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Recombinant human tartrate-resistant acid phosphatase
458
457
MACROPHAGECOLONY-STIMULATING FACTOR (M-CSF) RESTORESIN PRESENCE OF PARATHYROID HORMONE
(PTR) OR 1,2!j(OH)aVit.D3 BONE RESORPTION (BR) IN METATARSALS OF OSTEOPETROTIC (OP/OP) MICE IN VITRO. T. Morohashi, V. Antonioli Corboz. M. Cecchin+ R. Felix and Ii Fl i h Department
af'B*rne, !OlO Berm Switzerland.
CH-
The op/hp variant of murine osteopetrosis is
characterized by impairedBR due to a lack of osteoclasts. The defect originatesfrom a point mutationin the M-CSF gene. As shown previously M-CSF restoresboth the osteoclastpopulation and BR in this mutant in vlvo. This cytokine also stimulatesthe generationof new osteoclasts in vitro in normal embryonic bones and in cell cultures. The aim of this study was to investigatewhetherM-CSF restoresalso in vitro osteoclasto enesis in op/op bone. Metatarsals,obtaBned from newbornop/op mice, were culturedfor 9 or 15 da&s. BR was assessed measuring the releaseof Cs from prelabeled and by countingthe number of tartretetariff acid hosphatase(TRAP) osltive PTH, nuclearcaP1s. M-CSF, 1,25(OH saVit.D¶, given alone up to 15 and Interloukin-6(IL-6) ulata either. However, M-CSF ombinetionwith PTW OH)rVit.Da(10"M), enhanced and fha number of osteoclasts. nhlblltsdby celcitonin (0.1 day 9 to 15. The numberof 11s correlatedwith BR. IL-6 ), in combinationwith M-CSF, These resultsfurther effect. SF plays an important rol
RECOMBINANT HUMAN TARTRATE.RESISTAN’i.AClDPHOSPHATASE. A.R. Havman. V. Mumhv.’ T.M. COX, Univcrsir; of CambridgeCliniml School.Dcpxuncntof Mcdicinc.Addcnbrooke’s tlosPi;al,CambridgcCB22~. U.K.. ‘SmilhKlincBcccbamPharmaccuriwls.Gmat Burgh. Epsom. Surrey KTI
SXQ. U.K.
Type V acid phasphalase (E.C. X1.3.2.) is an iron-camaining lysosomal phosphohydrolasc lhar ac& on phosphopmlcins, nuclcotidcs and aryl phosphass. and is m&am 10 inhibidon by d(+)!aruale. It is expressed principally in diffcrcnlialcd cells of tic mononuclear pbagocy~c system cspccially oslcoclas~s. The cnzymcis pathologically incmascd in Ihe serum of paticnls with disorders when: bone rcsorplion is aimulalcd but iu rdlc in lhc ostcoclasl is unknown. Toobmin sufl7cicm nalivc human acid phosphatasc for molecular analysis, WC have used Ihe baculovirus vector ‘system for prolcin expression in insect cells. This syslcm is compclcnl in posWanslational processing and cxprcsscs foreign gcncs of inlcresl at a high Icvel. The full-lcng!h cDNA clone for Type V acid phosphalaa. isolalcd from a human placcnml library in Agl I I, wascloned inlo the Bum HI site of tic usnsplaccmcnt vccIor pAc CL29. Baculovirus hosl cells from Spodopferu jiugiperdo, IPLB-SRI. wcrc co-uansfecwd wilh !he rccombinanl vector DNA and lh6 wild ~ypc AcNP virus DNA. Insect cells containing rccombiMnt baculovirus lhat cxprcsscd lhc phosphalase were screened by lhc plaque assay for polyhcdrln-negative ~ransfcclanu and later by zymography in sifu. The presence of Ihe human cDNh conslrucL andexpression of phosphslasc mRNA was madily deeclpblc in tic e~r.ymc+ositivc clones and abundant Wlc-rcsismm acid pbosphsmx! aclivity was idcmificd in the cukuw medium. Wcsrcm-blot expcrimcnki using isorymc-specific antibodies confirmtic molculiuidcnriry of theexpressed cnzymo and P two-step purification proccdun has been set up. Large scale prcparalion of human Type V phosphatasc for 3. dimensional suuc~uml characlcridarion and onalysis of its courlylic mechanism in theosrcoclrs~ is now within reach.
ostaoclastgenerationand that, at least i Vitro, the combinationof M-CSF with PTH or 1,25(OH)zVit.D3 is necessary.
