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The role of the calcium-sensing receptor in cancer
Cell Calcium 35 (2004) 291–295
The role of the calcium-sensing receptor in cancer Karin D. Rodland Pacific Northwest National Laboratory, Biological Sciences Division, Richland, WA 99352, USA Received 20 October 2003; accepted 27 October 2003
Abstract The extracellular calcium-sensing receptor (CaR) is a versatile sensor of small, polycationic molecules ranging from Ca2+ and Mg2+ through polyarginine, spermine, and neomycin. The sensitivity of the CaR to changes in extracellular Ca2+ over the range of 0.05–5 mM positions the CaR as a key mediator of cellular responses to physiologically relevant changes in extracellular Ca2+ . For many cell types, including intestinal epithelial cells, breast epithelial cells, keratinocytes, and ovarian surface epithelial cells, changes in extracellular Ca2+ concentration over this range can switch the cellular behaviour from proliferation to terminal differentiation or quiescence. As cancer is predominantly a disease of disordered balance between proliferation, differentiation, and apoptosis, disruptions in the function of the CaR could contribute to the progression of neoplastic disease. Loss of the growth suppressing effects of elevated extracellular Ca2+ have been demonstrated in parathyroid hyperplasias and in colon carcinoma, and have been correlated with changes in the level of CaR expression. Activation of the CaR has also been linked to increased expression and secretion of PTHrP (parathyroid hormone-related peptide), a primary causal factor in hypercalcemia of malignancy and a contributor to metastatic processes involving bone. Although mutation of the CaR does not appear to be an early event in carcinogenesis, loss or upregulation of normal CaR function can contribute to several aspects of neoplastic progression, so that therapeutic strategies directed at the CaR could potentially serve a supportive function in cancer management. © 2003 Elsevier Ltd. All rights reserved. Keywords: Calcium-sensing receptor; Cancer; Cancer management
The decision of a cell to proliferate or quiesce, to differentiate or apoptosis, is the result of a regulated process of communication between the cell and its environment. One of the molecular hallmarks of cancer is the disruption of this normal regulatory process. Cells either fail to sense the appropriate environmental cues promoting differentiation over proliferation, or the response to those cues is inappropriate. Therefore, it is highly appropriate that, in the search for molecular mechanisms of cancer, each newly characterized receptor for environmental stimuli is examined for its potential role in carcinogenesis. The calcium-sensing receptor (CaR) is a seven transmembrane domain G-protein coupled receptor that was initially characterized as the sensor responsible for modulating parathormone and calcitonin release in response to changes in blood calcium levels [7]. However, the CaR is more than just a calcium sensor; it is a fairly broad spectrum sensor of small cationic molecules, capable of transducing signals in response to changes in the concentration of heavy metals, including lead and cadmium [21], as well as cationic amino acids [39]. Although plasma free calcium levels are tightly regulated, significant calcium gradients do exist in several physiological compartments, such as gut and skin (e.g. [37]), and these calcium gradients are known to af0143-4160/$ – see front matter © 2003 Elsevier Ltd. All rights reserved. doi:10.1016/j.ceca.2003.10.011
fect the behaviour of the resident cell types. It has been known for over two decades that human keratinocytes only proliferate in a very low calcium environment [22,23], and that even modest increase in calcium concentration trigger keratinocyte differentiation [44,47]. The crypts of the large intestine also display a gradient of calcium concentration, generally increasing from the base of the crypt to its lumen [4]. This calcium gradient corresponds with differences in cellular behaviour; proliferation of intestinal epithelial cells originates in the base of the crypts, and cells differentiate as they progress apically, eventually undergoing apoptosis at the distal end of the crypt [4,37].
