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Abstract

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. CASE REPORT
  5. MATERIALS AND METHODS
  6. RESULTS
  7. DISCUSSION
  8. Acknowledgements
  9. References

A patient with classic clinical and biochemical features of tumor-induced osteomalacia (hypophosphatemia, phosphaturia, and undetectable serum concentrations of 1,25-dihydroxyvitamin D [1,25(OH)2D]) was studied before and after resection of a benign extraskeletal chondroma from the plantar surface of the foot. Presurgical laboratory evaluation was notable for normal serum concentrations of calcium, intact parathyroid hormone (PTH), parathyroid hormone-related protein (PTHrP), and osteocalcin, increased serum alkaline phosphatase activity, and frankly elevated urinary cyclic adenosine monophosphate (cAMP) and pyridinium cross-link excretion. Quantitative histomorphometry showed severe osteomalacia and deep erosions of the cancellous surface by active osteoclasts. After resection, serum 1,25(OH)2D normalized within 24 h, while renal tubular phosphorus reabsorption and serum phosphorus did not normalize until days 2 and 3, respectively; serum Ca declined slightly, and serum intact PTH, osteocalcin, and urinary pyridinium cross-link excretion increased dramatically. Urinary cAMP excretion declined immediately after resection and then began to increase concomitant with the increase in serum intact PTH. A second bone biopsy taken 3 months after resection demonstrated complete resolution of the osteomalacia, increased mineral apposition rate (1.09 μ/day), resorption surface (9.2%), mineralizing surface (71%), and bone formation rate (0.83 mm3/mm2/day), and marked decreases in cancellous bone volume (13.1%) and trabecular connectivity compared with the first biopsy. Tumor extracts did not affect phosphate transport in renal epithelial cell lines or 1α-hydroxylase activity in a myelomonocytic cell line. The patient's course suggests that the abnormal 1,25(OH)2D and phosphorus metabolism is due to a tumor product that may be acting via stimulation of adenylate cyclase activity. Increased bone resorption prior to surgical resection suggests that the tumor may also produce an osteoclast activator. The rise in resorption surface and pyridinium cross-link excretion, increase in serum osteocalcin and bone mineralization, normalization of osteoid width, and fall in cancellous bone volume after resection are consistent with healing of osteomalacia by rapid remodeling.


INTRODUCTION

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. CASE REPORT
  5. MATERIALS AND METHODS
  6. RESULTS
  7. DISCUSSION
  8. Acknowledgements
  9. References

TUMOR-INDUCED OSTEOMALACIA is a rare disorder in which rickets or osteomalacia is associated with a tumor. Since the syndrome was first described by McCance in 1947,1 approximately 102 patients have been reported with this disease.2,3 Affected individuals are frequently afflicted with severe osteomalacia manifested by bone pain, fractures, and severe proximal muscle weakness. The syndrome is characterized biochemically by hypophosphatemia, renal phosphate wasting, and low circulating concentrations of 1,25-dihydroxyvitamin D (1,25(OH)2D), biochemical alterations that contribute to defective mineralization of osteoid. Successful resection of the tumors results in prompt resolution of the biochemical abnormalities and subsequent healing of the osteomalacia. A great deal of evidence supports the hypothesis that the tumors secrete one or more substances that interfere with normal renal phosphate handling and with the conversion of 25-hydroxyvitamin D (25(OH)D) to 1,25(OH)2D. However, the precise mechanisms by which these functions are altered remains unclear.

We had the opportunity to study a patient with tumor-induced osteomalacia. Similar to a case we reported previously,4 the tumor was of mesenchymal origin and was located on the plantar surface of the foot. A series of studies, documenting the biochemical, densitometric, and histomorphometric response to resection of this tumor, are the subject of this report. We also evaluated the effects of the patient's serum and of an extract prepared from the tumor on phosphate transport in both parathyroid hormone (PTH)-responsive and nonresponsive renal epithelial cell lines and the effects of the tumor extract on 25-hydroxyvitamin D-1α-hydroxylase activity.

