Clinical analysis and treatment progress of NUT carcinoma in the nasal cavity and sinuses: a retrospective study from a single institution.

OBJECTIVE: Nuclear protein in testis (NUT) carcinoma is characterized by NUT gene rearrangement on chromosome 15. The objective of this study was to investigate the clinical features, immunohistochemistry, treatment, diagnosis and prognosis of sinonasal NUT carcinoma specifically. METHODS: Clinical data for 10 cases of NUT cancer confirmed by pathology were retrospectively analyzed, and the relevant literature was reviewed. RESULTS: Among the 10 patients, 6 were males, and 4 were females. The median age was 34 years (15-69 years). Nine patients presented with locally advanced cT4a stage. The most common treatment was complete resection combined with radiotherapy, chemotherapy, and targeted therapy. All 10 patients had pathologically poorly differentiated or undifferentiated carcinoma. Furthermore, immunohistochemical staining showed that NUT protein was positive in all 10 patients, and most cases expressed p63, p40 and CK. The Ki-67 positive index of 8 patients ranged from 40 to 80%, with a median of 50%, and NUTM1 gene disruption was detected in both of the remaining cases by FISH. As of April, 2023, all patients were followed up with for 1-51 months, with a median follow-up time of 14 months. Three patients died due to widespread systemic metastasis, 3 relapsed, and 4 had no recurrence or metastasis. CONCLUSION: Sinonasal NCs (NUT carcinomas) is a rare and highly aggressive malignant tumor with rapid progression and a poor prognosis. Correct histopathological diagnosis is the primary prerequisite for determining appropriate treatment. There are currently no effective treatment options for NCs. Targeted therapy may become an effective method to treat NCs.

The BET degrader ZBC260 suppresses stemness and tumorigenesis and promotes differentiation in triple-negative breast cancer by disrupting inflammatory signaling.

BACKGROUND: Breast cancer stem cells (BCSCs) are resistant to standard therapies, facilitate tumor dissemination, and contribute to relapse and progression. Super-enhancers are regulators of stemness, and BET proteins, which are critical for super-enhancer function, are a potential therapeutic target. Here, we investigated the effects of BET proteins on the regulation of breast cancer stemness using the pan-BET degrader ZBC260. METHODS: We evaluated the effect of ZBC260 on CSCs in TNBC cell lines. We assessed the effect of ZBC260 on cellular viability and tumor growth and measured its effects on cancer stemness. We used RNA sequencing and stemness index to determine the global transcriptomic changes in CSCs and bulk cells and further validated our findings by qPCR, western blot, and ELISA. RESULTS: ZBC260 potently inhibited TNBC growth both in vitro and in vivo. ZBC260 reduced stemness as measured by cell surface marker expression, ALDH activity, tumorsphere number, and stemness index while increasing differentiated cells. GSEA analysis indicated preferential downregulation of stemness-associated and inflammatory genes by ZBC260 in ALDH+ CSCs. CONCLUSIONS: The BET degrader ZBC260 is an efficient degrader of BET proteins that suppresses tumor progression and decreases CSCs through the downregulation of inflammatory genes and pathways. Our findings support the further development of BET degraders alone and in combination with other therapeutics as CSC targeting agents.

Comprehensive analysis of the prognosis and immune infiltrates for the BET protein family reveals the significance of BRD4 in glioblastoma multiforme.

