Effect of isocaloric low-fat diet on prostate cancer xenograft progression to androgen independence

An isocaloric low-fat diet has been shown to slow androgen-sensitive Los Angeles Prostate Cancer-4 (LAPC-4) tumor growth in a mouse xenograft model. LAPC-4 cells were injected into male severe combined immunodeficient mice. After palpable tumors developed, the mice were divided into three groups, high-fat intact, high-fat castration, and low-fat castration. Tumor latency (18 versus 9 weeks; P < 0.001) and mouse survival (20.8 +/- 1.3 versus 13 +/- 0.7 weeks; P < 0.01) were significantly longer in the low-fat castration versus high-fat castration group. Reduced dietary fat intake delayed conversion from androgen-sensitive to -insensitive prostate cancer and significantly prolonged survival of severe combined immunodeficient mice bearing LAPC-4 xenografts.     

Growth inhibition of human prostate cancer cells in human adult bone implanted into nonobese diabetic/severe combined immunodeficient mice by a ligand-specific antibody to human insulin-like growth factors

Advanced prostate cancer frequently involves the bone that has the largest content of insulin-like growth factors (IGFs). However, the role of bone-derived IGFs in bone metastasis of prostate cancer has not been studied extensively because of the lack of a reliable animal model. Therefore, we investigated whether a novel antibody directed against human IGF-I and IGF-II (KM1468) could inhibit the development of new bone tumors and the progression of established bone tumors in nonobese diabetic/severe combined immunodeficient mice implanted with human adult bone. We first confirmed that KM1468 bound specifically to human IGF-I, human IGF-II, and mouse IGF-II but not to insulin. It also blocked autophosphorylation of the type I IGF receptor induced by the binding of IGFs in human-type I IGF receptor-overexpressing BALB/c 3T3 cells, and it inhibited the IGF-stimulated growth of MDA PCa 2b cells in vitro. Then mice were injected intraperitoneally with KM1468 once weekly for 4 weeks either immediately or 4 weeks after inoculation of MDA PCa 2b cells. KM1468 markedly and dose-dependently suppressed the development of new bone tumors and the progression of established tumor foci, as determined by histomorphometry, and it also decreased serum prostate-specific antigen levels, compared with the control. This is the first report of an IGF ligand-specific inhibitory antibody that suppresses the growth of human prostate cancer cells in human adult bone. These results indicate that the IGF signaling axis is a potential target for prevention and treatment of bone metastases arising from prostate cancer.     

Osteoprotegerin/osteoclastogenesis inhibitory factor decreases human prostate cancer burden in human adult bone implanted into nonobese diabetic/severe combined immunodeficient mice

Human prostate cancer frequently metastasizes to bone, where it gives rise to osteoblastic bone metastases with an underlying osteoclastic component and subsequent bone pain. However, the importance of osteoclastogenesis in the development of prostate cancer bone lesions in humans is unclear. Osteoprotegerin/osteoclastogenesis inhibitory factor (OCIF) is a member of the tumor necrosis factor receptor family and a novel secreted protein, and it is a negative regulator of osteoclast differentiation, activation, and survival both in vitro and in vivo. In the present study we used a model in which human LNCaP prostate cancer cells that give rise to osteoblastic bone tumors were injected directly into the intramedullary space of human adult bone implanted into nonobese diabetic/severe combined immunodeficient mice to investigate whether the new bone-resorption inhibitor osteoprotegerin/OCIF would inhibit the development of new bone tumors and the progression of established osteoblastic bone tumors. The mice were given consecutive daily s.c. injections of recombinant human OCIF (rhOCIF; 100 micro g/mouse/day) for 2 weeks starting either immediately or 2 weeks after injection of the LNCaP cells. In both protocols, rhOCIF markedly inhibited both the development of bone tumors and the progression of established bone tumor foci quantified by histological examination. Histomorphometrical analysis revealed that rhOCIF markedly reduced the number of osteoclasts and the size of the tumors at the bone sites, but that it had no effect on the local growth of s.c. LNCaP tumors or on LNCaP cell proliferation in culture. These findings demonstrate that osteoclasts play an important role in bone tumor by prostate cancer, and that rhOCIF decreases the LNCaP prostate cancer burden selectively in bone, suppresses the progression of established tumor lesions, and prevents the development of new lesions. These results suggest that inhibition of osteoclastic bone resorption may be an effective therapy for the treatment of prostate cancer that has colonized bone.     

