234 Boatright and Boden have reported on a pilot study of single-level anterior lumbar interbody fusions in humans comparing rhBMP-2/collagen sponge-filled cages with iliac crest autograft-filled cages. All 11 patients random- ized to the rhBMP group were fused at 6 mo postoperatively, while one of the three patients randomized to the control group receiving autograft in their cages was finally deemed a nonunion at 1 yr (82). Since that time over 350 patients have received that combination of cage/BMP-2/collagen sponge, and the extremely high success rate resulted in approval of rhBMP-2 (InFuse Bone Graft, Medtronic Sofamor Danek, Memphis TN) by the US Food and Drug Administration for use inside tapered fusion cages for anterior lumbar interbody fusion. Early pilot studies using rhBMP-2 with a hydroxyapatite/ tricalcium phosphate carrier matrix have yielded encouraging results for posterolateral lumbar spine fusions. Other bone-inducing growth factors that have been evaluated include rhBMP-7 and growth and differentiation factor-5. Cook et al. have investigated rhBMP-7, also known as osteogenic protein-1 (Stryker Biotech, Hopkinton, MA) extensively in long-bone defect models, where it has been found to be an effective bone generator in combination with collagen matrix (83,84). Further work by this group utilized a canine spinal fusion model to demonstrate successful rapid posterior spinal fusion when comparing rhBMP-7 to autograft (85). rhBMP-2 has also yielded a high rate of posterolateral spine fusions in the rabbit model. Early results from clinical trials in posterolateral spine have demon- strated fusion rates based only on plain radiographs (not CT scans) of 50–70%. Growth and differen- tiation factor 5 (GDF-5), another member of the transforming growth factor-β (TGF-β) superfamily, has also been shown to be effective in a long-bone defect model in rats and subsequently in a rabbit spinal fusion model. Spiro et al. used a rabbit posterolateral intertransverse process fusion model to compare rhGDF-5 delivered in a mineralized collagen osteoconductive bone graft matrix (Healos, Orquest, Mountainview, CA) with iliac crest autograft. The rhGDF-5/Healos combination functioned as a bone graft substitute performing as well as autograft alone (18). THE FUTURE IS HERE, BUT CHALLENGES REMAIN As described earlier, the ideal bone generator for clinical use in spinal surgery will function to induce the migration of cells capable of becoming bone-forming cells and then activate the system of signals necessary to affect these cells to differientiate into osteoblasts. This bone generator must also supply the proper spatial environment for these bone-forming cells to function in; this requires that neovascularization occur in proximity to surface areas that provide physiologically resorbable scaf- folding to act as a template for the various cells involved in bony remodeling. In this manner, the grafted material can be replaced by functional bone that can be maintained physiologically over the patient’s lifetime. As the necessary ingredients for a bone generator are better understood, it becomes clearer why no single substitute has been able to supplant autograft. It is also easier to explain why even autograft is not uniformly successful, because at times it fails to provide a sufficient quantity of osteoinductive substances over an appropriate time course once it has been devitalized by the grafting process. Focus has now shifted to synthesis of a composite that maximizes the potential of each ingredient. Growth factors and an adequate supply of progenitor cells are the key to osteoinductivity. As dis- cussed in the previous section, the glycoprotein molecules of the BMP family are effective bone-gen- erating growth factors. The challenge now lies in delivering a potent growth factor over the appropriate time course for each specific clinical need. The time course for many spinal fusion models appears to be protracted over several months, especially in larger animal models. The normal physiological half- life of glycoprotein molecules in the cellular environment is measured in hours and days, not the weeks or months necessary for spinal fusion in primates. In addition, it is necessary to find a “growth factor” that works early enough in the cascade of events leading to bone formation that all of the conditions for bone formation will be in place at a clinical This is trial version www.adultpdf.com Biology of Spine Fusion 235 site with appropriate physiological timing. The ideal factor will initiate bone formation by triggering the construction of the biochemical bone-forming environment, attracting and effecting differentia- tion of osteoprogenitor cells, and then potentiating the activity of those cells involved in physiologi- cal bone formation and remodeling. As more physiological environments are characterized, the complexity of each has become increas- ingly evident. It is likely that bone