Background: Glioblastoma multiforme (GBM) is the most common and invasive primary central nervous system tumor. The prognosis after surgery, radiation and chemotherapy is very poor. Bromodomain (BRD) proteins have been identified in oncogenic rearrangements, and play a key role in the development of multiple cancers. However, the relationship between BET proteins and prognosis of GBM are still worth exploring, and the distinct functions of BET proteins and tumor immunology in GBM have not been fully elucidated. Therefore, it is particularly important to develop effective biomarkers to predict the prognosis of GBM patients. Methods: Metascape, David, Kaplan-Meier Plotter, Oncomine, GEPIA, TCGA, TIMER, and LinkedOmics databases were used to assess the expression and prognosis for BET proteins in GBM. ROC analysis of risk model was established to identify the correlation between BET genes and overall survival in GBM patients. TIMER and GEPIA databases were used to comprehensively investigate the correlation between BET genes and tumor immune infiltration cells. Moreover, the image of immunohistochemistry staining of BET proteins in normal tissue and tumor tissue were retrived from the HPA database. In addition, differential analysis and pathway enrichment analysis of BRD4 gene expression profile were also carried out. Finally, immune-fluorescence and Western blot were used to clarify the expression of BRD4 in GBM cells. Results: Bioinformatics analysis showed that the expression levels of BET genes in GBM may play an important role in oncogenesis. Specifically, bioinformatic and immunohistochemistry analysis showed that BRD4 protein was more highly expressed in tumor tissues than that in normal tissues. The high expression of BRD4 was associated with poor prognosis in GBM. The expression of BET genes were closely related to the immune checkpoint in GBM. The correlation effect of BRD4 was significantly higher than other BET genes, which represented negative correlation with immune checkpoint. The expression of BRD4 was positively associated with tumor purity, and negatively associated with immune infiltration abundance of macrophage, neutrophil and CD8+ T-cell, respectively. Cox analysis showed that the model had good survival prediction and prognosis discrimination ability. In addition, the expression levels of BRD4 protein was significantly higher in U-251 MG cells than that in normal cells, which was consistent with the results of bioinformatics data. Conclusion: This study implied that BRD4 could be hopeful prognostic biomarker in GBM. The increased expression of BRD4 may act as a molecular marker to identify GBM patients with high-risk subgroups. BRD4 may be a valuable prognostic biomarker, and a potential target of precision therapy against GBM.

An efficient combination immunotherapy for antitumor immunity without accelerating cardiac allograft rejection.

Immunotherapy with immune checkpoint inhibitors (ICIs), including antibodies against programmed cell death protein-1 (PD-1) and its receptor programmed cell death ligand-1 (PD-L1), represents a promising systematic treatment for advanced human malignancies. Transplantation remains the ultimate therapy for end-stage organ diseases. However, the efficacy of ICI treatment in solid organ transplant (SOT) recipients remains controversial. We established a transgenic primary liver cancer mouse model and performed allogeneic heterotopic heart transplantation. Different treatments were performed and survival curves were calculated. Graft samples were collected, and immune cells and the cell surface expression of PD-L1 were analysed by flow cytometry. Inflammatory cytokine levels in the serum were measured by an inflammatory array. The specificity of the histochemical techniques was tested by staining sections. A combination immunotherapy comprising a BET protein inhibitor (JQ1) and an immune checkpoint inhibitor (anti-PD-L1 antibody) was administered to primary liver cancer model mice bearing cardiac allografts. Interestingly, the combination immunotherapy effectively suppressed the progression of primary liver cancer but did not accelerate allograft rejection. In accordance with our previous findings, BET protein inhibition enhances the expression of a putative membrane transporter (Rab8A), which upregulates the expression of PD-L1 on the plasma membrane in a transgenic primary liver cancer mouse model. This may be a crucial mechanism of tumour progression arrest. Our data showed that heart transplantation upregulated the expression of the proinflammatory factor IFN-γ and suggested that BET protein inhibition (with JQ1) decreased PD-L1 expression in heart tissues after cardiac transplantation. This phenomenon was accompanied by enhanced infiltration of inflammatory IFN-γ. Our study provides a novel and efficient therapeutic strategy for SOT recipients.

JQ1 attenuates neuroinflammation by inhibiting the inflammasome-dependent canonical pyroptosis pathway in SAE.