Antibody targeting of doxorubicin-loaded liposomes suppresses the growth and metastatic spread of established human lung tumor xenografts in severe combined immunodeficient mice

Beta1 integrins, expressed on the cell surface of human non-small cell lung carcinomas, are used here as a target for the selective delivery of anti-cancer drug-loaded liposomes. Fab' fragments of a monoclonal antibody specific for human beta1 integrins were conjugated to sterically stabilized liposomes. Confocal microscopy of beta1 integrin-positive lung tumor cells incubated with fluorescently labeled anti-beta1 Fab immunoliposomes revealed a tumor-specific binding and efficient internalization of the liposomes into the tumor cells. The ability of these liposomes to deliver cytotoxic drugs to the tumor and kill these cells was demonstrated in vitro by incubating tumor cells with doxorubicin-loaded anti-beta1 Fab' immunoliposomes. The drug-loaded immunoliposomes were >30-fold more cytotoxic to the tumor cells than drug-loaded liposomes without antibody, nonspecific Fab' control immunoliposomes with drug or immunoliposomes without drug. The therapeutic efficacy of doxorubicin-loaded immunoliposomes was also evaluated in a metastatic human lung tumor xenograft/severe combined immunodeficient (SCID) mouse model. SCID mice that received i.v. injections of human lung tumor cells developed primary tumor nodules in the lung that subsequently metastasized to the liver and adrenal gland. Treatment of SCID mice bearing established lung tumor xenografts with doxorubicin-loaded anti-beta1 Fab immunoliposomes resulted in a significant suppression of tumor growth (monitored periodically by quantifying serum levels of a tumor marker), whereas tumors grew progressively in mice treated with control formulations. In addition to suppressing the growth of the primary lung tumor nodules, the immunoliposomes prevented the metastatic spread of the tumor to the liver and adrenal glands and increased the median survival time of the tumor-bearing mice. We conclude that Fab' immunoliposomes directed to tumor-associated integrins represent a potentially viable approach clinically for the selective delivery of drugs to solid tumors and may be useful in preventing the metastatic spread of lung cancer.     

Oncolytic capacity of attenuated replicative semliki forest virus in human melanoma xenografts in severe combined immunodeficient mice

Oncolytic viruses have gained attention as a novel form of cancer treatment. Many viral vectors in use today have been rendered safe by deletion of genes encoding viral structural proteins, thus making them unable to spread beyond the first infected cells. Hence, such replication-deficient constructs may lack efficacy. Here, we analyzed the oncolytic potential of the replication-competent vector VA7-EGFP, based on the avirulent Semliki Forest virus (SFV) strain A7(74), to kill cancer cells in culture as well as to target s.c. human melanoma xenografts in severe combined immunodeficient (SCID) mice. VA7-EGFP was able to infect most cancer cell lines studied, leading to complete lysis of the cells within 72 hours after infection. In SCID mice grafted with A2058 human melanoma, marked regression of the xenografts was observed following a single injection of 10(6) plaque-forming units of virus given either i.p., i.v., or intratumorally. Histologic analysis revealed the presence of virus not only in all treated tumors but also in the brains of the treated mice, causing progressing neuropathology beginning at day 16 after infection. Following initial oncolysis, clusters of viable tumor cells were observed embedded in connective tissue, and at later stages, encapsulated tumor nodules had formed. Infection of melanoma cells from explant cultures of these nodules revealed that a portion of the cells were resistant to virus. To be eligible for use in virotherapy, the ability of avirulent SFV to spread within tumor tissue may have to be improved and the biological safety of the virus may have to be addressed thoroughly in higher animals.     

