WO2006124062A1 - Methode pour stimuler la formation et la preservation des os - Google Patents

Methode pour stimuler la formation et la preservation des os Download PDF

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Publication number
WO2006124062A1
WO2006124062A1 PCT/US2005/040032 US2005040032W WO2006124062A1 WO 2006124062 A1 WO2006124062 A1 WO 2006124062A1 US 2005040032 W US2005040032 W US 2005040032W WO 2006124062 A1 WO2006124062 A1 WO 2006124062A1
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Prior art keywords
bone
subject
calcitonin
pth
agent
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PCT/US2005/040032
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English (en)
Inventor
Agnes Vignery
Nozer M. Mehta
James P. Gilligan
Original Assignee
Unigene Laboratories, Inc.
Yale University
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Publication date
Priority claimed from US11/128,095 external-priority patent/US7531518B2/en
Application filed by Unigene Laboratories, Inc., Yale University filed Critical Unigene Laboratories, Inc.
Priority to AU2005331907A priority Critical patent/AU2005331907B2/en
Priority to CA002608016A priority patent/CA2608016A1/fr
Priority to CN2005800509720A priority patent/CN101217972B/zh
Priority to JP2008511101A priority patent/JP2008540522A/ja
Priority to EP05851371A priority patent/EP1879608A4/fr
Publication of WO2006124062A1 publication Critical patent/WO2006124062A1/fr

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/22—Hormones
    • A61K38/29—Parathyroid hormone (parathormone); Parathyroid hormone-related peptides
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/66—Phosphorus compounds
    • A61K31/665—Phosphorus compounds having oxygen as a ring hetero atom, e.g. fosfomycin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/22—Hormones
    • A61K38/225—Calcitonin gene related peptide
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/22—Hormones
    • A61K38/23—Calcitonins
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/43—Enzymes; Proenzymes; Derivatives thereof
    • A61K38/46—Hydrolases (3)
    • A61K38/48—Hydrolases (3) acting on peptide bonds (3.4)
    • A61K38/482—Serine endopeptidases (3.4.21)
    • A61K38/4846—Factor VII (3.4.21.21); Factor IX (3.4.21.22); Factor Xa (3.4.21.6); Factor XI (3.4.21.27); Factor XII (3.4.21.38)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P19/00—Drugs for skeletal disorders
    • A61P19/08—Drugs for skeletal disorders for bone diseases, e.g. rachitism, Paget's disease
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
    • Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
    • Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Definitions