Sepsis-associated encephalopathy (SAE) manifests clinically in hyperneuroinflammation. Pyroptosis, which can induce an inflammatory cascade response, has been considered to be a causative factor of SAE. Evidence has shown that the bromo- and extraterminal (BET) proteins (including BRD2, BRD3, BRD4 and BRDT) inhibitor JQ1 can inhibit inflammation and suppress pyroptosis in various diseases. Therefore, we examined the effect of JQ1 on inflammasome-induced pyroptosis in the hippocampus in a mouse model of sepsis induced by lipopolysaccharide (LPS) injection. The results showed that JQ1 treatment alleviated sepsis-related symptoms, protected the blood-brain barrier (BBB), as indicated by upregulated expression of the tight junction proteins occludin and ZO-1, and remarkably rescued neuronal damage in SAE mice. Mechanistically, we demonstrated that JQ1 intervention inhibited the expression of BRD proteins and decreased the expression of inflammasomes by blocking phospho-nuclear factor kappa B (p-NF-κB) signalling, attenuating the canonical pyroptosis (mediated by cleaved-Caspase1/11) pathway and the release of proinflammatory factors in the hippocampus of septic mice. Interestingly, we also found that JQ1 selectively suppressed the activation of hippocampal microglia in SAE mice. Thus, JQ1 protected the hippocampal BBB and neuronal damage through the attenuation of neuroinflammation by inhibiting the inflammasome-dependent canonical pyroptosis pathway induced by LPS injection in mice, and JQ1 may be a promising target for the prevention of SAE.

Inhibition of BET Protein Function Suppressed the Overactivation of the Canonical NF-κB Signaling Pathway in 6-OHDA-Lesioned Rat Model of Levodopa-Induced Dyskinesia.

Background: Neuroinflammation is involved in the mechanisms of levodopa-induced dyskinesia (LID). The canonical NF-κB activation signaling pathway plays a critical role in the neuroinflammation development and BET protein-induced NF-κB-mediated neuroinflammation. The inhibition of the BET protein function has been reported to alleviate LID; however, its association with the canonical NF-κB signaling pathway in the 6-OHDA-lesioned striatum of the LID rat model remains unknown. Accordingly, we identified the status of the canonical NF-κB signaling pathway in the 6-OHDA-lesioned striatum of the LID rat model and whether the anti-dyskinetic effect of the BET inhibitor JQ1 was associated with its suppression on NF-κB-mediated neuroinflammation. Methods: 6-OHDA PD rat models were treated with either L-dopa plus JQ1 or L-dopa alone. L-dopa treatment was given for 2 weeks, and the JQ1 treatment was given for 3 weeks and was initiated a week prior to L-dopa treatment. As a control, the sham rats were treated with JQ1 or Veh for 3 weeks. The ALO AIM assessment and cylinder test were performed during the treatment. Glial activation markers, pro-inflammatory substances, and critical proteins in the canonical NF-κB signaling pathway were tested in the lesioned striatum after the final treatment. Results: JQ1 effectively alleviated LID without influencing motor improvement. In the lesioned striatum, L-dopa triggered an overactivation of the canonical NF-κB signaling pathway, with an increase in the phospho-IKKα/ß, phospho-IκBα, and NF-κB nuclear translocation and its phosphorylation at Ser 536 and Ser 276 sites (p < 0.01 vs. sham group). L-dopa induced an overexpression of the pro-inflammatory substances of tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, IL-6, and inducible nitric oxide synthase (iNOS), and the glial activation markers CD68 and GFAP. All the molecular changes were greatly inhibited by JQ1. Conclusion: L-dopa triggered an overactivation of the canonical NF-κB signaling pathway, leading to an enhanced neuroinflammation response in the 6-OHDA-lesioned striatum of LID rat models. The inhibition of the BET protein function significantly suppressed the activation of the canonical NF-κB signaling pathway in the striatum, alleviating the neuroinflammation response and the severity of LID.

BET and CDK Inhibition Reveal Differences in the Proliferation Control of Sympathetic Ganglion Neuroblasts and Adrenal Chromaffin Cells.

