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Clinical Chemistry 44: 1621-1628, 1998;
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(Clinical Chemistry. 1998;44:1621-1628.)
© 1998 American Association for Clinical Chemistry, Inc.

Enzymes and Protein Markers

Differences of bone alkaline phosphatase isoforms in metastatic bone disease and discrepant effects of clodronate on different skeletal sites indicated by the location of pain

Per Magnusson1,a, Lasse Larsson1, Gunnar Englund2, Brita Larsson1, Peter Strang3, and Lena Selin-Sjögren4

1 Bone and Mineral Metabolic Unit, Division of Clinical Chemistry, Department of Biomedicine and Surgery, Linköping University Hospital, S-581 85 Linköping, Sweden.

2 Department of Mathematical Statistics, Royal Institute of Technology, S-100 44 Stockholm, Sweden.

3 Division of Oncology, Department of Biomedicine and Surgery, Linköping University Hospital, S-581 85 Linköping, Sweden.

4 Clinical Research, Medical Department, Astra Läkemedel AB, S-151 85 Södertälje, Sweden.
a Author for correspondence. Fax 46-13-223240; e-mail Per.Magnusson{at}klk.liu.se.


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
We compared clodronate with placebo administration in 42 primarily or secondarily hormone-refractory prostate cancer patients with skeletal metastases and persisting pain. Serum total alkaline phosphatase (ALP), bone ALP isoforms, osteocalcin, cross-linked carboxy-terminal telopeptide of type I collagen, and prostate-specific antigen were analyzed before and after 1 month of treatment. Six ALP isoforms were quantified by HPLC: one bone/intestinal, two bone (B1, B2), and three liver ALP isoforms. The most apparent difference compared with healthy males was observed for the bone ALP isoform B2. Patients and healthy males had a B2 activity corresponding to 75% and 35% of the total ALP activity, respectively (P <0.0001). We propose that the different bone ALP isoforms reflect different stages of osteoblast differentiation during the extracellular matrix maturation phase of osteogenesis. All bone markers except osteocalcin increased after 1 month of clodronate administration. These increases were associated with pain only in the upper part of the body. We suggest that the uptake of clodronate by the skeleton was not uniform during our treatment period.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
Patients with breast, lung, and prostate cancer are particularly prone to develop metastatic bone disease. Most of the prostate cancer patients, >60%, will develop skeletal metastases at some stage of the disease; consequently bone pain is a major problem in these patients (1)(2). The recent development of new biochemical markers of bone turnover has generated interest to use these markers for early detection of bone metastases and to assess efficacy and the response to antiresorptive treatment for patients with metastatic bone disease (3)(4)(5)(6). Two collagen markers of bone turnover, cross-linked carboxy-terminal telopeptide of type I collagen (ICTP)1 and carboxy-terminal propeptide of type I procollagen, give some information about the type and activity of bone metastases; moreover, ICTP has been reported to have a prognostic value in prostate cancer (7). Serum alkaline phosphatase (ALP, EC 3.1.3.1) and tartrate-resistant acid phosphatase (EC 3.1.3.2) are increased in prostate cancer patients with bone metastases, and the highest predictive value from a positive bone scintigraphy was obtained with bone ALP (0.88) (8).

ALP is the most frequently used biochemical marker of osteoblastic bone formation. Four human gene loci are encoding for the ALP isoenzymes: "tissue nonspecific", placental, germ cell, and small intestinal locus (9). ALP from the tissue nonspecific locus is expressed in tissues such as bone and liver and constitutes ~95% of the total ALP activity in serum with a ratio of approximately 1:1 during healthy conditions in adults (10). Because bone and liver ALP are encoded by the same gene locus, they are referred to as isoforms of the same isoenzyme. Different carbohydrate side chains or maybe remaining fragments of the in situ cell membrane glycosyl-phosphatidylinositol anchor, or both, yield "tissue specific" structures in the ALP isoforms from this gene locus (11)(12). In this study, we used a previously described HPLC method that can detect six different ALP isoforms in serum from healthy adults: one bone/intestinal (B/I), two bone (B1 and B2), and three liver ALP isoforms (L1, L2, and L3) (13)(14).

