Risk SNP-Mediated Promoter-Enhancer Switching Drives Prostate Cancer through lncRNA PCAT19.

The prostate cancer (PCa) risk-associated SNP rs11672691 is positively associated with aggressive disease at diagnosis. We showed that rs11672691 maps to the promoter of a short isoform of long noncoding RNA PCAT19 (PCAT19-short), which is in the third intron of the long isoform (PCAT19-long). The risk variant is associated with decreased and increased levels of PCAT19-short and PCAT19-long, respectively. Mechanistically, the risk SNP region is bifunctional with both promoter and enhancer activity. The risk variants of rs11672691 and its LD SNP rs887391 decrease binding of transcription factors NKX3.1 and YY1 to the promoter of PCAT19-short, resulting in weaker promoter but stronger enhancer activity that subsequently activates PCAT19-long. PCAT19-long interacts with HNRNPAB to activate a subset of cell-cycle genes associated with PCa progression, thereby promoting PCa tumor growth and metastasis. Taken together, these findings reveal a risk SNP-mediated promoter-enhancer switching mechanism underlying both initiation and progression of aggressive PCa.

Aberrant expression of GATA3 in metastatic adenocarcinoma of the prostate: an important pitfall.

AIMS: The distinction of high-grade prostate cancer (PCa) from poorly differentiated urothelial carcinoma (UC) can be somewhat challenging on clinical and morphological grounds alone, yet it is of great importance for prognostication and choice of treatment. GATA3 is a useful immunohistochemical marker to confirm urothelial origin. However, recent works report strong GATA3 immunoexpression in primary high-grade PCa. The aim of this study was to explore GATA3 expression specifically in metastatic PCa. METHODS AND RESULTS: The pathology databases of four tertiary institutions were queried for cases of metastatic PCa. Available slides and clinical records were reviewed by experienced genitourinary pathologists. Prostatic markers (PSA, PSAP, NKX3.1) and GATA3 immunohistochemistry were performed. A total of 163 metastatic PCa cases were included. At least one prostate marker was positive in each case of non-regional distant metastasis, confirming prostatic origin. GATA3 strong staining was found in four (2.5%) cases: two liver, one bone and one non-regional lymph-node metastases. All four patients had Grade Group 5 PCa at the initial diagnosis. The metastatic prostatic adenocarcinomas were solid, either with no gland formation (n = 3) or with only focal cribriforming (n = 1). CONCLUSIONS: To our knowledge, this is the first study exploring GATA3 expression specifically in metastatic PCa. Despite being infrequent, GATA3 positivity in high-grade PCa may lead to misdiagnosis, with clinical implications. We recommend a panel of immunohistochemical markers, both prostatic and urothelial, for ruling out UC, either in primary tumour samples or in the event of metastases of unknown primary, when a genitourinary origin is suspected.

Fibroblast-derived exosomal microRNA regulates NKX3-1 expression in androgen-sensitive, androgen receptor-dependent prostate cancer cells.

Androgen deprivation therapy (ADT) targeting androgen production and androgen receptor (AR) signaling is the primary antihormonal therapy in the treatment of advanced prostate cancer (PCa). However, no clinically established molecular biomarkers have been identified to predict the effectiveness of ADT before starting ADT. The tumor microenvironment of PCa contains fibroblasts that regulate PCa progression by producing multiple soluble factors. We have previously reported that AR-activating factor-secreted fibroblasts increase the responsiveness of androgen-sensitive, AR-dependent PCa cells to ADT. Thus, we hypothesized that fibroblast-derived soluble factors may affect cancer cell differentiation by regulating cancer-related gene expression in PCa cells and that the biochemical characteristics of fibroblasts may be used to predict the effectiveness of ADT. Here, we investigated the effects of normal fibroblasts (PrSC cells) and three PCa patient-derived fibroblast lines (pcPrF-M5, -M28, and -M31 cells) on the expression of cancer-related genes in androgen-sensitive, AR-dependent human PCa cells (LNCaP cells) and three sublines showing different androgen sensitivities and AR dependencies. The mRNA expression of the tumor suppressor gene NKX3-1 in LNCaP cells and E9 cells (which show low androgen sensitivity and AR dependency) was significantly increased by treatment with conditioned media from PrSC and pcPrF-M5 cells but not from pcPrF-M28 and pcPrF-M31 cells. Notably, no upregulation of NKX3-1 was observed in F10 cells (AR-V7-expressing, AR-independent cells with low androgen sensitivity) and AIDL cells (androgen-insensitive, AR-independent cells). Among 81 common fibroblast-derived exosomal microRNAs that showed 0.5-fold lower expression in pcPrF-M28 and pcPrF-M31 cells than in PrSC and pcPrF-M5 cells, miR-449c-3p and miR-3121-3p were found to target NKX3-1. In only LNCaP cells, the NKX3-1 mRNA expression was significantly increased by transfection of an miR-3121-3p mimic but not that of the miR-449c-3p mimic. Thus, fibroblast-derived exosomal miR-3121-3p may be involved in preventing the oncogenic dedifferentiation of PCa cells by targeting NKX3-1 in androgen-sensitive, AR-dependent PCa cells.