CASE REPORT

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. CASE REPORT
  5. MATERIALS AND METHODS
  6. RESULTS
  7. DISCUSSION
  8. Acknowledgements
  9. References

A 21-year-old man presented for evaluation of diffuse skeletal pain and inability to walk. The patient had been in excellent health until age 15 when he noted the onset of dull pain in his knees which was exacerbated by weightbearing. Over the next 6 years, there was gradual progression of the pain to involve the ankles, feet, and hips, and progressive difficulty with ambulation. At presentation, the patient was noted to have kyphosis, a wide-based antalgic gait, and signs of severe proximal muscle weakness.

A biochemical evaluation (Table 1) was noteworthy for severe hypophosphatemia associated with phosphaturia and reduced tubular reabsorption of phosphorus, undetectable serum 1,25(OH)2D, increased serum alkaline phosphatase (ALP) activity and urinary cyclic adenosine monophosphate (cAMP) excretion. The serum 25(OH)D level was at the low end of the normal range. Bone resorption markers, urinary hydroxyproline, and pyridinium cross-link excretion were elevated. The remainder of the chemistry profile was within normal limits, including the serum calcium, intact PTH, osteocalcin, and PTH-related protein (PTHrP) levels.

Table Table 1. SERUM AND URINE BIOCHEMISTRIES IN A PATIENT WITH TUMOR-INDUCED OSTEOMALACIA PRIOR TO TUMOR RESECTION
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Radiographs revealed diffuse osteopenia, coarsened trabecular markings, thoracic scoliosis, multiple rib and vertebral compression fractures, and bilateral insufficiency fractures of the femoral neck, triradiate pelvis, and delayed epiphyseal closure. Bone mineral density (BMD) by dual-energy X-ray absorptiometry (DEXA) showed markedly diminished BMD of the lumbar spine, femoral neck, and nondominant forearm. A transiliac crest bone biopsy, performed after double labeling with tetracycline antibiotics, demonstrated severe osteomalacia (Fig. 1A, B, C; Table 2). Cancellous bone volume was normal, although the trabeculae consisted predominantly of unmineralized osteoid, and intertrabecular connectivity was decreased. Active osteoblasts were absent. The extent of eroded surface was normal but erosions were abnormally deep, with active osteoclasts present in deep erosion pits.

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Figure FIG. 1. Magnetic resonance scanning of the right foot. A 4 × 2 × 2.5 cm mass, with signal characteristics of a fibrous lesion, abutts the proximal plantar fascia and slightly indents the adjacent musculature.

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Table Table 2. QUANTITATIVE HISTOMORPHOMETRY IN TUMOR-INDUCED OSTEOMALACIA BEFORE AND AFTER TUMOR RESECTION
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A diagnosis of hypophosphatemic osteomalacia due to a tumor or a sporadic form of X-linked hypophosphatemia (XLH) was made. An ill-defined soft tissue mass was detected on the plantar aspect of the right foot. Magnetic resonance scanning demonstrated a 4 × 2 × 2.5 cm mass, with signal characteristics of a fibrous lesion, that abutted and slightly breached the proximal plantar fascia, indenting the adjacent musculature (Fig. 2).

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Figure FIG. 2. Transiliac crest bone biopsies. (A, B, C) One month before surgical resection. The presurgical biopsy demonstrates marked hyperosteoidosis in (A) cortical and (B) cancellous bone. Tunneling erosion of mineralized bone undermining osteoid can be seen in (A, arrow) and disconnectivity of trabecular structure in (B). Goldner trichrome stain: Osteoid is stained orange-red and mineralized bone is stained green. (C) demonstrates absence of tetracycline uptake on unstained bone surfaces. Final magnifications: (A, B) 30.4×; (C) 76.6×. (D, E, F) Three months after surgical resection. There has been dramatic resolution of excess osteoid. The osteoid seams are now of normal thickness in (D), a low-power photomicrograph of cortical and cancellous bone, and in magnified trabecula (E). Severe loss of connectivity is apparent in (D). Erosion of bone surface (long arrows) and plump osteoblasts covering osteoid seams are visible in (E, short arrows). Golder trichrome stain. (F) Marked uptake of tetracycline with extensive double labels covering unstained cortical bone surfaces. Final magnifications: (D) 15.2×; (E, F) 76.6×.

