BAP1 is required prenatally for differentiation and maintenance of postnatal murine enteric nervous system.

Epigenetic regulatory mechanisms are underappreciated, yet are critical for enteric nervous system (ENS) development and maintenance. We discovered that fetal loss of the epigenetic regulator Bap1 in the ENS lineage caused severe postnatal bowel dysfunction and early death in Tyrosinase-Cre Bap1fl/fl mice. Bap1-depleted ENS appeared normal in neonates; however, by P15, Bap1-deficient enteric neurons were largely absent from the small and large intestine of Tyrosinase-Cre Bap1fl/fl mice. Bowel motility became markedly abnormal with disproportionate loss of cholinergic neurons. Single-cell RNA sequencing at P5 showed that fetal Bap1 loss in Tyrosinase-Cre Bap1fl/fl mice markedly altered the composition and relative proportions of enteric neuron subtypes. In contrast, postnatal deletion of Bap1 did not cause enteric neuron loss or impaired bowel motility. These findings suggest that BAP1 is critical for postnatal enteric neuron differentiation and for early enteric neuron survival, a finding that may be relevant to the recently described human BAP1-associated neurodevelopmental disorder.

PRAME induces genomic instability in uveal melanoma.

PRAME is a CUL2 ubiquitin ligase subunit that is normally expressed in the testis but becomes aberrantly overexpressed in many cancer types in association with aneuploidy and metastasis. Here, we show that PRAME is expressed predominantly in spermatogonia around the time of meiotic crossing-over in coordination with genes mediating DNA double strand break repair. Expression of PRAME in somatic cells upregulates pathways involved in meiosis, chromosome segregation and DNA repair, and it leads to increased DNA double strand breaks, telomere dysfunction and aneuploidy in neoplastic and non-neoplastic cells. This effect is mediated at least in part by ubiquitination of SMC1A and altered cohesin function. PRAME expression renders cells susceptible to inhibition of PARP1/2, suggesting increased dependence on alternative base excision repair pathways. These findings reveal a distinct oncogenic function of PRAME that can be targeted therapeutically in cancer.

Vitreous metastasis from cutaneous melanoma: diagnosis and management / Metástase vítrea de melanoma cutâneo: diagnóstico e tratamento

ABSTRACT Purpose: To report the clinical findings, treatments, and outcomes in a series of patients with vitreous metastasis from cutaneous melanoma. Methods: This single-center, retrospective, interventional case series included patients with biopsy-confirmed vitreous metastasis from cutaneous melanoma diagnosed between 1997 and 2020. Standard 23- or 25-gauge pars plana vitrectomy was performed for diagnostic sampling. Sclerotomies were treated with double or triple freeze-thaw cryotherapy. Perioperative intravitreal injections of melphalan (32 µg/0.075 mL) were administered, when indicated. Visual acuity, intraocular pressure, and systemic and ocular treatment responses were reported. Results: Five eyes of five patients with unilateral vitreous metastasis from cutaneous melanoma were identified. The median age at diagnosis was 84 (range, 37-88) years. The median follow-up after ophthalmic diagnosis was 28 (8.5-36) months; one patient did not have a follow-up. The initial visual acuity ranged from 20/30 to hand motions. Baseline clinical findings included pigmented or non-pigmented cellular infiltration of the vitreous (5/5), anterior segment (4/5), and retina (3/5). Four patients had secondary glaucoma. Systemic therapy included checkpoint inhibitor immunotherapy (n=3, all with partial/complete response), systemic chemotherapy (n=2), surgical resection (n=3), and radiation (n=2). The median time from primary diagnosis to vitreous metastasis was 2 (2-15) years. One patient had an active systemic disease at the time of vitreous metastasis. The final visual acuity ranged from 20/40 to no light perception. Ophthalmic treatment included vitrectomy in all five patients, intravitreal administration of melphalan in three, and intravitreal administration of methotrexate in one. One patient required enucleation, and histopathology revealed extensive invasion by melanoma cells. Conclusions: Vitreous metastasis from cutaneous melanoma can present as a diffuse infiltration of pigmented or non-pigmented cells into the vitreous and may be misdiagnosed as uveitis. Diagnostic pars plana vitrectomy and periodic intravitreal chemotherapy may be indicated.

