Circulating tumor cells (CTC) with a very-small-nuclear phenotype (vsnCTC) in prostate cancer are characterized by nuclei smaller than 8.5 μm. Our previous studies established an association between vsnCTCs and visceral metastasis. Reduction of emerin (EMD), a nuclear envelope protein, contributes to prostate cancer metastasis and nuclear shape instability. In this study, we investigated the correlation between EMD expression and the vsnCTC phenotype and its clinical impact.

We analyzed CTCs from 93 patients with metastatic castration-resistant prostate cancer and categorized them as either vsnCTC+ or vsnCTC- and compared overall survival and progression-free survival. C4-2B, 22Rv1, and DU145 with EMD knockdown were developed and characterized by nuclear size and gene expression by gene set enrichment analysis. Abiraterone- and enzalutamide-resistant C4-2B cells were also characterized by nuclear size and EMD expression.

Patients who were vsnCTC+ had significantly worse overall survival and progression-free survival compared with patients who were vsnCTC-. EMD expression was markedly reduced in CTCs from patients who were vsnCTC+ compared with patients who were vsnCTC-, with a significant positive correlation between EMD expression and CTC nuclear size. EMD knockdown in prostate cancer cells resulted in smaller nuclei, enhanced invasion, and the upregulation of genes associated with lineage plasticity. Additionally, abiraterone- and enzalutamide-resistant C4-2B cells had smaller nuclei and lower EMD expression. vsnCTC+ cells also showed enhanced platinum sensitivity.

The presence of vsnCTCs represents a novel hallmark of an aggressive subtype of metastatic castration-resistant prostate cancer closely linked to EMD loss and lineage plasticity. These findings highlight the importance of EMD dysregulation in the vsn phenotype, disease progression, and therapeutic resistance in patients with prostate cancer.

Circular RNA (circRNA) is a class of noncoding RNA with regulatory potentials. Its role in the transdifferentiation of prostate and lung adenocarcinoma into neuroendocrine prostate cancer (NEPC) and small cell lung cancer (SCLC) remains unexplored. Here, we identified circRMST as an exceptionally abundant circRNA predominantly expressed in NEPC and SCLC, with strong conservation between humans and mice. Functional studies using shRNA, siRNA, CRISPR-Cas13, and Cas9 consistently demonstrate that circRMST is essential for tumor growth and the expression of ASCL1, a master regulator of neuroendocrine fate. Genetic knockout of Rmst in NEPC genetic engineered mouse models prevents neuroendocrine transdifferentiation, maintaining tumors in an adenocarcinoma state. Mechanistically, circRMST physically interacts with lineage transcription factors NKX2-1 and SOX2. Loss of circRMST induces NKX2-1 protein degradation through autophagy-lysosomal pathway and alters the genomic binding of SOX2, collectively leading to the loss of ASCL1 transcription.

Treatment decisions in metastatic castration-resistant prostate cancer are mostly guided by clinical variables, but efforts to molecularly monitor the disease remain hampered by challenges in acquiring tumor tissue repeatedly. In this study, we simultaneously profiled the genome copy number and exome in longitudinal plasma circulating tumor DNA (ctDNA) acquired before, during, and upon progression to serial treatments with androgen signaling inhibitors and taxane chemotherapy from 60 patients with metastatic castration-resistant prostate cancer (2-10 samples per patient). The genomic data were used to delineate the clonal substructure and evolutionary dynamics of each patient, and an evolutionary dynamic index was developed to measure the longitudinal changes of the tumor subclones. Treatment with androgen signaling inhibitors resulted in greater subclonal selection and population structure changes than taxane treatment. The subclones that emerged in association with serial therapy resistance harbored recurrent aberrations in previously identified and new candidate genes, with particular enrichment in genes related to PI3K-AKT signaling. These findings indicate that the integration of detailed clinical and genomic data can provide a framework for future unbiased genomic applications for ctDNA in the clinic to enable precision medicine. Significance: Profiling of the genomic copy number changes and mutations in circulating tumor DNA collected longitudinally from prostate cancer patients receiving serial life-prolonging therapies elucidates evolutionary dynamics and identifies emerging resistant subclones.