Mucous Gland Adenoma of the Lung: A Neoplastic Counterpart of Mucinous Bronchial Glands.

Mucous gland adenoma (MGA) is a rare benign tumor that usually arises in the proximal airway and consists of mucus-secreting cells resembling bronchial glands. Here, we report 2 cases of MGAs and describe their morphologic, immunohistochemical, and molecular profiles in comparison with 19 pulmonary tumors of 5 other histologic types with mucinous cells (invasive mucinous adenocarcinoma, mucoepidermoid carcinoma, mixed squamous cell and glandular papilloma, bronchiolar adenoma/ciliated muconodular papillary tumor, and sialadenoma papilliferum). Two MGAs were found in 1 male patient and 1 female patient, located in the bronchus and trachea, respectively. One MGA was examined by RNA sequencing, and no putative driver mutations (including BRAF, KRAS, and AKT1 mutations) or gene fusions were identified. In another case of MGA, V600E mutations of BRAF and E17K mutations of AKT1 were not detected by allele-specific real-time PCR or digital PCR, respectively. However, a gene expression analysis revealed that the MGA presented a specific RNA expression profile with multiple genes enriched in the salivary gland. The gene expression of NKX3.1 was significantly higher in the MGA case in comparison to normal control lungs (P < .001). We then examined NKX3.1 immunohistochemistry for 2 MGAs and 19 tumors of 5 other histologic types. NKX3.1 was positive in MGA (2/2, 100%), whereas all constituent cells, including mucinous cells, were negative for NKX3.1 in other histologic types (0%, 0/19). In normal lung tissue, NKX3.1 was positive for mucinous acinar cells of the bronchial glands. In conclusion, the gene expression profile, taken together with the histologic similarity between MGA and bronchial glands, and the preferred location of the tumors (proximal airways with submucosal glands) suggest that MGA is a neoplastic counterpart of mucinous bronchial glands. NKX3.1 immunohistochemistry can be a sensitive and specific ancillary marker that distinguishes MGA from other histologic mimics.

TALEN-mediated generation of Nkx3.1 knockout rat model.

BACKGROUND: Recent developments in gene editing, using transcriptional activator-like effector nucleases (TALENs), have greatly helped the generation of genetically engineered animal models. The NK3 homeobox 1 (NKX3.1) protein plays important roles in prostate development and protein production, and functions as a tumor suppressor. Recently, NKX3.1 was shown to be associated with breast cancer in humans. METHODS: Our aim was to create a new rat model to elucidate the functions of NKX3.1. To that end, we generated Nkx3.1 knockout rats using TALENs and analyzed their phenotype. TALEN-mediated Nkx3.1 knockout was confirmed by T7 endonuclease I (T7E1) assay and DNA sequencing. Prostate weight and fertility were evaluated in the knockout rats, besides determining the proportion of epithelial cells and messenger RNA (mRNA) expression of genes associated with carcinogenesis. Breast tumors were examined by histopathology. RESULTS: Results suggested Nkx3.1 knockout rats have reduced fertility, decreased prostate weights, and increased epithelial cell layers. The mRNA expression of genes related to prostate carcinogenesis, namely Ar, Akt, and Pi3k, also increased. Moreover, the Nkx3.1 knockout rats often developed malignant breast tumors. CONCLUSIONS: We, therefore, successfully created the first Nkx3.1 knockout rat model, using TALEN-mediated gene targeting, and used it to identify defects associated with Nkx3.1 deficiency, not previously observed in mice. Loss of Nkx3.1 in rats led to lower reproductive capacity, and decreased prostate weights, apart from the risk of developing breast cancer. We, thus, proposed Nkx3.1 knockout rats as reliable models for studying the role of NKX3.1 in decreased prostate weights, fertility, and breast cancer, as well as in prostate cancer.