Clodronate has been reported to have multiple effects on human bone, such as reducing bone pain, hypercalcemia, hypercalciuria, formation of new osteolytic lesions, further growth of existing lesions, and development of vertebral fractures in patients with tumor bone disease, and to prevent bone loss (15)(16)(17)(18)(19)(20). The pharmacologically active bisphosphonates have indeed proved to be powerful inhibitors of bone resorption when tested in a variety of conditions both in vitro and in vivo; however, the precise mode of action is still not completely elucidated. In vitro studies of bisphosphonates have demonstrated powerful inhibitory effects on the function of existing osteoclasts with minor or conflicting effects on osteoclast recruitment (21)(22)(23). It has also been demonstrated that bisphosphonates affect osteoclasts not only directly but also indirectly via effects on cells of the osteoblast family (24)(25). The distribution of clodronate in bone of adult rats and its effects on trabecular and cortical bone has recently been described. The highest activity of C-clodronate was found in the primary spongiosa of the distal femoral metaphysis and in the cortical bone of the femoral diaphysis (26). The distribution of clodronate in human bone has not, to our knowledge, been reported. The human skeleton is a heterogeneous tissue with a wide morphologic, functional, and metabolic variety in which cortical bone mainly fulfills a mechanical and protective function and the trabecular bone a metabolic function (27). Trabecular bone has ~5-fold more surface area per unit of volume than cortical bone, and a higher rate of metabolic activity and remodeling. Despite all these known differences, reports are sparse on biochemical markers of bone turnover in relation to trabecular and cortical bone. Bone markers are thus far not able to distinguish metabolic events in trabecular from cortical bone (28)(29)(30), although some differences have been found (31).

The aim of this randomized, double-blind study (clodronate or placebo) was to examine and compare serum total ALP, bone ALP isoforms, osteocalcin, ICTP, and prostate-specific antigen (PSA) in 45 healthy men with those same markers in 42 patients with primarily or secondarily hormone-refractory prostate cancer with skeletal metastases and who were suffering from persisting pain. We also investigated these markers of bone turnover with respect to the extent of metastases according to Soloway score, I to IV, and their association to different skeletal sites indicated by the localization of pain.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
subjects
The study group was composed of 42 patients (mean age, 71 years; range, 54–86 years) with primarily or secondarily hormone-refractory prostate cancer, histopathologically confirmed evidence of skeletal metastases, and suffering from persisting pain despite analgesic treatment. Patients with other malignant diseases or who had a previous cancer that might interfere in the assessments were not included. No extreme hypercalcemic patients were included, defined as a corrected serum calcium concentration >2.85 mmol/L, corrected for serum albumin concentration. All patients had serum creatinine values <155 µmol/L, indicating no serious deterioration of the glomerular filtration. None of the patients were treated with any other drugs known to affect calcium metabolism within 30 days or palliative radiotherapy within 3 weeks before the start of study treatment. The analgesic effect and safety of clodronate as well as the effect on quality of life compared with placebo in this material is reported elsewhere (32).

Bone metastases were diagnosed by bone scintigraphy after intravenous administration of 99mTc-labeled methylene disphosphonate and x-ray examinations of selected painful areas. Bone scintigraphy was performed on all patients and evaluated at one center (Uppsala) according to Soloway score I to IV (33)(34). Pain intensity was assessed using a 10-cm-long graded Visual Analog Scale (35) on day 1 and after 1 month of treatment. The primary pain efficacy variable was the "main pain", which corresponded to mean pain during the last week and recorded at day 1 before treatment. A body picture was also filled in where the pain was localized and characterized as upper (U), lower (L), or upper and lower (UL) body pain. The pelvis and hip were classified as belonging to the lower body.

The control group for osteocalcin, ICTP, and bone ALP isoforms was composed of 45 healthy men (blood donors) with a mean age of 44 years (range, 25–65 years). This study was conducted in accordance with the Declaration of Helsinki and approved by the Ethics Committees of each study center: Danderyd, Eskilstuna, Göteborg, Linköping, Norrköping, and Uppsala. Written informed consent was obtained from each patient before participation in the study.

study protocol for clodronate or placebo administration
The patients were randomized either to the clodronate (n = 22) or the placebo arm (n = 20). The clodronate (Bonefos, Leiras Oy) group started with intravenous administration, 300 mg/day for 3 days, followed by oral administration of clodronate, 3200 mg/day (1600 mg twice a day) during 4 weeks. The placebo group started with isotonic saline solution for 3 days, followed by placebo capsules during 4 weeks. Thirty-nine of the 42 patients completed the treatment period. One patient in the placebo group stopped treatment because of severe pain, and two patients in the clodronate group ended because of severe diarrhea and a femur fracture, respectively. Samples for determination of serum total ALP, bone and liver ALP isoforms, osteocalcin, ICTP, and PSA were taken before treatment and after 1 month of treatment.

