Neuroendocrine Cell Differentiation Research Articles

Generation of ASCL1-mCherry knock-in reporter in human embryonic stem cell line, WAe001-A-2E, using CRISPR/Cas9-based gene targeting

Achaete-Scute Complex Homolog 1 (ASCL1) is a key regulator in the development and function of the nervous system, particularly in the process of neuronal and neuroendocrine cell differentiation. By employing the CRISPR/Cas9 system, we successfully established an ASCL1-mCherry knock-in human embryonic stem cell (hESC) line by inserting a P2A-mCherry fragment at the ASCL1 locus. The mCherry reporter effectively demonstrated the expression level of endogenous ASCL1 during the process of inducing pulmonary neuroendocrine cells (PNECs) from hESC. This reporter cell line holds significant value as a research tool for investigating the process of lung neuroendocrine cell differentiation, conducting drug screening, and exploring the underlying mechanisms of lung diseases associated with PNECs dysfunction.

Spontaneous intersibling polymorphism in the development of dopaminergic neuroendocrine cells in sea urchin larvae: impacts on the expansion of marine benthic species.

The plasticity of the nervous system plays a crucial role in shaping adaptive neural circuits and corresponding animal behaviors. Understanding the mechanisms underlying neural plasticity during development and its implications for animal adaptation constitutes an intriguing area of research. Sea urchin larvae offer a fascinating subject for investigation due to their remarkable evolutionary and ecological diversity, as well as their diverse developmental forms and behavioral patterns. We conducted immunochemical and histochemical analyses of serotonin-containing (5-HT-neurons) and dopamine-containing (DA-positive) neurons to study their developmental dynamics in two sea urchin species: Mesocentrotus nudus and Paracentrotus lividus. Our approach involved detailed visualization of 5-HT- and DA-positive neurons at gastrula-pluteus stages, coupled with behavioral assays to assess larval upward and downward swimming in the water column, with a focus on correlating cell numbers with larval swimming ability. The study reveals a heterochronic polymorphism in the appearance of post-oral DA-positive neuroendocrine cells and confirms the stable differentiation pattern of apical 5-HT neurons in larvae of both species. Notably, larvae of the same age exhibit a two- to four-fold difference in DA neurons. An increased number of DA neurons and application of dopamine positively correlate with larval downward swimming, whereas 5-HT-neurons and serotonin application induce upward swimming. The ratio of 5-HT/DA neurons determines the stage-dependent vertical distribution of larvae within the water column. Consequently, larvae from the same generation with a higher number of DA-positive neurons tend to remain at the bottom compared to those with fewer DA-positive neurons. The proportion of 5-HT and DA neurons within larvae of the same age underlies the different potentials of individuals for upward and downward swimming. A proposed model illustrates how coordination in humoral regulation, based on heterochrony in DA-positive neuroendocrine cell differentiation, influences larval behavior, mitigates competition between siblings, and ensures optimal population expansion. The study explores the evolutionary and ecological implications of these neuroendocrine adaptations in marine species.

Abstract 5599: Role of HOX gene expression in differentiation of colon cancer stem cells

Abstract HOX genes encode a highly conserved set of transcription factors critical for stem cell (SC) function and embryonic development. Several HOX genes have also been implicated in a myriad of cancers, including colorectal cancer (CRC). However, how HOX genes play a role in SC regulation and contributes to cancer development is unclear. HOX expression is regulated by WNT, FGF, and retinoic acid (RA) signaling - pathways which also become dysregulated in cancer. Goal: To determine how regulation of HOX genes specifies differentiation of colonic SCs into specialized cell lineages and how dysregulation of HOX genes contributes to CRC development. Hypotheses: i) Each of the different cell types within human colonic epithelium has a distinct HOX gene expression signature; ii) All-trans retinoic acid (ATRA) induced differentiation of CRC cells changes HOX expression that correlates with specific HOX expression signatures in normal colon and CRC cells. Results: Immunostaining shows that specific HOX proteins (e.g., HOXA9 & HOXC9) are selectively expressed in ALDH-positive SCs and expression of these HOX genes is increased in CRC tissues compared to normal epithelium. Bioinformatics analysis predicted that these HOX genes (plus HOXA4, HOXA10, HOXC8) are regulated by RA signaling. Accordingly, we studied how ATRA influences HOX expression in CRC cells (HT29 & SW480). We found ATRA treatment: 1) Decreases proliferation and increases neuroendocrine cell (NEC) differentiation; and 2). Leads to specific changes in HOX gene expression patterns. Using NanoString profiling, we also discovered that LGR5+ SCs, ALDH+ SCs and GLP2R+ NECs have unique HOX signatures. By defining how these HOX gene expression changes in response to ATRA treatment in comparison to the HOX gene signatures of different colonic cell lineages will help us understand the role that HOX genes play in SC differentiation and how dysregulation of HOX gene expression contributes to CRC development. Citation Format: Brian T. Osmond, Caroline O. Facey, Lynn O. Opdenaker, Chi Zhang, Bruce M. Boman. Role of HOX gene expression in differentiation of colon cancer stem cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5599.