459
EPPICACY OF ETIDROHATE (EHDP) OH BONEIN HULTIPLE WBLOHA.RESULTS OFAWLTXCEHTRICDOUELE BLINDSTUDY. A.DaRAGQN,
C. HUMEZ,C. MICHOT, X. LE LOET, B. GROOBBOIS, F. POUYOL, L. EUUER-ZIEGLER, I.AZAAtSr J.F. BERNARD,J.F. MENARD,M. UONCONDUIT. Srouped’Etude ot de Recherche sur le My6lome (GERM), FRANCE. FQW data have been published about the efficacy of biephosphonatos on bone in muktipk myeleme (M.M). In a multicontrie controlled study. 41 MM, aged 50 to 80, stage II end III of Durde and Salmon, without previous ehomothlerapy, were randomized : 22 received etidronete (10 mg/kg/d) and 19 a placebo, for 4 months (M4). During the study. all patients had the same chemotherapy (cylophosphamide IV, prednisone per OS). Iliac crest bone biopsy was performed at the beginning and at the end of the study. Histomorphometric parameters were compared to 49 controls. No difference was found between the two groups sex, and bone histology before treatment ; compared to controls, bone parameters showed : decrease of bone volume (p C 0.05). and an increase o: bone resorption evaluated by6eroded surface (p < 10”). oeteocla _s) surface (p c 10 ) and osteoclast number (p < 10 1. At M 4. bone volume was stable in the two groups : but bcne resorption parameters decreased significantly on etidronate (p < O.OS), whereas no change occurred on placebo. for
age,
We conclude that etidronate is able to inhibit the increased bone resorptionobserved in M.H.
460 PALLIATIVE PAMIDRONATE TREATMENT OF BONE METASTASE.S IN PATIENTS WlTH BREAST CANCER. A.T.M. van Holten, O.L.M. Bijvoet, F.J. Cleton, J. Herma2g.S.E. Paoarqu10s. Depts. of Endocrinology, Oncology and Medical StatisG, University Hospital, Leiden, Netherlands. This multicenter study was performed to investigate the effect of oral pamidronate treatment on the morbidity due to bone metastases From breast cancer. Breast. cancer patients with osteolytic metastases were randomly allocated to a pamidronate (n:81) or a control (n-80) group. Pamidronate was given life-long, in addition to unrestricted tu- mor treatment. The dose was 600 mg/d during the First phase of the study, but was changed to 300 mg/d because of gast.rointestinaS toxicity. The study therefore comprises 29 patients who shifted From 600 -L to 300 mg/d (pamfdrorrate HD/LDl and 52 patients who received 300 mg/d throughout (pamidronate LD) with their respective controls. Every 3 months events of skeletal morbidity and survival, and every 6 months the radiologic course of skeletal disease were monitored For a median of 18 (pamidronate) and 21 (control) months. In the pamidronate group the incidences of hypercalcaemia, bone pain and imminent Fractures were significantly reduced by 65%, 30% and 50%, respectively; therapeutic interventions For progressive bone disease (systemic and radiotherapy) decreased by 35% (p
190
457
MACROPHAGECOLONY-STIMULATING FACTOR (M-CSF) RESTORESIN PRESENCE OF PARATHYROID HORMONE
(PTR) OR 1,2!j(OH)aVit.D3 BONE RESORPTION (BR) IN METATARSALS OF OSTEOPETROTIC (OP/OP) MICE IN VITRO. T. Morohashi, V. Antonioli Corboz. M. Cecchin+ R. Felix and Ii Fl i h Department
af'B*rne, !OlO Berm Switzerland.
CH-
The op/hp variant of murine osteopetrosis is
characterized by impairedBR due to a lack of osteoclasts. The defect originatesfrom a point mutationin the M-CSF gene. As shown previously M-CSF restoresboth the osteoclastpopulation and BR in this mutant in vlvo. This cytokine also stimulatesthe generationof new osteoclasts in vitro in normal embryonic bones and in cell cultures. The aim of this study was to investigatewhetherM-CSF restoresalso in vitro osteoclasto enesis in op/op bone. Metatarsals,obtaBned from newbornop/op mice, were culturedfor 9 or 15 da&s. BR was assessed measuring the releaseof Cs from prelabeled and by countingthe number of tartretetariff acid hosphatase(TRAP) osltive PTH, nuclearcaP1s. M-CSF, 1,25(OH saVit.D¶, given alone up to 15 and Interloukin-6(IL-6) ulata either. However, M-CSF ombinetionwith PTW OH)rVit.Da(10"M), enhanced and fha number of osteoclasts. nhlblltsdby celcitonin (0.1 day 9 to 15. The numberof 11s correlatedwith BR. IL-6 ), in combinationwith M-CSF, These resultsfurther effect. SF plays an important rol
RECOMBINANT HUMAN TARTRATE.RESISTAN’i.AClDPHOSPHATASE. A.R. Havman. V. Mumhv.’ T.M. COX, Univcrsir; of CambridgeCliniml School.Dcpxuncntof Mcdicinc.Addcnbrooke’s tlosPi;al,CambridgcCB22~. U.K.. ‘SmilhKlincBcccbamPharmaccuriwls.Gmat Burgh. Epsom. Surrey KTI
SXQ. U.K.