biochemical determinations
The bone and liver ALP isoforms were separated and quantified by a previously described HPLC method (13)(14). A weak anion-exchange column, SynChropak AP300 (250 x 4.6 mm i.d.; MICRA Scientific, Inc.) was used instead of the referred SynChropak AX300. SynChropak AP300 is a modified SynChropak AX300, optimized for bone ALP isoform analysis (10). Serum total ALP activity was measured on a Hitachi 917 analyzer (Boehringer Mannheim GmbH) at 37 °C (36). The relation between the enzymatic activity units and µkat is 1/60, i.e., 1 U/L corresponds to 0.01667 µkat/L. Serum osteocalcin was determined by RIA with a double antibody technique (INCSTAR) (37), and serum ICTP was determined by RIA (Orion Diagnostica) (38). Serum PSA was determined by an IMx immunoassay (Abbott Laboratories) (39).

statistical analysis
All calculations were performed with the StatView® 4.5 program (Abacus Concepts, Inc.). Nonparametric statistics were used because the distributions were not gaussian according to the Kolmogorov–Smirnov test. The Mann–Whitney test was used to test for differences between the groups and subgroups: healthy men, prostate cancer patients with bone metastases, placebo, clodronate, Soloway score, main pain, and upper and lower pain of the body. To examine differences between the paired observations before treatment and 1 month after treatment, we used the Wilcoxon signed-rank test. To measure the association of linear relationship between variables before treatment and after 1 month, we calculated Kendall's tau rank correlation coefficient. For all statistical tests, a difference was considered significant at P <0.05.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
We were able to separate and quantify six ALP isoforms in each serum sample during the entire study period: three bone (B/I, B1, and B2), and three liver ALP isoforms (L1, L2, and L3). ALP isoform chromatographic profiles are presented from one healthy male and one prostate cancer patient with bone metastases (Fig. 1 ).



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  		Figure 1. Serum ALP isoform profiles from a healthy male (A) and a prostate cancer patient with bone metastases (B).

(A) Healthy male, 61 years of age, with a total ALP of 174 U/L. Peaks, retention times, and activities, in order of elution, are: B/I, 4.68 min, 4.8 U/L; B1, 6.65 min, 29 U/L; B2, 10.56 min, 63 U/L; L1, 14.12 min, 29 U/L; L2, 16.15 min, 33 U/L; and L3, 17.85 min, 15 U/L. (B) Prostate cancer patient with bone metastases, 77 years of age, with a total ALP of 354 U/L. Peaks, retention times, and activities, in order of elution, are: B/I, 4.65 min, 7.8 U/L; B1, 6.58 min, 44 U/L; B2, 10.68 min, 223 U/L; L1, 14.04 min, 37 U/L; L2, 16.09 min, 29 U/L; and L3, 17.74 min, 14 U/L.

baseline values in prostate cancer patients with bone metastases
Osteocalcin, ICTP, total ALP, and all three bone ALP isoforms, B/I, B1, B2, were increased in prostate cancer patients with bone metastases as compared with healthy men (Table 1 ). The most apparent increase was observed for B2. Patients and healthy men had a B2 activity corresponding to 75% and 35% of the total ALP activity, respectively. The calculated ratio B1/B2 was significantly decreased because of considerable large B2 isoform activities in the patient group with bone metastases (Table 1 ). PSA was increased in 37 patients (median, 348 µg/L; range, 2.4–6400 µg/L) compared with age-specific reference interval limits (40). As expected, the correlation coefficients were high between total ALP and all bone ALP isoforms (Table 2 ). ICTP correlated with all the other biochemical markers at baseline, including PSA (Table 2 ). We also found a positive correlation between PSA and the bone ALP isoform B2. However, no significant correlation was found between PSA and the bone ALP isoform B1, total ALP, or osteocalcin (Table 2 ).


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  		Table 1. Markers of bone turnover in prostate cancer patients with bone metastases compared with healthy males.a


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  		Table 2. Correlation coefficients between markers of bone turnover and PSA in prostate cancer patients with bone metastases.1

The patients were classified according to Soloway score and grouped into three categories: Soloway I (n = 7), Soloway II (n = 19), and Soloway III-IV (n = 11). Markers of bone turnover and PSA increased with the extent of bone metastases indicated by Soloway score from group I to group III–IV. No differences were found for markers of bone turnover and PSA between Soloway groups I and II. However, we found significant differences between Soloway groups II and III–IV for total ALP and the bone ALP isoforms, B1 and B2.

