PTHrP and Tumorigenesis Is There a Role in Prognosis? MINORU NISHIHARA,a,b TAKASHI KANEMATSU,b TAKASHI TAGUCHI,a AND MOHAMMED S. RAZZAQUEa a Department
of Pathology, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki, Japan b Department
of Surgery, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki, Japan
ABSTRACT: The role of parathyroid hormone-related peptide (PTHrP) in the regulation of hypercalcemia in patients with malignancies is well studied, but whether its expression in tumor tissue correlates with tumor progression is not clear at present. The majority of tumors that metastasize to the bone produce PTHrP, and PTHrP expression correlates with skeletal localization of tumors. About 95% of colorectal adenocarcinomas overexpress PTHrP mRNA and protein, and the expression level is higher in poorly differentiated than in well-differentiated adenocarcinomas. However, there is some controversy at present about the prognostic significance of PTHrP expression on primary tumor cells, and studies suggest that there might be tissue-specific responses. We will briefly present here existing evidences that suggest that the expression of PTHrP in the primary tumor tissue could have both positive and/or negative impact on tumor progression and clinical outcome of the disease. KEYWORDS: PTHrP; colorectal tumors; prognosis
INTRODUCTION Parathyroid hormone-related protein (PTHrP) shares structural similarities with the parathyroid hormone (PTH), particularly in the 34 amino acids of the amino-terminal. The PTHrP 1–34 fragment binds to the same PTH receptor as PTH 1–34,1,2 and usually intensifies the effects of PTH in tissues that express the receptors. PTHrP is actively involved in endochondral bone formation.3–5 During embryonic development, once the cartilage mold is formed, perichondral cells and Address for correspondence: Takashi Taguchi, M.D., Ph.D., Department of Pathology, Nagasaki University Graduate School of Biomedical Sciences, 1-12-4 Sakamoto, Nagasaki 852-8523, Japan. Voice: 81-958-497-053; fax: 81-958-497-056. taguchi@nagasaki-u.ac.jp C 2007 New York Academy of Sciences. Ann. N.Y. Acad. Sci. 1117: 385–392 (2007). doi: 10.1196/annals.1402.046 385
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chondrocytes at the ends of the mold start to produce PTHrP, which through PTH/PTHrP receptors on chondrocytes, induces proliferation and differentiation of these cells to hypertrophic chondrocytes. Such proliferation continues in the presence of PTHrP, while it delays further differentiation. Although the roles of PTHrP in chondrogenesis and eventual osteoclastogenesis are well documented, PTHrP was initially discovered in patients with malignancies associated with hypercalcemia.6–9 Subsequent studies found PTHrP to be expressed in many normal and neoplastic tissues, even without hypercalcemia, and to have diverse physiological effects in a paracrine/autocrine fashion.10–12 The results of recent studies also suggest “intracrine” effects for PTHrP, thus possibly bypassing the requirement for cell surface receptor binding. The intracrine action of PTHrP might facilitate nucleocytoplasmic trafficking to influence cell cycle progression, RNA transcription, and transport.13,14 Our knowledge of the structure, regulation, and function of PTHrP has significantly enhanced our understanding of the roles of PTHrP in normal physiology and in pathologic conditions, such as malignancies and skeletal metastasis.