Abstract 5430: Discovery of CYP26A1 as a link between WNT and retinoic acid signaling pathways that modulates retinoid-induced differentiation of APC-mutant colorectal cancer cells

Abstract APC mutation is the main driver mechanism of CRC development and leads to constitutively activated WNT signaling, overpopulation of ALDH+ stem cells (SCs), and incomplete differentiation. We previously reported that retinoic acid (RA) receptors are selectively expressed in ALDH+ SCs, which provides a way to target cancer SCs with retinoids to induce differentiation. We hypothesize that a functional link exists between WNT and RA pathways, and APC mutation generates a WNT:RA imbalance that decreases retinoid-induced differentiation and increases ALDH+ SCs. Accordingly, to restore parity in WNT:RA signaling, we induced wt-APC expression in APC-mutant CRC cells and assessed the ability of all-trans-retinoic acid (ATRA) to induce differentiation. Notably, ATRA substantially increased expression of the WNT-target gene, CYP26A1, and inducing wt-APC reduced this expression by 50%. Thus, RA and WNT pathways crosstalk to modulate CYP26A1 metabolism of retinoids and retinoid agents. We found that inducing wt-APC augments ATRA-induced cell differentiation by: i) decreasing cell proliferation; ii) suppressing ALDH1A1 expression; iii) decreasing ALDH+ SCs; and iv) increasing neuroendocrine cell differentiation. NanoString profiling and bioinformatics also identified a novel CYP26A1-based network that links WNT and RA signaling. Moreover, in an effort to translate manipulation of the WNT/RA imbalance as a therapeutic approach in vitro, we treated HT29, SW480, and HCT116 CRC cells with inhibitors against CYP26A1 enzyme activity and WNT signaling. We found that CYP26A1 inhibitors sensitized CRC cells to the anti-proliferative effect of drugs that downregulate WNT signaling. Furthermore, decreased CYP26A1 expression in tumor vs normal tissue was determined to be a predictor of patient survival in patients who have wt-APC. Thus, by inducing wt-APC expression and exogenously activating RA signaling, we found a way to increase differentiation of SCs along the NEC (and other) lineage(s). Translating this strategy in vivo might lead to new, more effective treatments involving retinoids for CRC patients. Citation Format: Caroline O. Facey, Victoria O. Hunsu, Chi Zhang, Brian T. Osmond, Lynn M. Opdenaker, Bruce M. Boman. Discovery of CYP26A1 as a link between WNT and retinoic acid signaling pathways that modulates retinoid-induced differentiation of APC-mutant colorectal cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5430.

CYP26A1 Links WNT and Retinoic Acid Signaling: A Target to Differentiate ALDH+ Stem Cells in APC-Mutant CRC.

APC mutation is the main driving mechanism of CRC development and leads to constitutively activated WNT signaling, overpopulation of ALDH+ stem cells (SCs), and incomplete differentiation. We previously reported that retinoic acid (RA) receptors are selectively expressed in ALDH+ SCs, which provides a way to target cancer SCs with retinoids to induce differentiation. Hypotheses: A functional link exists between the WNT and RA pathways, and APC mutation generates a WNT:RA imbalance that decreases retinoid-induced differentiation and increases ALDH+ SCs. Accordingly, to restore parity in WNT:RA signaling, we induce wt-APC expression in APC-mutant CRC cells, and we assess the ability of all-trans retinoic acid (ATRA) to induce differentiation. We found that ATRA increased expression of the WNT target gene, CYP26A1, and inducing wt-APC reduced this expression by 50%. Thus, the RA and WNT pathways crosstalk to modulate CYP26A1, which metabolizes retinoids. Moreover, inducing wt-APC augments ATRA-induced cell differentiation by: (i) decreasing cell proliferation; (ii) suppressing ALDH1A1 expression; (iii) decreasing ALDH+ SCs; and (iv) increasing neuroendocrine cell differentiation. A novel CYP26A1-based network that links WNT and RA signaling was also identified by NanoString profiling/bioinformatics analysis. Furthermore, CYP26A1 inhibitors sensitized CRC cells to the anti-proliferative effect of drugs that downregulate WNT signaling. Notably, in wt-APC-CRCs, decreased CYP26A1 improved patient survival. These findings have strong potential for clinical translation.