We found significant negative correlations between the primary pain efficacy variable main pain and total ALP, r = -0.22; P <0.05, and between main pain and B2, r = -0.23; P <0.05. The other markers of bone turnover and PSA showed no significant associations. The patients were also classified in terms of body pain according to where the pain was localized. Body pain was characterized as U (n = 11), L (n = 9), and UL (n = 22). No significant differences were found at baseline for markers of bone turnover and PSA between the body pain groups U and L (Table 3 ). However, a significant difference was found between the body pain groups L and UL for the bone ALP isoform B1 (P <0.05; Table 3 ).


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  		Table 3. Markers of bone turnover and PSA in prostate cancer patients with bone metastases classified according to localization of body pain.

effects of clodronate or placebo administration after 1 month
We found increased concentrations of PSA but no significant differences for the markers of bone turnover in the placebo group (Fig. 2 ). In the clodronate group, we found increased PSA, ICTP, total ALP, B/I, B1, and B2, whereas no change was observed for osteocalcin (Fig. 2 ). We found no differences in the placebo group between the body pain groups U and L for PSA and markers of bone turnover. However, after 1 month of clodronate treatment, significantly (P <0.05) higher activities were found for osteocalcin, ICTP, total ALP, B1, and B2 in the body pain group U (n = 6) compared with L (n = 6), despite the small number of patients in each group. No significant associations were observed between the primary pain efficacy variable main pain and the markers of bone turnover or PSA.



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  		Figure 2. Effects of 1 month placebo or clodronate administration on the median serum PSA, osteocalcin, ICTP, total ALP, B/I, B1, B2, and the median B1/B2 ratio (placebo, n = 19; clodronate, n = 20).

The medians for the differences between baseline values and after 1 month were: PSA, placebo 86 µg/L, clodronate 68 µg/L; osteocalcin, placebo 0.1 µg/L, clodronate 0.4 µg/L; ICTP, placebo 1.1 µg/L, clodronate 2.9 µg/L; total ALP, placebo 18 U/L, clodronate 240 U/L; B/I, placebo 0.6 U/L, clodronate 1.8 U/L; B1, placebo 3.0 U/L, clodronate 38 U/L; B2, placebo 19 U/L, clodronate 176 U/L; and B1/B2 ratio, placebo 0.00, clodronate 0.01. NS, not significant.


   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
The results of the present study demonstrate, as expected, that all the measured markers of bone turnover were higher in the prostate cancer patients with skeletal metastases compared with healthy men. However, we found significant differences between the markers of bone turnover. The largest increase was observed for the bone ALP isoform B2. Prostate cancer patients and healthy men had a B2 activity corresponding to 75% and 35% of the total ALP activity, respectively (P <0.0001). Increased B2 activities have previously been demonstrated for patients with Paget's disease of bone (41). An opposite finding has been observed in growth hormone-deficient adults who have increased B1 activities as compared with healthy adults (10). Other selective differences between the bone ALP isoforms have also been described in disease states such as hypophosphatasia and stress fractures (41). Moreover, we have previously shown that adolescent girls reach higher values than boys of the B1/B2 ratio in Tanner stage IV–V, because of a more rapid decline of the bone ALP isoform B2 compared with B1 after puberty (42). Taken together, we propose that the different bone ALP isoforms reflect different stages in osteoblast differentiation during osteogenesis, where one isoform is presented before the other during the extracellular matrix maturation phase. The bone ALP isoforms may indicate further osteoblast differentiation, i.e., when osteoblasts mature to bone lining cells, or osteocytes, or undergo apoptosis.

Osteogenesis involves three major events: proliferation with collagen synthesis, matrix maturation, and mineralization. In vitro studies have shown that gene expression of collagen type I occurs first, followed by ALP, and then osteocalcin (43). Reflecting the modest increase at baseline of osteocalcin in most of our patients, our results indicate that the bone metastases to a large extent consist of nonmineralized tissue, which also is in agreement with the morphological findings of metastatic bone. The same interpretation can be made in patients with osteomalacia in whom the ALP activities are strongly increased and the osteocalcin concentrations are within reference values or slightly raised (44). Previously described findings of slightly raised concentrations of the carboxy-terminal propeptide of type I procollagen and osteocalcin and grossly increased ALP activities in patients with skeletal metastases (44) indicate that osteoblast differentiation is disturbed during the mineralization phase of osteosclerotic lesions. During unaffected osteoblast differentiation, mineralization down-regulates expression of genes, such as ALP, associated with the matrix maturation period (43). One previously proposed function for ALP is that ALP is necessary for the initiation of mineralization but not for the continuation of mineralization of bone nodules in vitro (45)(46). Osteocalcin, the most abundant osteoblast-specific noncollagenous protein, has been reported to be 21-fold more abundant in cortical than trabecular bone on the basis of bone weight (31). Cortical bone is less metabolically active than trabecular bone; therefore, this could be one of the reasons why osteocalcin sometimes do not respond as well as other markers of bone turnover during certain pharmacological therapies and in different disease groups, such as in chronic renal failure, Paget's disease, multiple myeloma, skeletal metastases, and osteomalacia (44)(47)(48).