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Resection of the lesion revealed a multilobulated, highly vascular tumor, comprised of hypervascular, cellular, and cartilaginous components (Fig. 3). The predominant component of the lesion consisted of irregularly distributed nodules of immature chondroid with a granuloma-like appearance. The nodules were centrally myxoid with hypercellular margins and showed early signs of calcification. The vascular component formed stalks that separated chondroid nodules and was composed of numerous slit-like capillaries surrounded by small, uniform spindle cells without cellular atypia. Multinucleated giant cells were irregularly distributed within the stalks. Mitoses were rare. Blood-filled channels with frequent zones of hemorrhage were observed. The lesion was interpreted as an extraskeletal chondroma, probably benign.

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Figure FIG. 3. Extraskeletal chondroma. Photomicrograph of tissue section stained with Masson trichrome. Nodules of immature chondroid (stained blue) are separated by vascular and cellular tissue (stained red) containing numerous small uniform cells, multinucleated giant cells (white arrowheads) and numerous capillaries (white arrows). Final magnification 120×.

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MATERIALS AND METHODS

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. CASE REPORT
  5. MATERIALS AND METHODS
  6. RESULTS
  7. DISCUSSION
  8. Acknowledgements
  9. References

Serum calcium, phosphorus, and albumin were measured by standard automated techniques (Technicon Instruments, Tarrytown, NY, U.S.A.). All serum calcium concentrations were corrected for serum albumin.5 Serum osteocalcin was measured by radioimmunoassay6,7 and intact PTH(1–84) by immunoradiometric assay.8 Serum 25(OH)D and 1,25(OH)2D were measured after extraction, by radiobinding9 and radioreceptor assays,10 respectively. PTHrP was measured by a two-site immunoradiometric assay.11 Urinary calcium, creatinine, and phosphorus were measured by standard automated techniques. The maximal capacity of the renal tubules to reabsorb phosphate (TmPO4/GFR) was calculated by the method of Bijvoet.12 Hydroxyproline and pyridinium cross-link excretion were measured as previously described13,14 and total cAMP excretion by radioimmunoassay (NEN Life Science Products, Boston, MA, U.S.A.).

BMD of the lumbar spine, right proximal femur, and nondominant forearm were measured by DEXA utilizing a QDR-1000 Bone Densitometer (Hologic, Inc., Waltham, MA, U.S.A.) and expressed in grams per square centimeter and as T scores which compare individual BMD determinations to those of a young normal population of the same gender.15 In our laboratory, the in vitro reproducibility of this machine using an anthropomorphic spine phantom is 0.51%. The short-term in vivo coefficient of variation in a group of middle-aged women with low bone mass is 0.68% for the lumbar spine (L2–L4) and 1.36% for the femur.

Percutaneous transiliac crest bone biopsies were performed 1 month prior to and 3 months after tumor resection following double labeling with tetracycline and processed without decalcification. Nonserial sections (7 μ thick) were cut, stained, and analyzed using standard histomorphometric techniques and nomenclature recommended by the American Society of Bone and Mineral Research.16 The patient's biopsy was compared with previously published results in normal men.17,18