PTHrP AND TUMORIGENESIS Upregulated PTHrP expression is often found in tumors with skeletal metastasis.15 More importantly, its expression was found to correlate with differentiation and progression of the primary tumor, as well as with bone metastasis.15–17 Such expression often has a prognostic significance. PTHrP could play a role in the growth and differentiation of both neoplastic and nonneoplastic cells. Overexpression of PTHrP has been detected in advanced prostatic cancer 18,19 and in the skeletal metastases of breast cancer.15,20 Approximately 80% of patients with breast cancer develop bone metastases during the course of their disease. Osteolytic responses of breast cancer induce bone loss through stimulation of osteoclastic activities. Although the role of PTHrP in bone resorption is well established, whether it initiates or sustains such process is an unsettled issue. The discovery of overexpression of interleukin-8 (IL-8) in metastatic breast cancer cells prompted additional analysis of the role of IL-8 in osteolysis. Studies using recombinant IL-8 reported that this cytokine induces the expression of RANKL mRNA and protein in osteoblastic cells and stimulates the formation of bone resorbing osteoclasts. The ability of IL-8 to stimulate osteoclastogenesis directly via RANKL suggests that it may play an important role in the process of osteoclast formation and function.21,22 Using metastatic human breast cancer cells, Bendre et al.21 demonstrated that upregulation of IL-8 and not PTHrP renders the tumor cells phenotypically more invasive to metastasize the bone. One of the thoughts put forward is that IL-8 is involved in the early stages of breast cancer metastasis and that it initiates the process
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of osteoclastic bone resorption, while PTHrP expression is induced later to stimulate the vicious cycle of bone destruction.23 Whatever the molecular mechanism of osteolysis, there is agreement that PTHrP plays a role both in primary and secondary metastatic tumors in the bone. When nude mice were injected with human breast cancer cells (MDA-MB231), they developed osteolytic lesions without any change in peripheral blood levels of PTHrP or calcium, although elevated PTHrP level was described in the bone marrow.24 Inactivation of PTHrP in MDA-MB-231 cells using intracardially injected neutralizing antibody reduced the ability of these cells to grow in the bone.24,25 Enhanced bone tumor growth was also achieved with the less invasive MCF-7 cells that were transfected to overexpress PTHrP.26 In a separate study, metastatic pituitary tumor cells (mGH3) were found to express high levels of PTHrP. In vivo transfection of the antisense oligonucleotide for PTHrP in mGH3 cells reduced their tumor growth potential.27 Inoculation of mGH3 cells in the cerebral ventricle resulted in a rapid growth of tumor cells within 3 weeks and their dissemination throughout the entire ventricular system. In vivo administration of antisense PTHrP at 1 week after tumor cell implantation into the right lateral ventricle markedly reduced tumor size and suppressed the formation of metastases. Of relevance, treatment with sense or mismatched PTHrP oligonucleotide did not influence subsequent tumor growth. Furthermore, the survival rate of mGH3 tumor-injected rats improved following antisense PTHrP therapy. Considered together, these results point to the protumorigenic effects of PTHrP.
PTHrP EXPRESSION AND POSITIVE CORRELATION WITH TUMOR PROGRESSION Analysis of 108 colorectal tumors, including 12 adenomas, 6 adenocarcinomas in adenomas, and 90 adenocarcinomas showed lack of PTHrP expression in all adenomas and background nonneoplastic mucosal epithelia. In contrast to the adenomatous lesions, PTHrP was expressed in 85 out of 90 colorectal adenocarcinomas.16 Further analysis showed higher PTHrP immunoreactivity in poorly differentiated adenocarcinomas than in well-differentiated ones (FIG. 1). One of the important prognostic factors in colorectal carcinoma is the extension of tumor cells into the bowel wall and the presence of lymph node metastasis. PTHrP expression significantly correlated with differentiation, depth of invasion, lymphatic invasion, lymph node metastasis, and hepatic metastases. These findings clearly suggest that PTHrP is involved in carcinogenesis, tumor growth, differentiation, and progression, and might have prognostic value by determining the behavior of human colorectal adenocarcinomas. Similar observations are also noted in gastric tumors. Out of 92 cases of gastric adenocarcinoma, 71 (77%) cases of gastric tumors without humoral hypercalcemia showed PTHrP expression.28 In the same study, 5% of the adenomas
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FIGURE 1. PTHrP immunohistochemistry in normal colonic mucosa and colorectal tumors. Note lack of PTHrP expression in normal colonic mucosa (A), and in tubulo-villus adenoma cells (B). In adenocarcinoma within a representative adenoma; note the expression of PTHrP in the carcinomatous component (arrows) but not in the adenomatous component (C). PTHrP expression is weakly positive in a well-differentiated adenocarcinoma located in the submucosa (D). PTHrP is strongly expressed in the entire cytoplasm of tumor cells in a poorly differentiated adenocarcinoma (E). Strong expression of PTHrP is present in the subserosal invasive site of a moderately differentiated adenocarcinoma (F). Note that the expression of PTHrP is stronger in the advancing margins (arrows) than in the adjacent part of the moderately differentiated adenocarcinoma (G). Strong immunoreactivity is also found in the carcinoma cells invading the lymphatic vessel (H). Reproduced from the Journal of Pathology16 by permission of the Pathological Society of Great Britain and Ireland.
