulation of OxPhos in response to MEKi + CDK4/6i treatment in uveal melanoma and suggest that suppression of this metabolic state may improve the efficacy of MEKi plus CDK4/6i combinations.