The increased concentrations of total ALP, bone ALP isoforms, and ICTP after 1 month of clodronate treatment in metastatic bone disease are in conjunction with other reports (49)(50). However, a decrease in bone formation markers tends to occur 2–3 months later, reflecting the coupling between resorption and formation. The increase in markers of bone formation soon after bisphosphonate treatment has been attributed to increased recruitment of osteoblasts to sites undergoing bone resorption or increased activity of preexisting osteoblasts (51). Either mechanism might suggest the repair of metastatic bone. The development of osteomalacia because of hypocalcemia, hypophosphatemia, and secondary hyperparathyroidism has also been proposed as a reason for increased bone ALP activities during clodronate treatment (49). The increased markers of bone turnover were significantly associated with different skeletal sites indicated by the location of body pain. With respect to the small number of patients in each body pain group, further investigations must be made before any major conclusions can be drawn on the basis of our findings. We suggest, however, that the uptake of clodronate by the skeleton was not uniform and appeared to be dependent in part on bone blood flow, and it was particularly marked at sites of active bone formation and mineralization.

Despite almost exclusively osteosclerotic lesions in prostate cancer, we found increased concentrations of ICTP. This finding could to some extent be explained by coupling of bone formation to resorption induced by growth factors (52). ICTP reflects the degradation of mature type I collagen, i.e., type I collagen cross-linked by pyridinolines or pyrroles. Bone collagen degradation is tightly coupled with bone resorption. However, if the newly formed nonmineralized organic matrix of bone is not mineralized, it is possible that it will be degraded, and such a process is not bone resorption by definition. Thus, the breakdown of the collagenous matrix may give rise to an increased ICTP concentration despite the absence of an obvious bone resorption.

In conclusion, we found that prostate cancer patients with skeletal metastases have grossly increased activities of the bone ALP isoform B2. We propose that the different bone ALP isoforms, B1 and B2, reflect different stages of osteoblast differentiation during osteogenesis, where one isoform is presented before the other during the extracellular matrix maturation phase. The bone ALP isoforms may indicate further osteoblast differentiation, i.e., when osteoblasts mature to bone lining cells, or osteocytes, or undergo apoptosis. After 1 month of clodronate administration, we demonstrated increased concentrations for all markers of bone turnover, except for osteocalcin, which were significantly associated with pain located only in the upper part of the body. Therefore, we suggest that the uptake of clodronate by the skeleton was not uniform during our treatment period. Future investigations, such as determination of the different bone ALP isoforms with respect to trabecular and cortical bone, are necessary to clarify the clinical significance of the site-specific differences found in this study. The bone ALP isoforms B1 and B2 should also be further evaluated concerning their ability for early detection of bone metastases and to assess efficacy and the response to antiresorptive treatment for patients with metastatic bone diseases.


   Acknowledgments
 
This study was supported by grants from Leiras Oy, Finland; Astra Läkemedel AB, Sweden; and the County Council of Östergötland (Nos. 94/180 and 95/123). This multicenter study was performed in five Swedish cities. Principal investigators were as follows: Dr. Sten Nilsson and Dr. Peter Strang, Department of Oncology, Uppsala University Hospital, Sweden; Dr. Stephan Brändstedt and Dr. Jan Sehlin, Department of Urology, Danderyd Hospital, Sweden; Dr. Eberhard Varenhorst, Department of Urology, Norrköping Hospital, Sweden; Dr. Göran Borghede, Department of Oncology, Sahlgrenska University Hospital, Göteborg, Sweden; Dr. Ulf Bandmann, Department of Oncology; and Dr. Leif Borck, Department of Urology, Eskilstuna Hospital, Sweden.