Recurrent gastric amphicrine tumor with neuroendocrine and pancreatic acinar cell differentiation and somatic MEN1 inactivation arisen during immunotherapy.

Amphicrine neoplasms (ANs) are poorly understood epithelial malignancies composed of cells with co-existing exocrine-neuroendocrine features. Here, we report a recurrent mucin-producing gastric amphicrine tumor co-expressing neuroendocrine (chromogranin-A, synaptophysin, and CD56) and pancreatic acinar cell (BCL10 and trypsin) markers, arisen in a 64-year-old woman during adjuvant immunotherapy for melanoma. Ki-67 was < 2%. The gastric background context was atrophic gastritis. Next-generation sequencing showed MEN1 mutation (p.P71fs*42) coupled with loss of heterozygosity. The key lessons were as follows: (1) gastric ANs can show the co-existence of exocrine mucin-producing elements with neuroendocrine and pancreatic acinar differentiation; (2) they may represent a new entity arising in the context of atrophic gastritis and during immunotherapy; (3) they should be considered in the diagnostic workup of gastric neuroendocrine tumors; and (4) their molecular profile can show striking similarities with well-differentiated neuroendocrine tumors. These findings may be of help to improve the knowledge and the biological taxonomy of ANs.

Manipulating the composition of the culture medium promotes neuroendocrine cell differentiation in mouse and human gastric organoids

Manipulating the composition of the culture medium promotes neuroendocrine cell differentiation in mouse and human gastric organoids

Abstract 5434: Insulinoma-associated protein 1 (INSM1) expression in small cell neuroendocrine carcinoma of the urinary tract

Abstract Introduction Clinical guidelines including the American Urological Association (AUA) state that neoadjuvant chemotherapy should be administered prior to surgery in cases of muscle invasive urinary bladder small cell neuroendocrine carcinoma. Although the recently described marker insulinoma-associated protein 1 (INSM1) has been reported to be sensitive and specific for neuroendocrine differentiation in neuroendocrine tumors at several sites, its efficacy as a marker in urinary bladder small cell carcinoma has not been well established. This study aims to examine the immunohistochemical expression pattern of INSM1 on whole tissue sections of urinary bladder small cell neuroendocrine carcinoma and compare INSM1 expression to those of established markers of neuroendocrine differentiation. Methods Archived tissue from 32 cases of urinary bladder small cell neuroendocrine carcinoma were obtained from Rhode Island Hospital and The Miriam Hospital. Immunohistochemical stains for CD56, INSM1, synaptophysin, and chromogranin were performed. Staining was scored for intensity (0: no staining; 1: weak; 2: moderate; 3: strong) and proportion of cells stained (0: 0%; 1: &amp;gt;0 to 25%; 2: &amp;gt;25 to 50%; 3: &amp;gt;50 to 75%; 4: &amp;gt;75 to 100%). Results INSM1 was positive (intensity 1-3 or proportion 1-4) in 87% (28/32) of small cell neuroendocrine carcinoma cases (20 with intensity 2-3, 17 with proportion 3-4). CD56, synaptophysin, and chromogranin were positive in 75% (24/32), 60% (19/32), and 44% (14/32) of cases, respectively. While the rate of positivity was high, INSM1 was negative (n=4) or only showed weak intensity staining (n=7) in 34% (11/32) of cases. Conclusion INSM1 is a sensitive marker of small cell neuroendocrine differentiation of the urinary bladder. However, the findings in this study suggest that optimal utilization of INSM1 would be in a limited panel of stains rather than as a stand-alone screening marker given that it is negative or only shows weak intensity staining in a significant proportion of cases. StainNegative [n (%)]Positive [n (%)]INSM14 (13)28 (87)CD568 (25)24 (75)Synaptophysin13 (40)19 (60)Chromogranin18 (56)14 (44) Citation Format: Isaac E. Kim, Ali Amin, Li Juan Wang, Liang Cheng, Carmen M. Perrino. Insulinoma-associated protein 1 (INSM1) expression in small cell neuroendocrine carcinoma of the urinary tract [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5434.