Extraction of protein from tumor and normal human spleen was accomplished by homogenization of 500 mg of tissue in 2 ml of acid/ethanol solution (25 ml of 95% ethanol/500 ml of concentrated HCl) according to the method of Roberts et al.19 Recovery of PTH-like bioactivity was assessed by the addition of human PTH(1–84) to a sample of spleen tissue at the homogenization step. The effect of this tumor extract on sodium-dependent phosphate transport was assessed in both PTH-sensitive (OK/E)20 and PTH-insensitive (LLCPK-1)21 renal epithelial cell lines as described previously.22 Briefly, confluent layers of quiescent OK/E and LLCPK-1 cells were stimulated for 4 h with tumor or spleen extract (20–480 ng of protein) prior to increasing the rate of uptake of32P over a 5-minute period. In addition, serial dilutions (0.5–10%) of presurgical serum from the patient were compared with control serum alone and control serum to which bovine PTH(1–34) had been added to a final concentration of 10−10 M, with respect to their ability to influence phosphate transport in the PTH-sensitive OK/E renal cell line. To inactivate complement, serum samples were first heated to 56°C for 30 minutes then diluted in HEPES buffered Dulbecco's modified Eagle's medium (DMEM). Finally, the effect of multiple dilutions of tumor extract (15–150 μg of protein) upon 1α-hydroxylase activity was evaluated in a myelomonocytic cell line as previously described.23 Briefly, monolayer cultures of the v-myc–transformed chicken myelomonocytic cell line HD-11 were preincubated for 16 h in serum-free DMEM or Tris buffer and then exposed to unlabeled 25(OH)D (30 nM, solubilized in 0.01% ethanol) in the presence or absence of tissue extract for 3 h. The cells and conditioned medium were subjected to lipid extraction, chromatography, and assay of 1,25(OH)2D-like metabolites according to the method of Reinhardt et al.10 Metabolite yield in lipid extracts was determined by open-column sample purification and quantitation by 1,25(OH)2D-receptor (VDR) binding analysis. Extracts from normal human spleen and mouse liver were used as controls.

RESULTS

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. CASE REPORT
  5. MATERIALS AND METHODS
  6. RESULTS
  7. DISCUSSION
  8. Acknowledgements
  9. References

Tumor resection resulted in rapid normalization of serum 1,25(OH)2D, serum phosphorus levels, and TmPO4/GFR (Fig. 4). Serum 1,25(OH)2D had reached the midnormal range (31 pg/ml) by 24 h and had become supranormal (101 pg/ml) by 48 h. TmPO4/GFR rose into the normal range on the second day and serum phosphorus normalized on the third day. These biochemical changes were not associated with any change in serum intact PTH (data not shown).

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Figure FIG. 4. Serum 1,25(OH)2D and phosphorus and renal tubular phosphorus reabsorption after tumor resection. Shaded areas represent the normal ranges for each parameter.

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Throughout the 8-month observation period after tumor resection (Fig. 5), serum 1,25(OH)2D remained elevated. By week 3, both serum phosphorus and TmPO4/GFR were also elevated. Serum phosphorus peaked (6.2 mg/dl) by week 5 and remained above normal for 4 months. Despite supplementation with elemental calcium (2 g/day) and vitamin D (400 IU/day), serum calcium fell below normal during week 2, and thereafter remained in the low-normal range. This pattern was mirrored by the decline and subsequent recovery of urinary calcium excretion to the low-normal range. Serum intact PTH, repeatedly normal preoperatively, began to rise by day 4 (50 pg/ml), peaked by week 2, and remained elevated for 3 months. Urinary cAMP excretion, which was frankly elevated preoperatively despite normal serum PTH, fell to the upper normal range immediately after tumor resection and then began to increase again concomitant with rising serum intact PTH levels.

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Figure FIG. 5. Mineral homeostasis after tumor resection. Shaded areas represent the normal ranges for each parameter.