Reciprocal deregulation of NKX3.1 and AURKA axis in castration-resistant prostate cancer and NEPC models.

BACKGROUND: NKX3.1, a prostate-specific tumor suppressor, is either genomically lost or its protein levels are severely downregulated, which are invariably associated with poor prognosis in prostate cancer (PCa). Nevertheless, a clear disconnect exists between its mRNA and protein levels, indicating that its post-translational regulation may be critical in maintaining its protein levels. Similarly, AURKA is vastly overexpressed in all stages of prostate cancer (PCa), including castration-resistant PCa (CRPC) and neuroendocrine PCa (NEPC), although its transcripts are only increased in ~ 15% of cases, hinting at additional mechanisms of deregulation. Thus, identifying the upstream regulators that control AURKA and NKX3.1's levels and/or their downstream effectors offer an alternative route to inhibit AURKA and upregulate NKX3.1 in highly fatal CRPC and NEPC. AURKA and NKX3.1 have not linked to each other in any study to date. METHODS: A chemical genetic screen revealed NKX3.1 as a direct target of AURKA. AURKA-NKX3.1 cross-talk was analyzed using several biochemical techniques in CRPC and NEPC cells. RESULTS: We uncovered a reciprocal loop between AURKA and NKX3.1 in CRPC and NEPC cells. We observed that AURKA-mediated NKX3.1 downregulation is a major mechanism that drives CRPC pathogenesis and NEPC differentiation. AURKA phosphorylates NKX3.1 at three sites, which degrades it, but AURKA does not regulate NKX3.1 mRNA levels. NKX3.1 degradation drives highly aggressive oncogenic phenotypes in cells. NKX3.1 also degrades AURKA in a feedback loop. NKX3.1-AURKA loop thus upregulates AKT, ARv7 and Androgen Receptor (AR)-signaling in tandem promoting highly malignant phenotypes. Just as importantly, we observed that NKX3.1 overexpression fully abolished synaptophysin and enolase expression in NEPC cells, uncovering a strong negative relationship between NKX3.1 and neuroendocrine phenotypes, which was further confirmed be measuring neurite outgrowth. While WT-NKX3.1 inhibited neuronal differentiation, 3A-NKX3.1 expression obliterated it. CONCLUSIONS: NKX3.1 loss could be a major mechanism causing AURKA upregulation in CRPC and NEPC and vice versa. NKX3.1 genomic loss requires gene therapy, nonetheless, targeting AURKA provides a powerful tool to maintain NKX3.1 levels. Conversely, when NKX3.1 upregulation strategy using small molecules comes to fruition, AURKA inhibition should work synergistically due to the reciprocal loop in these highly aggressive incurable diseases.

A comparison of prostatic development in xenografts of human fetal prostate and human female fetal proximal urethra grown in dihydrotestosterone-treated hosts.

The goal of this paper is to explore the ability of the human female urogenital sinus immediately below the bladder (proximal urethra) to undergo prostatic development in response to dihydrotestosterone (DHT). To establish this idea, xenografts of human fetal female proximal urethra were grown in castrated nude mouse hosts receiving a subcutaneous DHT pellet. To verify the prostatic nature of the resultant glands, DHT-treated human fetal female urethral xenografts were compared with human fetal prostatic xenografts (derived from male specimens) grown in untreated and DHT-treated castrated mouse hosts and human fetal female proximal urethral xenografts grown in untreated castrated hosts. The resultant glands observed in DHT-treated human fetal female proximal urethral xenografts expressed 3 prostate-specific markers, NKX3.1, prostate specific antigen and prostatic acid phosphatase as well as the androgen receptor. Glands induced by DHT exhibited a protein expression profile of additional immunohistochemical markers (seven keratins, RUNX1, ESR2, TP63 and FOXA1) consistent with the unique spatial pattern of these proteins in prostatic ducts. Xenografts of human fetal female proximal urethra grown in DHT-treated hosts also expressed one of the salient features of prostatic development, namely androgen responsiveness. The experimental induction of prostatic differentiation from human fetal female proximal urethra makes possible future in-depth analysis of the molecular pathways directly involved in initiation of human prostatic development and subsequent epithelial differentiation, and more important whether the molecular pathways involved in human prostatic development are similar/identical versus different from that in murine prostatic development.