   Footnotes
 
1 Nonstandard abbreviations: ICTP, cross-linked carboxy-terminal telopeptide of type I collagen; ALP, alkaline phosphatase; B/I, bone/intestinal alkaline phosphatase; B1, bone 1 alkaline phosphatase; B2, bone 2 alkaline phosphatase; B1/B2 ratio, bone 1 alkaline phosphatase/bone 2 alkaline phosphatase ratio; L1, liver 1 alkaline phosphatase; L2, liver 2 alkaline phosphatase; L3, liver 3 alkaline phosphatase; PSA, prostate-specific antigen; U, upper body; and L, lower body.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

  1. Tofe AJ, Francis MD, Harvey WJ. Correlation of neoplasms with incidence and localization of skeletal metastases: an analysis of 1,355 diphosphonate bone scans. J Nucl Med 1975;16:986-989. [Abstract/Free Full Text]
  2. Kim SE, Kim DY, Lee DS, Chung J-K, Lee MC, Koh C-S. Absent or faint renal uptake on bone scan. Etiology and significance in metastatic bone disease. Clin Nucl Med 1991;16:545-549. [ISI][Medline] [Order article via Infotrieve]
  3. Demers LM, Costa L, Chinchilli VM, Gaydos L, Curley E, Lipton A. Biochemical markers of bone turnover in patients with metastatic bone disease. Clin Chem 1995;41:1489-1494. [Abstract/Free Full Text]
  4. Withold W, Georgescu G, Khakzad H, Vosberg H, Mueller-Gaertner HW, Reinauer H. Efficacy of simultaneous determination of bone alkaline phosphatase mass concentration in serum and urinary excretion of pyridinium cross-links for detection of bone metastases. Clin Biochem 1995;28:511-517. [ISI][Medline] [Order article via Infotrieve]
  5. Berruti A, Piovesan A, Torta M, Raucci CA, Gorzegno G, Paccotti P, et al. Biochemical evaluation of bone turnover in cancer patients with bone metastases: relationship with radiograph appearances and disease extension. Br J Cancer 1996;73:1581-1587. [ISI][Medline] [Order article via Infotrieve]
  6. Maeda H, Koizumi M, Yoshimura K, Yamauchi T, Kawai T, Ogata E. Correlation between bone metabolic markers and bone scan in prostatic cancer. J Urol 1997;157:539-543. [ISI][Medline] [Order article via Infotrieve]
  7. Kylmälä T, Tammela TLJ, Risteli L, Risteli J, Kontturi M, Elomaa I. Type I collagen degradation product (ICTP) gives information about the nature of bone metastases and has prognostic value in prostate cancer. Br J Cancer 1995;71:1061-1064. [ISI][Medline] [Order article via Infotrieve]
  8. Desoize B, Amico S, Larbre H, Coninx P, Jardillier J-C. Phosphatase isoenzymes as bone metastasis markers in prostatic carcinoma. Clin Biochem 1991;24:443-446. [ISI][Medline] [Order article via Infotrieve]
  9. Millán JL, Fishman WH. Biology of human alkaline phosphatases with special reference to cancer. Crit Rev Clin Lab Sci 1995;32:1-39. [ISI][Medline] [Order article via Infotrieve]
  10. Magnusson P, Degerblad M, Sääf M, Larsson L, Thorén M. Different responses of bone alkaline phosphatase isoforms during recombinant insulin-like growth factor-I (IGF-I) and during growth hormone therapy in adults with growth hormone deficiency. J Bone Miner Res 1997;12:210-220. [ISI][Medline] [Order article via Infotrieve]
  11. Price CP. Multiple forms of human serum alkaline phosphatase–detection and quantitation. Ann Clin Biochem 1993;30:355-372.
  12. Miura M, Sakagishi Y, Hata K, Komoda T. Differences between the sugar moieties of liver- and bone-type alkaline phosphatases: a re-evaluation. Ann Clin Biochem 1994;31:25-30.
  13. Magnusson P, Löfman O, Larsson L. Determination of alkaline phosphatase isoenzymes in serum by high-performance liquid chromatography with post-column reaction detection. J Chromatogr 1992;576:79-86. [ISI][Medline] [Order article via Infotrieve]
  14. Magnusson P, Löfman O, Larsson L. Methodological aspects on separation and reaction conditions of bone and liver alkaline phosphatase isoform analysis by high-performance liquid chromatography. Anal Biochem 1993;211:156-163. [ISI][Medline] [Order article via Infotrieve]
  15. Elomaa I, Blomqvist C, Gröhn P, Porkka L, Kairento A-L, Selander K, et al. Long-term controlled trial with diphosphonate in patients with osteolytic bone metastases. Lancet 1983;1:146-149. [ISI][Medline] [Order article via Infotrieve]