Combined inhibitory effects of curcumin and phenethyl isothiocyanate on the growth of human PC-3 prostate xenografts in immunodeficient mice

Earlier studies using prostate cancer cells in culture showed that phenethyl isothiocyanate (PEITC) and curcumin have significant chemopreventive and possibly chemotherapeutic effects. However, their in vivo effects are still lacking. Hence, this study was undertaken to determine the possible in vivo efficacy of prostate cancer-prevention as well as cancer-therapeutic treatment by PEITC and curcumin alone or in combination. We evaluated the effects on tumor growth in vivo, using NCr immunodeficient (nu/nu) mice bearing s.c. xenografts of PC-3 human prostate cancer cells. Molecular biomarkers representing proliferation and apoptosis were determined. Continued i.p. injection of curcumin or PEITC (6 and 5 mumol; thrice a week for 28 days), beginning a day before tumor implantation significantly retarded the growth of PC-3 xenografts. Combination of i.p. administration of PEITC (2.5 mumol) and curcumin (3 mumol) showed stronger growth-inhibitory effects. Next, we evaluated the cancer-therapeutic potential of curcumin and PEITC in mice with well-established tumors, and the results showed that PEITC or curcumin alone had little effect, whereas combination of curcumin and PEITC significantly reduced the growth of PC-3 xenografts. Immunohistochemistry staining and Western blot analysis revealed that the inhibition of Akt and nuclear factor-kappaB signaling pathways could contribute to the inhibition of cell proliferation and induction of apoptosis. Taken together, our results show that PEITC and curcumin alone or in combination possess significant cancer-preventive activities in the PC-3 prostate tumor xenografts. Furthermore, we found that combination of PEITC and curcumin could be effective in the cancer-therapeutic treatment of prostate cancers.     

Screen-detected prostate cancer and the insulin-like growth factor axis: results of a population-based case-control study

Higher circulating levels of IGF-I have been associated with increased risk of prostate and some other cancers. Most research on prostate cancer has been based on men with symptoms or identified following treatment of benign disease. However, increasing numbers of cancer cases are now detected in asymptomatic men following prostate-specific antigen (PSA) tests. We therefore used a population-based case-finding exercise using the PSA test to examine whether associations between the IGF axis and cancer risk were apparent in this population. A matched case-control study was conducted among 7,383 men (50-70 years) receiving a PSA test as part of a case-finding exercise. Assays of IGF-I, IGF-II, IGFBP-2 and IGFBP-3 were performed on cases and 2 controls matched on age, recruitment center and calendar time. Analyses were based on 176 cases and 324 matched controls. The risk of prostate cancer increased across quartiles of IGF-I (highest vs. lowest quartile, OR = 2.34; 95% CI = 1.26-4.34; p(trend) = 0.02) and IGF-II (OR = 1.78; 95% CI = 0.94-3.15; p(trend) = 0.09). Controlling for smoking history and IGFBP-3 strengthened associations with cancer for both IGF-I (OR = 3.00; 95% CI = 1.50-6.01; p(trend) 0.005) and IGF-II (OR = 2.02; 95% CI = 1.07-3.84; p(trend) = 0.04) Associations between the IGFs and cancer risk were stronger for advanced cases. Our findings suggest that both IGF-I and IGF-II are associated with an increased risk of screen-detected prostate cancer.     

Valproic Acid prolongs survival time of severe combined immunodeficient mice bearing intracerebellar orthotopic medulloblastoma xenografts.