NKX3.1 a useful marker for mesenchymal chondrosarcoma: An immunohistochemical study.

INTRODUCTION: Mesenchymal chondrosarcoma is a rare subtype of chondrosarcoma. The tumor has a characteristic bimorphic pattern with areas of poorly differentiated small round cell component and interspersed islands of well differentiated hyaline cartilage. Histological diagnosis of mesenchymal chondrosarcoma is very challenging especially in small biopsies when tumor presents with little cartilaginous component. In such cases, it is very difficult to distinguish mesenchymal chondrosarcoma from other round blue cell tumors like Ewing's sarcoma, rhabdomyosarcoma, small cell osteosarcoma and desmoplastic round blue cell tumor. Immunohistochemically, mesenchymal chondrosarcoma stains positive for NKX2.2, CD99, S100 and SOX9. This immunoprofile is non-specific and overlaps with other round blue cell tumors. Till recently, there was no reliable immunohistochemical marker to differentiate mesenchymal chondrosarcoma from other round blue cell tumors. NKX3.1, though widely used as a diagnostic biomarker for prostatic adenocarcinoma, has been recently proposed by Yoshida et al. (2020) as a unique marker of mesenchymal chondrosarcoma and EWSR1-NFATC2 sarcoma. OBJECTIVE: The aim of our study was to further explore utility of NKX3.1 as a diagnostic marker of mesenchymal chondrosarcoma. MATERIAL & METHODS: We applied NKX3.1 immunohistochemistry to 21 cases of mesenchymal chondrosarcoma and 32 cases of other round blue cell tumors. RESULTS: 14 out of 21 cases (66.7%) of mesenchymal chondrosarcoma stained positive for NKX3.1 with nuclear expression in small round component. Cartilaginous component was predominantly negative. All other round blue cell tumors showed negative results. CONCLUSION: Based on our study results we suggest that NKX3.1 is a useful immunohistochemical marker in differentiating mesenchymal chondrosarcoma from its histological mimics.

Pleomorphic giant cell carcinoma of prostate: Rare tumor with unique clinicopathological, immunohistochemical, and molecular features.

Pleomorphic giant cell carcinoma (PGCC) of the prostate is a rare entity categorized as a variant of prostatic acinar adenocarcinoma in the 2016 World Health Organization (WHO) classification system. PGCC differs from conventional prostatic adenocarcinoma by having bizarre, markedly enlarged, and pleomorphic cells. It differs from high grade urothelial carcinoma by negativity for urothelial differentiation markers, and can be distinguished from sarcomatoid carcinoma by lack of spindle cells. Including two new cases described herein, there have been 51 cases of prostate PGCC reported in the English literature. Clinical features shared by cases of prostate PGCC include poor prognosis, occurrence in older patients, and frequent association with prior therapy. Pathologic features common to cases of prostate PGCC include admixture with a high-grade conventional prostate carcinoma component and absent or reduced expression of prostate differentiation markers. More recent studies have begun to elucidate the molecular characteristics of PGCC, detecting specific mutations and chromosomal translocations, and showing evidence of a high degree of molecular instability in these tumors. We report novel findings in two cases of PGCC including a PIK3CA p.His1047Arg mutation not previously described. One of our cases is the first to clearly demonstrate chronological loss of prostate markers during dedifferentiation from prior conventional prostate carcinoma to PGCC. Herein, we present our two new cases and comprehensively review the literature on all reported cases of PGCC with critical commentary on findings in cases of this rare tumor.

Immunohistochemical Reactivity of Prostate-Specific Markers for Salivary Duct Carcinoma.