THE many faces of urothelial carcinoma; divergent villoglandular and small cell neuroendocrine differentiation in a 47-year-old man

THE many faces of urothelial carcinoma; divergent villoglandular and small cell neuroendocrine differentiation in a 47-year-old man

Insulinoma-associated Protein 1 (INSM1) Expression in Small Cell Neuroendocrine Carcinoma of the Urinary Tract.

Clinical guidelines state that neoadjuvant chemotherapy should be administered before surgery in muscle invasive urinary bladder small cell neuroendocrine carcinoma. Recently described marker insulinoma-associated protein 1 (INSM1) has been reported to be sensitive and specific for neuroendocrine differentiation, however, its efficacy in urinary tract small cell carcinoma is not well established. This study examines immunohistochemical expression of INSM1 on whole tissue sections of urinary tract small cell neuroendocrine carcinoma and compares INSM1 expression with established neuroendocrine markers. Immunohistochemical stains for CD56, INSM1, synaptophysin, and chromogranin were performed on 32 cases of small cell neuroendocrine carcinoma of the bladder. Staining was scored for intensity (0: no staining; 1: weak; 2: moderate; 3: strong) and proportion of cells stained (0: 0%; 1: >0% to ≤25%; 2: >25% to ≤50%; 3: >50% to ≤75%; 4: >75% to 100%). INSM1 was positive (intensity 1 to 3 or proportion 1 to 4) in 87% (28/32) of cases (20 with intensity 2 to 3, 17 with proportion 3 to 4). CD56, synaptophysin, and chromogranin were positive in 75% (24/32), 60% (19/32), and 44% (14/32) of cases, respectively. INSM1 was negative (n=4) or only showed weak intensity staining (n=7) in 34% (11/32) of cases. INSM1 is a sensitive marker of small cell neuroendocrine differentiation of the urinary tract. However, this study suggests that optimal utilization of INSM1 would be inclusion in a limited panel of stains rather than as a stand-alone screening marker given that it is negative or only shows weak intensity staining in a significant proportion of cases.

Dual-Specificity Phosphatases in Neuroblastoma Cell Growth and Differentiation.

Dual-specificity phosphatases (DUSPs) are important regulators of neuronal cell growth and differentiation by targeting proteins essential to neuronal survival in signaling pathways, among which the MAP kinases (MAPKs) stand out. DUSPs include the MAPK phosphatases (MKPs), a family of enzymes that directly dephosphorylate MAPKs, as well as the small-size atypical DUSPs, a group of low molecular-weight enzymes which display more heterogeneous substrate specificity. Neuroblastoma (NB) is a malignancy intimately associated with the course of neuronal and neuroendocrine cell differentiation, and constitutes the source of more common extracranial solid pediatric tumors. Here, we review the current knowledge on the involvement of MKPs and small-size atypical DUSPs in NB cell growth and differentiation, and discuss the potential of DUSPs as predictive biomarkers and therapeutic targets in human NB.

Pancreatic mixed acinar cell carcinoma: genomic analysis and characterization of a patient-derived organoid culture

Background: Mixed acinar cell carcinomas occur in <2% of all pancreatic cancers and express markers for both neuroendocrine and acinar cell differentiation. The clinical prognosis and molecular characterization of mixed acinar cell carcinomas are not well understood; therefore, we initiated a study to link genomic and transcriptomic analysis of mixed acinar cell carcinomas with pathogenesis. This case study represents a mixed acinar cell carcinoma in the pancreatic head that was surgically resected by a pancreaticoduedenectomy. The tumor was clinically staged pT3N1MX with a KI-67 labeling index of 5%. Methods: The macrodissected tumor specimen was subdivided for genomic analysis and explantation in organoid culture. After digestion with collagenase, the tissue was washed and plated in 50% Matrigel in Human Complete Organoid media. The cultures were subsequently established and characterized by immunolabeling for neuroendocrine and acinar lineage markers and by quantitative RT-PCR. After 3 passages of the organoid cultures, the cultures were then placed in 2D culture on 2% Matrigel coated dishes. DNA was isolated and whole exome sequencing was performed using the Nextera Rapid Capture Exome Kit by Illumina on an Illumina HiSeq 2000/2500. The following criteria were used to define genetic variants: bidirectional, non-synonymous, clean mapping in IGV, ≥ 15X coverage, and an alternate allele frequency of 0.3 ≤ x ≤ 0.7. Results: The primary tumor showed a mixed population of cell types on H&E. Tumor cells were positive for intracytoplasmic trypsin staining indicative of acinar differentiation. Tumor cells were also positive for the neuroendocrine markers synaptophysin and chromogranin A suggesting a population of cells that show both acinar and endocrine histology. The tumor was negative for gastrin, insulin, somatostatin, and glucagon. Cell growth in the initial organoid culture resulted in a mixed culture with 3D acinar structures and a second population of cells with multiple protrusions that invaded into the Matrigel. Whole exome sequencing of the dissected primary tumor showed mutations in several genes including MEN1, histone modification genes, and DNA repair pathway genes. Conclusion: Our approach is to combine patient-derived PanNET organoid cultures with cancer biology and molecular genetic analyses to understand their clinical significance.