Neuroblastoma arising from the adrenal differ from ganglionic neuroblastoma both genetically and clinically, with adrenal tumors being associated with a more severe prognosis. The different tumor properties may be linked to specific tumor founder cells in adrenal and sympathetic ganglia. To address this question, we first set up cultures of mouse sympathetic neuroblasts and adrenal chromaffin cells. These cultures were then treated with various proliferation inhibitors to identify lineage-specific responses. We show that neuroblast and chromaffin cell proliferation was affected by WNT, ALK, IGF1, and PRC2/EZH2 signaling inhibitors to a similar extent. However, differential effects were observed in response to bromodomain and extraterminal (BET) protein inhibitors (JQ1, GSK1324726A) and to the CDK-7 inhibitor THZ1, with BET inhibitors preferentially affecting chromaffin cells, and THZ1 preferentially affecting neuroblasts. The differential dependence of chromaffin cells and neuroblasts on BET and CDK signaling may indicate different mechanisms during tumor initiation in sympathetic ganglia and adrenal.

NUT carcinoma in children, adolescents and young adults.

BACKGROUND: NUT carcinoma (NC), defined by the presence of the NUTM1 rearrangement, is an aggressive tumour associated with poor prognosis. This rare cancer is underdiagnosed and difficult to treat. OBJECTIVE AND METHODS: The primary objective of this review is to describe the clinical, radiological and laboratory features of NC in young patients. The secondary objective is to propose a consensual strategy for the French very Rare Tumour group (FRACTURE group). RESULTS: NUT-specific antibody immunostaining in cases of undifferentiated or poorly differentiated carcinoma may demonstrate the specific NUT gene rearrangement. NCs are frequently advanced stage at diagnosis and the outcome remains poor despite a global strategy that generally includes conventional combination chemotherapy with wide local therapy (surgery, radiotherapy). Chemosensitivity is frequently only transient. CONCLUSION: Recent data have shown that new targeted drugs (histone deacetylase and bromodomain and extra-terminal protein inhibitors) are promising, but their role has yet to be evaluated in NC. Centralized data review is necessary to improve our knowledge of paediatric NC. We propose a multimodal strategy based on published data and their personal experience.

Inhibition of the bromodomain and extra-terminal family of epigenetic regulators as a promising therapeutic approach for gastric cancer.

PURPOSE: Epigenetic dysregulation is a common characteristic of cancers, including gastric cancer (GC), and contributes to cancer development and progression. Although the efficacy of BET (an epigenetic regulator) inhibition has been demonstrated in various cancer types, predictive genetic markers of its efficacy in GC are currently lacking. Therefore, we aimed to identify markers that predict the response of BET inhibition in GC and, suggest an effective treatment regimen through combined therapy. METHODS: The effect of BET inhibition was evaluated using iBET-151, a small-molecule inhibitor of BET proteins, in a large panel (n = 49) of GC cell lines and xenograft mouse models. Comprehensive genetic information was used to identify cell lines sensitive to iBET-151. Flow cytometry, Western blotting, and colony-formation and migration assays were used to evaluate the effects of iBET-151 and/or paclitaxel. The synergistic effect of iBET-151 and paclitaxel was evaluated using an organoid model. RESULTS: We found that iBET-151 showed a modest growth-inhibitory effect in GC cells (73%, 36/49). iBET-151 inhibited tumorigenicity in vitro and significantly promoted cell cycle arrest and apoptosis. Based on comprehensive genetic information analysis in relation to BET family expression, we found that BRD4 was highly expressed in the iBET-151-sensitive cell lines. We also identified WNT5B and IRS2 as potential biomarkers that are predictive for sensitivity to iBET-151. In GC xenograft model mice, iBET-151 significantly decreased tumor volumes and Ki-67 and BRD4 expression. Combination treatment showed that iBET-151 increased the sensitivity of GC cells to paclitaxel in approximately 70% of the cell lines (34/49) tested. iBET-151 plus paclitaxel significantly promoted cell cycle arrest and apoptosis and suppressed c-Myc, Bcl-2 and Bcl-xL expression. In GC organoids, iBET-151 and paclitaxel showed a synergistic effect. CONCLUSIONS: Collectively, our data suggest that iBET-151 is a potential therapeutic agent for GC, especially in combination with paclitaxel, and that WNT5B and IRS2 may predict iBET-151 sensitivity.