RESUMO Objetivo: Descrever os achados clínicos, tratamentos, e desfechos em uma série de pacientes com me tástases vítreas de melanoma cutâneo. Métodos: Série retrospectiva de casos de único centro com intervenção. Pacientes incluídos tiveram seu diagnóstico de MVMC confirmado por biópsia entre 1997 e 2020. Vitrectomia via pars plana com 23 ou 25 gauge foram realizadas para obter espécimens. Esclerotomias foram tratadas com crioterapia em duplo ou triplo congelamento. Injeção intravítrea perioperatória de melfalano (32 ug/0,075 mL) foi administrada quando necessário. Foram relatados acuidade visual, pressão intraocular, resposta terapêutica sistêmica e ocular. Resultados: Cinco olhos de 5 pacientes com metástases vítreas de melanoma cutâneo unilateral foram identificados. Idade média de diagnóstico foi 84 anos (variando de 37-88). Seguimento médio após diagnóstico oftalmológico foi 28 (8,5-36) meses; 1 paciente não teve acompanhamento. Acuidade visual inicial variou de 20/30 a movimentos de mão. Achados clínicos iniciais incluíram infiltração de células pigmentadas e não-pigmentadas no vítreo (5/5), segmento anterior (4/5), e retina (3/5). Quatro pacientes tiveram glaucoma secundário. Tratamento sistêmico incluiu imunoterapia com inibidores da via de sinalização (3 - todos com resposta parcial/completa), quimioterapia sistêmica (2), ressecção cirúrgica (3), e irradiação (2). Intervalo médio entre diagnóstico primário e metástases vítreas foi 2 (2-15) anos. Um paciente teve doença sistêmica ativa simultânea as metástases vítreas. Acuidade visual final variou entre 20/40 e SPL. Tratamento oftalmológico incluiu vitrectomia nos 5 pacientes, melfalano intravítreo em 3 e metotrexato intravítreo em 1. Um paciente precisou de enucleação. A histopatologia revelou invasão celular extensa de melanoma. Conclusões: Metástases vítreas de melanoma cutâneo pode se manifestar como uma infiltração difusa de células pigmentadas e não-pigmentadas no vítreo e erroneamente diagnosticada como uveites. Vitrectomia diagnóstica e quimioterapia intravítrea periódica podem estar indicadas.

Activation of NFAT by HGF and IGF-1 via ARF6 and its effector ASAP1 promotes uveal melanoma metastasis.

Preventing or effectively treating metastatic uveal melanoma (UM) is critical because it occurs in about half of patients and confers a very poor prognosis. There is emerging evidence that hepatocyte growth factor (HGF) and insulin-like growth factor 1 (IGF-1) promote metastasis and contribute to the striking metastatic hepatotropism observed in UM metastasis. However, the molecular mechanisms by which HGF and IGF-1 promote UM liver metastasis have not been elucidated. ASAP1, which acts as an effector for the small GTPase ARF6, is highly expressed in the subset of uveal melanomas most likely to metastasize. Here, we found that HGF and IGF-1 hyperactivate ARF6, leading to its interaction with ASAP1, which then acts as an effector to induce nuclear localization and transcriptional activity of NFAT1. Inhibition of any component of this pathway impairs cellular invasiveness. Additionally, knocking down ASAP1 or inhibiting NFAT signaling reduces metastasis in a xenograft mouse model of UM. The discovery of this signaling pathway represents not only an advancement in our understanding of the biology of uveal melanoma metastasis but also identifies a novel pathway that could be targeted to treat or prevent metastatic uveal melanoma.

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.

PRAME induces genomic instability in uveal melanoma.

PRAME is a CUL2 ubiquitin ligase subunit that is normally expressed in the testis but becomes aberrantly overexpressed in many cancer types in association with aneuploidy and metastasis. Here, we show that PRAME is expressed predominantly in spermatogonia around the time of meiotic crossing-over in coordination with genes mediating DNA double strand break repair. Expression of PRAME in somatic cells upregulates pathways involved in meiosis, chromosome segregation and DNA repair, and it leads to increased DNA double strand breaks, telomere dysfunction and aneuploidy in neoplastic and non-neoplastic cells. This effect is mediated at least in part by ubiquitination of SMC1A and altered cohesin function. PRAME expression renders cells susceptible to inhibition of PARP1/2, suggesting increased dependence on alternative base excision repair pathways. These findings reveal a distinct oncogenic function of PRAME than can be targeted therapeutically in cancer.

