Parathyroid hormone-related protein in human renal cell carcinoma

Parathyroid hormone-related protein (PTHrP), a polyprotein discovered in 1987, plays crucial roles not only in development and in various physiological events associated with normal life, but also in a number of pathological conditions such as cancer. PTHrP appears as the major causative agent in humoral hypercalcemia of malignancy (HHM) associated to a broad range of tumors. However, this is only one aspect of the multiple facets of PTHrP in cancer biology. Indeed, the complex growth factor-like properties of PTHrP has shed new light onto potential roles of this peptide in the regulation of tumor growth and invasion. Initial studies in breast, prostate and lung cancer and recent results in renal cell carcinoma (RCC) suggest such roles and highlight the therapeutic potential of PTHrP-targeting strategies in human cancer including RCC. In this review, the role of PTHrP in RCC tumorigenesis and its potential as a therapeutic target will be discussed.     

Parathyroid hormone-related protein is an independent prognostic factor for renal cell carcinoma

BACKGROUND: Parathyroid hormone-related protein (PTHrP) has been shown to be the principal cause of humoral hypercalcemia associated with renal cell carcinoma (RCC). Recent studies have demonstrated that the amino-terminal region of PTHrP has growth factor-like activities, suggesting it may play a role in the development of RCC. In this study, expression of the carboxy-terminal region of PTHrP was assessed immunohistochemically and its significance in predicting the prognosis of RCC was studied. METHODS: Forty radical nephrectomy specimens were immunostained with a murine monoclonal antibody (9H7) against the carboxy-terminal region (amino acids 109-141) of PTHrP using the streptavidin-peroxidase enzyme conjugate method. Staining intensity was evaluated semiquantitatively and compared with clinicopathologic features of the corresponding RCC. RESULTS: Immunoreactivity to 9H7 was observed to be localized to the cytosol of tumor cells at various staining intensities. There were 30 cases (75.0%) with strong staining and 10 cases (25.0%) in which staining was weak or nonexistent. Staining intensity showed no significant correlation with gender, tumor greatest dimension, stage, or grade. Tumors of the clear cell type expressed PTHrP to a significantly greater extent than tumors of the granular cell type. Tumor recurrence was significantly greater in the weakly stained or unstained group compared with the strongly stained group (P = 0.035). Multivariate analysis indicated that PTHrP expression and tumor stage were equally significant prognostic indicators in RCCs measuring <10 cm in greatest dimension. CONCLUSIONS: Evident PTHrP(109-141) expression is present in the majority of RCCs. The results of the current study indicate PTHrP(109-141) may be a possible marker of cellular differentiation and may be useful for predicting recurrence free survival in RCC patients after radical nephrectomy.     

Parathyroid hormone-related protein serves as a prognostic indicator in oral squamous cell carcinoma

Background

In our previous study, parathyroid hormone-like hormone (PTHLH) which encodes parathyroid hormone-related protein (PTHrP) was revealed to be up-regulated in oral squamous cell carcinoma (OSCC) compared with paired apparently normal surgical margins using microarray method. However, the function and prognostic indicators of PTHLH/PTHrP in OSCC remain obscure.

Methods

The mRNA levels of PTHLH and its protein levels were investigated in 9 OSCC cell lines and in 36 paired OSCC specimens by real-time PCR and western blotting. The biological function of PTHLH/PTHrP was investigated using small interfering RNA (siRNA) in 3 OSCC cell lines, and immunohistochemistry was used to estimate the prognostic value of PTHrP in 101 patients with head and neck squamous cell carcinoma (HNSCC), including OSCC and oropharyngeal squamous cell carcinoma. Cell cycle was tested by flow cytometry and cell cycle related genes were investigated by western blotting and immunocytochemistry assay.