Androgen deprivation therapy remains a cornerstone in managing prostate cancer. However, its recurrence often leads to the more aggressive castration-resistant prostate cancer (CRPC). Although second-line androgen receptor signaling inhibition treatments such as enzalutamide and abiraterone are available, their effectiveness against CRPC is only transient. High-dose testosterone (Hi-T) has recently emerged as a promising treatment for CRPC, primarily through the suppression of E2F and MYC signaling. However, the roles of Rb family proteins in influencing this therapeutic response remain debated. In this study, we utilized a CRPC patient-derived xenograft model that includes both Rb pathway-proficient and -deficient cell populations based on the positive or negative expression of RB family genes. Single-cell RNA sequencing analysis revealed that Rb-proficient cells displayed a robust response to Hi-T, whereas Rb-deficient cells exhibited significant resistance. Notably, our analysis indicated increased enrichment of the hypoxia signature in the Rb-deficient cell population. Further studies in RB1-silenced CRPC cell lines showed that treatment with a hypoxia-inducible factor-1α inhibitor can restore the sensitivity of Rb-deficient cells to high-dose dihydrotestosterone treatment. In conclusion, our research provides new molecular insights into CRPC tumor cell responses to Hi-T and proposes a new strategy to resensitize Rb-deficient CRPC cells to Hi-T treatment.

Colorectal cancer (CRC) progression and metastasis involve numerous regulatory factors. Among these, cellular retinoic acid-binding protein 2 (CRABP2) has been implicated as both a tumor activator and suppressor. Here, it is aimed to clarify the role of CRABP2 in CRC growth and metastasis and explore the underlying molecular mechanisms mediating its cellular functions. Using both in vitro and in vivo models, including a colonocyte-specific CRABP2 conditional knockout mouse model (Crabp2ΔIEC) and a subcutaneous tumorigenesis assay in BALB/c nude mice, it is shown that nuclear CRABP2 enhances tumor growth by interacting with and downregulating the tumor suppressor RB1, whereas cytoplasmic CRABP2 suppresses CRC liver metastasis by interacting with AFG3L2 and promoting mitophagy. In addition, the AFG3L2-SLC25A39 axis is identified as a distinct mechanism by which cytoplasmic CRABP2 increases mitochondrial glutathione stability to promote cell proliferation independent of the nuclear RB1 pathway. Notably, analysis of tissue from CRC patients reveals that CRABP2 protein has distinct prognostic implications and functional roles in the progression and metastasis of CRC dependent on its subcellular localization. Ultimately, by elucidating the role of CRABP2 in CRC, it is aimed to provide new insight into disease pathogenesis and inform the development of therapeutic interventions.

Double-negative prostate cancer (DNPC), characterized by an androgen receptor (AR)- and neuroendocrine-null phenotype, frequently emerges following androgen deprivation therapy (ADT). However, our understanding of the origins and regulatory mechanisms of DNPC remains limited. Here, we discover that tumors with KMT2C mutation or loss are highly susceptible to transitioning into DNPC following ADT. We clarify that DNPC primarily stems from luminal cell transdifferentiation rather than basal cell transformation. Antiandrogen treatment induces KMT2C binding at enhancers of a subset of AR-regulated genes, preserving the adenocarcinoma lineage. KMT2C maintains ASPP2 expression via enhancer-promoter communication post-AR inhibition, while its inactivation reduces ASPP2, triggering ΔNp63-dependent transdifferentiation. This DNPC transition maintains fatty acid (FA) synthesis through ΔNp63-mediated SREBP1c transactivation, fueling DNPC growth via HRAS palmitoylation and MAPK signaling activation. These findings highlight KMT2C as an epigenetic checkpoint against DNPC development and suggest the therapeutic potential of targeting fatty acid synthesis.

Intrinsic and acquired resistance to second-generation anti-androgens pose a significant clinical challenge in the treatment of metastatic castration-resistant prostate cancer (mCRPC). Novel biomarkers to predict treatment response and inform alternative treatment options are urgently needed.

Deep targeted sequencing, with a prostate cancer-specific gene panel, was performed on circulating tumor DNA (ctDNA) and germline DNA from blood of mCRPC patients recruited in Denmark (n = 53), prior to starting first-line treatment with enzalutamide or abiraterone acetate, and for a subset of patients also at progression (n = 18). Likely clonal hematopoietic variants were filtered out. Genomic findings were correlated to clinical outcomes (PSA progression-free survival (PFS), overall survival (OS)). Intrinsic resistance candidate biomarkers were considered by enrichment analysis of nonresponders vs. responders. Genomic alterations at progression were considered as possible drivers of acquired resistance. Clinical actionability was assessed based on OncoKB and ESCAT.