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Bone resorption markers,24 which were frankly elevated preoperatively, rose dramatically (Fig. 6), reaching peak values 30- to 40-fold above normal by the end of week 3 (pyridinoline, 2230 nmol/mmol creatinine; deoxypyridinoline, 680 nmol/mmol creatinine) and gradually declining thereafter. These increases were followed sequentially by further increases in serum ALP activity, which had peaked (494 U/I) by week 4. In contrast, serum osteocalcin was normal preoperatively and did not increase until 2 weeks after resection. All four indexes of bone turnover remained elevated for the entire observation period.

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Figure FIG. 6. Markers of bone turnover after tumor resection. Shaded areas represent the normal ranges for each parameter.

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A second transiliac bone biopsy taken from the opposite iliac crest 3 months after tumor resection demonstrated dramatic improvement in hyperosteoidosis (Table 2; Fig. 1D, E, F). Although osteoid surface remained increased, osteoid width had normalized and plump active osteoblasts covered most of the osteoid seams. Tetracycline uptake had improved markedly, as shown by the elevated mineralizing surface and mineral apposition rate. Eroded surface had also increased and there were deep erosions, perforation of trabecular plates, and a marked decrease in cancellous bone volume (22.4–13.1%). Although this difference is within the range of intraindividual variability, it is of note that trabecular plates were thinner and showed marked loss of connectivity in the second biopsy. However, BMD, measured twice during the 5 months after tumor resection, increased markedly (Fig. 7). Lumbar spine BMD rose by 78% from 0.53 to 0.81 g/cm2, femoral neck BMD increased from 0.21 to 0.98 g/cm2, and total hip BMD increased from 0.32 to 0.78 g/cm2. In contrast, radial bone mass did not change appreciably.

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Figure FIG. 7. Bone mineral density after tumor resection.

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The effect of the heat-treated serial dilutions of patient serum on phosphate transport was evaluated in the PTH-sensitive OK/E renal cell line (Table 3). As expected, bPTH(1–34) inhibited phosphate transport in this assay.22 Due to the presence of growth factors, serum will stimulate phosphate transport in this assay, and we did observe such an effect. Serum from the patient, however, stimulated phosphate transport to a greater extent than control serum at each concentration assessed. Stimulation of either PTH-sensitive (OK/E) or PTH-insensitive (LLCPK-1) renal cell lines with tumor or normal spleen extracts (20–480 μg of protein) had no significant effect on phosphate transport (data not shown), while a spleen homogenate containing 10−9 M bPTH(1–34) elicited the expected reduction in phosphate transport when assayed at a protein concentration of 120 μg. In a similar manner, tumor and spleen extracts (15–150 μg of protein) failed to inhibit 1α-hydroxylase activity in a myelomonocytic cell line (data not shown).

Table Table 3. EFFECT OF HEAT-TREATED SERUM ON PHOSPHATE TRANSPORT IN PTH-SENSITIVE OK/E RENAL CELL LINE
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DISCUSSION

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. CASE REPORT
  5. MATERIALS AND METHODS
  6. RESULTS
  7. DISCUSSION
  8. Acknowledgements
  9. References

This patient's clinical and biochemical presentation was typical of tumor-induced osteomalacia1–3 in an adolescent. Resection of a benign extraskeletal chondroma resulted in prompt resolution of the biochemical abnormalities and rapid healing of the osteomalacia. The biochemical responses to tumor resection in this patient were consistent with the generally held view that such tumors produce a humoral substance (or substances) that inhibits renal tubular reabsorption of phosphorus and conversion of 25(OH)D to 1,25(OH)2D.