  • the present invention relates generally to a method for fostering bone formation in a subject. More particularly the invention concerns a method for inducing rapid bone formation and then preserving the bone thus produced, e.g., by minimizing its resorption.
  • the invention permits specific targeting of particular bones of a subject for repair, strengthening, reshaping and/or modeling.
  • the invention is additionally directed to a kit for carrying out the method of the invention.
  • Bones are multi-purpose structures that play diverse, vital roles in vertebrates. They provide a framework that supports the body and gives it shape. Bone undergoes a continuous renewal or remodeling during the lifetime of an individual. Bone consists of living cells widely scattered within a non-living material known as matrix. Two main types of cells are responsible for bone remodeling: the osteoblasts involved in bone formation and the osteoclasts involved in bone resorption. The matrix is formed by the action of osteoblasts, that make and secrete bone matrix proteins such as collagen, which provide elasticity, as well as mineral salts formed from calcium and phosphorous, which impart hardness to bone.
  • osteoblasts As bone tissue matures, some osteoblasts are trapped in the bone matrix and differentiate into osteocytes, which are mature bone cells that carry out normal cellular activities. These osteocytes connect with other osteocytes through the bone matrix and can sense pressure or cracks in the bone. They therefore assist in directing where osteoclasts will act to resorb bone during the repair and/or regeneration of bone.
  • Osteoclasts are cells that resorb existing bone, thus facilitating bone growth, repair and regeneration. Osteoclasts are multinucleated cells that originate from the fusion of mononuclear phagocytes. Osteoclasts secrete protons that lower the pH of an extracellular compartment located between osteoclasts and bone. This low pH facilitates the dissolution of bone crystals and activates lysosomal enzymes that digest the bone matrix. Osteoclasts are therefore powerful and efficient bone resorbing cells that cover only 0.5% of the bone surface. With regard to bone formation, osteoblasts produce a structure, known as "osteoid", which is formed of bone collagen and other proteins.
  • the osteoblasts thereafter control the deposition of calcium and other minerals into the osteoid in order to produce the calcified bone tissue.
  • the osteoblasts flatten out and form a lining upon the surface of the bone.
  • These flattened osteoblasts known as "lining cells”, regulate passage of calcium into and out of the bone.
  • they produce, upon hormonal activation, proteins that promote osteoblast differentiation and activation. Making new bone is therefore a slow process that requires the lying down of the osteoid, its maturation and then its calcification. In contrast to osteoclasts, osteoblasts cover 30% of the bone surface.
  • the bones of the skeleton are not entirely solid throughout.
  • the outside, i.e., cortical, bone is substantially solid throughout, having only a few small canals.
  • spongy bone Located inwardly from the cortical bone, however, is spongy bone known as cancellous bone.
  • the cancellous bone is composed of a honeycomb network of trabecular bone defining a plurality of spaces or cavities filled with fluid bone marrow, stem cells and some fat cells. Existing within these bone marrow cavities are, inter alia, various highly specialized cells which assist in breaking down existing bone and correspondingly producing new bone to replace that which is broken down or which may be otherwise lost due to injury or illnesses such as osteoporosis.
  • the physical structure of bone may be compromised by a variety of factors, including disease and injury.
  • One of the most common bone diseases is osteoporosis, which is characterized by low bone mass and structural deterioration of bone tissue, leading to bone fragility and an increased susceptibility to fractures, particularly of the hip, spine and wrist.
  • Osteoporosis develops when there is an imbalance such that the rate of bone resorption exceeds the rate of bone formation. This is in part due to the fact that it requires six months for osteoblasts to rebuild the amount of bone destroyed by osteoclasts in three days.
  • Bone injury involves localized trauma to the bone.
  • a variety of methods are well-known in the art for fostering bone formation in individuals who (1) suffer from diminished bone mass due, for example, to illness, (2) are subjected to bone trauma causing injury such as bone fractures, and (3) need to strengthen bone, such as vertebral bones.
  • Such prior art methods for treating these disorders are typically systemic in nature, however. That is, they treat the whole skeleton as a single entity. These methods are therefore not commonly able to be targeted on one or more specific bones, e.g., those of the hip, shoulder, spine and/or wrist, which may require a more focused treatment due to bone losses due to disease effects caused by, e.g., osteoporosis or by bone trauma such as that due to a fracture.
  • prior art methods frequently require undesirably long treatment regimens, with accompanying patient compliance problems.
  • the method of the invention is utilized with a human subject.
  • the invention additionally contemplates veterinary applications.
  • the invention provides a method of inducing bone formation in a subject in need of such inducement comprising the steps of (a) mechanically inducing an increase in osteoblast activity in the subject; and (b) elevating blood concentration of at least one bone anabolic agent therein, e.g., by administering such an agent or by administering a compound which causes natural formation of such an agent.
  • the aforesaid steps may be performed in any order, but in sufficient time proximity that the elevated concentration of the anabolic agent and the mechanically induced increase in osteoblast activity at least partially overlaps.
  • the method comprises the steps of targeting for treatment at least one bone of the subject, wherein each such targeted bone defines a bone marrow cavity therein.
  • the bone marrow cavity contains, inter alia, a quantity of bone marrow and a plurality of osteoblasts.
  • the method further comprises mechanically altering the contents of the bone marrow cavity to thereby stimulate and thus increase osteoblast differentiation and/or activity therein.
  • the method additionally comprises administering to the subject at least one bone anabolic agent for a duration and at a concentration sufficient to raise blood levels of the anabolic agent within the subject above natural levels thereof and thereby prolong the mechanically induced osteoblast activity.
  • the mechanical alteration of the contents of the bone marrow cavity in bones wherein it is desired to foster such bone growth permits specific bones of the subject to be particularly targeted for inducing bone formation therein.
  • the invention further provides a method of inducing bone formation in a subject suffering from diminished bone mass which comprises the steps of targeting for treatment at least one bone of the subject, wherein each targeted bone defines a bone marrow cavity therein.
  • the bone marrow cavity contains a quantity of bone marrow and a plurality of osteoblasts.
  • the method of the invention further comprises mechanically altering the contents of the bone marrow cavity to thereby stimulate and thus increase osteoblast activity (e.g., increased differentiation or increased bone formation by stimulating osteoblasts) therein. Thereafter, bone mass is increased within the cavity due to the increased osteoblast activity.
  • the method additionally comprises administering to the subject at least one bone anabolic agent for a duration and at a concentration sufficient to raise blood levels of the anabolic agent within the subject above natural levels thereof and thereby prolong the mechanically induced osteoblast activity.
  • the method further comprises additionally administering, either (1) contemporaneous with, (2) overlapping with, or (3) subsequent to the administration of the bone anabolic agent, an antiresorptive agent for a duration and at a concentration sufficient to diminish resorption of new bone produced due to the osteoblast activity.
  • an antiresorptive agent for a duration and at a concentration sufficient to diminish resorption of new bone produced due to the osteoblast activity.
  • the invention further provides a method for inducing bone formation within a bone of a subject which comprises mechanically altering the contents of a bone marrow cavity within said bone to thereby stimulate, and thus increase, osteoblast differentiation activity therein so as to induce bone growth therein.
  • the method further comprises administering to said subject an amount of an anti-resorptive agent for a time and at a concentration sufficient to substantially prevent resorption of the new bone produced due to the osteoblast activity.
  • the anti-resorptive agent which may be but is not necessarily salmon calcitonin, may be administered contemporaneous with the mechanical alteration and/or subsequent thereto. This method does not include the administration of a bone anabolic agent
  • the invention additionally provides a kit for fostering bone formation in at least one targeted bone of a subject in need of such bone formation.
  • the kit includes at least one container having therein at least one bone anabolic agent, as well as a mechanical alteration device for altering contents of a bone marrow cavity in at least one targeted bone.
  • the kit may include at least one container having therein at least one bone anti-resorptive agent, as well as a mechanical alteration device for altering contents of the bone marrow cavity of a selected bone.
  • the kit may include multiple containers with at least one bone anabolic agent and at least one bone anti-resorptive agent, as well as the mechanical alteration device described above.