1. Parathyroid adenomas and the CaR The primary site of CaR action is in the chief cells of the parathyroid gland, where the CaR translates increases in blood calcium concentrations into decreased secretion of parathormone (PTH) [7,8]. CaR protein and mRNA is more abundantly expressed in the parathyroid than in most other tissues, therefore it is logical to examine CaR expression in parathyroid adenomas. Several investigators have surveyed human parathyroid adenomas for changes in either the level
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of CaR expression or for somatic mutation in the CaR gene [13,15,40]. In general, CaR protein levels appear to be decreased in parathyroid adenomas; at least three different groups have reported a decrease in CaR mRNA levels in parathyroid adenomas and hyperplasias [15,40,46]. Closer examination of mRNA transcripts has revealed an interesting discordance between the levels of two distinct CaR transcripts driven from distinct promoters. The CaR transcript which contains exon 1B is expressed at similar levels in normal and hyperplastic parathyroid; however, the exon 1A transcript is significantly suppressed in parathyroid adenomas [13]. At this point, no functional distinction between the two CaR transcripts is evident. However, this finding does suggest that CaR exon 1A is regulated differently than the exon 1B transcripts, and may be correlated with the ability of the CaR to inhibit proliferation in certain cell types such as the parathyroid [14] and intestinal epithelial cells [27]. Sequencing of the CaR from patients with hyperparathyroidism has led to the identification of a novel polymorphism that may be associated with increased activity of the CaR [3]. However, this polymorphism is also observed in normal tissues, and therefore is not likely to be causal for parathyroid hyperplasia. More extensive studies of genetic variability at the CaR locus in both parathyroid adenomas and pituitary tumours have failed to reveal any consistent link between mutation of the CaR and tumorigenesis [10].
2. Colon cancer and the CaR The colon carcinoma cell lines Caco-2 [26], HT-29 [16] and T-84 [16] express functional CaR and respond to addition of Ca2+ by releasing intracellular calcium. Kallay et al. [27] have shown that Caco-2 cells respond differentially to changes in Ca2+ depending on whether the stimulus was applied to basolateral or luminal surfaces. Reduction of Ca2+ to 0.025 mM on the luminal surface resulted in reduction of c-myc RNA expression, while a similar change confined to the basolateral surface had no effect [27]. These data correlate well with the observations that c-myc is a major factor regulating cell cycle progression in intestinal epithelial cells [26], and that proliferation of normal intestinal epithelial cells is highest in the base of crypts, were extracellular calcium concentrations are lowest [37]. Furthermore, immunohistochemical studies of human colon carcinomas indicated that CaR protein was most highly expressed in well-differentiated regions of the tumour and nearly lacking in poorly differentiated regions [29,43]. These data suggest that loss of a normal growth inhibitory response to elevated extracellular calcium, dependent on expression of functional CaR, may contribute to the progression of colon carcinomas. The possible linkage between loss of CaR function and cancer progression in the colon is particularly interesting in light of epidemiological data linking increased dietary calcium and Vitamin D with a decreased risk of colon cancer [17,31]. Thus, the CaR may be playing a physiologically
relevant role in regulating the proliferation and apoptosis of normal intestinal epithelial cells. Although it is highly unlikely that loss of CaR function is sufficient to initiate carcinogenesis in the colon or other organ sites, disruption of CaR function by dietary or other means may contribute significantly to tumour progression. Other epithelial cells may also respond to increases in extracellular calcium with a slowing of proliferation. Increased dietary calcium is associated with a decreased risk of intestinal carcinoma [25] and breast cancer [32]. Normal pancreatic islet cells express both CaR mRNA and protein [6,18,30], and functional CaR were observed in both insulinoma cells [30] and gastrinoma cells [18]. The level of CaR protein expression varied significantly within gastrinoma cells, and the correlation between levels of CaR expression and levels of gastrin release were not determined. However, it is possible that increased CaR expression may contribute to the increased sensitivity of both insulinoma cells and gastrinoma cells to extracellular calcium [18,30].