AMPK Activation of PGC-1α/NRF-1-Dependent SELENOT Gene Transcription Promotes PACAP-Induced Neuroendocrine Cell Differentiation Through Tolerance to Oxidative Stress.

Several cues including pituitary adenylate cyclase-activating polypeptide (PACAP), which acts through cAMP stimulation, specify the conversion of sympathoadrenal (SA) precursors toward different cell phenotypes by promoting their survival and differentiation. Selenoprotein T (SELENOT) is a PACAP-stimulated ER oxidoreductase that exerts an essential antioxidant activity and whose up-regulation is associated with SA cell differentiation. In the present study, we investigated the transcriptional cascade elicited by PACAP/cAMP to trigger SELENOT gene transcription during the conversion of PC12 cells from SA progenitor-like cells toward a neuroendocrine phenotype. Unexpectedly, we found that PACAP/cAMP recruits the canonical pathway that regulates mitochondrial function in order to elicit SELENOT gene transcription and the consequent antioxidant response during PC12 cell differentiation. This cascade involves LKB1-mediated AMPK activation in order to stimulate SELENOT gene transcription through the PGC1-α/NRF-1 complex, thus allowing SELENOT to promote PACAP-stimulated neuroendocrine cell survival and differentiation. Our data reveal that a PACAP and cAMP-activated AMPK-PGC-1α/NRF-1 cascade is critical for the coupling of oxidative stress tolerance, via SELENOT gene expression, and mitochondrial biogenesis in order to achieve PC12 cell differentiation. The data further highlight the essential role of SELENOT in cell metabolism during differentiation.

Frequency of NOTCH pathway mutation in primary tumor of SCLC compared to metastatic biopsies and association with better survival.

e20574Background: Small cell lung cancer (SCLC) is a neuroendocrine tumor characterized by early distant metastasis. NOTCH signaling controls neuroendocrine cell differentiation. In mouse models, N...

Transcriptional deregulation underlying the pathogenesis of small cell lung cancer.

The discovery of recurrent alterations in genes encoding transcription regulators and chromatin modifiers is one of the most important recent developments in the study of the small cell lung cancer (SCLC) genome. With advances in models and analytical methods, the field of SCLC biology has seen remarkable progress in understanding the deregulated transcription networks linked to the tumor development and malignant progression. This review will discuss recent discoveries on the roles of RB and P53 family of tumor suppressors and MYC family of oncogenes in tumor initiation and development. It will also describe the roles of lineage-specific factors in neuroendocrine (NE) cell differentiation and homeostasis and the roles of epigenetic alterations driven by changes in NFIB and chromatin modifiers in malignant progression and chemoresistance. These recent findings have led to a model of transcriptional network in which multiple pathways converge on regulatory regions of crucial genes linked to tumor development. Validation of this model and characterization of target genes will provide critical insights into the biology of SCLC and novel strategies for tumor intervention.

Aggressive large-cell neuroendocrine carcinoma of the sigmoid colon in a patient with ulcerative colitis

Colonic inflammation seen in inflammatory bowel disease (IBD) predisposes to the development of colorectal adenocarcinoma. In contrast, colorectal neuroendocrine carcinomas (NECs) have rarely been reported in the setting of IBD,...

Large Cell Neuroendocrine Carcinoma of Prostate: A Rare Interesting Case and Literature Review

: Prostate cancer (PC) is one of the most common forms of malignancies and the second cause of cancer death all around the world and the eight causes in Iran. The main portion of PC is adenocarcinoma, in some cases neuroendocrine differentiation occurs. Neuroendocrine prostate cancers (NePCs) incidence varies from 0.5% to 2% of all prostate cancers. Large cell neuroendocrine differentiation is very rare. In this study we presented a 71-year-old man with large cell neuroendocrine carcinoma of the prostate.