Results

This study showed that the mRNA and protein levels of PTHLH in 9 OSCC cell lines were much higher than that in normal epithelial cells (P < 0.0001). In 36 paired OSCC tissues, PTHLH mRNA expressions were found higher in 32 OSCC tissues than that of paired apparently normal surgical margins (P = 0.0001). The results revealed that the down-regulation of PTHLH/PTHrP by siRNAs could reduce cell proliferation and inhibit plate and soft agar colony formation as well as affect the cell cycle of OSCC cells. The key proteins related to the cell cycle were changed by anti-PTHLH siRNA. The results showed that cyclin D1 and CDK4 expressions were significantly reduced in the cells transfected with anti-PTHLH siRNA. On the other hand, the expression of p21 was increased. The results also showed that high PTHrP level was associated with poor pathologic differentiation (P = 0.0001) and poor prognosis (P = 0.0003) in patients with HNSCC.

Conclusions

This study suggests that PTHLH/PTHrP is up-regulated in OSCCs. Therefore, PTHLH/PTHrP could play a role in the pathogenesis of OSCC by affecting cell proliferation and cell cycle, and the protein levels of PTHrP might serve as a prognostic indicator for evaluating patients with HNSCCs.

     

Sex-specific survival advantage with parathyroid hormone-related protein in non-small cell lung carcinoma patients.

PURPOSE: Parathyroid hormone-related protein (PTHrP) is commonly expressed in non-small cell lung carcinomas (NSCLC). Expression of the protein could have implications for progression of the disease because it regulates cancer cell growth, apoptosis, and angiogenesis. However, its relationship with survival has not been evaluated in a large-scale investigation. EXPERIMENTAL DESIGN: PTHrP expression was assessed in paraffin-embedded tumor samples from 407 patients with NSCLC by immunohistochemistry. A pathologist unaware of the clinical history classified specimens as PTHrP positive or PTHrP negative. The log-rank test was used to compare survivals of PTHrP-positive and PTHrP-negative groups, and Cox regression was used to adjust for additional covariates. RESULTS: Median survival was 55 versus 22 months (P < 0.001) in female patients with and without tumor PTHrP, respectively. Male survival was 38 months independent of PTHrP status. Stage, histology, age, and smoking history were also associated with increased longevity. PTHrP remained a significant predictor of survival for female patients after controlling for stage, histology, and age. CONCLUSIONS: In this study, PTHrP expression was associated with a survival advantage in female patients. Additional investigations must be done to ascertain whether the result is reproducible and independent of potential confounding covariates. Sex-dependent effects of PTHrP in lung cancer would open new avenues of research into the role of sex in cancer progression.     

Parathyroid hormone-related protein varies with sex and androgen status in nonsmall cell lung cancer

BACKGROUND: In nonsmall cell lung cancer, tumor parathyroid hormone-related protein (PTHrP) expression predicts longer survival in women but not in men. To explain the sex-dependent survival effect, the authors proposed that hormonal influences decrease PTHrP in men versus women, that PTHrP inhibits tumor growth, and that the effect is greater in women than in men. The objectives of this study were to compare lung carcinoma PTHrP expression and carcinoma growth in male and female mice and to determine whether gonadal steroids regulate PTHrP in lung cancer cells. METHODS: Tumor PTHrP content was measured by immunoassay, and tumor burden was assessed with multiple measures in BEN squamous cell orthotopic lung carcinomas in athymic mice. In addition, lung adenocarcinoma PTHrP messenger RNA (mRNA) values determined by microarray analyses were compared between men and women. Cultured lung cancer cells were assayed for PTHrP after treatment with estradiol or R1881, a synthetic androgen. RESULTS: Lung carcinomas contained approximately 3 times more PTHrP in female mice than in male mice. Similarly, levels of PTHrP mRNA were significantly greater in adenocarcinomas from patients who were women than from patients who were men. Male mice had greater tumor burden than female mice. Androgen treatment reduced PTHrP in 3 lung cancer lines. Estradiol had no effect. Testosterone treatment also reduced lung carcinoma PTHrP in female mice. CONCLUSIONS: Lung carcinomas in females expressed more PTHrP than in males possibly because of negative regulation by androgens in males. Female mice with higher tumor PTHrP content had significantly less tumor burden than male mice, supporting the hypothesis that PTHrP inhibits tumor growth.     