Are BET Inhibitors yet Promising Latency-Reversing Agents for HIV-1 Reactivation in AIDS Therapy?

AIDS first emerged decades ago; however, its cure, i.e., eliminating all virus sources, is still unachievable. A critical burden of AIDS therapy is the evasive nature of HIV-1 in face of host immune responses, the so-called "latency." Recently, a promising approach, the "Shock and Kill" strategy, was proposed to eliminate latently HIV-1-infected cell reservoirs. The "Shock and Kill" concept involves two crucial steps: HIV-1 reactivation from its latency stage using a latency-reversing agent (LRA) followed by host immune responses to destroy HIV-1-infected cells in combination with reinforced antiretroviral therapy to kill the progeny virus. Hence, a key challenge is to search for optimal LRAs. Looking at epigenetics of HIV-1 infection, researchers proved that some bromodomains and extra-terminal motif protein inhibitors (BETis) are able to reactivate HIV-1 from latency. However, to date, only a few BETis have shown HIV-1-reactivating functions, and none of them have yet been approved for clinical trial. In this review, we aim to demonstrate the epigenetic roles of BETis in HIV-1 infection and HIV-1-related immune responses. Possible future applications of BETis and their HIV-1-reactivating properties are summarized and discussed.

Selective inhibition mechanism of nitroxoline to the BET family: Insight from molecular simulations.

Although the proteins in bromodomain and extra-terminal domain (BET) family are promising therapy drug targets for numerous human diseases, the binding effectiveness is interfered by the competition from non-BET protein BRD9. In this study, molecular docking, molecular dynamics simulations, binding free energy calculations and per-residue energy decomposition methods were employed to clarify the selective inhibition mechanism of nitroxoline. The results showed that the different cavity volume of effective embedding inhibitor and the changes in conserved residues were associated with the significant higher selectivity of inhibitor nitroxoline for BET family than non-BET protein (BRD9). In addition, the non-polar interactions occurred in Phe83, Val87 at ZA loop, and the polar interaction appeared in Met132, Asn135 at BC loop. Therefore, when designing a new inhibitor, it could better improve the inhibitor activity by introducing the heteroatom conjugated pyridine-like moiety and the strong electron-withdrawing nitro-like moiety. Overall, this study not only clarified the molecular mechanism of the selective inhibition of nitroxoline, but also provided insight into designing more effective BET inhibitors in next step.

BRD4 regulates self-renewal ability and tumorigenicity of glioma-initiating cells by enrichment in the Notch1 promoter region.

Bromodomain and extraterminal domain (BET) family proteins are considered to be epigenetic readers that regulate gene expression by recognizing acetyl lysine residues on histones and nonhistone chromatin factors and have been classified as curative targets for a variety of cancers. Glioma-initiating cells (GICs), which commit self-renewal, perpetual proliferation, multidirectional differentiation, and vigorous tumorigenicity, sustain the peculiar genetic and epigenetic diversification in the GBM patients, thus, GICs result in tumor recurrence. Abundant evidence demonstrates that BET proteins regulate differentiation of stem cells. However, it endures ambiguous how individual BET proteins take part in GIC advancement, and how do small molecule inhibitors like I-BET151 target functional autonomous BET proteins. Here, we validated that BRD4, not BRD2 or BRD3, has value in targeted glioma therapy. We announce a signaling pathway concerning BRD4 and Notch1 that sustains the self-renewal of GICs. Moreover, in-depth mechanistic research showed that BRD4 was concentrated at the promoter region of Notch1 and may be involved in the process of tumor metabolism. Furthermore, in intracranial models, I-BET151 eliminated U87 GICs' tumorigenicity. The outcomes of this research could be conducive to design clinical trials for treatment of glioma based on BRD4.

Effect of TNIK upregulation on JQ1-resistant human colorectal cancer HCT116 cells.