PURPOSE: To develop novel orthotopic xenograft models of medulloblastoma in severe combined immunodeficient mice and to evaluate the in vivo antitumor efficacy of valproic acid. EXPERIMENTAL DESIGN: Orthotopic xenografts were developed by injecting 10(3) to 10(5) tumor cells from four medulloblastoma cell lines (D283-MED, DAOY, MHH-MED-1, and MEB-MED-8A) into the right cerebellum of severe combined immunodeficient mice. Animals were then examined for reproducibility of tumorigenicity, cell number-survival time relationship, and histopathologic features. Tumor growth was monitored in vivo by serially sectioning the xenograft brains at 2, 4, 6, and 8 weeks postinjection. Valproic acid treatment, administered at 600 microg/h for 2 weeks via s.c. osmotic minipumps, was initiated 2 weeks after injection of 10(5) medulloblastoma cells, and treated and untreated animals were monitored for differences in survival. Changes in histone acetylation, proliferation, apoptosis, differentiation, and angiogenesis in xenografts were also evaluated. RESULTS: Tumorigenicity was maintained at 100% in D283-MED, DAOY, and MHH-MED-1 cell lines. These cerebellar xenografts displayed histologic features and immunohistochemical profiles (microtubule-associated protein 2, glial fibrillary acidic protein, and vimentin) similar to human medulloblastomas. Animal survival time was inversely correlated with injected tumor cell number. Treatment with valproic acid prolonged survival time in two (D283-MED and MHH-MED-1) of the three models and was associated with induction of histone hyperacetylation, inhibition of proliferation and angiogenesis, and enhancement of apoptosis and differentiation. CONCLUSION: We have developed intracerebellar orthotopic models that closely recapitulated the biological features of human medulloblastomas and characterized their in vivo growth characteristics. Valproic acid treatment of these xenografts showed potent in vivo anti-medulloblastoma activity. These xenograft models should facilitate the understanding of medulloblastoma pathogenesis and future preclinical evaluation of new therapies against medulloblastoma.     

Immunohistochemical demonstration of epidermal growth factor in human gastric cancer xenografts of nude mice.

Thirty-two surgical specimens and three cell lines of human gastric cancers were used for subcutaneous transplantation into nude mice, resulting in the establishment of eight (25%) xenografts from the surgical specimens and two (67%) from the cell lines. The localization of epidermal growth factor (EGF) in the surgical specimens and cell lines of the gastric cancers and their xenografts in nude mice was then investigated immunohistochemically. Epidermal growth factor was stained in the cytoplasm of the cancer cells, being detected in 16 (50%) of the 32 surgical specimens and in all of the cell lines. Seven (44%) of the sixteen EGF-positive surgical specimens and one (6%) of the 16 EGF-negative ones were tumorigenic in nude mice. All of the xenografts in nude mice were positive for EGF. The tumorigenicity of human gastric cancer xenografts in nude mice may, therefore, be correlated with the presence of EGF in cancer cells.     

Docetaxel followed by castration improves outcomes in LNCaP prostate cancer-bearing severe combined immunodeficient mice.

PURPOSE: Androgen ablation is the standard initial treatment for advanced prostate cancer; however, tumors eventually develop androgen independence and become incurable. Chemotherapy is commonly used after hormone treatment fails but has not shown significant survival benefit. Studies suggest that androgen ablation can select for a population of hormone-independent cells that are also relatively chemotherapy resistant. Thus, it may be therapeutically advantageous to target prostate cancer with chemotherapy before hormone ablation. This study was undertaken to determine the relative efficacy of such an approach in a preclinical model of prostate cancer. EXPERIMENTAL DESIGN: Severe combined immunodeficient mice bearing human LNCaP prostate tumors were treated with docetaxel and/or surgical castration applied singly, concurrently, or in different sequences. Treatment efficacy was determined by tumor volume and growth delay measurements. The extent of apoptosis in tumors in response to treatments was assessed via terminal deoxynucleotidyl transferase-mediated nick-end labeling (TUNEL) assays. In addition, Western blots were done to study the relative expression of Bcl-2 and Bax in the tumors. RESULTS: Docetaxel followed by castration showed the most potent antitumor effects. In contrast, with the exception of castration alone, castration followed by docetaxel produced the least antitumor activity. TUNEL assays confirmed that the density of apoptotic tumor cells was significantly greater for docetaxel followed by castration than for any other treatment. In tumors of mice treated with single modality therapies, Bax to Bcl-2 ratios decreased significantly after castration, whereas this ratio remained high after docetaxel treatment. CONCLUSION: A treatment sequence of docetaxel followed by hormone ablation may be more effective in treating prostate cancer than concurrent docetaxel/hormone therapy or hormone ablation followed by docetaxel.     