Parathyroid hormone-related protein expression is correlated with clinical course in patients with glial tumors

BACKGROUND: Parathyroid hormone-related protein (PTHrP) expression modulates cell survival in a number of human solid tumors. Although PTHrP is expressed in normal developing and neoplastic central nervous system tissue, clinical data indicating the importance of this protein with respect to local control and/or survival in patients with glial tumors are scarce. METHODS: Using a standard immunoperoxidase technique, the authors examined PTHrP expression in a population of 51 patients with Daumas-Duport Grade II-IV astrocytomas over a 15-year period. Both local control and survival were calculated from the date of definitive irradiation to the last time of known follow-up examination using the actuarial method. PTHrP expression was scored on examination under 40x magnification, with the incidence of cellular staining averaged over 10 high-power fields. The intensity and extent of staining were characterized semiquantitatively using the standard World Health Organization classification criteria. The median follow-up duration was approximately 5.5 years. Multivariate analyses were performed to ascertain the statistical significance of several standard clinicohistopatholgic factors (Karnofsky functional status, age, gender, extent of surgical resection, radiotherapy dose, grade, and PTHrP expression) with respect to local control and survival. P < 0.05 was considered indicative of statistical significance. RESULTS: Patients with high levels of PTHrP expression had significantly lower glial tumor local control rates and corresponding decreases in progression-free and overall actuarial survival after definitive irradiation (P < 0.01). In a Cox 3-variable model, the PTHrP staining score was independent of tumor grade or Karnofsky functional status. It is notable that the strongest predictor of survival was tumor grade (P < 0.001). CONCLUSIONS: PTHrP may be an important adjunct to standard immunopathologic criteria in the determination of glial tumor responses. A number of mechanisms were explored to derive a more mechanistic understanding of these translational results. Subsequent prospective studies involving larger patient populations will be necessary before findings can be translated to clinical practice.     

Parathyroid hormone-related protein and hypercalcemia in patients with metastatic melanoma: case report and review

Hypercalcemia associated with malignancy has been attributed to osteolytic processes secondary to bony metastases and to humoral factors causing increased bone resorption and decreased renal excretion of calcium. Parathyroid hormone-related protein (PTH-rP) is a humoral factor that has been associated with hypercalcemia in renal cell carcinoma, squamous cell carcinoma, and bladder carcinoma. Hypercalcemia does occur in patients with melanoma; however, few studies have reported on hypercalcemia in these patients, and even fewer have described a direct connection to PTH-rP. We here report a patient with stage IV malignant melanoma presenting with severe hypercalcemia associated with elevated PTH-rP levels. Immunohistochemistry showed strong expression of PTH-rP in biopsy of the patient's subcutaneous masses. In addition, we found a 4.9% incidence of hypercalcemia in 1,146 consecutive patients treated for metastatic melanoma at the Surgery Branch of the National Cancer Institute between January 1, 1988 and March 31, 2000. Thus, PTH-rP may play a significant role in severe hypercalcemia in patients with metastatic melanoma. The discovery of PTH-rP and relevant literature will also be reviewed.     

Parathyroid hormone-related protein in tumor tissues obtained from patients with humoral hypercalcemia of malignancy

We assessed the relationship between the parathyroid hormone-related protein (PTHrP) and the development of humoral hypercalcemia of malignancy (i.e., hypercalcemia due to the production by solid tumors of hypercalcemic factors) by assaying tumor extracts from hypercalcemic and normocalcemic patients with cancer for immunoreactive PTHrP contents. Immunoreactive PTHrP was demonstrated in extracts of 21 of 22 tumor tissues obtained from 22 patients with humoral hypercalcemia of malignancy. Immunoreactive PTHrP was rarely seen in tumor tissue extracts obtained from 34 normocalcemic patients with cancer and two hypercalcemic patients with cancer with severe bone metastases. Gel filtration studies of tumor extracts revealed a molecular-size heterogeneity of immunoreactive PTHrP. These radioimmunoassay data indicate a close relationship between detection of PTHrP in tumor tissues and the development of humoral hypercalcemia of malignancy.     