  • the kit may additionally be provided with an evacuation device for evacuating at least a portion of the contents from the bone marrow cavity.
  • FIG. 1 is a graphical representation of the results of a PDGMUS analysis of the femur distal marrow cavity bone mineral density (“BMD”) of groups of laboratory rats which were sacrificed at one-week intervals during a three-week test regimen.
  • BMD femur distal marrow cavity bone mineral density
  • the PDQMUS analyzer is available from Lunar Corp., Palo Alto, CA and provides Dual Energy X-Ray Absorptiometry (“DEXA”) data for small animals such as mice and rats.
  • EXA Dual Energy X-Ray Absorptiometry
  • the various groups were subjected to the following treatment modalities: Group A - left femur control; Group B - left femur sham; Group C - mechanical bone marrow ablation (bmx) to alter various groups were subjected to the following treatment modalities: Group A - left femur control; Group B - left femur sham; Group C - mechanical bone marrow ablation (bmx) to alter the contents of the bone marrow cavity of the left femur; Group D - mechanical bone marrow ablation treatment (bmx) of the left femur, coupled with the administration of an amidated parathyroid hormone truncate (PTH[I -34] -NH 2 ) during days 1-21; and Group E - a right (non- bmx) femur BMD measurement of rats treated during days 1-21 with an amidated parathyroid hormone truncate (PTH[I -34] -NH 2 ) wherein the rat
  • FIG.2 is a graphical representation of the results of a PIXIMUS analysis of the femur distal marrow cavity BMD of a second set of groups of male Sprague-Dawley rats (5 animals per group) aged 9 weeks which were sacrificed and tested at time 0 and after 1, 2 and 3 weeks.
  • the various groups were subjected to the following treatment modalities: Group F - left femur control; Group G - left femur sham; Group H - mechanical bone marrow ablation (bmx) to alter the contents of the bone marrow cavity of the left femur; Group I - mechanical bone marrow ablation (bmx) of the left femur, coupled with administration of an amidated parathyroid hormone truncate (PTH[ l-34]-NH 2 ) during days 1-7, followed by the administration of a salmon calcitonin (sCT) antiresorptive agent during days 7-21; and Group J - a right femur BMD measurement of rats treated with an amidated parathyroid hormone truncate (PTH[I -34]-NH 2 ) wherein the rat's left femur had undergone a bone marrow ablation (bmx) treatment;
  • PTH[I -34]-NH 2 an amidated parathyroid
  • FIGS. 3 present high resolution radiograms of, respectively, the left (ablated) and right (non-ablated) femurs of some of the rats described with regard to FIGS. 1 and 2 above. (See discussion below of Tables 1 and 2) The radiographs were taken three weeks (i.e., 21 days) following commencement of treatment.
  • the rats were treated as follows: (a) control (i.e., no surgery); (b) sham (i.e., surgery without marrow ablation); (c) mechanical bone marrow ablation (bmx) + phosphate buffer saline (PBS) administration daily for 21 days; (d) bmx followed by parathyroid hormone (PTH[I -34] -NH 2 ) administered daily for 21 days; and (e) bmx followed by parathyroid hormone (PTH[l-34]-NH 2 ) administered daily for 7 days and then salmon calcitonin given daily from days 7-21.
  • control i.e., no surgery
  • sham i.e., surgery without marrow ablation
  • PBS phosphate buffer saline
  • FIGS 4 (a-d) are graphical representations of the results of Peripheral Quantitative Computerized Tomography (PQCT) analyses of, respectively, the left (ablated) and right (non-ablated) femoral shafts of the rats which are the subject of FIG. 3 above.
  • FIGS. 1-10 are graphical representations of the results of Peripheral Quantitative Computerized Tomography (PQCT) analyses of, respectively, the left (ablated) and right (non-ablated) femoral shafts of the rats which are the subject of FIG. 3 above.
  • FIGS. 4(a) and 4(b) illustrate the trabecular bone density observed in respectively, the left and right femoral shafts of these rats, whereas, FIGS .4(c) and 4(d) illustrate the total bone density of the respective shafts.
  • the meanings of the abbreviations used in FIGS. 4a-4d are identical to those described above with regard to FIGS. 3 (a-e);
  • FIG. 5 provides a series of measurements of the serum osteocalcin concentration (in ng/ml) in the blood of the rats which are the subjects of FIGS. 3-4 on the day of sacrifice.
  • the meaning of the abbreviations is the same as that in FIGS. 3-4;
  • FIG. 6 illustrates the results of Micro Computerized Tomography (MicroCT) analyses of the left (ablated) femoral shafts of the rats which are the subjects of FIGS. 3-4.
  • the abbreviations mean the same as in FIGS. 3-5;
  • FIG. 7 is similar to FTG. 6 but illustrates the results of a Micro Computerized Tomography (MicroCT) analysis of the right (non-ablated) femoral shafts of the same rats.
  • MicroCT Micro Computerized Tomography
  • FIG. 8 illustrates sections (at 4X magnification) of undecalcified left femurs, subjected to bone marrow ablation (bmx), wherein the bone sections were stained with Von Kossa stain;
  • FIG. 9 illustrates the new bone formed in the marrow cavity of groups of rats who underwent the following treatments: control at day 0; control at day 21 ; sham at day 21 ; bmx + PBS daily for 7 days; bmx + PBS daily for 14 days; bmx + PBS daily for 21 days; bmx + parathyroid hormone (PTH[l-34]-NH 2 ) daily for 7 days; bmx + parathyroid hormone (PTH[I- 34]-NH 2 ) dailyfor 14days; bmx + parathyroid hormone (PTH[l-34]-NH 2 ) daily for 21 days; bmx + salmon calcitonin (CT) daily from days 7-14; bmx + CT daily from days 7-21; bmx + parathyroid hormone (PTH[I -34] -NH 2 ) daily from days 1-7 and CT daily from days 7-14; and bmx + parathyroid hormone (PTH[I -34] -
  • FIG. 9 presents high resolution radiograms of the left and right femurs, respectively, from an additional series of experiments on additional groups of male Sprague- Dawley rats (6 rats per group) aged 9 weeks which were sacrificed and tested after 21 days (3 weeks).
  • the various groups were subjected to the following treatment modalities: Group K - control day 0; Group L - control day 21; Group M - bone marrow ablation (bmx) day 0; Group N - sham day 21; Group O - bmx + phosphate buffer saline (PBS) daily for 21 days; Group P - bmx + PTH[l-34]-NH 2 daily for 21 days; Group Q - bmx + parathyroid hormone related peptide (PTHRP) daily for 21 days; and Group R - bmx + rat ⁇ -calcitonin gene related peptide (CGRP) daily for 21 days;
  • FTG. 11 (a-d) are graphical representations of the results of Peripheral Quantitative Computerized Tomography (PQCT) analyses of, respectively, the left (ablated) and right (non-ablated) femoral shafts of the rats from Groups K-R (see description of Fig. 10).
  • FIGS . 11 (a) and 11 (b) illustrate the trabecular bone density observed in, respectively the left and right femoral shafts at day 21; whereas, FIGS. 1 l(c) and ll(d) illustrate the total bone density of, respectively, the left and right femoral shafts at the same interval.
  • FIGS. 1 l(c) and ll(d) illustrate the total bone density of, respectively, the left and right femoral shafts at the same interval.
  • FIG. 12 provides a measurement of the serum osteocalcin concentration (in ng/ml) in the blood of the rats in Groups K-R when the rats were sacrificed 21 days after commencement of treatment.
  • FIG. 13 (a-b) illustrate the results of Micro Computerized Tomography (MicroCT) analysis of the left (ablated) and right (non-ablated) femoral shafts of the rats of Groups K-R.
  • the abbreviations are the same as those used in FIGS. 10-12;
  • FIGS. 14A-14D illustrate a representative series of steps for carrying out a left femur bone marrow ablation in a rat from one of Groups A-J discussed above.
  • FIG. 14A accessing the distal femur
  • FIG. 14B drilling into the bone marrow cavity of the distal femur
  • FIG. 14C washing the bone marrow cavity
  • FIG. 14D suturing the incision.
  • Many alternate methods of ablation are possible and thus these figures do not limit the method of the invention.
  • FIG. 15 is a bar graph comparing the Lumbar Vertebral Bone Mineral Density obtained in osteoporotic female Sprague-Dawley rats wherein the osteoporosis was induced via an ovariectomy.
  • the rats were treated with (1)PTH[1-31]-NH 2 ; (2)PTH[l-34]-NH 2 ; andPTH[l- 34]-OH; and
  • FIG. 16 is a bar graph comparing the Lumbar Vertebral Bone Formation Rate obtained in osteoporotic female Sprague-Dawley rats wherein the osteoporosis was induced via an ovariectomy.
  • the rats were treated with (I)PTH[I -31]-NH 2 ; (2) PTH[l-34]-NH 2 ; andPTH[l- 34]-OH.
  • Li a first embodiment the invention provides a method of inducing new bone formation in a subject in need of such inducement.
  • the method comprises the steps of mechanically inducing an increase in osteoblast activity in the subject and elevating blood concentration of at least one bone anabolic agent in the subject.
  • the above steps may be performed in any order, but are to be carried out in sufficient time proximity that the elevated concentration of the anabolic agent and the mechanically induced increase in osteoblast activity at least partially overlap.
  • Inducement of bone growth may include, for example, generating new or additional bone at locations where such bone growth is not presently taking place and/or stimulating the growth (i.e., increasing the rapidity thereof) of bone which is already in the process of formation.
  • the inducement of bone growth takes place due to the combined effects of (1) a mechanical inducement of osteoblast activity in the subject coupled with (2) an elevation in the blood concentration of at least one bone anabolic agent therein.
  • the bone described as being formed by the process of the invention is not limited solely to trabecular bone and should also be taken to include any one or more of the following additional 'types' of bone: compact, cortical and /or lamellar bone.