OBJECTIVES: NKX3.1, a transcription factor related to androgen expression, has recently been introduced as a diagnostic marker of prostate adenocarcinoma. Salivary duct carcinoma (SDC) is typically positive for androgen receptor (AR). Therefore, we hypothesized that NKX3.1 is a new immunohistochemical marker for SDC and aimed to investigate whether NKX3.1 staining in combination with other immunomarkers of prostate carcinoma could have a diagnostic or prognostic value in SDC. METHODS: Materials obtained from 42 resected SDCs were examined by immunohistochemistry using antibodies against AR, NKX3.1, α-methylacyl-CoA racemase (AMACR), prostatic acid phosphatase (PAP), and prostate-specific antigen (PSA). RESULTS: In immunoreactivity among SDC cases, 81.0, 35.7, 58.5, 33.3, and 0% were positive for AR, NKX3.1, AMACR, PAP, and PSA, respectively. AMACR and PAP immunoreactivity rates were higher in recurrence cases than in cases with no recurrence. CONCLUSIONS: NKX3.1 expression is useful for SDC diagnosis, but decreased NKX3.1 expression was not correlated with SDC progression. The immunoreactivity of AMACR and PAP could be useful for assessing prognosis in SDC, but immunohistochemical staining of prostate-specific markers should be interpreted with caution when determining whether a metastatic tumor is of prostate origin, especially when patients have a history of SDC.

Utility of NKX3.1 immunohistochemistry in the differential diagnosis of seminal vesicles versus prostatic tissue in needle biopsy.

NKX3.1 is considered a reliable immunohistochemical marker of prostatic origin with high specificity and sensitivity. However, NKX3.1 positivity has been described in other neoplastic and non-neoplastic tissues, such as mesenchymal chondrosarcoma, sex-cord stromal tumors, rete testis adenocarcinoma, lobular and ductal carcinoma of the breast, salivary glands, peribronchial submucosal glands, and Sertoli cells. We analyzed expression of two antibodies (mono and polyclonal) of NKX3.1 in a total of 63 non-neoplastic seminal vesicles. We used 52 resection materials (12 seminal vesicles without prostatic adenocarcinoma, 26 seminal vesicles with prostatic adenocarcinoma infiltration, and 14 cases of seminal vesicles infiltrated by urothelial carcinoma) and 11 prostatic core needle biopsies with incidentally sampled fragment of seminal vesicles. In all cases, tissues from seminal vesicles were completely negative for NKX3.1, despite using polyclonal and monoclonal NKX3.1 antibodies, and regardless of the detection system utilized (diaminobenzidine (DAB) versus alkaline phosphatase (AF)). However, prostatic adenocarcinoma was negative in several cases (n = 6), when AF detection system was used. Reaction with DAB was strong and robust in all cases. Based on our data, we can recommend NKX3.1 as a negative immunohistochemical marker of seminal vesicles.

Immunohistochemistry in prostate pathology.

The diagnosis of prostatic adenocarcinoma is based on a combination of histological features, none of which is absolutely sensitive and specific. Immunohistochemical examination is therefore sometimes necessary in difficult cases for confirmation of the diagnosis and distinction of mimickers. The second major indication of immunohistochemical staining in prostatic pathology is metastatic prostatic adenocarcinoma.

Oxidative DNA Damage-Mediated Genomic Heterogeneity Is Regulated by NKX3.1 in Prostate Cancer.

The 8-hydroxy-2'-deoxyguanosine (8-OHdG) damages are base damages induced by reactive oxygen species. We aimed to investigate the role of Androgen Receptor and NKX3.1 in 8-OHdG formation and repair activation by quantitating the DNA damage using Aklides.NUK system. The data demonstrated that the loss of NKX3.1 resulted in increased oxidative DNA damage and its overexpression contributes to the removal of menadione-induced 8-OHdG damage even under oxidative stress conditions. Moreover, 8-oxoguanine DNA glycosylase-1 (OGG1) expression level positively correlates to NKX3.1 expression. Also in this study, first time a reliable cell-based quantitation method for 8-OHdG damages is reported and used for data collection.

Transcriptional regulation of the Nkx3.1 gene in prostate luminal stem cell specification and cancer initiation via its 3' genomic region.