The majority of cases of tumor-induced osteomalacia are caused by mesenchymal tumors of various types, including giant-cell tumors of bone. Recent observations suggest that tumor-induced osteomalacia may also occur in association with a variety of other tumors, including breast, prostate, small cell carcinoma of the lung, multiple myeloma, and chronic lymphocytic leukemia and also in association with widespread fibrous dysplasia of bone, neurofibromatosis, and linear nevus sebaceous syndrome.2,3 The connective tissue tumors, which may be small and inconspicuous, have recently been classified by Weidner and Cruz.25 The largest category, namely the primitive-appearing, mixed connective tissue tumors, accounts for approximately 80% of cases and likely includes the tumor in this patient. These tumors consist of sheets of primitive stromal cells admixed with a proliferating vascular component containing giant cells, and in some cases, multiple foci of immature cartilage, osteoid, and bone. Three other categories, osteoblastoma-like tumors, nonossifying fibroma-like tumors, and ossifying fibroma-like tumors, occur in and morphologically resemble bone.

Serum PTH levels were repeatedly normal in this patient, as has been the case in most,2,3 although not all, cases26,27 of tumor-induced osteomalacia. Despite the normal serum concentrations of both PTH and PTHrP, urinary cAMP excretion was not only frankly elevated before tumor resection but fell promptly to the upper end of the normal range immediately afterward. Three other patients with tumor-induced osteomalacia and elevated urinary cAMP have been reported,28–30 one with coexistent hyperparathyroidism.30 The association of elevated urinary cAMP excretion with normal serum PTH and PTHrP suggests that the tumor products may have been acting via stimulation of adenylate cyclase. This observation is notable in view of the recent report of a factor, partially purified from a sclerosing hemangioma, that inhibited renal phosphate transport independent of cAMP.31 Other possible explanations for the increased urinary cAMP excretion in our patient include elevated calcitonin (which was not measured), that normal serum PTH may not have reflected its biological activity, or that sensitivity of target tissues to PTH was increased.

There was considerable evidence for increased bone resorption in this patient prior to tumor resection. Although quantitative histomorphometry revealed that the extent of resorption surface was not increased as in our previously reported patient,4 there were deep erosion pits under the osteoid; the pits contained osteoclasts which perforated osteoid seams, burrowed into the trabeculae, resorbing the core of mineralized bone, and leaving the trabeculae hollowed out. In addition, urinary excretion of pyridinium cross-links and hydroxyproline, which to our knowledge have not been previously reported in tumor-induced osteomalacia, were markedly elevated. Pyridinoline and deoxypyridinoline excretion reflect degradation of mature collagen24 and are considered to be specific markers of bone resorption. The elevated excretion of these markers prior to tumor resection could represent resorption of mineralized bone, unmineralized osteoid, or both. The increased bone resorption was unlikely to be due to PTH and PTHrP in view of their normal circulating concentrations. Another conceivable explanation includes the possibility that the tumor also produced a factor that stimulates osteoclast activity. Alternatively, there may have been a direct effect of hypophosphatemia to mobilize calcium from bone, as has been reported in early studies.32,33

Tumor resection was followed by hypocalcemia secondary to rapid remineralization and secondary hyperparathyroidism that probably developed in response to the rapid deposition of calcium into osteoid. This postoperative hyperparathyroidism in some respects resembled the bone hunger often observed after successful parathyroidectomy in patients with severe primary or secondary hyperparathyroidism. After resection, excretion of bone resorption markers increased dramatically. By the second biopsy, only 3 months later, the osteomalacia had completely resolved and the resorption surface was frankly elevated. The increase in remodeling was probably driven by the secondary increase in PTH. The decrease in cancellous bone volume and connectivity observed on the second biopsy suggests that the unmineralized osteoid was at least partially resorbed rather than completely mineralized. It is possible that the inability of osteoclasts to resorb osteoid under normal conditions may have been overcome by the marked increase in serum PTH concentrations, as previously described.34 However, it should be noted that although cancellous bone volume was lower in the second biopsy, the amount of mineralized bone increased from 6.4 to 11.6%, consistent with the increase in BMD during the same period.