Ectopic Gastric and Intestinal Phenotypes, Neuroendocrine Cell Differentiation, and SOX2 Expression Correlated With Early Tumor Progression in Colorectal Laterally Spreading Tumors

IntroductionThe significance of the ectopic gastric phenotype remains unclear in patients with colorectal laterally spreading tumors (LSTs). We investigated clinicopathologic differences among LST subtypes, aiming to identify factors indicative of malignant transformation and invasion that are linked to ectopic gastric phenotype and tumor progression. Materials and MethodsWe analyzed the morphologic characteristics of 105 colorectal LSTs resected by endoscopic submucosal dissection. LSTs were classified into 2 subtypes: granular (G-LST) and nongranular (NG-LST). Resected LSTs were analyzed histologically and were immunohistochemically stained for MUC5AC, MUC6, chromogranin A, CD10, and SOX2. ResultsThe 105 LSTs included 60 G-LSTs and 45 NG-LSTs. By histology, G-LSTs comprised 5 adenomas with low-grade dysplasia (LAs), 45 adenomas with high-grade dysplasia (HAs), and 10 adenocarcinomas invading the submucosa (SMs). NG-LSTs comprised 8 LAs, 25 HAs, and 12 SMs. MUC5AC positivity was significantly higher in G-LSTs compared to NG-LSTs (P = .002), and MUC5AC positivity in HA lesions was significantly higher than in LA lesions (P = .01). MUC6 and SOX2 positivity in SM G-LSTs, and chromogranin A positivity in SM NG-LSTs were significantly higher than in HAs (P = .01, .01, and .03, respectively). CD10 positivity in SM NG-LSTs was significantly higher than in HAs and LAs (P = .02 and .01, respectively). ConclusionEctopic gastric and intestinal phenotypes, neuroendocrine cell differentiation, and SOX2 expression differ according to tumor grade in colorectal LSTs, and these markers are correlated with early tumor progression in each LST subtype.

Up-Regulated Expression of LAMP2 and Autophagy Activity during Neuroendocrine Differentiation of Prostate Cancer LNCaP Cells.

Neuroendocrine (NE) prostate cancer (PCa) is a highly aggressive subtype of prostate cancer associated with resistance to androgen ablation therapy. In this study, we used LNCaP prostate cancer cells cultured in a serum-free medium for 6 days as a NE model of prostate cancer. Serum deprivation increased the expression of NE markers such as neuron-specific enolase (NSE) and βIII tubulin (βIII tub) and decreased the expression of the androgen receptor protein in LNCaP cells. Using cDNA microarrays, we compared gene expression profiles of NE cells and non-differentiated LNCaP cells. We identified up-regulation of 155 genes, among them LAMP2, a lysosomal membrane protein involved in lysosomal stability and autophagy. We then confirmed up-regulation of LAMP2 in NE cells by qRT-PCR, Western blot and confocal microscopy assays, showing that mRNA up-regulation correlated with increased levels of LAMP2 protein. Subsequently, we determined autophagy activity in NE cells by assessing the protein levels of SQSTM/p62 and LC3 by Western blot and LC3 and Atg5 mRNAs content by qRT-PCR. The decreased levels of SQSTM/p62 was accompanied by an enhanced expression of LC3 and ATG5, suggesting activation of autophagy in NE cells. Blockage of autophagy with 1μM AKT inhibitor IV, or by silencing Beclin 1 and Atg5, prevented NE cell differentiation, as revealed by decreased levels of the NE markers. In addition, AKT inhibitor IV as well as Beclin1 and Atg5 kwockdown attenuated LAMP2 expression in NE cells. On the other hand, LAMP2 knockdown by siRNA led to a marked blockage of autophagy, prevention of NE differentiation and decrease of cell survival. Taken together, these results suggest that LAMP2 overexpression assists NE differentiation of LNCaP cells induced by serum deprivation and facilitates autophagy activity in order to attain the NE phenotype and cell survival. LAMP2 could thus be a potential biomarker and potential target for NE prostate cancer.

Sa1792 Ectopic Gastric Phenotype, Neuroendocrine Cell Differentiation, and CD10 Expression Correlated With Early Tumor Progression in Colorectal Laterally Spreading Tumors

Sa1792 Ectopic Gastric Phenotype, Neuroendocrine Cell Differentiation, and CD10 Expression Correlated With Early Tumor Progression in Colorectal Laterally Spreading Tumors