Characterization of PacMetUT1, a recently isolated human prostate cancer cell line

BACKGROUND: Existing prostate cancer cell lines have limitations. METHODS: Cells were characterized using Western blotting, immunohistochemistry, invasion into Matrigel, and by studying xenograft tumors. RESULTS: We describe a cell line (PacMetUT1) isolated from a lymph node of a 57-year-old male with prostate cancer. Compared to existing prostate cancer cell lines, the growth rate of PacMetUT1 xenograft tumors is slower with tumors occurring at injection sites and with metastases to lung and liver. Androgen receptor (AR) was detected in vivo by Western blotting and the cells responded to methyltrienolone (R1881). PacMetUT1 cells are more invasive in Matrigel than DU-145, PC-3, and LNCaP cells, and showed greater anchorage-independent growth in soft agar. The cells do not express prostate specific antigen (PSA) in vitro or in xenografts. However, the green fluorescent protein (GFP) gene was introduced and stably expressed in PacMetUT1 cells, allowing tumor imaging in vivo. Xenograft tumors show epithelial features and are positive for keratin, epithelial membrane antigen, EGF receptor, and E cadherin. In contrast, fibroblast markers vimentin, desmin, and Factor VIII, were negative. Karyotyping showed losses of 6p, 7q, 8p, 18q, and 22q, and gains of 8q and 9q; additional genetic material was observed at 2q and 12p. CONCLUSION: The PacMetUT1 cell line allows metastases to be assessed using a single animal model. Because of its slower growth, PacMetUT1 more closely mimics the human disease. Studies of tumor progression or metastasis can be conducted over a longer period of time.     

人食管鳞癌细胞系RJEC-2的建立及其生物学特性分析

目的:建立新的人食管鳞状细胞癌细胞系并分析其生物学特性,为食管癌分子机制和治疗干预的研究提供新的实验模型。方法:采用组织块培养法,从病人食管癌组织中分离纯化鳞状上皮癌细胞并建立细胞系。对细胞系的形态特点、细胞角蛋白表达、生长特征、细胞周期分布、细胞遗传特征和致瘤能力进行了研究分析。结果:建立了食管鳞状细胞癌细胞系RJEC-2,已在体外持续传代4个多月,传至46代,生长稳定;该细胞系具有鳞状上皮细胞的形态和特点:单层贴壁生长,免疫组化显示细胞角蛋白表达阳性,电镜下可见明显的胞质内张力纤维束和细胞间桥粒;群体倍增时间为46.5 h,细胞培养至90%汇合时,细胞周期分析显示G0/G1期平均占56.72%,S期平均占33.96%,G2/M期平均占9.32%;细胞染色体结构和数量异常,呈现肿瘤细胞特性;细胞呈克隆性生长,平板克隆平均形成率为13.93%,裸鼠移植瘤实验表明细胞具有致瘤能力,病理分析显示移植瘤与病人肿瘤病理形态特征相似,均为中、高分化鳞状细胞癌。结论:成功建立的人食管鳞癌细胞系RJEC-2,为食管癌发病机制和治疗方案的研究提供了新的研究实验模型。     

Human primary prostate tumor cell line,ALVA-31: A new model for studying the hormonal regulation of prostate tumor cell growth

A new human prostate tumor cell line (ALVA-31) has been established from a biopsy specimen of primary tumor obtained during prostatectomy. The cell line has been maintained for more than 48 months in stable growth. The in vitro doubling time was determined to be approximately 26 hr. The chromosome number ranged from 24–112, with a modal number of 59 tested over several time points throughout continuous culture. Karyotypic analysis of late-passaged cells demonstrated approximately 70 human chromosomes, 8–14 markers, and two × chromosomes without a Y chromosome. Prostatic origin was confirmed by the expression of both prostate specific antigen and prostatic acid phosphatase, using specific antisera and immunoradiolabelling techniques. Prostate tumor xenografts were grown in intact male, castrate male, and female athymic mice; however, the rate of tumor growth was clearly dependent upon serum testosterone levels. © 1993 Wiley-Liss, Inc.     