Somatic alterations in PTEN, cell cycle regulators (CCND1, CDKN1B, CDKN2A, and RB1) and chromatin modulators (CHD1, ARID1A) were associated with significantly shorter PFS and OS, also after adjusting for ctDNA% in multivariate Cox regression analysis. The associations with poorer outcomes for alterations in PTEN and chromatin modulators were validated in an external dataset. Patients with primary resistance to enzalutamide/abiraterone had enrichment for BRAF amplification and CHD1 loss, while responders had enrichment for TMPRSS2 fusions. AR resistance mutations emerged in 22% of patients at progression. These were mutually exclusive with other alterations that may confer resistance (i.e., activating CTNNB1 mutations, combined TP53/RB1 loss). Clinically actionable alterations, primarily in homologous recombination repair genes, were found in 54.7% and 49.0% of patients (OncoKB and ESCAT, respectively), with few additional alterations detected at progression. Level I alterations were identified in 41.5% of patients employing OncoKB, however only in 13.2% based on ESCAT.

Our study identifies known and novel prognostic and predictive biomarker candidates in patients with mCRPC undergoing first-line treatment with enzalutamide or abiraterone acetate. It further provides real-world evidence of the significant potential of genomic profiling of ctDNA to inform treatment in this setting. Clinical trials are warranted to advance the implementation of ctDNA-based biomarkers into clinical practice.

Transcription factors regulate a variety of events and maintain cellular homeostasis. Several transcription factors involved in embryonic development, has been shown to be closely associated with carcinogenesis when deregulated. Sry-like high mobility group box (SOX) proteins are potential transcription factors which are evolutionarily conserved. They regulate downstream genes to determine cell fate, via various signaling pathways and cellular processes essential for tissue and organ development. Dysregulation of SOXs has been reported to promote or suppress tumorigenesis by modulating cellular reprogramming, growth, proliferation, angiogenesis, metastasis, apoptosis, immune modulation, lineage plasticity, maintenance of the stem cell pool, therapy resistance and cancer relapse. This review provides a crucial understanding of the molecular mechanism by which SOXs play multifaceted roles in embryonic development and carcinogenesis. It also highlights their potential in advancing therapeutic strategies aimed at targeting SOXs and their downstream effectors in various malignancies.

The transdifferentiation from adenocarcinoma to neuroendocrine prostate cancer (NEPC) in men confers antiandrogen therapy resistance. Here our analysis combining CRISPR‒Cas9 screening with single-cell RNA sequencing tracking of tumor transition demonstrated that antiandrogen-induced zinc finger MYND-type containing 8 (ZMYND8)-dependent epigenetic programming orchestrates NEPC transdifferentiation. Ablation of Zmynd8 prevents NEPC development, while ZMYND8 upregulation mediated by achaete-scute homolog 1 promotes NEPC differentiation. We show that forkhead box protein M1 (FOXM1) stabilizes ZMYND8 binding to chromatin regions characterized by H3K4me1-H3K14ac modification and FOXM1 targeting. Antiandrogen therapy releases the SWI/SNF chromatin remodeling complex from the androgen receptor, facilitating its interaction with ZMYND8-FOXM1 to upregulate critical neuroendocrine lineage regulators. We develop iZMYND8-34, a small molecule designed to inhibit ZMYND8's histone recognition, which effectively blocks NEPC development. These findings reveal the critical role of ZMYND8-dependent epigenetic programming induced by androgen deprivation therapy in orchestrating lineage fate. Targeting ZMYND8 emerges as a promising strategy for impeding NEPC development.

Treatment-emergent neuroendocrine prostate cancer (NEPC) is aggressive and lethal. As androgen receptor signaling inhibitors (ARSIs) are increasingly used in earlier disease settings, treatment-emergent NEPC becomes more prevalent, and effective therapies are urgently needed. The purpose of this review was to summarize recent progress on emerging therapeutic targets of NEPC.

A multitude of therapeutic targets have emerged in NEPC over recent years. These targets may represent drivers of treatment-emergent lineage plasticity or simply be overexpressed on the surface of NEPC cells. Multiple modalities have been employed to drug these targets, with promising preclinical and clinical results. Treatment-emergent NEPC represents a distinct and clinically significant subset of castration-resistant prostate cancer (CRPC). Emerging therapeutic approaches have demonstrated encouraging efficacy and safety profiles, offering the potential to improve patient outcomes.