generation requires a molecular milieu that is provided at specific phases of the wound-healing process. During each phase, a different milieu of permissive factors is available. These factors are substances such as transforming growth factor-β and fibroblast growth factor. It is important that these permissive and/or potentiating factors be present within the bone- forming environment for factors such as the BMPs to be maximally effective (86,87). Thus exogenous growth factors must be delivered appropriately in both a spatial and a temporal sense. Strategies for accomplishing this have included the utilization of differing doses and/or carriers with different breakdown rates, in the hope that some of the growth factor will remain and be avail- able at the appropriate times. Pilot studies by Boden et al. have proved that it is possible for BMP to induce bone consistently in humans, but both NeOsteo and rhBMP-2 require higher dosing and take longer for osteoinduction in primates than in smaller animals (70,82). These data prove that these sub- stances can be effective in primates, but the high doses necessary and the length of time to fusion dem- onstrate the need to refine these systems before they will be clinically practical. One major strategy is to develop a better delivery system for the growth factor. Multiple alterna- tives have been explored, which utilize the various available osteoconductive substances soaked with growth factors. These synthetic bone-graft substitute materials integrated with rhBMP have been explored in several posterolateral canine fusion models. Sandhu et al. found that rhBMP-2 in a polylac- tic acid carrier was superior to autogenous iliac crest bone graft for inducing transverse process arthrod- esis (73). Also in a canine posterolateral spine fusion model, Muschler et al. reported that rhBMP-2 in a similar biodegradable copolymer carrier of polylactic acid and glycolic acid had equivalent fusion rates and strength to autograft (74). Gene therapy is a more sophisticated delivery system for growth factors. Utilizing various molec- ular strategies, genes encoding for factors of the bone formation cascade are inserted into the patient’s own cells that exist at the site for fusion (in vivo) or that have been removed and will be reimplanted at the site of fusion (ex vivo) (88). Once these cells are in place, they will then produce a protein product from the transfected gene that leads to bone formation. In this manner, the half-life of the cell or the gene within the cell and not the actual glycoprotein is the limiting temporal factor for presence of a specific growth factor at the fusion site. This strategy has been used in a rat posterolateral spine fusion model with excellent results. Boden et al. have reported on the use of a novel protein that was isolated via molecular methods and appears to function very early in the cascade of events leading to bone formation (89). This intracellular sig- naling protein, named LIM mineralization protein-1 (LMP-1), has been isolated and its gene identified. This gene was then transfected into the harvested bone marrow cells of rats and reimplanted at sites for posterolateral spine fusion. Nine of nine (100%) sites implanted with cells containing the LMP-1 gene fused solidly, while 0/9 (0%) sites implanted with control cells fused (90) (Fig. 6). This study validates the feasibility of local gene therapy to induce bone formation and spinal fusion. A more recent study has demonstrated that ordinary white blood cells from venous blood can be used to deliver the LMP-1 gene with a low dose of adenovirus to achieve successful spine fusion (91). Optimizing gene therapy introduces even more challenges to the search for an ideal bone genera- tor. Vectors for the delivery of genes into cells, the types of cells transfected, and the control of gene expression are all areas to be explored. As knowledge of each growth factor and its mechanism of action is elucidated, the most potent factor can be identified and exploited. 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Friedlaender © Humana Press Inc., Totowa, NJ 13 Bone Allograft Transplantation Theory and Practice Henry J. Mankin, MD, Francis J. Hornicek, MD, PhD, Mark C. Gebhardt, MD, and William W. Tomford, MD INTRODUCTION An amazing change has occurred in the last 30 yr in our ability to care for children and adults with bone sarcomas. Because of better imaging technology and vast improvements in our systems for treat- ing sarcoma with chemotherapy, we no longer use amputations as our first line of care and now perform limb-sparing surgery for most of our patients. The success of metallic implants is very attractive, and many centers use these technologies (1–14) in the treatment of high-grade tumors involving a joint, but in addition there is a long history in orthopedics of the use of allograft implants (15–42). The latter system is intriguing in many ways and may in