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were positive for PTHrP expression, while the background nonneoplastic epithelium was negative for PTHrP expression. Furthermore, PTHrP expression was higher in moderately differentiated and poorly differentiated gastric adenocarcinomas, suggesting that PTHrP might be involved in the malignant transformation and progression of gastric adenocarcinoma.28 Furthermore, association between PTHrP expression and growth and differentiation of gastrointestinal stromal tumors, leiomyomas, and schwannomas has also been proposed.29 In 177 surgically resected breast carcinoma tissues stained immunohistochemically with a monoclonal antibody against PTHrP, showed positive staining in 64% of cases. Such PTHrP expression correlated significantly with the histological grade of breast tumors. Moreover, PTHrP expression is also reported to be associated with bone metastasis; statistical analysis suggested a significantly shorter overall survival of patients harboring PTHrP-expressing tumors.30 Similarly, a separate study of 176 surgically excised early breast cancers and 43 bone metastases found that 68% of cases of early breast cancer showed PTHrP expression, while 100% of cases of bone metastases showed PTHrP expression. Moreover, 88 out of the 176 (50%) cases of early breast cancer also expressed PTHrP receptors, while 35 out of 43 (81%) cases of bone metastases expressed PTHrP receptors. The investigators found that the expression of either PTHrP or PTHrP receptor correlated with poor diseasefree survival, while coexpression of both PTHrP and receptor predicted a worse clinical outcome at 5 years, with a mortality rate of 32% compared with the ligand and receptor-negative group with a mortality rate of 6%.31 Although there are numerous reports favoring the role of PTHrP in tumor progression, there are also studies that suggest opposing effects.17
PTHrP EXPRESSION AND NEGATIVE CORRELATION WITH TUMOR PROGRESSION In a recent study using tumor samples from 407 patients with nonsmall cell lung carcinoma (NSCLC), PTHrP expression was found to be associated with a survival advantage in female patients.17 The authors indicated that the expression of PTHrP was a significant predictor of survival in female patients, independent of tumor stage, histology, and age.17 Of relevance, no such relationship with survival was noted among the male patients.17 Their results suggest possible involvement of sex-related factors in the pathogenesis of lung tumors, and the positive impact of expression of PTHrP on survival. In a separate study of primary breast tumors, no difference in the PTHrP status was documented among three different groups of patients: (1) the favorable outcome group (patients had a favorable prognosis and minimum 3 years disease-free follow up), (2) the unfavorable outcome group (all patients presented with localized breast cancer but developed distant disease within 3 years), and (3) the unfavorable presentation group (all had distant disease at first presentation),
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although increased level of PTHrP in the primary tumor was associated with metastasis.32 Similarly, a relationship between intensity of PTHrP expression and survival time was documented in ductal mammary carcinoma; patients with high expression of PTHrP showed longer survival than patients with low expression of PTHrP.33 Moreover, in the group with low PTHrP expression, a negative relationship was detected between expression of PTHrP and survival time. The investigators did not find any relationship between PTHrP expression and the development of distant metastases.33 In a prospective study of a relatively large number of patients (526 breast cancer patients), the significance of PTHrP expression examined by immunohistology was evaluated over a median follow-up of 10 years. The investigators found improved survival of 79% of patients with PTHrP-positive tumors.34 Moreover, the authors indicated that patients with PTHrP-positive primary tumors were less likely to develop bone metastases, and speculated that increased production of PTHrP by breast tumor cells might render the tumor cells less invasive.34
CONCLUDING REMARKS Despite the notion that increased expression of PTHrP is associated with the development of bone metastasis, a number of studies have indicated that the expression of PTHrP in the primary tumor site improves prognosis. Such inconsistency cannot be explained always by related factors, such as patient population, their selection, site, and stage of the disease and follow-up period. Whether certain genetic background has a differential effect on the expression of PTHrP and its functions needs further studies. Moreover, PTHrP might exert distinct effects on different microenvironments, and the bone resorption effect that favors bone metastasis might be influenced also by the intrinsic microenvironment of the bone.34 Despite a significant number of studies that detailed the role of PTHrP in tumorigenesis, such studies generated as many questions as they answered. Since the prognostic significance of PTHrP expression on primary tumor cells is an unsettled issue, any clinical utilities that manipulate PTHrP therapeutically need thoughtful consideration.
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