Prostate cancer is one of the most common malignancies in men, with most cases initially responding to androgen deprivation therapy. However, a significant number of patients eventually develop castration-resistant prostate cancer, an aggressive form of the disease. Although androgen receptor (AR) pathway inhibitors target AR signaling, and have extended survival in patients with castration-resistant prostate cancer, prolonged treatment can lead to the emergence of neuroendocrine prostate cancer (NEPC), a lethal subtype characterized by the expression of neuroendocrine markers and reduced AR activity. The transition from adenocarcinoma to NEPC is driven by lineage plasticity, wherein cancer cells adopt a neuroendocrine phenotype to evade treatment. Consequently, NEPC patients face poor clinical outcomes and limited effective treatment options. To improve outcomes, it is crucial to understand the molecular mechanisms driving NEPC development. In this review, we highlight the role of transcription factors in this process and explore their potential as therapeutic targets.

The mitochondrial unfolded protein response (UPRmt) maintains mitochondrial quality control and proteostasis under stress conditions. However, the role of UPRmt in aggressive and resistant prostate cancer is not clearly defined. We show that castration-resistant neuroendocrine prostate cancer (CRPC-NE) harbored highly dysfunctional oxidative phosphorylation (OXPHOS) Complexes. However, biochemical and protein analyses of CRPC-NE tumors showed upregulation of nuclear-encoded OXPHOS proteins and UPRmt in this lethal subset of prostate cancer suggestive of compensatory upregulation of stress signaling. Genetic deletion and pharmacological inhibition of the main chaperone of UPRmt heat shock protein 60 (HSP60) reduced neuroendocrine prostate cancer (NEPC) growth in vivo as well as reverted NEPC cells to a more epithelial-like state. HSP60-dependent aggressive NEPC phenotypes was associated with upregulation of β-catenin signaling both in cancer cells and in vivo tumors. HSP60 expression rendered enrichment of aggressive prostate cancer signatures and metastatic potential were inhibited upon suppression of UPRmt. We discovered that UPRmt promoted OXPHOS functions including mitochondrial bioenergetics in CRPC-NE via regulation of β-catenin signaling. Mitochondrial biogenesis facilitated cisplatin resistance and inhibition of UPRmt resensitizes CRPC-NE cells to cisplatin. Together, our findings demonstrated that UPRmt promotes mitochondrial health via upregulating β-catenin signaling and UPRmt represents viable therapeutic target for NEPC.

Neuroendocrine prostate cancer (NEPC), an aggressive subtype induced by hormone therapy, lacks effective treatments. This study explored the role of E26 transformation-specific variant 5 (ETV5) in NEPC development. Analysis of multiple prostate cancer datasets revealed that NEPC is characterized by significantly elevated ETV5 expression compared to other subtypes. ETV5 expression increased progressively under hormone therapy through epigenetic modifications. ETV5 induced neural stem-like features in prostate cancer cells and facilitated their differentiation into NEPC under hormone treatment conditions, both in vitro and in vivo. Our molecular mechanistic study identified PBX3 and TLL1 as target genes of ETV5 that contribute to ETV5 overexpression-induced castration resistance and stemness. Notably, obeticholic acid, identified as an ETV5 inhibitor in this study, exhibited promising efficacy in suppressing NEPC development. This study highlights ETV5 as a key transcription factor that facilitates NEPC development and underscores its potential as a therapeutic target for this aggressive cancer subtype.

Vaccinia-related kinase 1 (VRK1) is a serine-threonine kinase involved in the proliferation and migration of various cancer cells. However, its role in prostate cancer (PCa), particularly in the development of therapeutic resistance, remains unclear.

We established an androgen-independent PCa cell line derived from LNCaP prostate cancer cells and conducted transcriptome and proteome sequencing together with bioinformatic analyses of large clinical sample databases to investigate the potential role of VRK1 in PCa progression. The correlation between VRK1 and androgen receptor (AR) signaling was evaluated under simulated clinical treatment conditions. The effects of VRK1 on cell proliferation were assessed in vitro and in vivo using Cell Counting Kit-8 and colony formation assays. Additionally, proteome and transcriptome sequencing, combined with rescue experiments were performed to explore VRK1-regulated signaling pathways related to cell proliferation and therapeutic resistance.

VRK1 expression was elevated during the progression of androgen-dependent prostate cancer to castration-resistant prostate cancer under therapeutic conditions, and high VRK1 expression was associated with a poor prognosis in patients with PCa. VRK1 was regulated by AR signaling, and its silencing suppressed PCa cell proliferation both in vitro and in vivo. VRK1 drove cell proliferation and therapeutic resistance in PCa by modulating yes-associated protein 1 (YAP1).