Evaluation of MENT on primary cell cultures from benign prostatic hyperplasia and prostate carcinoma

7-alpha-Methyl-19-Nortestosterone (MENT) is a synthetic androgen more potent than testosterone (T) and cannot be reduced at 5-alpha position. No important effects of MENT on prostate growth have been reported. However, little is known about the effect of MENT on benign prostatic hyperplasia (BPH) or prostate carcinoma (CaP). We evaluate the effect of MENT, T and dihydrotestosterone (DHT) on secretion, proliferation and gene expression of primary cell cultures from human BPH and CaP. Moreover, the effect of these androgens was examined in the presence of finasteride to determine the influence of the 5-alpha reductase (5-AR) activity on the androgenic potency. BPH and CaP primary cultures were treated with 0, 1, 10 and 100 n m of T, MENT or DHT during 24 and 48 h. Prostate-specific antigen (PSA) was measured by micro particles immunoassay and proliferation rate by spectrophotometric assay (MTT) and by the immunochemical detection of the proliferation marker Ki-67. Gene expression of FGF8b (androgen sensitive gene) was evaluated by semi-quantitative RT-PCR. Results showed that MENT treatments increased PSA secretion and proliferation rate with a potency ranged between T and DHT. Similar effects of MENT were observed in both BPH and CaP cultures. The studies with finasteride showed that in BPH and CaP cells, the conversion of T into DHT significantly contributes to its effect on the proliferation and PSA secretion, and corroborated the resistance of MENT to the 5-AR. The effect of MENT on the gene expression of FGF8b in CaP cells was similar to T and lower than DHT. It is concluded that MENT increases proliferative and secretory activities and gene expression on pathological prostate cells although in less extent than the active metabolite DHT. Furthermore, the fall of endogenous concentration of T during MENT treatment anticipates that this androgen will be of low impact for the prostate.     

Establishment and characterization of a primary androgen-responsive African-American prostate cancer cell line, E006AA

BACKGROUND: Establishment of human prostate cancer cell lines is essential to advance our understanding of complex processes associated with the initiation and progression of the disease. In the present study, we report the establishment of a primary African-American prostate cancer cell line (E006AA) as well as its associated stromal cells (S006AA). METHODS: E006AA cell line was established as a spontaneously immortalized cells from a patient with a clinically localized prostate cancer. Extensive characterization of the cells was accomplished using androgen-dependent growth and sensitivity assays, Western analyses, RT-PCR/real-time PCR, cytogenetic analyses, and tumorigenicity in nude mice. RESULTS: E006AA cell line shows androgen-dependent growth, expresses PSA and the androgen receptor (AR) with 26 CAG repeats in exon 1 of AR. Cytogenetic analyses revealed a hypertriploid karyotype with additional numerical gains in chromosomes 5, 6, 8, 10, 17, 20, 21 and a marker chromosome of unknown origin as well as structural abnormalities in chromosomes 4, 5, 8, 9, 11-14, 18, and 20. This cell line is not tumorigenic in nude mice. S006AA cell line, isolated from the same tumor specimen, also expresses AR and shows the morphological characteristics of smooth muscle cells of prostatic stroma. CONCLUSIONS: These cell lines are the first available primary epithelial and stromal cells derived from an African-American patient with organ-confined prostate cancer and in conjunction with other established cell lines, could provide an in vitro model system to investigate early transforming events in prostate cancer.     

Establishment and characterization of androgen-independent human prostate cancer LNCaP cell model

BACKGROUND: The acquisition of an androgen-independent phenotype is the most serious issue of prostate cancer treatment. Although several experimental cell models have been reported for studying androgen independence, they have limited applications related to hormone-refractory prostate cancer. To investigate the molecular mechanism of androgen-independent growth of prostate cancer, we established a useful LNCaP cell model that resembles the clinical scenario of hormone-refractory prostate cancer. METHODS: Androgen-sensitive LNCaP parental cells were continuously maintained in a regular cell-culture medium, that is, phenol red-positive RPMI 1640 medium supplemented with 5% fetal bovine serum and 1% glutamine. Upon passage, the androgen responsiveness of those cells decreased, to a level lower than that of parental cells. We examined the growth properties and androgen responsiveness of these different LNCaP cells in vitro and in vivo. Cytogenetic characteristics and expression of androgen receptors (ARs) and prostate-specific antigen (PSA) were determined. RESULTS: Upon continuous passage, the biological behavior of parental C-33 cells (passage number less than 33) was altered. C-81 cells (passage number higher than 81) clearly exhibited more aggressive growth and lower androgen responsiveness than C-33 and C-51 cells (passage number between 35 and 80) in vitro and in vivo. Nevertheless, all these cells expressed a similar level of functional AR protein as well as a similar genetic profile. Moreover, in a steroid-reduced culture condition, C-81 cells secreted a higher level of PSA than C-33 cells. CONCLUSIONS: Our LNCaP cell model closely recapitulates the progression of human prostate cancer from the androgen-responsive to the hormone-refractory state under the androgen nondeprived condition. This cell model may provide the opportunity to understand the molecular mechanisms associated with the acquisition of androgen independence during human prostate cancer progression.     