3. CaR and cancer of the breast and ovary Functional CaR are expressed by both normal mammary epithelial cells [12] and normal ovarian surface epithelial cells of the human and rhesus monkey [35,45]. Immunohistochemical analysis of normal breast showed membrane-associated CaR staining in ductal epithelial cells, as well as in the cells lining cysts in patients with fibrocystic disease of the breast [12]. This abundant expression of CaR protein is consistent with the role of the mammary gland in concentrating calcium in milk, as well as the known sensitivity of cultured primary human mammary epithelial cells to undergo terminal differentiation in high calcium media [34,38]. The possibility that the CaR may have a function in breast carcinoma is supported by the observation of CaR protein in patient samples from two cases of ductal carcinoma of the breast [12], and in the breast cancer cell lines MCF-7 and MDA-MB-231 [41]. In neither of these studies was CaR expression quantified nor were immunoblots of cell lysates shown, therefore it is not possible to determine whether malignant transformation altered either the level of CaR protein or its processing or modification. However, both the MCF-7 and MDA-MB-231 cells showed an increase in PTHrP secretion in response to CaR agonists, either Ca2+ above 3 mM or spermine or neomycin [41]. Thus, it would appear the CaR expressed in these breast cancer cell lines are still functional, and most interestingly, may contribute to some of the systemic effects of advanced breast cancer by stimulating an increase in secreted PTHrP, a major contributing factor to hypercalcemia of malignancy [19]. The surface epithelium of the ovary is contiguous with the mesothelial lining of the peritoneum, and thus the ovarian surface epithelial (OSE) cells are actually derived from a mesodermal lineage [1,2]. Cultured OSE cells display a
K.D. Rodland / Cell Calcium 35 (2004) 291–295
combination of epithelial and mesenchymal markers, and changes in the ratio of E-cadherin to N-cadherin expression are associated with the early characteristics of premalignant changes [1,33]. Interestingly, the biological response of cultured human, rat and rhesus OSE cells to changes in extracellular calcium levels resembles that of mesenchymally derived cells. Since the early days of tissue culture studies, it has been known that fibroblasts require high calcium media (1.8–2.0 mM CaCl2 ) for optimal proliferation, while epithelial cells proliferate in low calcium media and differentiate upon exposure to calcium levels above 1 mM [22,23]. In Rat-1 fibroblasts, this proliferative response is dependent upon the presence of functional CaR [36]. SV40-immortalized human OSE cells show a sharp increase in thymidine incorporation when shifted from 0.8 mM Ca2+ to 1.8 mM Ca2+ [35], and this proliferative response is dependent on CaR-mediated activation of the MAP kinase cascade and PI3 kinase [24]. CaR protein and mRNA expression is still evident in ovarian cancer cell lines [35]. However, the cancer cell lines have become insensitive to changes in extracellular calcium, displaying a constant (and high) rate of thymidine incorporation at extracellular calcium concentrations from 0.3 to 4.0 mM [35]. Thus, it would appear that malignant transformation of OSE cells is accompanied by a loss of sensitivity to the quiescence-inducing qualities of low Ca2+ , just as malignant transformation of intestinal epithelial cells is accompanied by loss of sensitivity to the differentiation-inducing effects of elevated Ca2+ in the intestinal crypts [27,28]. These observations suggest that the CaR, while neither a potent oncogene nor tumour suppressor, does play an important, and cell type-specific, role in modulating the balance between proliferation and differentiation in response to changes in extracellular calcium concentration. Malignant progression is accompanied by a loss of this normal homeostatic mechanism, regardless of whether the cell type-specific effect of CaR activation is proliferation (OSE cells) or differentiation (intestinal epithelial cells).