  • Mechanical inducement of an increase in osteoblast activity may be obtained, in a preferred embodiment of the invention, by a process of bone marrow irrigation and ablation. Again, without being bound in any way by theory, applicants believe that the bone marrow irrigation and ablation process leads to the formation of a clot within the bone marrow cavity which, through a cascade of biochemical reactions, contributes to increasing osteoblast activity in the subject.
  • the increased osteoblast activity may alternately be obtained by coupling the mechanical inducement with an additional form of inducement such as biochemical inducement.
  • biochemical inducement may be obtained by administering to the subject, for example, a quantity of a blood factor such as Factor ("F") VH, Factor Vila or a combination thereof.
  • F Factor
  • tissue or vascular injury clotting is initiated by the binding of plasma FVDTFVIIa to tissue factor (tissue thromboplastin).
  • tissue thromboplastin tissue factor
  • This complex (FVnTFVIIa + Thromboplastin) initiates a sequence of events which leads to activation of the coagulation cascade ultimately leading to fibrin deposition and platelet activation.
  • This complex sequence of events may contribute in part to the stimulation of osteoblasts in the bone marrow.
  • Factors V ⁇ and Vila are commercially available from Novo Nordisk.
  • the increase in osteoblast activity obtained with the use of the method of the invention may be due to a variety of factors including, but not necessarily limited to (1) osteoblast differentiation, i.e., the production of additional osteoblasts, (2) increasing the activity and/or effectiveness of osteoblasts which are already present in inducing bone formation in the subject, and (3) a combination thereof.
  • the increase in osteoblast activity would include all of the above-noted functions.
  • the blood concentration of the at least one bone anabolic agent may be elevated by direct administration of one or more bone anabolic agents to the subject.
  • the method additionally comprises "targeting" one or more specific bones of the subject for inducement of bone growth. This targeting is accomplished by mechanically altering the contents of a bone marrow cavity within each targeted bone so as to induce the increased osteoblast activity therein.
  • the method of the invention is thus useful not only for bone repair, i.e., as in the case of a bone fracture due to trauma, but also for strengthening bone in a site-specific manner in the case of individuals shown by Dual Energy X-Ray Absorptiometry ("DEXA") or other techniques to require an increase in bone mass and/or density to prevent bone fractures, or who suffer due to bone weakness from chronic pain attributable to conditions such as vertebral crush.
  • the method of the invention additionally serves to provide (and retain) new bone needed to serve as an anchor for prostheses such as artificial hips, knees and shoulders and/or for implants such as dental implants.
  • the method of the invention additionally comprises providing the subject with an elevated blood concentration of at least one antiresorptive agent, wherein the elevated concentration is sufficient to diminish resorption of new bone growth produced due to the mechanically induced enhanced osteoblast activity according to the invention.
  • the invention provides a method of inducing bone formation in a subject in need of the same, wherein the method comprises (a) mechanically inducing an increase in osteoblast activity in the subject; and (b) administering to the subject at least one agent that causes elevated blood levels of an endogenous bone anabolic agent within the subject.
  • the method steps may be performed in any order, but in sufficient time proximity that the elevated concentration of the anabolic agent and the mechanically induced increase in osteoblast activity at least partially overlaps.
  • the agent causing an increased expression of the endogenous bone anabolic agent may be a calcilytic agent.
  • Calcilytic agents useful with the method of the invention include, but are not limited to any agent that limits the binding of calcium to its receptor and thereby triggers the release of endogenous PTH. Examples of such calcilytic compounds are set forth in United States Patents Nos. 6,362,231; 6,395,919; 6,432,656 and 6,521,667, the contents of which are incorporated herein by reference.
  • the invention additionally provides a method of inducing bone formation in a subject suffering from diminished bone mass or bone trauma.
  • the method includes the step of targeting for treatment at least one bone of the subject, wherein each targeted bone defines a bone marrow cavity therein.
  • the bone marrow cavity contains, inter alia, a quantity of bone marrow and a plurality of osteoblasts.
  • the method of the invention further comprises mechanically altering the contents of the bone marrow cavity to thereby stimulate and thus increase osteoblast activity therein.
  • the method additionally comprises administering to the subject at least one bone anabolic agent for a duration and at a concentration sufficient to raise blood levels of the anabolic agent within the subject above natural levels thereof and thereby prolong the mechanically induced osteoblast activity.
  • a bone anabolic agent endogenously produced in the human body is PTH[I -84] in the free acid form which is naturally found in levels of less than about 8 picomoles (pmoles) per liter in blood of a human subject.
  • PTH[I -84] in the free acid form which is naturally found in levels of less than about 8 picomoles (pmoles) per liter in blood of a human subject.
  • the practice of the invention would involve, as indicated above, raising the blood levels of the bone anabolic agent within the subject to levels correspondingly above such natural level.
  • the mechanical alteration of the contents of the bone marrow cavity thus permits specific bones of the subject to be targeted for enhanced bone formation.
  • bone formation may be induced at a location of a long bone fracture in bone of the subject to increase the rapidity of healing of the fracture.
  • the method further comprises reshaping or modeling at least one targeted bone of a subject by inducing additional bone formation in a controlled manner thereon.
  • an initial burst of osteoblast activity can be sustained at unexpectedly high levels by elevating blood levels of a bone anabolic agent during a time that at least partially overlaps with the initial burst of osteoblast activity. This effect is demonstrated by comparing the curves obtained with Groups C and D in FIG. 1. Absent the bone anabolic agent, the initial burst of bone formation is quickly followed by the resorption of newly formed bone by osteoclasts (see, e.g., the Group C curve in FIG. 1). The curve obtained with Group E of FIG.
  • Table 1 (below) sets forth the values upon which the curves obtained with Groups A-E in FIG. 1 are based. As no readings were taken for the Control (Group A) and Sham (Group B) groups at Week 1 and Week 2, these values were estimated for purposes of preparing the curves for these two Groups. Table 1, moreover, provides the standard deviations for the values obtained in each Group, which standard deviations are also indicated in FIG. 1.
  • the bmx treatment was always administered to the rats' left femur and never to the right femur.
  • the non-ablated contralateral femur was analyzed, however, throughout the testing protocols described herein as a control used for detecting the systemic effect of the bone anabolic agent. That is, the results achieved with the contralateral (right) femur permit one to exclude the effect attributable to the bmx treatment.
  • analysis of the rats' right femur control also enabled the inventors to determine whether the bmx treatment of the left femur had any effect on other skeletal sites located within the body of the rat.
  • FIG.2 is provided to illustrate, inter alia, the further beneficial effect achieved due to the overlapping (i.e., with PTH administration) or subsequent administration of the antiresorptive agent salmon calcitonin (sCT).
  • sCT antiresorptive agent salmon calcitonin
  • the curve obtained with Group I also demonstrates that the administration of the antiresorptive agent sCT following the initial burst of osteoblast activity which occurs due the coupling of bmx and PTH administration, significantly diminishes the proportion of the newly formed bone which is resorbed due to the action of the osteoclasts.
  • Table 2 (below) sets forth the values upon which the curves of FIG. 2 are based. As no readings were taken for the Control (Group F) and Sham (Group G) during Weeks 1 and 2, these values were estimated for purposes of preparing the curves. Table 2 provides the standard deviations for the values obtained in each Group, which standard deviations are also indicated in FIG. 2.
  • Groups F, G and H are the same as Groups A, B and C (see Table 1).
  • FIG. 3 presents a set of high resolution radiograms of the entire left and right femurs from rats taken from some of the groups A-J described above.
  • the treatment modality is indicated in the lower left corner of the radiogram.
  • Control no surgery
  • Sham surgery without any bone marrow ablation
  • BMX bone marrow ablation
  • PBS phosphate buffer saline
  • PTH parathyroid hormone (PTH[l-34]-NH 2 )
  • CT salmon calcitonin.
  • the midshaft portion of the bone was chosen because, in contrast to measurements taken at the epiphysis portion of the bone, it is relatively simple at this alternate location to measure the increase in trabecular bone formed within the shaft marrow cavity because the bone marrow cavity normally does not contain trabecular bone, only bone marrow cells, unless and until the bone is treated by the method of the present invention, as described herein.
  • the bone marrow ablation (bmx) technique was practiced, in all cases, only on the left femur of the test animals. That is to say, there was no ablation of the marrow portion in the right femur.