VRK1 serves as a prognostic marker in PCa, regulated by AR signaling. VRK1 depletion inhibited cell proliferation both in vitro and in vivo, while elevated VRK1 upregulated YAP1, promoting cell proliferation and therapeutic resistance.

Prostate cancer (PCa) stands as a leading cause of cancer-related mortality among men, with treatment-induced neuroendocrine prostate cancer (NEPC) posing a challenge as an ARPI-resistant subtype. The role of transcription factors (TFs) in PCa progression and NEPC transdifferentiation remains inadequately understood, underscoring a critical gap in current research. In this study, an internal Z score-based approach is developed to identify lineage-specific TF profiles in prostatic adenocarcinoma and NEPC for a nuanced understanding of TF expression dynamics. Distinct TF profiles for adenocarcinoma and NEPC are unveiled, identifying 126 shared TFs, 46 adenocarcinoma-TFs, and 56 NEPC-TFs, validated across multiple cohorts. Gene Ontology is employed to validate their biological and functional roles in PCa progression. Implications are revealed in cell development, differentiation, and lineage determination. Knockdown experiments suggest that lineage-TFs are functionally important in maintaining lineage-specific cell proliferation. Additionally, a longitudinal study on NE transdifferentiation highlights dynamic TF expression shifts, proposing a three-phases hypothesis for PCa progression mechanisms. This study introduces a groundbreaking approach for deciphering the TF landscape in PCa, providing a molecular basis for adenocarcinoma to NEPC progression, and paving the way for innovative treatment strategies with potential impact on patient outcomes.

Three-dimensional (3D) cell culture models are increasingly utilized in cancer research to better replicate in vivo tumor microenvironments. This study examines the effects of different 3D scaffolding materials, including Matrigel, GelTrex, and the plant-based GrowDex, on prostate cancer cell lines, with a particular emphasis on neuroendocrine prostate cancer (NEPC). Four cell lines (LNCaP, LASCPC-01, PC-3, and KUCaP13) were cultured in these scaffolds to evaluate spheroid formation, cell viability, and gene expression. The results revealed that while all scaffolds supported cell viability, spheroid formation varied significantly: Matrigel promoted the most robust spheroids, especially for LASCPC-01, whereas GrowDex exhibited limitations for certain cell lines. Gene expression analysis indicated a consistent reduction in androgen receptor (AR) expression in LNCaP cells across all scaffolds, suggesting a potential shift towards a neuroendocrine phenotype. However, the expression of neuroendocrine markers varied depending on the scaffold and culture method, with the mini-domes method in Matrigel leading to decreased expression of both castration-resistant prostate cancer (CRPC) and NEPC markers. These findings highlight the scaffold-dependent variability in 3D culture outcomes and emphasize the need for standardized methodologies to ensure consistency and relevance in prostate cancer research.

Neuroendocrine prostate cancer (NEPC) arises primarily through neuroendocrine transdifferentiation (NEtD) as an adaptive mechanism of therapeutic resistance. Models to define the functional effects of putative drivers of this process on androgen receptor (AR) signaling and NE cancer lineage programs are lacking. We adapted a genetically defined strategy from the field of cellular reprogramming to directly convert AR-active prostate cancer (ARPC) to AR-independent NEPC using candidate factors. We delineated critical roles of the pioneer factors ASCL1 and NeuroD1 in NEtD and uncovered their abilities to silence AR expression and signaling by remodeling chromatin at the somatically acquired AR enhancer and global AR binding sites with enhancer activity. We also elucidated the dynamic temporal changes in the transcriptomic and epigenomic landscapes of cells undergoing acute lineage conversion from ARPC to NEPC which should inform future therapeutic development. Further, we distinguished the activities of ASCL1 and NeuroD1 from the inactivation of RE-1 silencing transcription factor (REST), a master suppressor of a major neuronal gene program, in establishing a NEPC lineage state and in modulating the expression of genes associated with major histocompatibility complex class I (MHC I) antigen processing and presentation. These findings provide important, clinically relevant insights into the biological processes driving NEtD of prostate cancer.