Targeting the insulin-like growth factor axis as a cancer therapy

The insulin-like growth factor (IGF) axis has been implicated in malignant transformation and in tumor cell biology. Human population studies have demonstrated that high levels of circulating IGF-I are associated with an increased risk of certain malignancies. Many model systems show that IGFs stimulate tumor cell proliferation, survival and metastasis. In a new era of anticancer treatments aimed at tumor-specific targets, efforts are in progress for the development of novel anti-IGF therapies. Disrupting type I IGF-receptor (IGF-IR) function in vitro and in vivo results in tumor growth inhibition in several model systems. Antireceptor therapies in particular have provided encouraging results leading to the approval of the first Phase I human clinical trial targeting IGF-IR. Additional methods to decrease levels of circulating IGF-I and II have also been developed. In principle, a benefit of targeted therapies could be their relative lack of toxicity compared with conventional chemotherapy. Anti-IGF-IR therapies, however, raise theoretical concerns for the development of serious side effects, including diabetes. As targeted therapies against the IGF axis continue to be developed, efforts will need to be made to minimize the side effects that result from blocking normal ligand and receptor-induced functions.     

A prospective study of the insulin-like growth factor axis in relation with prostate cancer in the SU.VI.MAX trial.

Several epidemiologic studies have examined with diverging results the relationships between circulating levels of insulin-like growth factors (IGF) and of IGF-binding proteins (IGFBP) and prostate cancer risk. We assessed the association of prediagnostic plasma levels of IGF-I, IGF-II, IGFBP-2, and IGFBP-3 and subsequent occurrence of prostate cancer in a case-control study nested in the SU.VI.MAX trial. The SU.VI.MAX study was a primary prevention trial testing a daily supplementation with low-dose antioxidant vitamins and minerals in male and female middle-aged volunteers in France. One hundred prostate cancer cases were diagnosed among 4,855 SU.VI.MAX participants over a 9-year follow-up period. For each case, four age-matched controls were selected randomly. Frozen baseline plasma samples were used to measure IGF-I, IGF-II, IGFBP-2, and IGFBP-3. Conditional logistic regression was used to assess the association between these four biochemical markers and prostate cancer risk. After controlling for the intervention group in the trial and the other IGF axis variables, the odds ratios and 95% confidence interval (95% CI) comparing the upper quartile to the baseline quartile were 1.83 (95% CI, 0.85-3.95), 1.05 (95% CI, 0.35-3.18), 0.79 (95% CI, 0.39-1.58), and 0.42 (95% CI, 0.12-1.52) for IGF-I, IGF-II, IGFBP-2, and IGFBP-3, respectively. More suggestive associations for IGF-I and IGFBP-3 were observed with advanced and aggressive cancers. Our results are consistent with those of some previous prospective studies and suggest that IGF axis variables are not long-term predictors of the occurrence of prostate cancer.     

The insulin-like growth factor axis and prostate cancer: lessons from the transgenic adenocarcinoma of mouse prostate (TRAMP) model

We have characterized the temporal expression of the insulin-like growth factor (IGF) axis in the transgenic adenocarcinoma of mouse prostate (TRAMP) model as prostate cancer progression in this model closely mimics that observed in the human disease, and the model provides samples representing the earliest stages of prostate cancer that are clinically the most difficult to obtain. We report that prostate-specific IGF-I mRNA expression increased during prostate cancer progression in TRAMP mice and was elevated in the accompanying metastatic lesions, whereas prostatic IGF-I mRNA remained at nontransgenic levels in androgen-independent disease. Expression of IGF-II mRNA, however, was reduced in primary prostate cancer, metastatic lesions, and androgen-independent disease. Expression of type-1 IGF receptor (IGF1R) mRNA, encoding the cognate receptor for both IGF-I and IGF-II, as well as type-2 IGF receptor (IGF2R) mRNA was not found to be altered during primary prostate cancer progression in intact TRAMP mice but was dramatically reduced in metastatic lesions and in androgen-independent disease. Similar to reports from clinical disease, serum IGF-I levels were observed to increase precociously in TRAMP mice early in disease progression but remained at nontransgenic levels after castration. Elevated serum levels of IGF-binding protein 2 were observed to correlate with advanced prostate cancer in the TRAMP model. Together these observations implicate IGF-I as an important factor during the initiation and progression of primary prostate cancer and provide evidence that there is a strong selection against expression of IGF1R and IGF2R in metastatic and androgen-independent disease.     