Advances in the clinical management of uveal melanoma.

Melanomas arising in the uveal tract of the eye are a rare form of the disease with a biology and clinical phenotype distinct from their more common cutaneous counterparts. Treatment of primary uveal melanoma with radiotherapy, enucleation or other modalities achieves local control in more than 90% of patients, although 40% or more ultimately develop distant metastases, most commonly in the liver. Until January 2022, no systemic therapy had received regulatory approval for patients with metastatic uveal melanoma, and these patients have historically had a dismal prognosis owing to the limited efficacy of the available treatments. A series of seminal studies over the past two decades have identified highly prevalent early, tumour-initiating oncogenic genomic aberrations, later recurring prognostic alterations and immunological features that characterize uveal melanoma. These advances have driven the development of a number of novel emerging treatments, including tebentafusp, the first systemic therapy to achieve regulatory approval for this disease. In this Review, our multidisciplinary and international group of authors summarize the biology of uveal melanoma, management of primary disease and surveillance strategies to detect recurrent disease, and then focus on the current standard and emerging regional and systemic treatment approaches for metastatic uveal melanoma.

IRIS METASTASIS FROM BREAST CANCER SUCCESSFULLY TREATED WITH ABEMACICLIB AND LETROZOLE.

PURPOSE: To describe a patient with an unusual presentation of iris metastasis from breast cancer and her response to systemic therapy. METHODS: Retrospective chart review of one patient. RESULTS: A 57-year-old woman presented with a superonasal translucent vascularized iris stromal mass with fish egg-like structures budding from the surface. High-frequency anterior segment ultrasonography demonstrated a solid iris stromal mass measuring 6.0 mm × 3.3 mm × 1.9 mm. On optical coherence tomography, the egglike structures appeared as hyperreflective spheres, some of which were detached from the main iris stromal tumor. Oncologic evaluation revealed metastatic breast cancer involving the brain and lung. She was treated with oral abemaciclib and letrozole, as well as external beam radiotherapy to the brain. The iris mass had completely regressed within 4 months and remained undetectable through the 8-month follow-up. The other metastatic lesions responded well to therapy. CONCLUSION: A case of iris metastasis was reported as the presenting sign of cancer dissemination that was successfully treated with targeted systemic therapy without ocular radiotherapy.

Early Mechanisms of Chemoresistance in Retinoblastoma.

Retinoblastoma is the most common eye cancer in children and is fatal if left untreated. Over the past three decades, chemotherapy has become the mainstay of eye-sparing treatment. Nevertheless, chemoresistance continues to represent a major challenge leading to ocular and systemic toxicity, vision loss, and treatment failure. Unfortunately, the mechanisms leading to chemoresistance remain incompletely understood. Here, we engineered low-passage human retinoblastoma cells to study the early molecular mechanisms leading to resistance to carboplatin, one of the most widely used agents for treating retinoblastoma. Using single-cell next-generation RNA sequencing (scRNA-seq) and single-cell barcoding technologies, we found that carboplatin induced rapid transcriptomic reprogramming associated with the upregulation of PI3K-AKT pathway targets, including ABC transporters and metabolic regulators. Several of these targets are amenable to pharmacologic inhibition, which may reduce the emergence of chemoresistance. We provide evidence to support this hypothesis using a third-generation inhibitor of the ABCB1 transporter.

Clinical characteristics and postoperative complications as predictors of radiation toxicity after treatment with I125 Eye Plaque Brachytherapy for Uveal Melanomas.