The discrepancy between serum ALP activity (elevated) and serum osteocalcin (normal) prior to tumor resection differs from a recent report in which both were increased.35 After resection, ALP activity rose further before it began to decline while serum osteocalcin did not increase until after ALP activity started to fall. It is well-established that serum osteocalcin does not move in parallel with total or bone-specific ALP activity in some metabolic bone diseases. This may be because ALP expression appears before expression of osteocalcin in osteoblast differentiation36–38 or because serum osteocalcin may reflect mineralization rather than synthesis of osteoid. However, it is also possible that the low serum osteocalcin concentrations observed prior to tumor resection in this patient could be related to the extremely low concentrations of 1,25(OH)2D which controls synthesis of osteocalcin by binding of 1,25(OH)2D to a vitamin D response element in the osteocalcin gene. Thus, inhibition of renal 25-hydroxyvitamin D-1α-hydroxylase activity by a tumor product may secondarily affect osteoblast synthesis of osteocalcin. Alternatively, it is possible that tumors causing tumor-induced osteomalacia may produce factors that directly inhibit osteocalcin production by osteoblasts. In this regard, a patient with tumor-induced osteomalacia has been reported with low serum osteocalcin concentrations that did not increase after administration of 4 μg of 1,25(OH)2D, but rose into the normal range after tumor resection.39 Conditioned medium from both the original tumor grown in cell culture and SV-40–transformed cells inhibited 1,25(OH)2D-stimulated osteocalcin production in a human osteoblastic cell line.39

While the precise pathogenesis of the hypophosphatemia levels in tumor-induced osteomalacia has not been elucidated completely, considerable evidence points to production by such tumors of one or more humoral factors that inhibit renal phosphate reabsorption. Tumors from several patients with tumor-induced osteomalacia have been shown to stimulate phosphaturic activity when injected into animals.40 Hypophosphatemia and phosphaturia have been shown to develop in athymic nude mice heterotransplanted with tumors41,42 from affected patients. Conditioned medium from a cultured desmoid tumor inhibited phosphate reabsorption in opossum kidney cells.43 Recently, Cai et al.31 demonstrated that supernatant from cultured sclerosing hemangioma cells inhibited phosphate transport in opossum kidney cells through a cAMP-independent mechanism. Partial purification of the factor revealed it to be heat-labile, with a molecular weight between 8000 and 25,000 D. The factor, which has been called “phosphatonin,” bore immunologic similarity to PTH but was neither PTH nor PTHrP. Its activity was not blocked by a PTH receptor antagonist, and interestingly, in view of our results, did not affect skeletal mobilization of calcium or phosphorus or influence bone remodeling. In contrast, we did not observe an effect of this patient's tumor extract on phosphate transport in either PTH-responsive or PTH-unresponsive renal cell lines. Activity was unlikely to have been lost during processing because the procedure was designed for extraction of small peptide growth factors,19 and PTH(1–84) activity was well preserved. It is possible that supernatants from cultured tumor cells rather than tumor extracts are more likely to demonstrate such activity. An alternative explanation for the lack of bioactivity could be that the biologic effects of the tumor factor on phosphate transport may be time dependent. Longer incubation (16–48 h) of the cell cultures with tumor extract might have been associated with inhibition of phosphate transport, similar to the report of Lajeunesse et al. of the effects of Hyp mouse serum in this system.44 In this regard, however, an effect of medium from the sclerosing hemangioma cultures reported by Cai et al.31 to inhibit sodium-dependent phosphate transport in opossum kidney cells was apparent after a 3-h incubation period (R. Kumar, personal communication).

Unlike bPTH(1–34), which elicited the expected inhibition of phosphate transport in OK/E cells, serial dilutions of serum from the patient produced an anomalous, dose-dependent stimulation of phosphate transport compared with control serum under the assay conditions employed. These results are in agreement with those of Tenenhouse and Martel who observed similar stimulation of phosphate transport by normal and Hyp mouse serum under identical assay conditions.45 While we cannot explain these observations, it is possible that the heat treatment necessary to inactivate complement could have destroyed the factor(s) which other investigators have shown to be heat-labile.31,42 Alternately, this young man with an immature skeleton and delayed epiphyseal closure may have had increased levels of IGF-1, a known stimulant of renal phosphate reabsorption.