Cyclooxygenase-2 modulates the insulin-like growth factor axis in non-small-cell lung cancer

Constitutive overexpression of cyclooxygenase-2 (COX-2) occurs frequently in several different malignancies, including lung, colon, breast, and prostate cancer. Clinical studies have established elevated serum insulin-like growth factor (IGF-I) content and IGF-I:IGF-binding protein 3 (IGFBP-3) ratio as risk factors for these same malignancies. Therefore, we sought to determine the link between COX-2 expression and the IGF axis in COX-2 gene-modified human non-small-cell lung cancer (NSCLC) cells. Overexpression of COX-2 in NSCLC cells enhanced the antiapoptotic and mitogenic effects of IGF-I and IGF-II, facilitated the autophosphorylation of the type 1 IGF receptor, increased class IA phosphatidylinositol 3'-kinase activity, and decreased expression of IGFBP-3. Thus, these findings show that COX-2 augments the stimulatory arm of the IGF axis.     

脂联素对高脂饮食诱导的胰岛素抵抗子宫内膜癌裸鼠移植瘤生长的影响

目的 探讨脂联素对高脂饮食诱导裸鼠胰岛素抵抗子宫内膜癌移植瘤生长的影响。方法 40只裸鼠随机分成高脂组和普食组,每组20只,分别喂养高脂饲料（high-fat diet,HFD）和普通饲料（normal diet,ND）,10周后测定裸鼠空腹血糖（fasting blood glucose,FBG）和空腹血胰岛素（fasting serum insulin,FINS）水平,计算胰岛素抵抗指数（HOMA-IR）,建立胰岛素抵抗裸鼠模型。第11周接种子宫内膜癌HEC-1B细胞,待瘤体长至0.5 cm时,两组随机选取10只裸鼠腹腔注射脂联素（adiponectin,APN）分为HFD+APN组、ND+APN组;其余裸鼠分别注射0.9%氯化钠分为HFD组、ND组;14周后,测定各组血糖和血脂的代谢情况。结果 高脂组裸鼠10周后的平均体重、体长、FBG、FINS、HOMA-IR均大于普食组,糖耐量试验和胰岛素耐量试验结果显示血糖水平亦高于普食组（均P＜0.05）;接种子宫内膜癌HEC-1B细胞后均成瘤,建模成功。HFD+APN组和ND+APN组移植瘤平均重量和体积增长速度均分别低于HFD组和ND组（均P<0.05）。HFD组和HFD+APN组14周后的FINS、HOMA-IR、TC和TG均较ND组和ND+APN明显升高（均P<0.01）,且HFD组明显高于HFD+APN组（均P<0.01）;HFD+APN组的血脂联素水平低于ND+APN组（P<0.01）,HFD组则低于ND组（P<0.05）。结论 脂联素可改善高脂饮食导致的胰岛素抵抗,抑制子宫内膜癌移植瘤生长。     

Effect of low-fat diet on development of prostate cancer and Akt phosphorylation in the Hi-Myc transgenic mouse model