Parathyroid hormone-related protein and human papillomavirus in gynecological tumors

The presence of parathyroid hormone-related protein (PTHrP) and human papillomavirus (HPV) in a series of gynecological tumors from 131 unselected patients was examined. PTHrP was localized immunohistochemically using a highly specific rabbit polyclonal anti-serum against PTHrP(1–16). The results confirmed that gynecological malignancies, although rarely associated with humoral hypercalcemia of malignancy (HHM), stained for PTHrP in a majority of the squamous-cell carcinomas (SCC) at all sites, but only in a minority of adenocarcinomas, and then in areas of squamous metaplasia. This included a series of endometrial tumors. Detection of HPV types was achieved using a polymerase-chain-reaction (PCR) detection system enabling the detection of HPV types 6, 11, 16, 18,31,33 and 45. PTHrP production was not directly related to HPV infection, but correlated with the type of tumor.     

Decreased serum adiponectin levels in patients with metastatic renal cell carcinoma

Background: Low levels of serum adiponectin are associated with increasedrisk and aggressiveness of obesity-related cancer. The purposeof the study reported here was to investigate the associationbetween serum adiponectin levels and clinicopathological parametersof renal cell carcinoma. Methods: Preoperative serum total and high-molecular-weight (HMW) adiponectinlevels were measured in 118 patients with renal cell carcinoma,and their association with clinicopathological parameters wasanalysed. Results: There were no statistically significant associations betweentotal adiponectin and HMW adiponectin and pathological stage,regional lymph node involvement, histological grade, histologicaltype (clear cell carcinoma versus other types) or presence ofvenous invasion. Total and HMW adiponectin levels in patientswith metastasis, however, were significantly lower than in patientswithout metastasis (P = 0.044 for total adiponectin and P =0.041 for HMW adiponectin). Low total and HMW adiponectin levelswere significantly associated with metastasis in patients witha normal BMI (<25 kg/m²) (P = 0.034 for total adiponectinand P = 0.028 for HMW adiponectin) but not in overweight andobese patients (P = 0.652 for total adiponectin and P = 0.489for HMW adiponectin). Multivariate logistic regression analysisshowed that total adiponectin level was an independent predictorof metastasis of renal cell carcinoma in all patients (P = 0.024,95% CI = 1.031–1.560) and in patients with a normal BMI(P = 0.040, 95% CI = 1.043–6.534). Conclusions: Serum total and HMW adiponectin levels were decreased in patientswith metastatic renal cell carcinoma. Adiponectin might be amolecular link between obesity and the progression of renalcell carcinoma.     

Hypercalcemia in patients with esophageal carcinoma. The pathophysiologic role of parathyroid hormone-related protein

To elucidate the actual incidence of hypercalcemia in patients with esophageal carcinoma, 382 consecutive cases admitted to the National Cancer Center Hospital (Tokyo, Japan) from 1983 to 1988 were investigated. Hypercalcemia was detected in 5 patients of 376 (1.3%) at the time of primary detection of cancer, and in 45 patients of 120 (38%) patients with recurrent or unresectable cancer who were monitored within 2 months of death. These observations demonstrated that this electrolyte imbalance is a frequent paraneoplastic syndrome observed in patients with advanced esophageal carcinoma. With regard to the etiology, bone metastases were detected in 13 of 49 patients with hypercalcemia; the remaining 36 patients were assumed to be induced by the production of hypercalcemic substance(s) by tumor tissues. Parathyroid hormone-related protein (PTHrP) is a newly discovered factor which increases serum calcium in vivo. The detection of PTHrP mRNA in tumor tissues as well as the production of PTHrP-like immunoreactivity by tumor tissues were closely associated with the development of hypercalcemia, suggesting that PTHrP is the major cause of hypercalcemia in patients with esophageal carcinoma.     