  • FIGS .4 (a-d) graphically illustrate the formation of both trabecular bone and, to some extent, total bone, within the left femoral shaft in rats from the Groups described in Tables 1 and/or 2 above, wherein the left femur was subjected to a bmx (bone marrow ablation) treatment coupled with a concurrent and/or subsequent administration of one or more bone anabolic agents, including parathyroid hormone (PTH[I -34] -NH 2 ) and one or more anti- resorptive agents such as salmon calcitonin.
  • PTH[I -34] -NH 2 parathyroid hormone
  • anti- resorptive agents such as salmon calcitonin.
  • the least effective combination i.e., in improving bone density, the effect of which also diminished most rapidly over time, was bmx + a buffer (phosphate buffer saline or "PBS") wherein the buffer, taken by itself, has no known anabolic effect.
  • PBS phosphate buffer saline
  • FIGS. 4(b) and 4(d) are provided to illustrate the results obtained with the same animals' corresponding right femoral shafts, i.e, in which bone marrow ablation (bmx) was not performed. Little or no new bone formation was observed, i.e, the density remained substantially constant, since the only treatment received by the right femurs was due to the systemic administration of the bone anabolic agent. The agent(s) was administered for too short a period to have, by itself, any real positive effect on the bone density. In fact, the total bone density curve for the right femoral shaft presents a substantially straight line, which is attributable to the fact that the total density measurement includes a measurement of cortical bone. Furthermore, since the anabolic agent was not administered for a period sufficient to increase cortical bone density, the total density remained unchanged.
  • Table 3 sets forth the values upon which the curves of FIGs. 4 (a-d) are based.
  • the table additionally includes the standard deviations ("sd") for the values obtained for each analysis, which standard deviations are also indicated in FIGs. 4 (a-d).
  • FIG. 5 provides a graphical representation of the osteocalcin levels determined, at the time of sacrifice, in the serum of rats subjected to the following treatment modalities: bmx + PBS; bmx + parathyroid hormone (PTH[l-34]-NH 2 ); bmx + salmon calcitonin (CT); and bmx + parathyroid hormone (PTH[l-34]-NH 2 ) + CT.
  • bmx + PBS bmx + parathyroid hormone
  • CT salmon calcitonin
  • bmx + parathyroid hormone (PTH[l-34]-NH 2 ) + CT It is well known by those of ordinary skill in this art that osteocalcin is a biochemical marker related to the number of osteoblasts, i.e, a measure of new bone growth, which may be measured in the blood serum of the animal.
  • Table 4 (below) sets forth the values upon which the curves of FIG. 5 are based. Table 4 additionally provides the standard deviations for the values obtained during each of the indicated analyses, which standard deviations are also indicated in Figure 5.
  • FIG. 6 illustrates the results of a Micro Computerized Tomography (MicroCT) analysis of representative bones taken from the left femoral shaft of control and treated rats.
  • the modality of treatment is set forth in the lower left corner of each of the views, and the abbreviations used therein are the same as those which appear in FIGS. 3-5.
  • the Figure provides a view through a section of each such bone shaft.
  • FIG. 7 provides the results of MicroCT analysis on femoral sections obtained from the right femoral shaft of the same rats tested as in FIG. 6. As may be seen in the figure, there was substantially no additional newly formed bone within the interior marrow cavity of the bone shaft. This is due to the fact that the right femur did not undergo ablation treatment. Again, it is to be noted that any right femur view labeled with "bmx" should be taken to mean that it was the corresponding left femur which underwent bone marrow ablation. Any additional growth occurring within the right femoral shaft, therefore, is attributable to the systemic effect of the anabolic agent administered to the animal in question.
  • FIG. 8 provides views of stained and decalcified bone sections of the left femur of representative rats subjected to the following treatment modalities: Control day 0; Control day 21; Sham day 21; bmx day 0; bmx +PBS daily for 7 days; bmx + PBS daily for 14 days; bmx + PBS daily for 21 days; bmx + parathyroid hormone (PTH[1-34]NH 2 ) daily for 7 days; bmx + PTH daily for 14 days, and bmx + PTH daily for 21 days.
  • PTH[1-34]NH 2 parathyroid hormone
  • FIG. 9 is a view, taken at a higher magnification than the views in FIG. 8, within the left femoral bone marrow cavity of rats subjected to the following treatment modalities: a) control day 0, b) control day 21, c) sham day 21, d) bone marrow ablation (bmx) + phosphate buffer saline (PBS) daily for 7 days, e) bmx + PBS daily for 14 days, f) bmx + PBS daily for 21 days, g) bmx + parathyroid hormone (PTH[1-34]NH 2 ) daily for 7 days, h) bmx + PTH daily for 14 days, i) bmx + PTH daily for 21 days, j) bmx + salmon calcitonin (CT) daily from days 7-14, k) bmx + salmon calcitonin daily for days 7-21, 1) bmx + PTH daily for days 1-7, followed by salmon calcit
  • any views of the right femur in the figures described below indicating bone marrow ablation (bmx) as a treatment modality should be taken to mean that such treatment was applied to the corresponding left femur.
  • the samples described below with regard to FIGS. 10-13 were taken from the femur shafts, rather than the corresponding distal epiphysis as in the case of FIGS. 1 and 2.
  • the figures discussed below depict the results of end-point studies wherein the measurements were taken only at the 21 day end-point.
  • FIG. 10 (a-b) presents a set of high resolution radiograms of the entire left and right femurs of additional (to those described above) groups of rats, injected daily for a designated term with one or more of the following agents: phosphate buffer saline (PBS); parathyroid hormone (PTH[I -34] -NH 2 ); parathyroid hormone related peptide (PTHrP); and calcitonin gene related peptide (CGRP).
  • PBS phosphate buffer saline
  • PTH[I -34] -NH 2 parathyroid hormone related peptide
  • PTHrP parathyroid hormone related peptide
  • CGRP calcitonin gene related peptide
  • the rats chosen for test subjects were subjected to the following treatment modalities (as indicated in the lower left corner of each of the radiograms): control day 0; control day 21; bone marrow ablation (bmx) day 0; sham day 21; bmx followed by injection of PBS buffer daily for 21 days; bmx followed by injection of PTH daily for 21 days; bmx followed by injection of PTHrP daily for 21 days; and bmx followed by injection of CGRP daily for 21 days.
  • treatment modalities as indicated in the lower left corner of each of the radiograms: control day 0; control day 21; bone marrow ablation (bmx) day 0; sham day 21; bmx followed by injection of PBS buffer daily for 21 days; bmx followed by injection of PTH daily for 21 days; bmx followed by injection of PTHrP daily for 21 days; and bmx followed by injection of CGRP daily for 21 days.
  • the best results i.e., the greatest bone density
  • the lighter, i.e, more radio- opaque, areas indicate the locations of the greatest additional bone growth.
  • the best results were once again obtained with the use of the parathyroid hormone as the anabolic agent, but satisfactory results were also returned with the use of, respectively, PTHrP and CGRP as the anabolic agent.
  • the left femur showed a correspondingly greater improvement than did the right, which is again attributable to the bmx treatment accorded the left femur, in contrast to the right femur which underwent no such treatment.
  • FIG. 11 (a-d) graphically illustrates, by means of a bar graph, the improvements in trabecular bone density and, to some extent, total bone density, in the left femoral shaft of rats subjected to the treatment modalities described above with regard to FIG. 10 (a-b); that is, control day 0; bmx day 0; control day 21; sham day 21; bmx + PBS on day 21; bmx + PTH on day 21; bmx + PTHrP on day 21; and bmx + CGRP on day 21.
  • FIG. 1 l(a) the trabecular bone density of the animals' left femoral shaft was significantly improved following bone marrow ablation, in conjunction with which the animals were treated with, respectively, parathyroid hormone, PTHrP and CGRP.
  • FIG ll(c) also shows some increase in total bone density as determined in the left femoral shaft following bmx treatment linked with anabolic agent administration.
  • Table 5 (below) sets forth the values upon which the height of the bars in FIGs. 11 (a-d) are based.
  • the table additionally includes the standard deviations for the values obtained for each analysis, which standard deviations are also indicated in FlGs. 11 (a-d).
  • FIG. 12 is a bar graph which sets forth the osteocalcin levels determined, at the time of sacrifice, in the blood serum of the rats which were the subject of the analyses illustrated in FIGS. 10 and 11. As indicated above, it is well known that osteocalcin is a biochemical marker indicative of new bone growth. As can be seen in FIG. 12, the highest osteocalcin level, i.e, indicating the greatest bone growth, occurs due to the combination of bmx + the administration of parathyroid hormone (PTH[l-34]-NH 2 ) on a daily basis for 21 days.
  • PTH[l-34]-NH 2 parathyroid hormone
  • Table 6 (below) sets forth the values upon which the height of the bars in FIG. 12 are based.
  • the table additionally includes the standard deviations for the values obtained for each analysis, which standard deviations are also included in FIG. 12.
  • FIG. 13 (a-b) provides a comparison between Micro Computerized Tomography (MicroCT) analyses of bone samples taken from the left femoral shaft (FIG. 13 a) and the right femoral shaft (FlG. 13b) of rats treated in the manner indicated in FIGs. 10- 12.
  • MicroCT Micro Computerized Tomography
  • FIGs. 14A-14D The results set forth in FIGs. 1, 2, 4, 5, 11 and 12 and the corresponding Tables, Nos. 1, 2, 3, 4, 5 and 6 discussed above were obtained with the use of a method for mechanically inducing increased osteoblast activity as illustrated in FIGs. 14A-14D.