fact outlive the metallic implants over time. The aim of this chapter is to review the history of allografting, describe the current state of knowledge, present our series of over 1000 cases and their complications, and then try to establish some rules and approaches to alloimplants of the future. HISTORY In the entire world of orthopedics, there has never been a more wished-for “dream” or sought after “holy grail,” than osteochondral allograft implantation. When a limb is grossly diseased, a bone badly deformed, or a joint totally disabled, both the patient and physician fervently wish that they could start over with a new part, anatomically identical to the old but disease-free and completely functional. The concept of a well-accepted, low-complication, fully functional bone and cartilage alloimplant is a hope that has prevailed for centuries and remains at the present still not quite in reach. Grafts are available in appropriate sizes and shapes, the tissue is accepted with minimal problem in many cases, but still the perfect graft eludes us and remains a “dream” or perhaps may be described as the “holy grail of reconstructive orthopedics.” The dream is ancient, presumably occurring in many caretakers over the centuries but recogniz- ably remembered as the “miracle” performed by Saints Cosmas and Damian in the sixth century AD (43–45). The saints were twin physicians born in the third century AD in the town of Egea in Cilicia in Asia Minor. They were the sons of a physician and then became physicians, traveling widely in Greece, Turkey, and Rome, treating ailments and refusing payment for their services. They somehow angered Lysia, the Roman governor of Cilicia during the persecution of the Christians by the emperor Diocletian, and after a variety of attempts at killing the twins, they and their three brothers were beheaded and buried in a grave in Egea on September 27, 287 AD (43–45). They were returned, however, in the fifth century to a basilica in the Roman Forum, which now bears their name, where Deacon Justinian, a faithful church retainer with a cancerous limb was so exhausted by the pain that he fell asleep during his prayers. There came to him in a dream the twin physicians, who, after amputating the limb of a This is trial version www.adultpdf.com 242 Mankin et al. Moor who had died that morning, replaced the diseased part with the obtained allograft implant. The procedure, known as the “Miracle of the Black Leg,” was reportedly successful, and because of that, the twins were subsequently beatified, receiving their sainthood in approximately the year 550 AD. Of note is the fact that the occasion and drama associated with the procedure was so extraordinary that it captured the imagination of first the painter Fra Angelico and then many other artists; and literally hundreds of some of the most extraordinary paintings depicting the procedure can now be found in many of the world’s museums (43) (Fig. 1). In his exhaustive report on the history of allografting, Burwell (46) records several attempts by individuals over the many years that followed, but the world recognizes the first report of a successful alloimplant to be that of Macewen in 1881 (47). In that procedure, Macewen transplanted segments of bone from a rachitic patient to the humerus of a 3-yr-old child who had lost a portion of the shaft as a result of osteomyelitis. The major effort, however, in the early part of the 20th century was that Fig. 1. Painting by Pedro de Berruguete in the 15th century hanging in the Collegiate Church in Covarrubias. Note that the saints are performing the surgical procedure on the right lower extremity and that Damian, the surgeon in the foreground, is using his left hand to suture the host–donor junction site. This is trial version www.adultpdf.com Bone Allograft Transplantation 243 of Lexer, who in 1908 reported on four such procedures about the knee (48) and in 1925 described a reasonable success rate on 11 half joints and 23 whole joints using fresh cadaveric tissue (49). Sporadic case reports and short series were presented over the next 20 or so years, but it was a Russian group under the direction of Volkov (50) who reported a large series of successful procedures using processed but not frozen cadaveric bone. On the basis of a sophisticated group of experimental studies, Curtiss, Chase, and Herndon (51,52) proposed the concept that freezing the cadaveric bony parts would reduce immunological activity and thus reduce the rejection rate. This also made it possible to develop bone banks in which bony parts obtained at surgery or autopsy (or subsequently at harvest) were stored in a freezer at −20 to −70°C and thawed prior to implantation (16,46,53). Following World War II, the US Navy became interested in preservation of allograft tissue, and in 1950 founded the Navy Tissue Bank under the direction of George Hyatt (54). Subsequently, when Kenneth Sell became head, he recruited a