Serum antibody against Tamm-Horsfall protein in patients with renal cell carcinoma

We assayed sera from 24 patients with metastatic renal cell carcinoma for class specific antibody against urinary Tamm-Horsfall protein (THP) using indirect solid-phase radioimmunoassay techniques. Overall, 16 patients had elevated antibody levels (greater than 2 SD above the mean levels of control subjects). IgG against THP was elevated in eight of the patients, IgA in 12, and IgM in five. However, THP was not identifiable by immunohistochemical techniques in the sections of 15 of 24 tumors available for study. Nine of the 15 tumors were from patients with elevated antibody levels. Serum antibody against THP in patients with renal cell carcinoma appears to result from antigenic stimulation by THP derived from the urine rather than from the neoplasm.     

Antitumor effect of parathyroid hormone-related protein neutralizing antibody in human renal cell carcinoma in vitro and in vivo

Functional inactivation of the von Hippel-Lindau (VHL) tumor suppressor gene occurs in 40-80% of human conventional renal cell carcinomas (RCCs). We showed recently that VHL-deficient RCCs expressed large amounts of parathyroid hormone-related protein (PTHrP), and that PTHrP, acting through the PTH1 receptor (PTH1R), plays an essential role in tumor growth. We also showed that PTHrP expression is negatively regulated by the VHL gene products (pVHL). Our goal was to determine whether blocking the PTHrP/PTH1R system might be of therapeutic value against RCC, independent of VHL status and PTHrP expression levels. The antitumor activity of PTHrP neutralizing antibody and of PTH1R antagonist were evaluated in vitro and in vivo in a panel of human RCC lines expressing or not pVHL. PTHrP is upregulated compared with normal tubular cells. In vitro, tumor cell growth and viability was decreased by up to 80% by the antibody in all cell lines. These effects resulted from apoptosis. Exogenously added PTHrP had no effect on cell growth and viability, but reversed the inhibitory effects of the antibody. The growth inhibition was reproduced by a specific PTH1R antagonist in all cell lines. In vivo, the treatment of nude mice bearing the Caki-1 RCC tumor with the PTHrP antibody inhibited tumor growth by 80%, by inducing apoptosis. Proliferation and neovascularization were not affected by the antiserum. Anti-PTHrP treatment induced no side effects as assessed by animal weight and blood chemistries. Current therapeutic strategies are only marginally effective against metastatic RCC, and adverse effects are common. This study provides a rationale for evaluating the blockade of PTHrP signaling as therapy for human RCC in a clinical setting.     

Increased serum levels of vascular endothelial growth factor in patients with renal cell carcinoma.

Neovascularization, an essential event for the growth of solid tumors, is regulated by a number of angiogenic factors. One such factor, vascular endothelial growth factor (VEGF), is considered to exert a potent angiogenic activity, as indicated by immunohistochemical and molecular evidence. In this study we investigated the serum VEGF level (s-VEGF) in patients with renal cell carcinoma (RCC). s-VEGF in peripheral blood samples was analyzed in 40 RCC patients and 40 patients without cancer (controls) using a sandwich enzyme-linked immunoassay. In 20 RCC patients, serum samples were obtained separately from the bilateral renal veins. s-VEGF was also measured before, 4 and 8 weeks after nephrectomy in 11 patients. There were significant differences in s-VEGF between the RCC patients and the controls (207.3+/-32.9 vs. 71.5+/-9.1 pg/ml, mean+/-SE) (P<0.005), between the tumor-bearing renal veins and the contralateral ones (P<0.01), between the pre- and post-nephrectomy situations (P<0.01) and among the various parameters of tumor status such as tumor extent (P<0.001) and existence of metastasis (P<0.001). s-VEGF significantly correlated with the tumor volume obtained by a three-dimensional measurement (r=0.802, P<0.0001). The sensitivity and specificity of s-VEGF at the cut-off level of 100 pg/ml, as determined by the receiver-operating-characteristics curve, were 80.0% and 72.5%, respectively. The results indicate that tumor tissue of RCC liberates VEGF into the systemic blood flow and that s-VEGF is a possible marker for RCC.     