  16. Adami S, Mian M. Clodronate therapy of metastatic bone disease in patients with prostatic carcinoma. Recent Results Cancer Res 1989;116:67-72. [Medline] [Order article via Infotrieve]
  17. Lahtinen R, Laakso M, Palva I, Virkkunen P, Elomaa I. Randomised, placebo-controlled multicentre trial of clodronate in multiple myeloma. Lancet 1992;340:1049-1052. [ISI][Medline] [Order article via Infotrieve]
  18. Rotstein S, Glas U, Eriksson M, Pfeiffer P, Hansen J, Söderqvist J, et al. Intravenous clodronate for the treatment of hypercalcaemia in breast cancer patients with bone metastases–a prospective randomised placebo-controlled multicentre study. Eur J Cancer 1992;28A:890-893.
  19. Paterson AHG, Powles TJ, Kanis JA, McCloskey E, Hanson J, Ashley S. Double-blind controlled trial of oral clodronate in patients with bone metastases from breast cancer. J Clin Oncol 1993;11:59-65. [Abstract]
  20. Saarto T, Blomqvist C, Välimäki M, Mäkelä P, Sarna S, Elomaa I. Clodronate improves bone mineral density in post-menopausal breast cancer patients treated with adjuvant antioestrogens. Br J Cancer 1997;75:602-605. [ISI][Medline] [Order article via Infotrieve]
  21. Kanis JA, McCloskey EV. The use of clodronate in disorders of calcium and skeletal metabolism. Prog Basic Clin Pharmacol 1990;4:89-136.
  22. Plosker GL, Goa KL. Clodronate. A review of its pharmacological properties and therapeutic efficacy in resorptive bone disease. Drugs 1994;47:945-982. [ISI][Medline] [Order article via Infotrieve]
  23. Parfitt AM, Mundy GR, Roodman GD, Hughes EE, Boyce BF. A new model for the regulation of bone resorption, with particular reference to the effects of bisphosphonates. J Bone Miner Res 1996;11:150-159. [ISI][Medline] [Order article via Infotrieve]
  24. Sahni M, Guenther HL, Fleisch H, Collin P, Martin TJ. Bisphosphonates act on rat bone resorption through the mediation of osteoblasts. J Clin Investig 1993;91:2004-2011.
  25. Fleisch HA. Bisphosphonates: preclinical aspects and use in osteoporosis. Ann Med 1997;29:55-62. [ISI][Medline] [Order article via Infotrieve]
  26. Österman T, Virtamo T, Kippo K, Laurén L, Pasanen I, Hannuniemi R, Sellman R.. Distribution of clodronate in the bone of adult rats and its effects on trabecular and cortical bone. J Pharmacol Exp Ther 1997;280:1051-1056. [Abstract/Free Full Text]
  27. Parfitt AM. Osteonal and hemi-osteonal remodeling: the spatial and temporal framework for signal traffic in adult human bone. J Cell Biochem 1994;55:273-286. [ISI][Medline] [Order article via Infotrieve]
  28. Delmas PD. What do we know about biochemical bone markers?. Baillière Clin Obstet Gynaecol 1991;5:817-830.
  29. Withold W. Monitoring of bone turnover. Biological, preanalytical and technical criteria in the assessment of biochemical markers. Eur J Clin Chem Clin Biochem 1996;34:785-799. [ISI][Medline] [Order article via Infotrieve]
  30. Eastell R, Colwell A, Hampton L, Reeve J. Biochemical markers of bone resorption compared with estimates of bone resorption from radiotracer kinetic studies in osteoporosis. J Bone Miner Res 1997;12:59-65. [ISI][Medline] [Order article via Infotrieve]
  31. Ninomiya JT, Tracy RP, Calore JD, Gendreau MA, Kelm RJ, Mann KG. Heterogeneity of human bone. J Bone Miner Res 1990;5:933-938. [ISI][Medline] [Order article via Infotrieve]
  32. Strang P, Nilsson S, Brändstedt S, Sehlin J, Borghede G, Varenhorst E, et al. The analgesic efficacy of clodronate compared with placebo in patients with painful bone metastases from prostatic cancer. Anticancer Res 1997;17:4717-4721. [ISI][Medline] [Order article via Infotrieve]
  33. Soloway MS, Hardeman SW, Hickey D, Raymond J, Todd B, Soloway S, Moinuddin M. Stratification of patients with metastatic prostate cancer based on extent of disease on initial bone scan. Cancer 1988;61:195-202. [ISI][Medline] [Order article via Infotrieve]
  34. Rana A, Chisholm GD, Khan M, Sekharjit SS, Merrick MV, Elton RA. Patterns of bone metastasis and their prognostic significance in patients with carcinoma of the prostate. Br J Urol 1993;72:933-936. [ISI][Medline] [Order article via Infotrieve]
  35. Sriwatanakul K, Kelvie W, Lasagna L, Calimlim JF, Weis OF, Mehta G. Studies with different types of visual analog scales for measurement of pain. Clin Pharmacol Ther 1983;34:234-239. [ISI][Medline] [Order article via Infotrieve]