4. CaR function and PTHrP secretion in cancer Hypercalcemia of malignancy is a phenomenon that is consistently observed in several types of cancer, including prostate, breast, and bone cancers [5]. The proximal cause of hypercalcemia of malignancy is the increased circulating levels of parathyroid hormone related peptide, PTHrP [19,20]. Elevated levels of PTHrP also contribute to increased resorption of bone, and to the osteolysis observed in association with metastasis of epithelial-derived tumours to bone [20]. The potential role of the CaR in modulating PTHrP secretion has been investigated in breast cancer [41], prostate cancer [42], glial cell tumours [11], and Leydig cell tumours [9]. In each of these cases, the CaR has been shown to play an active role in stimulating the increased secretion of PTHrP. Use of polycationic agonists of the CaR such as
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neomycin and poly-spermine provoke dose-dependent secretion of PTHrP from astrocytomas, meningiomas [11], breast cancer cell lines [42], prostate cancer cell lines [43], and Leydig tumour cell lines. CaR mRNA has been identified in all of these cell types, and protein expression verified in the astrocytoma, breast cancer, prostate cancer, and Leydig cell tumour cell lines [9,11,41,42]. Moreover, in both breast and prostate, exposure of the cells to TGF prior to stimulation of the CaR causes an increase in both basal and CaR-mediated PTHrP release [41,42]. As TGF is released from bone during resorption, this positive interaction between CaR activation and TGF release provides a potential mechanism for a self-sustaining feedback loop that promotes continuous elevated bone resorption.
5. The CaR and cancer The primary function of the CaR is the regulation of homeostasis in response to changes in the extracellular concentration of polycationic small molecules, from calcium through polyarginine. Homeostasis can be defined as the balanced release of PTH, PTHrP, and calcitonin to maintain plasma calcium levels within tight physiological limits, or homeostasis can be defined as the appropriate balance between proliferation, differentiation, and apoptosis, depending on the overall cellular environment. Just as activation of the CaR stimulates PTH and PTHrP release while inhibiting calcitonin release, so activation of the CaR stimulates differentiation in most epithelial cells (intestinal, mammary and epidermal) while stimulating proliferation in mesenchymally-derived cells (fibroblasts and ovarian surface epithelial cells). The role of the CaR in neoplasia appears to be homeostatic as well; loss of normal CaR-induced response to extracellular calcium is observed in cancers of the intestine and ovary, while increased release of PTHrP is observed in cancers of the breast, prostate, Leydig cell, and glial cells. While disruption of CaR function may not be a primary event in carcinogenesis, disruption of CaR function clearly contributes to altered physiology of the neoplastic cell. Thus, therapeutic strategies directed at restoring normal CaR function may contribute to lessening the sequelae of cancer, particularly the hypercalcemia of malignancy, and could play a supporting role in cancer treatment.
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The role of the calcium-sensing receptor in cancer Karin D. Rodland Pacific Northwest National Laboratory, Biological Sciences Division, Richland, WA 99352, USA Received 20 October 2003; accepted 27 October 2003
Abstract The extracellular calcium-sensing receptor (CaR) is a versatile sensor of small, polycationic molecules ranging from Ca2+ and Mg2+ through polyarginine, spermine, and neomycin. The sensitivity of the CaR to changes in extracellular Ca2+ over the range of 0.05–5 mM positions the CaR as a key mediator of cellular responses to physiologically relevant changes in extracellular Ca2+ . For many cell types, including intestinal epithelial cells, breast epithelial cells, keratinocytes, and ovarian surface epithelial cells, changes in extracellular Ca2+ concentration over this range can switch the cellular behaviour from proliferation to terminal differentiation or quiescence. As cancer is predominantly a disease of disordered balance between proliferation, differentiation, and apoptosis, disruptions in the function of the CaR could contribute to the progression of neoplastic disease. Loss of the growth suppressing effects of elevated extracellular Ca2+ have been demonstrated in parathyroid hyperplasias and in colon carcinoma, and have been correlated with changes in the level of CaR expression. Activation of the CaR has also been linked to increased expression and secretion of PTHrP (parathyroid hormone-related peptide), a primary causal factor in hypercalcemia of malignancy and a contributor to metastatic processes involving bone. Although mutation of the CaR does not appear to be an early event in carcinogenesis, loss or upregulation of normal CaR function can contribute to several aspects of neoplastic progression, so that therapeutic strategies directed at the CaR could potentially serve a supportive function in cancer management. © 2003 Elsevier Ltd. All rights reserved. Keywords: Calcium-sensing receptor; Cancer; Cancer management