JQ1 disrupts the binding of bromodomain and extra-terminal (BET) family of proteins to acetylated histones, modulates the expression of various genes, and inhibits the proliferation of cancer cells. We established two JQ1-resistant sublines from human colorectal cancer HCT116 cells. These resistant cells showed an 8- to 9-fold higher resistance to JQ1, and a 2- to 4-fold higher resistance to various anti-cancer agents, such as doxorubicin, etoposide, mitoxantrone, SN-38, cisplatin, and methotrexate than the parental HCT116 cells. The JQ1-resistant cells expressed higher levels of TRAF2 and NCK-interacting protein kinase (TNIK), cyclin D1 (CCND1), cyclin E1 (CCNE1), and their corresponding mRNAs than the parental cells. TNIK is a regulator of Wnt/ß-catenin signaling and is known to transactivate CCND1. Transient transfection of HCT116 cells with a TNIK expression plasmid resulted in the upregulation of cyclin D1, cyclin E1, and their corresponding mRNAs, as well as an increase in CCNE1 promoter activity. Furthermore, luciferase assay revealed that the JQ1-resistant cells showed high CCNE1 promoter activity. These results suggest that TNIK also transactivates CCNE1. Three stable TNIK transfectant clones of HEK293 cells expressed 1.5- to 2-fold higher levels of TNIK, cyclin D1, and cyclin E1 than the parental cells. The 293/TNIK-6 cells, which expressed the highest level of TNIK among the transfectants, showed a 2.3-fold higher resistance to JQ1 than the parental cells. These results suggest the possible involvement of TNIK in cellular resistance to JQ1.

Bromodomain and Extraterminal (BET) protein inhibition suppresses tumor progression and inhibits HGF-MET signaling through targeting cancer-associated fibroblasts in colorectal cancer.

OBJECTIVE: Colorectal cancer (CRC) is one of the leading causes of cancer-related mortality. The bromodomain and extra-terminal domain (BET) inhibitors suppresses the gene expressions of various oncogenes and shows a good efficacy in the preclinical CRC models. We investigate the mechanism of action of BET inhibitors in CRC. METHODS: The effect of BET inhibitor (JQ1) on the HGF-MET signaling was assessed by qPCR, western blot and immunohistochemical staining in CRC and cancer-associated fibroblasts (CAFs). The effect of JQ1 on the CAFs was investigated using the primary CAFs derived from CRC tissues and induced-CAFs derived from isolating foreskin fibroblasts. The effect of JQ1 on the gene expression profile of CAFs was explored by RNA-sequence, qPCR and bioinformatic analysis. RESULTS: JQ1 decreased the mRNA and protein levels of MET in CRC cells and downregulated the mRNA and protein levels of HGF in both CRC cells and CAFs. JQ1 attenuated the pro-migratory activity of CAFs through downregulation of HGF expression in CAFs. Meanwhile, JQ1 also reduced the ability of contracting collagen gels, decreased the cell proliferation, induced G1 arrest and repressed the pro-inflammatory gene expressions in CAFs. MYC expression was suppressed by JQ1 in CAFs. Knockdown of MYC induced G1 arrest in CAFs. CONCLUSION: Our results demonstrate the inhibitory effect of BET inhibition on the HGF-MET signaling and the pro-tumor activity of CAFs, revealing a new mechanism by which BET inhibition suppresses CRC progression.