Neuroendocrine tumors (NETs) are a diverse group of malignancies that can occur in various organs, with a notable prevalence in the lungs and gastrointestinal tract, which are the focus of this Review. Although NETs are rare in individual organs, their incidence has increased over recent decades, highlighting the urgent need for current classification systems to evolve by incorporating recent advances in the understanding of NET biology. Several omics studies have revealed molecular subtypes, which, when integrated into existing classification frameworks, may provide more clinically relevant insights for patients with NETs. This Review examines recent progress in elucidating the biology of NETs, with a particular emphasis on the tumor microenvironment and cells of origin. The existence of different cells of origin, which may contribute to distinct molecular groups, along with profiles of immune infiltration - despite being generally low - could explain the emergence of more aggressive cases and the potential for metastatic progression. Given the molecular heterogeneity of NETs and the diversity of their microenvironments and different cells of origin, there is an urgent need to develop morphomolecular classification systems. Such systems would make it possible to better characterize tumor progression, identify new therapeutic targets, and, ultimately, guide the development of personalized therapies.

An 87-year-old man presented with worsening urinary dysfunction with urinary retention. The prostate-specific antigen was 1.23 ng/mL. Prostate biopsy confirmed adenocarcinoma with a Gleason score of 5+5; the final diagnosis was T4N1M0. Androgen deprivation therapy and immediate whole-pelvic radiotherapy (80 Gy) were administered. Nadir prostate-specific antigen of 0.40 ng/mL two months later increased to 35.92 ng/mL as the disease metastasized to the corpus cavernosum penis, causing malignant priapism. The best supportive care was provided, and the patient died 11 months later. High-grade prostate cancers with low prostate-specific antigen levels are rare but highly aggressive. Androgen deprivation therapy and radiotherapy alone may be insufficient in such cases, highlighting the need for more effective treatment strategies.

Prostate cancer (PC) growth is hormone-dependent and it frequently develops distant metastases as disease progresses. Patients with metastatic castration-sensitive prostate cancer (mCSPC) initially respond to androgen deprivation therapy (ADT) but eventually become refractory and develop metastatic castration-resistant prostate cancer (mCRPC). Castration-resistance is associated with high lethality and metastases confer poor prognosis, therefore unmet needs in treatment for mCSPC remain high. So far, improvements in survival in mCSPC have been achieved by doublet combination therapy such as docetaxel or an androgen-receptor signaling inhibitor (ARSI) in addition to ADT. Further, recent phase 3 trials have shown that triplet therapy-a combination of ARSI, docetaxel, and ADT improves prognosis compared with docetaxel plus ADT in mCSPC. PC tumors manifest intra- and inter-tumoral heterogeneity at both the genetic and phenotypic level. As heterogeneity increases during sequential treatment and disease progression, it is reasonable to initiate combination therapy using drugs with different mechanisms of action early in the course of disease, such as mCSPC. Previous research about tumor heterogeneity and drug resistant mechanism support this rationale, as well as preclinical studies and real-world data provide the scientific evidence of benefit by combining ARSI and docetaxel. Here, we review the rationale and clinical evidence for triplet therapy in patients with mCSPC.

Tumor lineage plasticity (LP) is an emerging hallmark of cancer progression. Through pharmacologically probing the function of epigenetic regulators in prostate cancer cells and organoids, we identified bromodomain protein BRD4 as a crucial player. Integrated ChIP-seq and RNA-seq analysis of tumors revealed, for the first time, that BRD4 directly activates hundreds of genes in the LP programs which include neurogenesis, axonogenesis, EMT and stem cells and key drivers such as POU3F2 (BRN2), ASCL1/2, NeuroD1, SOX2/9, RUNX1/2 and DLL3. Interestingly, BRD4 genome occupancy is reprogrammed by anti-AR drugs from facilitating AR function in CRPC cells to activating the LP programs and is facilitated by pioneer factor FOXA1. Significantly, we demonstrated that BRD4 inhibitor AZD5153, currently at clinical development, possesses potent activities in complete blockade of tumor growth of both de novo neuroendocrine prostate cancer (NEPC) and treatment-induced NEPC PDXs and that suppression of tumor expression of LP programs through reduction of local chromatin accessibility is the primary mechanism of action (MOA) by AZD5153. Together, our study revealed that BRD4 plays a fundamental role in direct activation of tumor LP programs and that its inhibitor AZD5153 is highly promising in effective treatment of the lethal forms of the diseases.