The decision of a cell to proliferate or quiesce, to differentiate or apoptosis, is the result of a regulated process of communication between the cell and its environment. One of the molecular hallmarks of cancer is the disruption of this normal regulatory process. Cells either fail to sense the appropriate environmental cues promoting differentiation over proliferation, or the response to those cues is inappropriate. Therefore, it is highly appropriate that, in the search for molecular mechanisms of cancer, each newly characterized receptor for environmental stimuli is examined for its potential role in carcinogenesis. The calcium-sensing receptor (CaR) is a seven transmembrane domain G-protein coupled receptor that was initially characterized as the sensor responsible for modulating parathormone and calcitonin release in response to changes in blood calcium levels [7]. However, the CaR is more than just a calcium sensor; it is a fairly broad spectrum sensor of small cationic molecules, capable of transducing signals in response to changes in the concentration of heavy metals, including lead and cadmium [21], as well as cationic amino acids [39]. Although plasma free calcium levels are tightly regulated, significant calcium gradients do exist in several physiological compartments, such as gut and skin (e.g. [37]), and these calcium gradients are known to af0143-4160/$ – see front matter © 2003 Elsevier Ltd. All rights reserved. doi:10.1016/j.ceca.2003.10.011
fect the behaviour of the resident cell types. It has been known for over two decades that human keratinocytes only proliferate in a very low calcium environment [22,23], and that even modest increase in calcium concentration trigger keratinocyte differentiation [44,47]. The crypts of the large intestine also display a gradient of calcium concentration, generally increasing from the base of the crypt to its lumen [4]. This calcium gradient corresponds with differences in cellular behaviour; proliferation of intestinal epithelial cells originates in the base of the crypts, and cells differentiate as they progress apically, eventually undergoing apoptosis at the distal end of the crypt [4,37].
1. Parathyroid adenomas and the CaR The primary site of CaR action is in the chief cells of the parathyroid gland, where the CaR translates increases in blood calcium concentrations into decreased secretion of parathormone (PTH) [7,8]. CaR protein and mRNA is more abundantly expressed in the parathyroid than in most other tissues, therefore it is logical to examine CaR expression in parathyroid adenomas. Several investigators have surveyed human parathyroid adenomas for changes in either the level
292
K.D. Rodland / Cell Calcium 35 (2004) 291–295
of CaR expression or for somatic mutation in the CaR gene [13,15,40]. In general, CaR protein levels appear to be decreased in parathyroid adenomas; at least three different groups have reported a decrease in CaR mRNA levels in parathyroid adenomas and hyperplasias [15,40,46]. Closer examination of mRNA transcripts has revealed an interesting discordance between the levels of two distinct CaR transcripts driven from distinct promoters. The CaR transcript which contains exon 1B is expressed at similar levels in normal and hyperplastic parathyroid; however, the exon 1A transcript is significantly suppressed in parathyroid adenomas [13]. At this point, no functional distinction between the two CaR transcripts is evident. However, this finding does suggest that CaR exon 1A is regulated differently than the exon 1B transcripts, and may be correlated with the ability of the CaR to inhibit proliferation in certain cell types such as the parathyroid [14] and intestinal epithelial cells [27]. Sequencing of the CaR from patients with hyperparathyroidism has led to the identification of a novel polymorphism that may be associated with increased activity of the CaR [3]. However, this polymorphism is also observed in normal tissues, and therefore is not likely to be causal for parathyroid hyperplasia. More extensive studies of genetic variability at the CaR locus in both parathyroid adenomas and pituitary tumours have failed to reveal any consistent link between mutation of the CaR and tumorigenesis [10].