  • the method shown in FIGs. 14A-14D is provided only for the purpose of illustration, however, and the invention is not limited to the procedure(s) illustrated in the indicated Figures since a number of alternate techniques, which would be well known to those of ordinary skill in the art, may be used to obtain the necessary mechanical induction of osteoblast activity.
  • FIGs. 14A-14D The method shown in FIGs. 14A-14D is provided only for the purpose of illustration, however, and the invention is not limited to the procedure(s) illustrated in the indicated Figures since a number of alternate techniques, which would be well known to those of ordinary skill in the art, may be used to obtain the necessary mechanical induction of osteoblast activity.
  • the contents of the bone marrow cavity were back-flushed by injecting 5 ml of normal saline solution into the femur using a syringe attached to a 26-gauge needle (see FIG. 14C).
  • the ablation treatment may be carried out at any location on the bone, e.g., through the epiphysis, the midshaft portion, etc., at any angle, e.g, parallel to the shaft starting from the epiphysis, perpendicular to the bone at the midshaft, etc., and to any depth of penetration, according to the results which the treating physician desires to achieve, i.e, depending upon a particular application chosen for the technique.
  • the next step involved suturing the medial ligamentous structures with 4-0 Dexon thread and closing the skin incision with surgical metal clips (see FIG. 14D). Thereafter, each rat was injected with a 5ml bolus of saline and tagged for identification. During recovery, the rats were given Tylenol® solution (300 mg/kg/day) for the first 24 hours after surgery and then checked on a daily basis for the first 5 days.
  • each rat was euthanized in a CO 2 chamber.
  • the rat's blood was then collected via a cardiac puncture.
  • the femurs were removed and fixed in a 4% formalin solution, after which they were dehydrated in a graded series of ethanol solutions.
  • the bones were thereafter subjected to a variety of sample analysis techniques. These included X-ray analysis wherein the femurs were X-rayed in individually sealed plastic bags; PrXTMUS analysis using a Lunar PIXIMUS scanner to measure bone mineral density in the bone marrow cavity of the femurs; MicroCT analysis; and blood analysis for osteocalcin, PTH, CT, NTX and growth factors.
  • the bone anabolic agent may be administered to the subject contemporaneous with the mechanical inducement of osteoblast activity (whether by increased osteoblast formation and/or by increased bone formation by pre-existing osteoblasts), which mechanical inducement may be achieved, e.g., by alteration of the bone marrow cavity.
  • the mechanical inducement of osteoblast activity may be achieved, e.g., by alteration of the bone marrow cavity.
  • marrow and/or other components of the marrow cavity is/are removed under pressure (e.g., by altering the relative pressure within versus without the marrow cavity).
  • the bone anabolic agent is administered subsequent to such mechanical inducement.
  • the bone anabolic agent may be administered prior to mechanical inducement such that elevated levels of bone anabolic agent are already present at the time of mechanical inducement, which levels may then be maintained or continued intermittently for an extended period thereafter.
  • the bone anabolic agent may be administered orally, intravenously, intramuscularly, subcutaneously, via implant, transmucosally, transdermally, rectally, nasally, by depot injection or by inhalation and pulmonary absorption.
  • the bone anabolic agent may be administered once as a time release formulation, a plurality of times, or over one or more extended periods. It is preferred that elevated blood levels of the anabolic agent be maintained at least intermittently for between about 14-365 days, and more preferably for between about 30-180 days, post-mechanical induction.
  • parathyroid hormone e.g., PTH[l-34]-NH 2
  • PTH[l-34]-NH 2 Intermittent administration of parathyroid hormone, e.g., PTH[l-34]-NH 2 , could occur once daily or once weekly resulting in peaks of blood concentration that return to baseline levels between doses, but nevertheless result in periodic elevated blood levels of a bone anabolic agent in a manner that overlaps the elevated osteoblast activity that is initially induced mechanically, although thereafter sustained, at least in part, by the anabolic agent.
  • the anabolic agent is selected from the group consisting of a parathyroid hormone (PTH), anabolic Vitamin D analogs, a low-density lipoprotein receptor-related protein 5 (LRP5), an activator of non-genomic estrogen-like signaling (ANGELS), a bone morphogenic protein (BMP), an insulin-like growth factor (IGF), a fibroblast growth factor (FGF), sclerostin, leptin, a prostaglandin, a statin, strontium, a growth hormone, a growth hormone releasing factor (GHRF), hepatocyte growth factor (HGF), calcitonin gene related peptide (CGRP), parathyroid hormone related peptide (PTHrP), transforming growth factor (TGF)- ⁇ l and combinations thereof.
  • the term parathyroid hormone includes, but is not limited to natural parathyroid hormone, a truncate of natural parathyroid hormone, an amidated truncate of natural parat
  • the bone anabolic agent is truncated PTH[I -34] in the free acid form.
  • This material is commercially available in an FDA-approved pharmaceutical formulation from Eli Lilly & Co. under the trade name Forteo® (teriparatide).
  • Other useful bone anabolic agents for use with the invention include, but are not limited to, an amidated truncate of natural parathyroid hormone, PTH[l-30]NH 2 , PTH[l-31]NH 2 , PTH[1-32]NH 2 , PTH[I -33]NH 2 , PTH[I -34]NH 2 and combinations thereof.
  • the bone anabolic agent is PTH[I -34]NH 2 .
  • a sufficient amount of the preferred truncated parathyroid hormone as discussed herein is administered to the subject to achieve, and thereafter maintain, a pulsatile blood concentration thereof in the subject of between about 50 and 350 pg/ml, preferably between about 100 and 200 pg/ml, and most preferably about 150 pg/ml.
  • the blood concentration of the parathyroid hormone in the subject is raised to its preferred level by no later than 7 days following mechanical alteration of the contents of the bone marrow cavity.
  • an appropriate dosage of the PTH bone anabolic agent must be calculated to achieve the above- indicated blood concentrations.
  • the dose (in pure weight of the active hormone) given to, for example, a human subject may be that taught in the literature relating to the bone anabolic activity of these various agents.
  • Such dose may, but does not necessarily, range between about 10-200 ⁇ g, given once per day, more preferably between about 20-100 ⁇ g per dose and most preferably between about 20-50 ⁇ g per dose.
  • Dosage levels of injectable formulations comprising bone anabolic agents other than the above-described parathyroid hormone-based agents would be consistent with those noted above for the PTH agents.
  • the rats were randomized into the following groups: sham OVX, OVX + vehicle, OVX + PTH [1-3I]-NH 2 or PTH [1-34J-NH 2 (obtained from Unigene Laboratories, Inc.) at 2.5, 10 or 40 ⁇ g/kg/day subcutaneous, or PTH[l-34]-OH (obtained from Bachem) at 10 ⁇ g/kg/day subcutaneous.
  • the right femur of each animal was analyzed by DEXA and bone histomorphometry.
  • HG. 16 is a bar graph comparing the lumbar vertebral formation rates achieved with these same truncates. Based on the results of these experiments it was determined that the bone anabolic activity of each of these analogs of PTH is substantially equivalent. Because of the similarity of anabolic action among the above- discussed parathyroid hormones, it is therefore reasonable to expect that all known PTH analogs will advantageously function in the desired manner in the method of the present invention.
  • the mechanical induction of osteoblast activity is accomplished by inserting, into a bone marrow cavity of a bone targeted for enhanced bone formation, an object configured or adapted to physically alter the contents of the cavity and thereby to stimulate the osteoblast activity within the cavity.
  • the mechanical alteration may include removal of at least a portion of the cavity contents.
  • the method of the invention additionally comprises administering to the subject an antiresorptive agent for a time and at a concentration sufficient to substantially prevent resorption of the new bone produced due to the osteoblast activity.
  • the antiresorptive agent may be administered contemporaneous with the administration of the bone anabolic agent.
  • the antiresorptive agent is administered subsequent to the administration of the bone anabolic agent.
  • the administration of the antiresorptive agent may be commenced during administration of the bone anabolic agent and such administration may then be continued beyond the termination of administration of the bone anabolic agent.
  • a single agent may by administered having both bone anabolic and antiresorptive properties.
  • examples of such materials include, but are not limited to estrogen, strontium ranalate and selective estrogen receptor modulators (SERMS).
  • the antiresorptive agent may be a calcitonin selected from the group consisting of human calcitonin, salmon calcitonin ("sCT"), eel calcitonin, elkatonin, porcine calcitonin, chicken calcitonin, calcitonin gene related peptide (CGRP) and combinations thereof.
  • sCT salmon calcitonin
  • eel calcitonin el calcitonin
  • CGRP calcitonin gene related peptide
  • the antiresorptive agent is salmon calcitonin.
  • Blood levels of calcitonin, when used as the antiresorptive agent preferably range between about 5-500 pg/ml, more preferably between about 10-250 pg/ml and most preferably 20-50 pg/ml.
  • human dosage levels of the subject calcitonin agents necessary to achieve the above blood levels may be those taught in the literature relating to the use of these materials as anabolic agents. Such dose may, but does not necessarily, range between about 5- 200 /xg given once per day, more preferably between about 5-50 ⁇ g and most preferably 8-20 /xg by weight of the pure drug, administered daily. Salmon calcitonin (sCT) administered by alternate routes, i.e., by nasal or oral administration, would require higher dosages than those discussed above.