NK3 homeobox 1 (Nkx3.1), a transcription factor expressed in the prostate epithelium, is crucial for maintaining prostate cell fate and suppressing tumor initiation. Nkx3.1 is ubiquitously expressed in luminal cells of hormonally intact prostate but, upon androgen deprivation, exclusively labels a type of luminal stem cells named castration-resistant Nkx3.1-expressing cells (CARNs). During prostate cancer initiation, Nkx3.1 expression is frequently lost in both humans and mouse models. Therefore, investigating how Nkx3.1 expression is regulated in vivo is important for understanding the mechanisms of prostate stem cell specification and cancer initiation. Here, using a transgenic mouse line with destabilized GFP, we identified an 11-kb genomic region 3' of the Nkx3.1 transcription start site to be responsible for alterations in Nkx3.1 expression patterns under various physiological conditions. We found that androgen cell-autonomously activates Nkx3.1 expression through androgen receptor (AR) binding to the 11-kb region in both normal luminal cells and CARNs and discovered new androgen response elements in the Nkx3.1 3' UTR. In contrast, we found that, in Pten-/- prostate tumors, loss of Nkx3.1 expression is mediated at the transcriptional level through the 11-kb region despite functional AR in the nucleus. Importantly, the GFP reporter specifically labeled CARNs in the regressed prostate only in the presence of cell-autonomous AR, supporting a facultative model for CARN specification.

Morphoregulatory pathways in prostate ductal development.

The mouse prostate is a male sex-accessory gland comprised of a branched ductal network arranged into three separate bilateral lobes: the anterior, dorsolateral, and ventral lobes. Prostate ductal development is the primary morphogenetic event in prostate development and requires a complex regulation of spatiotemporal factors. This review provides an overview of prostate development and the major genetic regulators and signaling pathways involved. To identify new areas for further study, we briefly highlight the likely important, but relatively understudied, role of the extracellular matrix (ECM). Finally, we point out the potential importance of the ECM in influencing the behavior and prognosis of prostate cancer. Developmental Dynamics 246:89-99, 2017. © 2016 Wiley Periodicals, Inc.

Regulated expression of the TPß isoform of the human T prostanoid receptor by the tumour suppressors FOXP1 and NKX3.1: Implications for the role of thromboxane in prostate cancer.

The prostanoid thromboxane (TX)A2 signals through the TPα and TPß isoforms of T Prostanoid receptor (TP) that are transcriptionally regulated by distinct promoters termed Prm1 and Prm3, respectively, within the TBXA2R gene. We recently demonstrated that expression of TPα and TPß is increased in PCa, differentially correlating with Gleason grade and with altered CpG methylation of the individual Prm1/Prm3 regions within the TBXA2R. The current study sought to localise the sites of CpG methylation within Prm1 and Prm3, and to identify the main transcription factors regulating TPß expression through Prm3 in the prostate adenocarcinoma PC-3 and LNCaP cell lines. Bisulfite sequencing revealed extensive differences in the pattern and status of CpG methylation of the individual Prm1 and Prm3 regions that regulate TPα and TPß expression, respectively, within the TBXA2R. More specifically, Prm1 is predominantly hypomethylated while Prm3 is hypermethylated across its entire sequence in PC-3 and LNCaP cells. Furthermore, the tumour suppressors FOXP1 and NKX3.1, strongly implicated in PCa development, were identified as key transcription factors regulating TPß expression through Prm3 in both PCa cell lines. Specific siRNA-disruption of FOXP1 and NKX3.1 each coincided with up-regulated TPß protein and mRNA expression, while genetic-reporter and chromatin immunoprecipitation (ChIP) analyses confirmed that both FOXP1 and NKX3.1 bind to cis­elements within Prm3 to transcriptionally repress TPß in the PCa lines. Collectively these data identify Prm3/TPß as a bona fide target of FOXP1 and NKX3.1 regulation, providing a mechanistic basis, at least in part, for the highly significant upregulation of TPß expression in PCa.

Skene's gland adenocarcinoma with intestinal differentiation: A case report and literature review.