Type V acid phasphalase (E.C. X1.3.2.) is an iron-camaining lysosomal phosphohydrolasc lhar ac& on phosphopmlcins, nuclcotidcs and aryl phosphass. and is m&am 10 inhibidon by d(+)!aruale. It is expressed principally in diffcrcnlialcd cells of tic mononuclear pbagocy~c system cspccially oslcoclas~s. The cnzymcis pathologically incmascd in Ihe serum of paticnls with disorders when: bone rcsorplion is aimulalcd but iu rdlc in lhc ostcoclasl is unknown. Toobmin sufl7cicm nalivc human acid phosphatasc for molecular analysis, WC have used Ihe baculovirus vector ‘system for prolcin expression in insect cells. This syslcm is compclcnl in posWanslational processing and cxprcsscs foreign gcncs of inlcresl at a high Icvel. The full-lcng!h cDNA clone for Type V acid phosphalaa. isolalcd from a human placcnml library in Agl I I, wascloned inlo the Bum HI site of tic usnsplaccmcnt vccIor pAc CL29. Baculovirus hosl cells from Spodopferu jiugiperdo, IPLB-SRI. wcrc co-uansfecwd wilh !he rccombinanl vector DNA and lh6 wild ~ypc AcNP virus DNA. Insect cells containing rccombiMnt baculovirus lhat cxprcsscd lhc phosphalase were screened by lhc plaque assay for polyhcdrln-negative ~ransfcclanu and later by zymography in sifu. The presence of Ihe human cDNh conslrucL andexpression of phosphslasc mRNA was madily deeclpblc in tic e~r.ymc+ositivc clones and abundant Wlc-rcsismm acid pbosphsmx! aclivity was idcmificd in the cukuw medium. Wcsrcm-blot expcrimcnki using isorymc-specific antibodies confirmtic molculiuidcnriry of theexpressed cnzymo and P two-step purification proccdun has been set up. Large scale prcparalion of human Type V phosphatasc for 3. dimensional suuc~uml characlcridarion and onalysis of its courlylic mechanism in theosrcoclrs~ is now within reach.
ostaoclastgenerationand that, at least i Vitro, the combinationof M-CSF with PTH or 1,25(OH)zVit.D3 is necessary.
459
EPPICACY OF ETIDROHATE (EHDP) OH BONEIN HULTIPLE WBLOHA.RESULTS OFAWLTXCEHTRICDOUELE BLINDSTUDY. A.DaRAGQN,
C. HUMEZ,C. MICHOT, X. LE LOET, B. GROOBBOIS, F. POUYOL, L. EUUER-ZIEGLER, I.AZAAtSr J.F. BERNARD,J.F. MENARD,M. UONCONDUIT. Srouped’Etude ot de Recherche sur le My6lome (GERM), FRANCE. FQW data have been published about the efficacy of biephosphonatos on bone in muktipk myeleme (M.M). In a multicontrie controlled study. 41 MM, aged 50 to 80, stage II end III of Durde and Salmon, without previous ehomothlerapy, were randomized : 22 received etidronete (10 mg/kg/d) and 19 a placebo, for 4 months (M4). During the study. all patients had the same chemotherapy (cylophosphamide IV, prednisone per OS). Iliac crest bone biopsy was performed at the beginning and at the end of the study. Histomorphometric parameters were compared to 49 controls. No difference was found between the two groups sex, and bone histology before treatment ; compared to controls, bone parameters showed : decrease of bone volume (p C 0.05). and an increase o: bone resorption evaluated by6eroded surface (p < 10”). oeteocla _s) surface (p c 10 ) and osteoclast number (p < 10 1. At M 4. bone volume was stable in the two groups : but bcne resorption parameters decreased significantly on etidronate (p < O.OS), whereas no change occurred on placebo. for
age,
We conclude that etidronate is able to inhibit the increased bone resorptionobserved in M.H.
460 PALLIATIVE PAMIDRONATE TREATMENT OF BONE METASTASE.S IN PATIENTS WlTH BREAST CANCER. A.T.M. van Holten, O.L.M. Bijvoet, F.J. Cleton, J. Herma2g.S.E. Paoarqu10s. Depts. of Endocrinology, Oncology and Medical StatisG, University Hospital, Leiden, Netherlands. This multicenter study was performed to investigate the effect of oral pamidronate treatment on the morbidity due to bone metastases From breast cancer. Breast. cancer patients with osteolytic metastases were randomly allocated to a pamidronate (n:81) or a control (n-80) group. Pamidronate was given life-long, in addition to unrestricted tu- mor treatment. The dose was 600 mg/d during the First phase of the study, but was changed to 300 mg/d because of gast.rointestinaS toxicity. The study therefore comprises 29 patients who shifted From 600 -L to 300 mg/d (pamfdrorrate HD/LDl and 52 patients who received 300 mg/d throughout (pamidronate LD) with their respective controls. Every 3 months events of skeletal morbidity and survival, and every 6 months the radiologic course of skeletal disease were monitored For a median of 18 (pamidronate) and 21 (control) months. In the pamidronate group the incidences of hypercalcaemia, bone pain and imminent Fractures were significantly reduced by 65%, 30% and 50%, respectively; therapeutic interventions For progressive bone disease (systemic and radiotherapy) decreased by 35% (p
190