Bromo- and extraterminal domain protein inhibition improves immunotherapy efficacy in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) represents the majority of liver cancer and is the fourth most common cause of cancer-related death. Although advances in molecular targeted therapy have shown promise, none of these agents has yet demonstrated significant clinical benefit. Bromo- and extraterminal domain (BET) protein inhibitors have been considered potential therapeutic drugs for HCC but the biological activity remains unclear. This study found that BET protein inhibition did not effectively suppress the progression of HCC, using a transgenic HCC mouse model. Mechanistically, the BET protein inhibitor JQ1 upregulated the expression of programmed cell death-ligand 1 (PD-L1) on the plasma membrane in vivo and in vitro. Moreover, JQ1 enhanced the expression of Rab8A, which upregulated the expression of PD-L1 on the plasma membrane. This study also showed that JQ1 combined with anti-PD-L1 Ab effectively suppressed HCC progression, and this benefit was obtained by enhancing the activation and cytotoxic capabilities of CD8 T cells. These results revealed the crucial role and regulation of BET protein inhibition on the expression of PD-L1 in HCC. Thus, combining BET protein inhibition with immune checkpoint blockade offers an efficient therapeutic approach for HCC.

Apabetalone - BET protein inhibition in cardiovascular disease and Type 2 diabetes.

Apabetalone is the first selective BET protein inhibitor in the field of cardiovascular diseases (CVD). BET proteins are epigenetic regulators that link upstream epigenetic modifications to downstream gene expression. Inhibition of BET proteins by apabetalone has been shown to modulate reverse cholesterol transport, coagulation, inflammation and vascular calcification. Furthermore, apabetalone reduces circulating markers of CVD risk and plaque vulnerability. Post-hoc pooled analyses suggest a potential reduction in risk of major adverse cardiac events (MACE) in patients with Type 2 diabetes (T2D) and stable CVD. However, the current cardiovascular outcomes trial BET-on-MACE failed to detect the assumed 30% reduction of MACE by apabetalone in patients with T2D after an acute coronary syndrome.

Long non-coding RNA MANCR is a target of BET bromodomain protein BRD4 and plays a critical role in cellular migration and invasion abilities of prostate cancer.

Androgen receptor (AR)-negative castration-resistant prostate cancer (CRPC) is highly aggressive and is resistant to most of the current therapies. Bromodomain and extra terminal domain (BET) protein BRD4 binds to super-enhancers (SEs) that drive high expression of oncogenes in many cancers. A BET inhibitor, JQ1, has been found to suppress the malignant phenotypes of prostate cancer cells, however, the target genes of JQ1 remain largely unknown. Here we show that SE-associated genes specific for AR-negative CRPC PC3 cells include genes involved in migration and invasion, and that JQ1 impairs migration and invasion of PC3 cells. We identified a long non-coding RNA, MANCR, which was markedly down-regulated by JQ1, and found that BRD4 binds to the MANCR locus. MANCR knockdown led to a significant decrease in migration and invasion of PC3 cells. Furthermore, RNA sequencing analysis revealed that expression of the genes involved in migration and invasion was altered by MANCR knockdown. In summary, our data demonstrate that MANCR plays a critical role in migration and invasion of PC3 cells.

De Novo Variants in SPOP Cause Two Clinically Distinct Neurodevelopmental Disorders.

Recurrent somatic variants in SPOP are cancer specific; endometrial and prostate cancers result from gain-of-function and dominant-negative effects toward BET proteins, respectively. By using clinical exome sequencing, we identified six de novo pathogenic missense variants in SPOP in seven individuals with developmental delay and/or intellectual disability, facial dysmorphisms, and congenital anomalies. Two individuals shared craniofacial dysmorphisms, including congenital microcephaly, that were strikingly different from those of the other five individuals, who had (relative) macrocephaly and hypertelorism. We measured the effect of SPOP variants on BET protein amounts in human Ishikawa endometrial cancer cells and patient-derived cell lines because we hypothesized that variants would lead to functional divergent effects on BET proteins. The de novo variants c.362G>A (p.Arg121Gln) and c. 430G>A (p.Asp144Asn), identified in the first two individuals, resulted in a gain of function, and conversely, the c.73A>G (p.Thr25Ala), c.248A>G (p.Tyr83Cys), c.395G>T (p.Gly132Val), and c.412C>T (p.Arg138Cys) variants resulted in a dominant-negative effect. Our findings suggest that these opposite functional effects caused by the variants in SPOP result in two distinct and clinically recognizable syndromic forms of intellectual disability with contrasting craniofacial dysmorphisms.