Acquired resistance to androgen receptor (AR)-targeted therapies underscores the need to identify alternative therapeutic targets for treating lethal prostate cancer. In this study, we evaluated the prognostic significance of 1635 human transcription factors (TFs) by analyzing castration-resistant prostate cancer (CRPC) datasets from the West and East Stand Up to Cancer (SU2C) cohorts. Through this screening approach, we identified E2F8, a putative transcriptional repressor, as a TF consistently associated with poorer patient outcomes in both cohorts. Notably, E2F8 is highly expressed and active in AR-negative CRPC compared to AR-positive CRPC. Integrative profiling of E2F8 cistromes and transcriptomes in AR-negative CRPC cells revealed that E2F8 directly and non-canonically activates target oncogenes involved in cancer-associated pathways. To target E2F8 in CRPC, we employed the CRISPR/CasRx system to knockdown E2F8 mRNA, resulting in effective and specific downregulation of E2F8 and its target oncogenes, as well as significant growth inhibition in AR-negative CRPC in both cultured cells and xenograft models. Our findings identify and characterize E2F8 as a targetable transcriptional activator driving CRPC, particularly the growth of AR-negative CRPC.

Loss of RB1 function represents a defining characteristic of triple-negative breast cancer (TNBC) and is intricately associated with resistance to therapeutic interventions. In this study, we investigate the epigenetic mechanisms governing RB1 expression in TNBC. Employing a combination of bioinformatics analyses and experimental validations, we identified lysine histone methyltransferase EZH2 as a key upstream regulator of RB1 expression. EZH2 primarily mediates trimethylation of lysine 27 on histone H3 as the catalytic subunit of the Polycomb repressive complex 2 (PRC2) complex. Furthermore, our findings demonstrate that pharmacological inhibition of EZH2 leads to a significant upregulation of RB1 expression levels, mediated by enhanced enrichment of the activating histone marker H3K27ac at the RB1 enhancer region, as evidenced by ATAC-sequencing and ChIP-qPCR assays. These insights unveil a promising clinical avenue for combating RB1-mediated drug resistance in TNBC through the strategic integration of epigenetic-targeting agents.

Gallbladder neuroendocrine carcinomas (GB-NECs) are a rare subtype of malignant gallbladder cancer (GBC). The genetic and molecular characteristics of GB-NECs are rarely reported. This study aims to assess the frequency of microsatellite instability (MSI) in GB-NECs and characterize their clinicopathologic and molecular features in comparison with gallbladder adenocarcinomas (GB-ADCs). Data from six patients with primary GB-NECs and 13 with GB-ADCs were collected and reevaluated. MSI assay, immunohistochemistry for mismatch repair proteins (MLH1, MSH2, MSH6, and PMS2), comprehensive genomic profiling (CGP) via next-generation sequencing (NGS), and evaluation of tumor mutation burden (TMB) were conducted on these samples. The six GB-NEC cases were all female, with a mean age of 62.0±9.2 years. Of these, two cases were diagnosed as large cell neuroendocrine carcinomas (LCNECs), while the remaining four were small cell neuroendocrine carcinomas (SCNECs). Microsatellite states observed in both GB-NECs and GB-ADCs were consistently microsatellite stable (MSS). Notably, TP53 (100%, 6/6) and RB1 (100%, 6/6) exhibited the highest mutation frequency in GB-NECs, followed by SMAD4 (50%, 3/6), GNAS (50%, 3/6), and RICTOR (33%, 2/6), with RB1, GNAS, and RICTOR specifically present in GB-NECs. Immunohistochemical (IHC) assays of p53 and Rb in the six GB-NECs were highly consistent with genetic mutations detected by targeted NGS. Moreover, no statistical difference was observed in TMB between GB-NECs and GB-ADCs (p=0.864). Although overall survival in GB-NEC patients tended to be worse than in GB-ADC patients, this difference did not reach statistical significance (p=0.119). This study has identified the microsatellite states and molecular mutation features of GB-NECs, suggesting that co-mutations in TP53 and RB1 may signify a neuroendocrine inclination in GB-NECs. The IHC assay provides an effective complement to targeted NGS for determining the functional status of p53 and Rb in clinical practice.

Despite incredible progress in describing the molecular underpinnings of prostate cancer over the last decades, pathologic examination remains indispensable for predicting aggressive behavior in the localized setting. Beyond pathologic grade, specific histologic findings have emerged as critical prognostic or predictive indicators. Here, the authors review molecular correlates of aggressive histologic subtypes of prostate cancer in the localized setting, demonstrating that many of the signature molecular alterations found in metastatic disease-such as tumor suppressor gene loss and DNA repair defects-are enriched in primary disease with adverse histologic features, presaging aggressive behavior, and presenting opportunities for earlier germline screening or targeted therapies.