  36. . Scandinavian Society for Clinical Chemistry and Clinical Physiology. Committee on enzymes. Recommended methods for the determination of four enzymes in blood. Scand J Clin Lab Investig 1974;33:291-306. [ISI][Medline] [Order article via Infotrieve]
  37. Gundberg CM, Wilson PS, Gallop PM, Parfitt AM. Determination of osteocalcin in human serum: results with two kits compared with those by a well-characterized assay. Clin Chem 1985;31:1720-1723. [Abstract]
  38. Risteli J, Elomaa I, Niemi S, Novamo A, Risteli L. Radioimmunoassay for the pyridinoline cross-linked carboxy-terminal telopeptide of type I collagen: a new serum marker of bone collagen degradation. Clin Chem 1993;39:635-640. [Abstract/Free Full Text]
  39. Vessella RL, Noteboom J, Lange PH. Evaluation of the Abbott IMx automated immunoassay of prostate-specific antigen. Clin Chem 1992;38:2044-2054. [Abstract]
  40. Löfman O, Lindahl T, Varenhorst E. Age-specific reference values for serum prostate-specific antigen in a community-based population of healthy Swedish men. Scand J Clin Lab Investig 1997;57:225-232. [ISI][Medline] [Order article via Infotrieve]
  41. Magnusson P, Löfman O, Toss G, Larsson L. Determination of bone alkaline phosphatase isoforms in serum by a new high-performance liquid chromatography assay in patients with metabolic bone disease. Acta Orthop Scand 1995;66(Suppl 266):203-204.
  42. Magnusson P, Häger A, Larsson L. Serum osteocalcin and bone and liver alkaline phosphatase isoforms in healthy children and adolescents. Pediatr Res 1995;38:955-961. [ISI][Medline] [Order article via Infotrieve]
  43. Stein GS, Lian JB, Stein JL, Van Wijnen AJ, Montecino M. Transcriptional control of osteoblast growth and differentiation. Physiol Rev 1996;76:593-629. [Abstract/Free Full Text]
  44. Seibel MJ, Pols HAP. Clinical application of biochemical markers of bone metabolism. Bilezikian JP Raisz LG Rodan GA eds. Principles of bone biology 1996:1293-1311 Academic Press San Diego. .
  45. Bellows CG, Aubin JE, Heersche JNM. Initiation and progression of mineralization of bone nodules formed in vitro: the role of alkaline phosphatase and organic phosphate. Bone Miner 1991;14:27-40. [ISI][Medline] [Order article via Infotrieve]
  46. Harrison G, Shapiro IM, Golub EE. The phosphatidylinositol-glycolipid anchor on alkaline phosphatase facilitates mineralization initiation in vitro. J Bone Miner Res 1995;10:568-573. [ISI][Medline] [Order article via Infotrieve]
  47. Duda RJ, Jr, O'Brien JF, Katzmann JA, Peterson JM, Mann KG, Riggs BL. Concurrent assays of circulating bone Gla-protein and bone alkaline phosphatase: effects of sex, age, and metabolic bone disease. J Clin Endocrinol Metab 1988;66:951-957. [Abstract]
  48. Parthemore JG, Burton DW, Deftos LJ. Associations and dissociations between serum bone Gla protein and alkaline phosphatase in skeletal metabolism. J Orthop Res 1993;11:671-676. [ISI][Medline] [Order article via Infotrieve]
  49. Taube T, Kylmälä T, Lamberg-Allardt C, Tammela TLJ, Elomaa I. The effect of clodronate on bone in metastatic prostate cancer. Histomorphometric report of a double-blind randomised placebo-controlled study. Eur J Cancer 1994;30A:751-758.
  50. Vinholes J, Coleman R, Eastell R. Effects of bone metastases on bone metabolism: implications for diagnosis, imaging and assessment of response to cancer treatment. Cancer Treat Rev 1996;22:289-331. [ISI][Medline] [Order article via Infotrieve]
  51. O'Rourke N, McCloskey E, Houghton F, Huss H, Kanis JA. Double-blind, placebo-controlled, dose-response trial of oral clodronate in patients with bone metastases. J Clin Oncol 1995;13:929-934. [Abstract]
  52. Baylink DJ, Finkelman RD, Mohan S. Growth factors to stimulate bone formation. J Bone Miner Res 1993;8(Suppl 2):S565-S572.



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Home page
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B. Fohr, C. R. Dunstan, and M. J. Seibel
Markers of Bone Remodeling in Metastatic Bone Disease
J. Clin. Endocrinol. Metab., November 1, 2003; 88(11): 5059 - 5075.
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