This study evaluated the effect of dietary fat on prostate cancer development by using the Hi-Myc mouse transgenic prostate cancer model. Hi-Myc mice develop murine prostatic intraepithelial neoplasia (mPIN) as early as 2 to 4 weeks and invasive adenocarcinoma between 6 and 9 months due to the overexpression of human c-Myc in the mouse prostate. Three-week-old male Hi-Myc mice were placed on high-fat (HF; 42% Kcal) or low-fat (LF; 12% Kcal) diets, and equal caloric intake was maintained until euthanasia at 7 months. The number of mice that developed invasive adenocarcinoma at 7 months was 27% less in the LF diet group (12/28) compared with the HF diet group (23/33, P < 0.05). Epithelial cells in mPIN lesions in the LF group had a significantly lower proliferative index compared with epithelial cells in the HF group (21.7% versus 28.9%, P < 0.05). During the mPIN phase of carcinogenesis (4 months), the LF group had higher serum insulin-like growth factor (IGF) binding protein-1 levels (21.0 +/- 8.9 ng/mL versus 3.2 +/- 0.8 ng/mL, P < 0.05) relative to the HF group. Akt (Ser(473)) phosphorylation, Akt kinase activity, and phosphorylation of downstream targets of Akt in prostates were significantly reduced in the LF diet group compared with the HF group. We conclude that dietary fat reduction delays transition from mPIN to invasive cancer in this Myc-driven transgenic mouse model, possibly through suppression of the IGF-Akt pathway and decreased proliferation of mPIN epithelial cells.     

Highly metastatic human prostate cancer growing within the prostate of athymic mice overexpresses vascular endothelial growth factor.

Angiogenesis is essential for tumor progression and metastasis. It is mediated by the release of angiogenic factors by the tumor or host. We analyzed the expression of angiogenic factors by the prostate cancer cell line LNCaP and two derived variants, in vitro and in vivo, to determine whether metastatic cell lines express higher levels of these factors. The production of three angiogenic factors, vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), and interleukin 8 (IL-8), by LNCaP and its variants, LNCaP-LN3 (highly metastatic) and LNCaP-Pro5 (slightly metastatic), was measured by ELISA. VEGF, bFGF, and IL-8 mRNA expression was determined in vitro by Northern blot analysis. VEGF mRNA expression was determined in vivo by in situ hybridization. VEGF and flk-1 protein expression and microvessel density of LNCaP cell tumors were quantified by immunohistochemistry. In vitro, VEGF production by LNCaP-LN3 (3.15+/-0.04 pg/ml/10(3) cells) was significantly higher than those of both LNCaP (2.38+/-0.34 pg/ml/10(3) cells) and LNCaP-Pro5 (1.67+/-0.37 pg/ml/10(3) cells; P = 0.049 and 0.001, respectively). None of the three cell lines produced detectable levels of bFGF or IL-8 in vitro. In vivo, LNCaP-LN3 tumors exhibited higher levels of VEGF mRNA and protein (152.2+/-28.5 and 200.5+/-28.3) and of flk-1 protein (156.5+/-20.6) and had higher microvessel density (16.4+/-4.2) than either LNCaP tumors (89+/-17.5, 173.3+/-23.0, 124.6+/-21.6, and 12.4+/-3.5, respectively) or LNCaP-Pro5 tumors (63+/-14.7, 141.2+/-38.1, 126.1+/-20, and 5.8+/-2.2, respectively). In conclusion, metastatic human prostate cancer cells exhibited enhanced VEGF production and tumor vascularity compared with prostate cancer cells of lower metastatic potential. Thus, VEGF may play an important role in prostate cancer metastasis.     

Morphology and function of human benign tumors and normal thyroid tissues maintained in severe combined immunodeficient mice.

In the improved SCID (severe combined immunodeficient) mice, various human benign tumors of the head and neck region were well maintained morphologically and functionally for 3 years until the experiments were terminated, e.g. transplanted parathyroid adenoma secreted parathyroid hormone (PTH) in the SCID mice for more than 1 year. Normal human thyroid tissue was also well maintained in the SCID mice for 3 years. Rapid and high uptake of radioiodine into the transplanted human thyroid tissue was observed. Furthermore, transplanted human thyroid tissue secreted thyroid hormone (T3) and T3 secretion was stimulated by the injection of human thyroid stimulating hormone (TSH). These findings suggest that the improved SCID mice will provide an invaluable experimental system for investigating the function of normal human tissues and the influence of endogenous and exogenous factors on human tissues.