PEAZ-1: a new human prostate neoplastic epithelial cell line.

BACKGROUND: The generation of prostatic cell lines provides in vitro models for experimental studies of the pathogenesis of prostate carcinoma. Therefore, we established and characterized a new human prostate epithelial cell line, PEAZ-1 (prostate epithelial Arizona-1). METHODS: The PEAZ-1 cells were grown from a primary human prostate carcinoma specimen obtained from radical prostatectomy. The isolated cells were characterized by immunobiochemistry, immunohistochemistry, and tumorigenicity studies. RESULTS: PEAZ-1 cells are near diploid, tumorigenic, and androgen independent for cell growth. PEAZ-1 cells express N-cadherin, alpha- and beta-catenins, and p120 at cell-cell contacts, cytoplasmic laminin 5, vinculin, paxillin, and phosphotyrosine at focal adhesions, vimentin, and cytokeratins 8 and 18. They do not express plakoglobin, E-cadherin, and PSA, and do not form desmosomes and hemidesomomes. PEAZ-1 respond to ocadaic acid, a pro-apoptotic agent, by expression of p53. CONCLUSIONS: PEAZ-1 cells is a human prostate cancer cell line that has a number of mesenchymal characteristics.     

Patterns of metastasis by the human prostate cancer cell line PC-3 in athymic nude mice

Cells from the PC-3 human prostate cancer cell line were evaluated in athymic nude mice in order to determine the influence of size of the primary tumor and site inoculation on the incidence and pattern of metastasis. At autopsy, all organs, including the skeleton, were evaluated for metastasis. Subcutaneous injections resulted in metastases to the draining axillary lymph node and lungs (56% and 13%, respectively), and were correlated with size of the primary tumor. Tail vein injection resulted in a high incidence of lung metastasis, while injection into the peritoneal space, spleen, and seminal vesicles resulted in intraabdominal tumor growth, liver metastasis, and large tumors within the seminal vesicles, respectively. Skeletal metastases were not observed in any of the animals studied. We conclude that injection of PC-3 cells into various sites results in different patterns of metastasis, but may not constitute an entirely suitable animal model of human prostate cancer due to the lack of metastasis to the skeleton.     

Molecular characterization of human prostate carcinoma cell lines

BACKGROUND: This study presents a comprehensive survey and characterization of available prostate carcinoma cell lines, most of which have been widely used but are incompletely characterized. METHODS: A total of 21 cell lines were investigated, including three "classical" (DU 145, LNCaP, and PC-3) and 18 "non-classical" lines (1013L, 22Rv1, ALVA-55, ALVA-101, ARCaP, CWR-R1, DuCaP, DuPro-1, LAPC-4, MDA PCa 1, MDA PCa 2a, MDA PCa 2b, NCI-H660, PC-346C, PC-93, PSK-1, UM-SCP-1, and VCaP). Cytogenetics, DNA profiling, expression of basal, luminal, and neuroendocrine differentiation markers, and mutation analyses of the TP53 and androgen receptor (AR) genes were performed. RESULTS: Based on cytogenetics and DNA profiling analyses, out of the 18 "non-classical" lines, six were confirmed to be unique, eight (in four pairs) were confirmed to be related in origin, and four lines were identified as cross-contaminants. Of this latter group, PC-93 was found to be a derivative of HeLa, whereas DuPro-1, ALVA-55, and ALVA-101 were derivatives of PC-3. The 17 genuine prostate cell lines expressed keratin 8 (K8) and K18. Nine showed AR expression, of which five harbored mutations in the AR gene. Prostate-specific antigen and DD3 were exclusively detected in AR expressing cell lines. Seven lines expressed the basal cell marker K5, three of these lines showed co-expression of AR. CONCLUSIONS: This study defines a collection of 17 genuine prostate carcinoma cell lines. This collection, although small, constitutes a variety of different types and stages of prostate cancer, while it also partly reflects the heterogeneous nature of this malignancy.     