number of Fellows to rotate through the system and perform research on graft technology. The list of graduates of Kenneth Sell’s program included some very distinguished investigators, such as Andrew Bassett, Gary Friedlaender, Theodore Malinin, William Tomford, and Michael Strong, all of whom started their own banks and also performed very competent research (34,55–69). Their work, along with Sell’s, not only advanced the field in terms of improved success of the implant, but also added greatly to the safety of the recipient in relation to infectious bacterial and especially virus transmission (55,64,69–72). On the basis of these pioneer efforts, two major sets of experimentation started. The first of these was clinical. Frank Parrish in Houston, acting in part on the reported success of Volkov, performed a series of surgical procedures in which frozen allografts were implanted after removal of a bone tumor (73,74). He carefully followed the patients and reported the complications of the procedure (73). Carlos Ottolenghi in Buenos Aires started a similar series and reported on successes, and most importantly, on the causes of failures (75). Stimulated by these efforts, several other groups began to look at allograft- ing as a possibly better solution than metallic implants and further advanced the search for the “holy grail” (18,20,21,25,27,29,31,33,34,76–81). During this same period, several investigators recognized that the complications, including infec- tion, fracture, and nonunion, that compromised the results in the clinical series were probably based on the immune response and began seeking a greater understanding of this phenomenon (57,77,82–85). A group in Canada headed by Langer demonstrated that the response to allografting in animals was markedly reduced by freezing the graft, suggesting that a blocking antibody was produced by the tem- perature reduction (86). Similar attempts to define the immune response in animal systems were reported by Burchardt (82,83), Elves (87,88), and Stevenson (84,89–92), but it seemed that these data were really not as applicable to humans (93). More recently, the studies of Friedlaender and Strong and their group showed that the clinical result was significantly improved in patients who achieved a match with MHC Class II antigens than with MHC Class I or with mismatch (65,94). Simultaneously, the rules regarding bone banking were being established in a number of centers. Methods of testing the donor for bacterial or viral diseases were established, as well as approaches to the optimal rules for freezing and thawing (most believe that slow freezing and rapid thawing is the most successful [62,66,68,95]) and the value and drawbacks of radiation to the graft (71,96). It seemed sensible to maintain cartilage at least partially alive during the freezing and thawing process, and the use of glycerol or dimethylsulfoxide (DMSO) was proposed to achieve this important goal (97,98). Establishing the Bone section of the American Association of Tissue Banks and promulgating Guidelines and Standards were major steps forward and allowed safe bone banks to spring up through- out the United States and Europe (67,68). CURRENT STATUS It is possible to summarize the current status of our understanding of the issues surrounding allo- graft transplantation as follows. The response to allograft implantation appears to be species-dependent This is trial version www.adultpdf.com [...]... Orthop 2 27, 666– 677 34 Mnaymneh, W and Malinin, T (1989) Massive allografts in surgery of bone tumors Orthop Clin N Am 20, 455–4 67 35 Ortiz-Cruz, E., Gebhardt, M C., Jennings, L C., Springfield, D S., and Mankin, H J (19 97) The result of transplantation of intercalary allografts after resection of tumors A long term follow-up study J Bone Joint Surg 79 A, 97 106 36 Mnaymneh, W A., Temple, H T., and Malinin,... 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M and Regazzoni, P, eds.), Springer-Verlag, Berlin, pp 145–150 68 Tomford, W W and Mankin, H J (1999) Bone banking: update on methods and materials Orthop Clin N Am 30, 553–565 69 Tomford, W W., Thongphasuk, J., Mankin, H J., and Ferraro, M J (1990) Frozen musculoskeletal allografts A study of the clinical incidence and causes of infection associated with their use J Bone Joint Surg 72 A, 11 37 1143 70 . include rhBMP -7 and growth and differentiation factor-5. Cook et al. have investigated rhBMP -7 , also known as osteogenic protein-1 (Stryker Biotech, Hopkinton, MA) extensively in long -bone defect. (15,20,24,25,30,32,34,35, 38,40–42,69 ,71 ,72 ,77 ,102,104–106). In similar fashion, cartilage is known to be highly antigenic and has been shown to evoke a pro- found cellular and humoral antibody response (18,28,63, 87, 97, 98) mass and composition of fracture callus and bone. Scand. J. Rheumatol. 9, 1 67 171 . 30. Tornkvist, H., Lindholm, T. S., Netz, P., Stromberg, L., and Lindholm, T. C. (1984) Effect of ibuprofen and