PURPOSE: I125 Eye Plaque brachytherapy is the standard treatment for medium-sized uveal melanomas (UM). Patients develop radiation toxicities (RTT), including radiation maculopathy (RM), radiation neovascular glaucoma/iris neovascularization (RNGI) and radiation optic neuropathy (RON). We aim to investigate demographics, pretreatment tumor characteristics and posttreatment complications as predictors of RTT. METHODS AND MATERIALS: An IRB-approved single-institution retrospective chart review was performed from 2011 to 2019 for patients with posterior UM treated with brachytherapy. We collected demographics, pretreatment tumor characteristics and posttreatment complications. Univariate analysis (UVA) and multivariate analysis (MVA) were performed using logistic regression model. Hazard ratios (HR) and corresponding p-values were reported. All tests were two-sided; statistical significance was considered when p<0.05. RESULTS: Two hundred and fifty eight patients were evaluated. Median follow-up was 33.50 months (range 3.02-97.31). 178 patients (69.0%) had RTT. 131 patients (50.8%) developed RM. Fifty-six patients (21.7%) developed RON. Nineteen patients (7.4%) developed RNGI. UVA found shorter distance to fovea (DF) (p = 0.04), posttreatment exudative retinal detachment (PERD) (p = 0.001) and posttreatment vitreous hemorrhage (PVH) (p = 0.001) are associated with RTT. MVA found shorter DF (HR=1.03, p = 0.04), PERD (HR=2.52, p = 0.01) and PVH (HR=3.34, p = 0.006) are associated with RTT. MVA found female sex (HR=1.731, p = 0.031) and tumor height (HR=1.13, p = 0.013) are associated with RM and pretreatment retinal detachment (HR=3.41, p<0.001) is associated with RON. CONCLUSIONS: Shorter DF, PERD and PVH are associated with RTT; female sex and tumor height are associated with RM and tumor height is associated with RON. These findings serve as prognostic tools to counsel patients and promote early intervention in management of RTT.

Adjuvant crizotinib in high-risk uveal melanoma following definitive therapy.

Introduction: Approximately 40% of patients with uveal melanoma (UM) will develop metastatic disease. Tumors measuring at least 12mm in basal diameter with a class 2 signature, as defined by a widely used gene expression-profiling test, are associated with significantly higher risk of metastasis, with a median time to recurrence of 32 months. No therapy has been shown to reduce this risk. Materials and Methods: This was a single-arm, multicenter study in patients with high-risk UM who received definitive treatment of primary disease and had no evidence of metastasis. Patients were consecutively enrolled to receive 12 four-week cycles of adjuvant crizotinib at a starting dose of 250mg twice daily and were subsequently monitored for 36 months. The primary outcome of this study was to assess recurrence-free survival (RFS) of patients with high-risk UM who received adjuvant crizotinib. Results: 34 patients enrolled and received at least one dose of crizotinib. Two patients were unevaluable due to early withdrawal and loss to follow-up, leaving 32 patients evaluable for efficacy. Eight patients (25%) did not complete the planned 48-week course of treatment due to disease recurrence (n=5) or toxicity (n=3). All patients experienced at least one adverse event (AE), with 11/34 (32%) experiencing a Common Terminology Criteria for Adverse Events (CTCAE) grade 3 or 4 AE. After a median duration of follow up of 47.1 months, 21 patients developed distant recurrent disease. The median RFS was 34.9 months (95% CI (Confidence Interval), 23-55 months), with a 32-month recurrence rate of 50% (95% CI, 33-67%). Analysis of protein contents from peripheral blood extracellular vesicles in a subset of patient samples from baseline, on-treatment, and off-treatment, revealed a change in protein content associated with crizotinib exposure, however without a clear association with disease outcome. Conclusions: The use of adjuvant crizotinib in patients with high-risk UM did not result in improved RFS when compared to historical controls. Analysis of blood extracellular vesicles revealed changes in protein content associated with treatment, raising the possibility of future use as a biomarker. Further investigation of adjuvant treatment options are necessary for this challenging disease.

BAP1 Loss Promotes Suppressive Tumor Immune Microenvironment via Upregulation of PROS1 in Class 2 Uveal Melanomas.