Low serum 1,25(OH)2D concentrations have been recognized as part of tumor-induced osteomalacia since the first observation of this association by Drezner et al.46 In this regard, tumors causing tumor-induced osteomalacia have been shown to inhibit renal 25-hydroxyvitamin D-1α-hydroxylase activity in heterotransplanted athymic nude mice47 and in renal cell cultures exposed to tumor extracts.41 In contrast to these observations, we did not detect any effect of serial dilutions of tumor extract from this patient on 1α-hydroxylase activity.

Recent progress in understanding the physiology of both renal phosphate handling and X-linked hypophosphatemia (XLH)48–50 have provided insights into the pathophysiology of tumor-induced osteomalacia. XLH, an inherited disorder of phosphate homeostasis, biochemically resembles tumor-induced osteomalacia.50 Studies in the murine homolog of XLH have identified a specific defect in a sodium-phosphorus cotransporter in the proximal renal tubule as the cause of phosphaturia and hypophosphatemia. The mutant gene in patients with XLH has recently been identified and is designated PEX (a phosphate-regulating gene with homologies to endopeptidases located on the X chromosome).51 The predicted PEX gene product exhibits homology to a family of neutral endopeptidases involved in activation or degradation of peptide hormones. It has been speculated that PEX is involved in the inactivation or processing of a peptide hormone that affects renal phosphate handling and that loss of PEX results in either decreased production of a phosphate conserving hormone or decreased clearance of a phosphaturic hormone.51 Moreover, recent data suggest that the defect in renal phosphate transport in XLH is not intrinsic to the kidney but is secondary to a circulating hormone or factor, which is as yet unidentified.45,52–54 Prompted by the similarities between tumor-induced osteomalacia and XLH, we examined PEX expression in tumor tissue from this patient and found it to be increased.55 This observation suggests that the physiological function of PEX is the inactivation of a factor that promotes phosphaturia, present in excess in patients with tumor-induced osteomalacia. Whether the factors that cause tumor-induced osteomalacia and XLH are the same or similar remains to be established.

In conclusion, our results confirm those previously published in patients with tumor-induced osteomalacia with respect to increased renal phosphate excretion and impaired renal production of 1,25(OH)2D that resolve promptly after tumor resection. Additionally, despite normal PTH and PTHrP concentrations, both urinary cAMP excretion and osteoclastic bone resorption were increased prior to tumor resection, suggesting that, in this patient, the factor stimulated activity of both the adenylate cyclase enzyme and osteoclasts. This patient also provides insight into both the rapidity and the processes by which tumor-induced osteomalacia heals after tumor resection. Further research is essential to identify the factor(s) causing tumor-induced osteomalacia and their relationships both to the physiology of normal phosphorus homeostasis and to the pathophysiology of XLH.

Acknowledgements

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. CASE REPORT
  5. MATERIALS AND METHODS
  6. RESULTS
  7. DISCUSSION
  8. Acknowledgements
  9. References

The authors thank Dr. Harriet S. Tenenhouse for the OK/E cell line and Dr. David Goltzman for the LLCPK-1 cell line used in these studies, and Michele Schnitzer Rosenberg for expert technical assistance with preparation of the bone biopsy specimens. This work was supported in part by Grants RR-006645 and AR-39191 from the National Institutes of Health, by the Medical Research Council of Canada (MRCC) and Fonds de la Recherche en Sante du Quebec (FRSQ). J.E.H. is Chercheuse Boursiere of the FRSQ and ACK holds a scholarship award from the MRCC.

References

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. CASE REPORT
  5. MATERIALS AND METHODS
  6. RESULTS
  7. DISCUSSION
  8. Acknowledgements
  9. References
  • 1
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  • 2
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  • 3
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