2. Colon cancer and the CaR The colon carcinoma cell lines Caco-2 [26], HT-29 [16] and T-84 [16] express functional CaR and respond to addition of Ca2+ by releasing intracellular calcium. Kallay et al. [27] have shown that Caco-2 cells respond differentially to changes in Ca2+ depending on whether the stimulus was applied to basolateral or luminal surfaces. Reduction of Ca2+ to 0.025 mM on the luminal surface resulted in reduction of c-myc RNA expression, while a similar change confined to the basolateral surface had no effect [27]. These data correlate well with the observations that c-myc is a major factor regulating cell cycle progression in intestinal epithelial cells [26], and that proliferation of normal intestinal epithelial cells is highest in the base of crypts, were extracellular calcium concentrations are lowest [37]. Furthermore, immunohistochemical studies of human colon carcinomas indicated that CaR protein was most highly expressed in well-differentiated regions of the tumour and nearly lacking in poorly differentiated regions [29,43]. These data suggest that loss of a normal growth inhibitory response to elevated extracellular calcium, dependent on expression of functional CaR, may contribute to the progression of colon carcinomas. The possible linkage between loss of CaR function and cancer progression in the colon is particularly interesting in light of epidemiological data linking increased dietary calcium and Vitamin D with a decreased risk of colon cancer [17,31]. Thus, the CaR may be playing a physiologically
relevant role in regulating the proliferation and apoptosis of normal intestinal epithelial cells. Although it is highly unlikely that loss of CaR function is sufficient to initiate carcinogenesis in the colon or other organ sites, disruption of CaR function by dietary or other means may contribute significantly to tumour progression. Other epithelial cells may also respond to increases in extracellular calcium with a slowing of proliferation. Increased dietary calcium is associated with a decreased risk of intestinal carcinoma [25] and breast cancer [32]. Normal pancreatic islet cells express both CaR mRNA and protein [6,18,30], and functional CaR were observed in both insulinoma cells [30] and gastrinoma cells [18]. The level of CaR protein expression varied significantly within gastrinoma cells, and the correlation between levels of CaR expression and levels of gastrin release were not determined. However, it is possible that increased CaR expression may contribute to the increased sensitivity of both insulinoma cells and gastrinoma cells to extracellular calcium [18,30].
3. CaR and cancer of the breast and ovary Functional CaR are expressed by both normal mammary epithelial cells [12] and normal ovarian surface epithelial cells of the human and rhesus monkey [35,45]. Immunohistochemical analysis of normal breast showed membrane-associated CaR staining in ductal epithelial cells, as well as in the cells lining cysts in patients with fibrocystic disease of the breast [12]. This abundant expression of CaR protein is consistent with the role of the mammary gland in concentrating calcium in milk, as well as the known sensitivity of cultured primary human mammary epithelial cells to undergo terminal differentiation in high calcium media [34,38]. The possibility that the CaR may have a function in breast carcinoma is supported by the observation of CaR protein in patient samples from two cases of ductal carcinoma of the breast [12], and in the breast cancer cell lines MCF-7 and MDA-MB-231 [41]. In neither of these studies was CaR expression quantified nor were immunoblots of cell lysates shown, therefore it is not possible to determine whether malignant transformation altered either the level of CaR protein or its processing or modification. However, both the MCF-7 and MDA-MB-231 cells showed an increase in PTHrP secretion in response to CaR agonists, either Ca2+ above 3 mM or spermine or neomycin [41]. Thus, it would appear the CaR expressed in these breast cancer cell lines are still functional, and most interestingly, may contribute to some of the systemic effects of advanced breast cancer by stimulating an increase in secreted PTHrP, a major contributing factor to hypercalcemia of malignancy [19]. The surface epithelium of the ovary is contiguous with the mesothelial lining of the peritoneum, and thus the ovarian surface epithelial (OSE) cells are actually derived from a mesodermal lineage [1,2]. Cultured OSE cells display a