Uveal melanoma (UM) is the most common primary cancer of the eye and is associated with a high rate of metastatic death. UM can be stratified into two main classes based on metastatic risk, with class 1 UM having a low metastatic risk and class 2 UM having a high metastatic risk. Class 2 UM have a distinctive genomic, transcriptomic, histopathologic, and clinical phenotype characterized by biallelic inactivation of the BAP1 tumor-suppressor gene, an immune-suppressive microenvironment enriched for M2-polarized macrophages, and poor response to checkpoint-inhibitor immunotherapy. To identify potential mechanistic links between BAP1 loss and immune suppression in class 2 UM, we performed an integrated analysis of UM samples, as well as genetically engineered UM cell lines and uveal melanocytes (UMC). Using RNA sequencing (RNA-seq), we found that the most highly upregulated gene associated with BAP1 loss across these datasets was PROS1, which encodes a ligand that triggers phosphorylation and activation of the immunosuppressive macrophage receptor MERTK. The inverse association between BAP1 and PROS1 in class 2 UM was confirmed by single-cell RNA-seq, which also revealed that MERTK was upregulated in CD163+ macrophages in class 2 UM. Using ChIP-seq, BAP1 knockdown in UM cells resulted in an accumulation of H3K27ac at the PROS1 locus, suggesting epigenetic regulation of PROS1 by BAP1. Phosphorylation of MERTK in RAW 264.7 monocyte-macrophage cells was increased upon coculture with BAP1-/- UMCs, and this phosphorylation was blocked by depletion of PROS1 in the UMCs. These findings were corroborated by multicolor immunohistochemistry, where class 2/BAP1-mutant UMs demonstrated increased PROS1 expression in tumor cells and increased MERTK phosphorylation in CD163+ macrophages compared with class 1/BAP1-wildtype UMs. Taken together, these findings provide a mechanistic link between BAP1 loss and the suppression of the tumor immune microenvironment in class 2 UMs, and they implicate the PROS1-MERTK pathway as a potential target for immunotherapy in UM.

Cyst-like recurrence of retinoblastoma diagnosed by multimodal imaging.

Purpose: To illustrate the diagnosis and management of an atypical, cyst-like recurrence of retinoblastoma. Observations: A 4-month-old boy was diagnosed with Group B retinoblastoma in the right eye, consisting of a 10 x 9 × 3.6 mm retinal tumor temporal to the macula. He was treated with one session of intra-arterial ophthalmic artery chemotherapy using melphalan, followed by three sessions of diode laser transpupillary thermotherapy (TTT), after which complete tumor regression was achieved. 45 weeks after initial treatment, a cystic lesion was detected adjacent to superior margin of the regressed tumor scar. The differential diagnosis included pigment epithelial detachment, retinal gliosis, secondary retinoschisis, and local tumor recurrence. Multimodal imaging including OCT angiography confirmed the diagnosis of local recurrence manifesting as a vascularized cyst-like lesion. Two additional sessions of TTT achieved sustained tumor regression through 16 months of additional follow-up. Conclusions and Importance: Recurrence of retinoblastoma following chemotherapy typically manifests as enlargement of a previously regressed tumor, or seeding into the vitreous or subretinal space. An unusual cyst-like recurrence of retinoblastoma at the margin of a previously regressed tumor was diagnosed by multimodal imaging. Focal diode laser transpupillary thermotherapy was curative.

RB1 loss triggers dependence on ESRRG in retinoblastoma.

Retinoblastoma (Rb) is a deadly childhood eye cancer that is classically initiated by inactivation of the RB1 tumor suppressor. Clinical management continues to rely on nonspecific chemotherapeutic agents that are associated with treatment resistance and toxicity. Here, we analyzed 103 whole exomes, 20 whole transcriptomes, 5 single-cell transcriptomes, and 4 whole genomes from primary Rb tumors to identify previously unknown Rb dependencies. Several recurrent genomic aberrations implicate estrogen-related receptor gamma (ESRRG) in Rb pathogenesis. RB1 directly interacts with and inhibits ESRRG, and RB1 loss uncouples ESRRG from negative regulation. ESRRG regulates genes involved in retinogenesis and oxygen metabolism in Rb cells. ESRRG is preferentially expressed in hypoxic Rb cells in vivo. Depletion or inhibition of ESRRG causes marked Rb cell death, which is exacerbated in hypoxia. These findings reveal a previously unidentified dependency of Rb cells on ESRRG, and they implicate ESRRG as a potential therapeutic vulnerability in Rb.

Pseudoangiomatous retinal gliosis (PARG) treated with iodine plaque in patient with chronic retinal detachment.