  • sCT Salmon calcitonin
  • additional antiresorptive agents i.e., other than the calcitonins
  • HRT agents include, generally, hormone replacement therapy (HRT) agents such as selective estrogen receptor modulators (SERMS), bisphosphonates, cathepsin-K inhibitors, strontium ranalate and various combinations thereof.
  • HRT agents include, but are not limited to, (1) Premarin® available from Wyeth Laboratories, which includes estrogen as the active ingredient. A typical accepted dosage is one 0.625 mg tablet daily; (2) Actonel® available from Proctor & Gamble, which includes, as its active ingredient, risedronate sodium.
  • a typical accepted dosage is one 5 mg tablet daily or one 35 mg tablet weekly; (3) Evista® sold by Eli Lilly & Co. which includes raloxifene HCl as the active ingredient.
  • a typical accepted dosage of this formulation is one 60 mg tablet taken daily; and (4) Fosamax® available from Merck Pharmaceuticals, which includes alendronate as the active ingredient. Typical dosages of this material are 10 mg/day or 70 mg/week.
  • the invention additionally provides a method for inducing bone formation within a bone of a subject which comprises mechanically altering the contents of a bone marrow cavity within a bone wherein such formation is to be induced to thereby stimulate, and thus increase osteoblast differentiation activity therein.
  • the method further comprises administering to such subject an amount of an antiresorptive agent for a time and at a concentration sufficient to substantially prevent resorption of the new bone produced due to the osteoblast activity.
  • the antiresorptive agent may be any of those identified above, but is preferably salmon calcitonin. It may be administered contemporaneous with the mechanical alteration of the marrow cavity and/or subsequent thereto. In this manner, the new bone growth attributable to the mechanical alteration treatment is substantially prevented from being resorbed.
  • This method does not include the administration of a bone anabolic agent.
  • dosages herein refer to the weight of the active compounds unaffected by pharmaceutical excipients, diluents, carriers or other ingredients, although such other ingredients are typically included in the variety of dosage forms useful in the method of the invention.
  • Any dosage form i.e., capsule, tablet, injection or the like
  • the terms "excipient”, “diluent” or “carrier” include such non- active ingredients as are typically included, together with active ingredients, in the industry.
  • typical capsules, pills, enteric coatings, solid or liquid diluents or excipients, flavorants, preservatives, or the like are included.
  • the attending clinician should monitor individual patient response, and adjust the dosage accordingly.
  • the antiresorptive agent may be administered orally, intravenously, intramuscularly, subcutaneously, via implant, transmucosally, rectally, nasally, by depot injection, by inhalation and pulmonary absorption or transdermally. Moreover, the antiresorptive agent may be administered once, a plurality of times, or over one or more extended periods.
  • the invention additionally provides a method of inducing bone formation in a subject suffering from diminished bone mass which comprises the steps of targeting for treatment at least one bone of the subject, wherein each targeted bone defines a bone marrow cavity therein.
  • the bone marrow cavity contains a quantity of bone marrow and a plurality of osteoblasts.
  • the method of the invention further comprises mechanically altering the contents of the bone marrow cavity to thereby stimulate and thus increase osteoblast activity therein. Thereafter, bone mass is increased within the cavity due to the increased osteoblast activity.
  • the method additionally comprises administering to the subject at least one bone anabolic agent for a duration and at a concentration sufficient to raise blood levels of the anabolic agent within the subject above natural levels thereof and thereby prolong the mechanically induced osteoblast activity.
  • the method then further comprises additionally administering, either (1) contemporaneous with, (b) overlapping with, or (3) subsequent to the administration of the bone anabolic agent, an antiresorptive agent for a duration and at a concentration sufficient to substantially prevent resorption of new bone produced due to the increased osteoblast activity achieved in accordance with the invention.
  • an antiresorptive agent for a duration and at a concentration sufficient to substantially prevent resorption of new bone produced due to the increased osteoblast activity achieved in accordance with the invention.
  • the bone anabolic agent may be selected from the group consisting of natural parathyroid hormone, a truncate of natural parathyroid hormone, an amidated truncate of natural parathyroid hormone, an amidated natural parathyroid hormone and combinations thereof.
  • the agent may be PTH- [1-34] in the free acid form, sold by Eli Lilly & Co. under the trade name Forteo® (teriparatide).
  • the bone anabolic agent is an amidated truncate of natural parathyroid hormone and may be selected from among PTH[l-30]NH 2 , PTH[1-31]NH 2 , PTH[1-32]NH 2 , PTH[1-33]NH 2 , PTH[1-34]NH 2 and combinations thereof.
  • a preferred choice for the bone anabolic agent is PTH[1-34]NH 2 .
  • a sufficient amount of an amidated truncate of natural parathyroid hormone is administered to the subject to achieve a pulsatile blood concentration thereof in the subject of between about 50 and 500 pg/ml, preferably between about 100-200 pg/ml and most preferably about 150 pg/ml.
  • the dose (in pure weight of the active hormone) given to a human subject may be that taught in the literature relating to the bone anabolic activity of these various agents.
  • Such dose may, but does not necessarily, range between about 10-200 ⁇ g per dose, more preferably between about 20-100 ⁇ g and most preferably between about 20-50 ⁇ g.
  • the dose may, but does not necessarily, range between about lO ⁇ g-lOmg.
  • the antiresorptive agent is a calcitonin selected from among human calcitonin, salmon calcitonin, eel calcitonin, elkatonin, porcine calcitonin, chicken calcitonin gene related peptide (CGRP) and combinations thereof.
  • the antiresorptive agent is salmon calcitonin. It is preferred to achieve blood levels of salmon calcitonin, when it is used as the antiresorptive agent, of between about 5- 500 pg/ml, more preferably between about 10-250 pg/ml and most preferably between about 20-50 pg/ml.
  • daily dosages of the antiresorptive agent used in, e.g., injectable formulations may range between about 5-200 ⁇ g (pure weight of the drug), more preferably between about 5-50 ⁇ g and most preferably between about 8-20 ⁇ g. When alternate delivery methods, i.e., other than injection, are used, the dose may range between about 5 ⁇ g-5mg.
  • Administration of the antiresorptive agent preferably continues for at least 3 months and more preferably between 12-24 months.
  • One embodiment of the invention comprises the use of any of the above described methods to form a sufficient amount of additional bone in a jaw region of the subject to provide an anchor for a dental implant implanted therein.
  • any of the methods described above may be utilized to form a sufficient amount of additional bone in one or more targeted bones of the subject to permit secure anchoring of a prosthetic device thereto.
  • Such prosthetic devices may include, but are not limited to, a prosthetic knee, shoulder or hip.
  • any of the methods of the invention may be utilized to form a sufficient amount of additional bone at any location in the subject to serve as a secure anchor for a hollow, adjustable insert anchored thereto.
  • any of the methods of the invention may be used in targeting at least one vertebra for additional bone formation, wherein a sufficient amount of bone is added to the at least one vertebra such that the subject is substantially freed from chronic pain caused due to vertebral crush.
  • the invention provides a kit for fostering bone formation in at least one targeted bone of a subject in need of such bone formation.
  • the kit comprises at least one container having therein at least one bone anabolic agent and a mechanical alteration device for altering contents of a bone marrow cavity in at least one targeted bone of a subject.
  • the kit may include at least one container having therein at least one bone anti-resorptive agent, as well as a mechanical alteration device for altering contents of the bone marrow cavity of a selected bone.
  • the kit may include multiple containers with at least one bone anabolic agent and at least one bone anti-resorptive agent, as well as the mechanical alteration device described above.
  • the kit may additionally be provided with an evacuation device for evacuating at least a portion of the contents from the bone marrow cavity.
  • the bone anabolic agent is selected from among natural parathyroid hormone, a truncate of natural parathyroid hormone, an amidated truncate of natural parathyroid hormone, an amidated natural parathyroid hormone, and combinations thereof.
  • the bone anabolic agent is a truncate of natural parathyroid hormone.
  • a preferred truncate for use as the agent is PTH[I- 34] in the free acid form.
  • Other preferred truncates include amidated truncates.
  • the bone anabolic agent may thus be selected from among PTH[l-30]NH 2 , PTH[l-31]NH 2 , PTH[I- 32]NH 2 , PTH[1-33]NH 2 , PTH[1-34]NH 2 and combinations thereof.
  • the bone anabolic agent is PTH[I -34]NH 2 .
  • the antiresorptive agent is a calcitonin selected from the group consisting of human calcitonin, salmon calcitonin, eel calcitonin, elkatonin, porcine calcitonin, chicken calcitonin, calcitonin related gene peptide (CGRP) and combinations thereof.
  • the antiresorptive agent is salmon calcitonin.