Adenocarcinoma of Skene's gland (the female homolog to the male prostate) is extremely rare, with only a few cases reported. We present a case of Skene's gland adenocarcinoma with intestinal differentiation. The patient was a 69-year-old Japanese woman who was operated on for a recurrent tumor of the external ostium of the urethra. Histopathologically, the tumor showed glandular and cribriform patterns with a signet-ring cell component in a mucus lake. Immunohistochemically, the tumor cells were positive for prostate specific acid phosphatase (PSAP), and AMACR, and negative for Nkx3.1 or prostate specific antigen (PSA). Although in situ lesion could not be discovered, positive immunostainings for Nkx3.1, PSAP, and androgen receptor in the remaining paraurethral glands around the tumor indirectly but strongly suggest that the tumor had originated from Skene's gland. This tumor also showed intestinal differentiation as suggested histologically and by positive immunostainings for CDX2, MUC2, and CK20, along with negative immunostaining for CK7. It is often very difficult to identify the origin of a female urethral carcinoma. In such cases, immunohistochemical features can be an essential clue to the origin. We therefore present this instructive case with a literature review.

Sensitivity of HOXB13 as a Diagnostic Immunohistochemical Marker of Prostatic Origin in Prostate Cancer Metastases: Comparison to PSA, Prostein, Androgen Receptor, ERG, NKX3.1, PSAP, and PSMA.

AIMS: Determining the origin of metastases is an important task of pathologists to allow for the initiation of a tumor-specific therapy. Recently, homeobox protein Hox-B13 (HOXB13) has been suggested as a new marker for the detection of prostatic origin. The aim of this study was to evaluate the diagnostic sensitivity of HOXB13 in comparison to commonly used immunohistochemical markers for prostate cancer. MATERIALS AND METHODS: Histologically confirmed prostate cancer lymph node metastases from 64 cases were used to test the diagnostic value of immunohistochemical markers: prostate specific antigen (PSA), Prostatic acid phosphatase (PSAP), prostate specific membrane antigen (PSMA), homeobox gene NKX3.1, prostein, androgen receptor (AR), HOXB13, and ETS-related gene (ERG). All markers were evaluated semi-quantitatively using Remmele's immune reactive score. RESULTS: The detection rate of prostate origin of metastasis for single markers was 100% for NKX3.1, 98.1% for AR, 84.3% for PSMA, 80.8% for PSA, 66% for PSAP, 60.4% for HOXB13, 59.6% for prostein, and 50.0% for ERG. CONCLUSIONS: Our data suggest that HOXB13 on its own lacks sensitivity for the detection of prostatic origin. Therefore, this marker should be only used in conjunction with other markers, preferably the highly specific PSA. The combination of PSA with NKX3.1 shows a higher sensitivity and thus appears preferable in this setting.

An NKX3.1 binding site polymorphism in the l-plastin promoter leads to differential gene expression in human prostate cancer.

The L-plastin gene is involved in the invasion and metastasis of prostate cancer. However, the molecular mechanisms underlying L-plastin transcription are unclear. We hypothesize that the occurrence of polymorphic genetic variations in the L-plastin promoter might affect an individual's susceptibility to prostate cancer. In this study, we identified a single nucleotide polymorphism (SNP) at position -1,687 in the L-plastin promoter by genotype sequencing. The SNP -1,687 showed different transcriptional activity in the luciferase assay in vitro. The TRANSFAC software was applied to predict the multiple cis-elements, and luciferase assay was used to further identify the L-plastin regulatory region. We performed EMSAs, supershift assays and ChIP-qPCR demonstrated that the transcriptional suppressor NKX3.1 binds to the SNP site of the L-plastin promoter. SNP -1,687 (T/T) led to an increase in the affinity of NKX3.1 for L-plastin promoter, resulted in lower levels of L-plastin RNA and protein expression. Furthermore, we collected and sequenced samples from 640 individuals (372 prostate cancer patients and 268 healthy controls) from 2000 to 2013. The results showed that SNP -1,687 (T/T) occurred more frequently in the healthy individuals than that in the prostate cancer patients compared to SNP -1,687 (C/C). Similarly, SNP -1,687 (T/T) genotype occurred more frequently compared to SNP -1,687 (C/C) genotype in the patients with low and moderately differentiated tumors. In conclusion, SNP -1,687, located in the NKX3.1 binding site within the L-plastin promoter, might reduce the expression of L-plastin and potentially decrease the tumorigenesis and progression of prostate cancer. This SNP could be a potential prognostic factor for prostate cancer.