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combination of epithelial and mesenchymal markers, and changes in the ratio of E-cadherin to N-cadherin expression are associated with the early characteristics of premalignant changes [1,33]. Interestingly, the biological response of cultured human, rat and rhesus OSE cells to changes in extracellular calcium levels resembles that of mesenchymally derived cells. Since the early days of tissue culture studies, it has been known that fibroblasts require high calcium media (1.8–2.0 mM CaCl2 ) for optimal proliferation, while epithelial cells proliferate in low calcium media and differentiate upon exposure to calcium levels above 1 mM [22,23]. In Rat-1 fibroblasts, this proliferative response is dependent upon the presence of functional CaR [36]. SV40-immortalized human OSE cells show a sharp increase in thymidine incorporation when shifted from 0.8 mM Ca2+ to 1.8 mM Ca2+ [35], and this proliferative response is dependent on CaR-mediated activation of the MAP kinase cascade and PI3 kinase [24]. CaR protein and mRNA expression is still evident in ovarian cancer cell lines [35]. However, the cancer cell lines have become insensitive to changes in extracellular calcium, displaying a constant (and high) rate of thymidine incorporation at extracellular calcium concentrations from 0.3 to 4.0 mM [35]. Thus, it would appear that malignant transformation of OSE cells is accompanied by a loss of sensitivity to the quiescence-inducing qualities of low Ca2+ , just as malignant transformation of intestinal epithelial cells is accompanied by loss of sensitivity to the differentiation-inducing effects of elevated Ca2+ in the intestinal crypts [27,28]. These observations suggest that the CaR, while neither a potent oncogene nor tumour suppressor, does play an important, and cell type-specific, role in modulating the balance between proliferation and differentiation in response to changes in extracellular calcium concentration. Malignant progression is accompanied by a loss of this normal homeostatic mechanism, regardless of whether the cell type-specific effect of CaR activation is proliferation (OSE cells) or differentiation (intestinal epithelial cells).
4. CaR function and PTHrP secretion in cancer Hypercalcemia of malignancy is a phenomenon that is consistently observed in several types of cancer, including prostate, breast, and bone cancers [5]. The proximal cause of hypercalcemia of malignancy is the increased circulating levels of parathyroid hormone related peptide, PTHrP [19,20]. Elevated levels of PTHrP also contribute to increased resorption of bone, and to the osteolysis observed in association with metastasis of epithelial-derived tumours to bone [20]. The potential role of the CaR in modulating PTHrP secretion has been investigated in breast cancer [41], prostate cancer [42], glial cell tumours [11], and Leydig cell tumours [9]. In each of these cases, the CaR has been shown to play an active role in stimulating the increased secretion of PTHrP. Use of polycationic agonists of the CaR such as
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neomycin and poly-spermine provoke dose-dependent secretion of PTHrP from astrocytomas, meningiomas [11], breast cancer cell lines [42], prostate cancer cell lines [43], and Leydig tumour cell lines. CaR mRNA has been identified in all of these cell types, and protein expression verified in the astrocytoma, breast cancer, prostate cancer, and Leydig cell tumour cell lines [9,11,41,42]. Moreover, in both breast and prostate, exposure of the cells to TGF prior to stimulation of the CaR causes an increase in both basal and CaR-mediated PTHrP release [41,42]. As TGF is released from bone during resorption, this positive interaction between CaR activation and TGF release provides a potential mechanism for a self-sustaining feedback loop that promotes continuous elevated bone resorption.
5. The CaR and cancer The primary function of the CaR is the regulation of homeostasis in response to changes in the extracellular concentration of polycationic small molecules, from calcium through polyarginine. Homeostasis can be defined as the balanced release of PTH, PTHrP, and calcitonin to maintain plasma calcium levels within tight physiological limits, or homeostasis can be defined as the appropriate balance between proliferation, differentiation, and apoptosis, depending on the overall cellular environment. Just as activation of the CaR stimulates PTH and PTHrP release while inhibiting calcitonin release, so activation of the CaR stimulates differentiation in most epithelial cells (intestinal, mammary and epidermal) while stimulating proliferation in mesenchymally-derived cells (fibroblasts and ovarian surface epithelial cells). The role of the CaR in neoplasia appears to be homeostatic as well; loss of normal CaR-induced response to extracellular calcium is observed in cancers of the intestine and ovary, while increased release of PTHrP is observed in cancers of the breast, prostate, Leydig cell, and glial cells. While disruption of CaR function may not be a primary event in carcinogenesis, disruption of CaR function clearly contributes to altered physiology of the neoplastic cell. Thus, therapeutic strategies directed at restoring normal CaR function may contribute to lessening the sequelae of cancer, particularly the hypercalcemia of malignancy, and could play a supporting role in cancer treatment.
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