Purpose: To describe a case of a chronic retinal detachment complicated by the development of pre and subretinal hemorrhage secondary to a large pseudoangiomatous retinal gliosis (PARG) that interfered with retinal reattachment. After the lesion was regressed following plaque radiotherapy retinal reattachment was successfully completed. Observations: A 56y.o healthy man with known history of a chronic inferior rhegmatogenous retinal detachment (RD) of the left eye (OS) presented to the Bascom Palmer Eye Institute (BPEI) emergency department (ED) complaining of new floaters OS. On examination, the patient had a visual acuity of 20/30 right eye (OD) and 20/200 OS. Fundoscopic examination showed a treated tear in OD and dense vitreous hemorrhage OS. Initial B-scan ultrasonography OS showed an inferior RD with diffuse hyperechoic material in the vitreous cavity, preretinal and subretinal space most consistent with hemorrhage. Three days later the patient presented with further vision loss and a repeat B scan showed total RD and increasing subretinal hemorrhage with a solid mass like lesion. At this point, decision was made to proceed with retinal detachment repair, removal of the vitreous hemorrhage, and retina evaluation. During surgery, a total retinal detachment was encountered with poor view of the inferior retina due to a large round vascular lesion in the subretinal space with surrounding hemorrhage and clots. The retina was reattached during surgery, however, the postop was complicated by recurrence of VH, dense hyphema, increased IOP, recurrence of retinal detachment, and growth of the mass like lesion noted during surgery. Consultation with ocular oncology diagnosed the patient with secondary PARG lesion and plaque radiotherapy was given achieving remarkable regression of the lesion. After the lesion had regressed, successful retinal reattachment was achieved, and the patient had excellent visual recovery. Conclusion and importance: PARG lesions are uncommon in particular when associated to chronic retinal detachments. This case highlights the importance of having a high clinical suspicion for the development of these lesions to diagnose them correctly and treat them aggressively with plaque radiotherapy in order to be able to manage the underlying complex retinal detachment.

Hematologic Complications Associated With Intra-arterial Chemotherapy for Retinoblastoma Treatment: A Single Institution Experience.

Retinoblastoma (RB) is the most common intraocular pediatric malignancy. Advancements in intra-arterial chemotherapy (IAC) for treatment of RB have resulted in dramatic improvement in eye salvage rates. Data regarding IAC outcomes and associated hematologic toxicities are limited. The objective of this retrospective study was to analyze baseline characteristics, efficacy, and hematologic complications associated with IAC treatment in children with RB at a single international referral institution. Ninety-five sessions of IAC were performed in 28 patients. Mean age at RB diagnosis was 12.5 months (SD, 9.2 mo). Fourteen patients had bilateral RB. IAC agents included melphalan, carboplatin, and topotecan. The most common regimens were triple-agent IAC and single-agent melphalan (66.3% and 15.8%, respectively). Median number of IAC sessions was 3 (mean: 3.39, range: 1 to 9). Eye salvage rate was 83.7% with an overall survival rate of 100% at a median follow-up of 29 months (mean: 29.8 mo, range: 1 to 63 mo). A total of 26 patients (92.9%) experienced at least 1 hematologic toxicity during their treatment course Prevalence of neutropenia, anemia, and thrombocytopenia were 89.3%, 85.7%, and 25%, respectively. While IAC is effective in salvaging most eyes with advanced intraocular RB, over half of patients experienced clinically significant neutropenia and anemia. Clinicians should be vigilant in monitoring patients for IAC-related complications.

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.

HDAC11 activity contributes to MEK inhibitor escape in uveal melanoma.

We previously demonstrated that pan-HDAC inhibitors could limit escape from MEK inhibitor (MEKi) therapy in uveal melanoma (UM) through suppression of AKT and YAP/TAZ signaling. Here, we focused on the role of specific HDACs in therapy adaptation. Class 2 UM displayed higher expression of HDACs 1, 2, and 3 than Class 1, whereas HDACs 6, 8, and 11 were uniformly expressed. Treatment of UM cells with MEKi led to modulation of multiple HDACs, with the strongest increases observed in HDAC11. RNA-seq analysis showed MEKi to decrease the expression of multiple HDAC11 target genes. Silencing of HDAC11 significantly reduced protein deacetylation, enhanced the apoptotic response to MEKi and reduced growth in long-term colony formation assays across multiple UM cell lines. Knockdown of HDAC11 led to decreased expression of TAZ in some UM cell lines, accompanied by decreased YAP/TAZ transcriptional activity and reduced expression of multiple YAP/TAZ target genes. Further studies showed this decrease in TAZ expression to be associated with increased LKB1 activation and modulation of glycolysis. In an in vivo model of uveal melanoma, silencing of HDAC11 limited the escape to MEKi therapy, an effect associated with reduced levels of Ki67 staining and increased cleaved caspase-3. We have demonstrated a novel role for adaptive HDAC11 activity in UM cells, that in some cases modulates YAP/TAZ signaling leading to MEKi escape.

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.