We established a robust and sensitive androgen response element luciferase reporter assay to monitor androgen receptor (AR) activity using KUCaP13 cells, a novel human-derived treatment-related neuroendocrine prostate cancer (t-NEPC) cell line. A high-throughput screening using a chemical library to identify potential compounds that induce AR re-expression in KUCaP13 cells revealed 30 candidate molecules potentially enhancing luciferase luminescence; however, subsequent validation steps demonstrated these signals to be false-positives. Despite not achieving the goal of AR re-expression, this study stands as a significant proof-of-concept for the application of high-throughput screening approaches in t-NEPC research.

Identifying effective therapies targeting multi-protein complexes that lack catalytic sites or cofactor pockets remains a long-standing challenge. The proto-oncogene, ubiquitin E3 ligase SCFSkp2, is one such target. SCFSkp2 promotes the proteasomal degradation of the cyclin-dependent kinase inhibitor p27, which controls cell cycle progression. Targeted knockout of Rb1/Trp53 causes metastatic prostate cancer in mice; additional knockout of Skp2 completely blocks tumorigenesis. We compared gene-edited mice that carried two different single amino acid changes in the SCFSkp2 complex, structurally predicted to inhibit the degradation of p27. Mutation of the SCFSkp2 accessory protein Cks1 (Cks1N45R) completely blocked Rb1/Trp53-driven prostate tumorigenesis, phenocopying Skp2 knockout, whereas a mutation directly stabilizing p27 (p27T187A) did not. This was consistent with structural models that predicted the binding of both p27 and p27T187A to the SCFSkp2/Cks1/Cdk2/CyclinA/p27 complex, and their subsequent ubiquitination and degradation, albeit at different rates. Two binding modes, which differ in their dependence on phosphorylated T187, are predicted by the model. Studies confirmed the role of p27 in mediating tumorigenesis in Rb1/Trp53 mutant tumors and revealed a mutually destabilizing Skp2 and p27 feedback loop. The integration of gene editing, drug-surrogate mutations, and mouse tumor models offers a blueprint for studying SCFSkp2 and other multi-subunit biomedical targets.

Metastatic castration-resistant prostate cancer (mCRPC) is a heterogeneous disease. Several studies have identified transcriptional subtypes of mCRPC, but comprehensive analysis of prognostic gene expression pathways has been limited. Therefore, we aggregated a cohort of 1012 mCRPC tissue samples from 769 patients and investigated the association of gene expression-based pathways with clinical outcomes and intrapatient and intratumor heterogeneity. Survival data were obtained for 272 patients. Pathway-level enrichment was evaluated using gene set variation analysis. scRNA-seq datasets from mCRPC tissue biopsies and circulating tumor cells were used to investigate heterogeneity of adverse pathways. We identified five pathway clusters: (a) Immune response/WNT/TGF-beta signaling, (b) AR signaling/luminal signatures, (c) mTOR signaling and glycolysis, (d) cell proliferation, and (e) neuroendocrine differentiation. Proliferation, AR signaling loss, and glycolysis/mTOR signaling were independently prognostic. Adverse prognostic pathway scores decreased on treatment with AR signaling inhibitors, but not at progression, suggesting failure to permanently target these pathways. scRNA-seq datasets from mCRPC tissue biopsies and circulating tumor cells were used to investigate heterogeneity of adverse pathways. Our results suggest loss of AR signaling, high proliferation, and a glycolytic phenotype as adverse prognostic pathways in mCRPC that could be used in conjunction with clinical factors to prognosticate for treatment decisions.

Neuroendocrine carcinoma (NEC) represents a category of malignant tumors originating from neuroendocrine cells. Given that NEC cells exhibit characteristics of both neural and endocrine cells, they can hijack neuronal signaling pathways and dynamically regulate the expression of neuronal lineage markers during tumor metastasis, thereby constructing a microenvironment conducive to tumor growth and metastasis. Conversely, alterations in the tumor microenvironment can enhance the interactions between neurons and tumor cells, ultimately synergistically promoting the metastasis of NEC. This review highlights recent advancements in the field of cancer neuroscience, uncovering neuronal lineage markers in NEC that facilitate tumor dissemination through mediating crosstalk, bidirectional communication, and synergistic interactions between tumor cells and the nervous system. Consequently, the latest findings in tumor neuroscience have enriched our understanding of the biological mechanisms underlying tumor metastasis, opening new research avenues for a deeper comprehension of the complex biological processes involved in tumor metastasis, particularly brain metastasis. This review provides a comprehensive review of the crosstalk between tumor cells and neural signaling in the metastasis of NEC.
.