Abstract

L'invention porte sur une méthode visant à induire la formation des os chez un sujet, cette méthode consistant à induire mécaniquement une augmentation de l'activité des ostéoblastes chez le sujet et à élever la concentration sanguine d'au moins un agent anabolique des os du sujet. Les étapes de cette méthode peuvent être effectuées dans n'importe quel ordre, mais sur une durée suffisante pour que la concentration élevée de l'agent anabolique et l'augmentation, induite mécaniquement, de l'activité des ostéoblastes se chevauchent. Cette méthode peut, additionnellement, consister à doter le sujet d'une concentration sanguine élevée d'au moins un inhibiteur de la résorption osseuse, la concentration sanguine élevée étant suffisante pour prévenir la résorption d'une nouvelle croissance osseuse due à l'activité des ostéoblastes. L'utilisation de cette méthode permet de cibler des os spécifiques du sujet pour la formation et la préservation des os, une formation plus rapide des os et une interruption précoce des produits pharmaceutiques anaboliques. L'invention porte également sur des kits adaptés pour réaliser cette méthode.
PCT/US2005/040032 2004-05-14 2005-11-07 Methode pour stimuler la formation et la preservation des os WO2006124062A1 (fr)

Priority Applications (5)

Application Number Priority Date Filing Date Title
AU2005331907A AU2005331907B2 (en) 2004-05-14 2005-11-07 Method for fostering bone formation and preservation
CA002608016A CA2608016A1 (fr) 2005-05-11 2005-11-07 Methode visant a ameliorer la formation et la preservation des os consistant a induire chirurgicalement une augmentation de l'activite des osteoblastes couplee aux effets de l'administration de la pthrp ou du cgrp
CN2005800509720A CN101217972B (zh) 2005-05-11 2005-11-07 促进骨形成和保持的方法
JP2008511101A JP2008540522A (ja) 2005-05-11 2005-11-07 骨の形成および保存を促進するための方法
EP05851371A EP1879608A4 (fr) 2005-05-11 2005-11-07 Methode pour stimuler la formation et la preservation des os

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US11/128,095 2005-05-11
US11/128,095 US7531518B2 (en) 2004-05-14 2005-05-11 Method for fostering bone formation and preservation

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WO2006124062A1 true WO2006124062A1 (fr) 2006-11-23

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JP (1) JP2008540522A (fr)
CN (1) CN101217972B (fr)
CA (1) CA2608016A1 (fr)
WO (1) WO2006124062A1 (fr)

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CN111511900A (zh) * 2017-11-09 2020-08-07 南加利福尼亚大学阿尔弗雷德·E·曼恩生物医学工程研究所 用于骨再生的干细胞和装置

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CN101663043A (zh) * 2007-04-27 2010-03-03 尤尼基因实验室公司 促进和保持骨生长的方法和组合物
CN101663043B (zh) * 2007-04-27 2014-03-12 尤尼基因实验室公司 促进和保持骨生长的方法和组合物
JP2010032502A (ja) * 2008-06-26 2010-02-12 Okayama Univ 顎顔面インプラント治療のための骨質検査方法
CN111511900A (zh) * 2017-11-09 2020-08-07 南加利福尼亚大学阿尔弗雷德·E·曼恩生物医学工程研究所 用于骨再生的干细胞和装置
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Also Published As

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EP1879608A4 (fr) 2009-11-11
EP1879608A1 (fr) 2008-01-23
JP2008540522A (ja) 2008-11-20
CA2608016A1 (fr) 2006-11-23
CN101217972B (zh) 2013-05-29
CN101217972A (zh) 2008-07-09

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