Establishment of a prostatic small-cell carcinoma cell line (SO-MI)

BACKGROUND: Prostatic small-cell carcinoma is an extremely rare, highly aggressive disease. We established a cell line from this tumor. MATERIALS AND METHODS: Tumor tissue obtained from a 24-year-old Japanese man was used to establish the cell line. Cultured cells and tumors transplanted into nude mice were characterized by histologic, immunohistologic, immunocytologic, and molecular biologic methods. RESULTS: An immortal culture cell line (SO-MI) was successfully established. SO-MI cells adhered weakly to plastic surfaces in vitro, showing a 52- to 72-hr doubling time. SO-MI cells were heterotopically and orthotopically transplantable in nude mice. The cells were immunoreactive for NSE, chromogranin A, and NCAM, but not for ACTH, calcitonin, serotonin, gastrin, insulin, glucagons, LCA, EMA, PAP, PSA, androgen receptor, and p53. SO-MI cells secreted NSE in vitro and in vivo. SO-MI cells at passage 30 contained 50-59 chromosomes with a modal number of 55. PCR suggested that the p53 gene was deleted in SO-MI cells. RT-PCR detected no mRNA encoding androgen receptor in these cells. CONCLUSIONS: SO-MI cells retain the neuroendocrine nature of the original tumor, and should be useful in studying possible etiologies and new treatments.     

Neoplastic transformation of a human prostate epithelial cell line by the v-Ki-ras oncogene

Investigations of mechanisms of human prostate carcinogenesis are limited by the unavailability of a suitable in vitro model system. We have demonstrated that an immortal, but nontumorigenic, human epithelial cell line (267B1) established from fetal prostate tissue can be malignantly transformed by a biological carcinogen, and can serve as a useful model for investigations of the progression steps of carcinogenesis. Activated Ki-ras was introduced into 267B1 cells by infection with the Kirsten murine sarcoma virus. Morphological alterations and anchorage-independent growth were observed; when cells were injected into nude mice, poorly differentiated adenocarcinomas developed. These findings represent the first evidence of malignant transformation of human prostate epithelial cells in culture, and support a role for Ki-ras activation in a multistep process for prostate neoplastic transformation. © 1993 Wiley-Liss, Inc.     

Murine androgen-independent neuroendocrine carcinoma promotes metastasis of human prostate cancer cell line LNCaP

BACKGROUND: Although neuroendocrine (NE) cells in prostate cancer have been speculated to accelerate the growth and progression of surrounding cancer cells, the evidence is as yet inconclusive. We investigated the effect of an NE allograft (NE-10) and its cell line, NE-CS, which were established from the prostate of the LPB-Tag 12T-10 transgenic mouse, on human prostate cancer cell line LNCaP. METHODS: The proliferation and pulmonary metastasis of LNCaP xenografts in athymic mice with and without NE-10 allografts were evaluated. Boyden chamber assay and microarray analysis were performed to investigate changes in invasion/migration and mRNA of LNCaP cells under the influence of the NE cells, respectively. RESULTS: NE-10 did not influence the proliferation of LNCaP. The pulmonary metastasis of LNCaP with NE-10 significantly increased compared to mice without it. The NE-CS cells accelerated the in vitro invasion/migration of adenocarcinoma cells. Increased expression of mRNA of gelsolin was observed in LNCaP cells incubated with the supernatant of NE-CS cells. CONCLUSIONS: The NE-10 allograft promotes pulmonary metastasis of subcutaneously inoculated LNCaP cells by facilitating cell invasion. Secretions from NE cells upregulate the expression of gelsolin, which is an actin-binding protein, resulting in acceleration of the migration of LNCaP cells.