Parathyroid hormone-related protein localization in breast cancers predict improved prognosis

In a prospective study of 526 consecutive patients with operable breast cancer, the significance of positive parathyroid hormone-related protein (PTHrP) staining by immunohistology has been evaluated for a median of 10-year follow-up. Improved survival was observed for the 79% of tumors which stained positively for PTHrP [estimated univariate hazard ratio, 0.43; 95% confidence interval (95% CI), 0.30-0.62; P < 0.001]. Adjustments for N stage, progesterone receptor status, and log tumor size changed this estimate only slightly to 0.47 (95% CI, 0.63-0.69; P = 0.001). Patients with PTHrP-positive primary tumors were less likely to develop bone metastases (hazard ratio, 0.63; 95% CI, 0.41-0.98; P = 0.04). PTHrP status was associated with estrogen receptor (P = 0.01), progesterone receptor (P = 0.03), and menopausal status (P = 0.006) but was not significantly associated with tumor size, vascular invasion, tumor grade, or patient age. Of 19 patients requiring surgery for bone metastases, the primary cancers were PTHrP negative in seven, all but one of whom had PTHrP-positive bone metastases. All 12 patients with PTHrP-positive primary cancers also had positive bone metastases. We conclude that increased production of PTHrP by breast cancers confers on them a less invasive phenotype, an effect distinct from the bone resorption-stimulating action that favors bone metastasis. It is likely that the latter property is influenced by factors in the bone microenvironment.     

术前外周血淋巴细胞与单核细胞比值和白蛋白在肾透明细胞癌预后评估中的价值

背景与目的：越来越多的证据提示炎症在肿瘤的发展演进中起着重要的作用,术前外周血淋巴细胞与单核细胞比值（lymphocyte-to-monocyte ratio,LMR）和白蛋白被证明是各种肿瘤的独立预后因素。探讨LMR和白蛋白在传统临床预后预测模型的基础上对肾透明细胞癌（clear cell renal cell carcinoma,ccRCC）预后评估的应用价值。方法：回顾性分析2012—2015年在广西医科大学第一附属医院行RCC根治术或肾部分切除的147例ccRCC患者的临床资料。外周血LMR和白蛋白于术前1周收集,LMR采用中位数3.42,白蛋白用40 g/L作为截点。采用单因素分析和COX回归模型,分析LMR和白蛋白及其他临床因素与患者预后的关系。将LMR和白蛋白结合传统预测指标TNM分期及Fuhrman分级进行分析。采用C指数预测LMR和白蛋白对RCC预后的准确性;同时,采用LMR、白蛋白、TNM分期和Fuhrman分级构建列线图,预测患者的生存率。结果：术前低LMR和低白蛋白是患者总生存期（overall survival,OS）的独立危险因素（P=0.001和P＜0.001）。低LMR与高Fuhrman分级（P=0.006）和肿瘤坏死（P=0.039）密切相关;低白蛋白与高Fuhrman分级（P＜0.001）和高Mayo临床分期、大小、分级和坏死（stage, size, grade and necrosis,SSIGN）分数（P=0.001）有关。单因素分析提示LMR、白蛋白、TNM分期、Fuhrman分级、肿瘤大小、肿瘤坏死与OS相关;COX回归模型提示LMR和白蛋白是OS的独立预后因素（HR=0.370,95% CI：0.145~0.942,P=0.037;HR=0.325,95% CI：0.136~0.775,P=0.011）。将LMR和白蛋白结合传统预测指标TNM分期及Fuhrman分级进行分析后,C指数上升;列线图构建结果发现可以预测RCC患者术后3和5年的生存率。结论：术前外周血LMR和白蛋白是ccRCC患者术后的独立预后因素,将其纳入常规的临床病理学参数中,可以提高ccRCC患者术后预后评估的精准性。