Faculty Bibliography

BACKGROUND: Systemic administration of sclerostin neutralizing antibody has led to increased bone formation in animal models of osteoporosis. The purpose of this study was to determine if systemic administration of sclerostin neutralizing antibody could increase the healing response in a critical-sized femoral defect in rats. METHODS: Critical-sized femoral defects were created in Lewis rats, and the rats were randomized into four groups. The sclerostin antibody (Scl-Ab) treatment groups included the continuous Scl-Ab group (twenty-one animals), the early Scl-Ab group (fifteen animals), and the delayed Scl-Ab group (fifteen animals), which received sclerostin antibody (25 mg/kg) twice weekly for weeks 0 through 12; weeks 0 through 2; and weeks 2 through 4; respectively. Twenty-one animals in the control group received vehicle from weeks 0 through 12. In a subsequent study, bone turnover markers were measured at zero, two, six, and twelve weeks after surgery in rats receiving vehicle or sclerostin neutralizing antibody for twelve weeks (fifteen rats per group). The quality of bone formed was evaluated with radiographs, microcomputed tomography, biomechanical testing, and histologic and histomorphometric analysis. RESULTS: In the primary study, four of fifteen defects in the continuous (zero to twelve-week) Scl-Ab group, three of fifteen defects in the early (zero to two-week) Scl-Ab group, and four of fifteen defects in the delayed (two to four-week) Scl-Ab group healed at twelve weeks, while none of the defects healed in the control group. In both studies, treatment with sclerostin antibody for twelve weeks demonstrated a significant increase in new bone formation (p < 0.05) compared with the control group. The three treatment groups did not differ significantly with respect to the healing rates and the quality of new bone formed in the defect. The serum markers of bone formation were significantly elevated in the animals in the continuous Scl-Ab group (p < 0.05) compared with the controls. CONCLUSIONS: Administration of sclerostin neutralizing antibody led to increased bone formation, resulting in complete healing of femoral defects in a small subset of rats, with a majority of the animals not healing the defect by twelve weeks.

Cadaver experiments and clinical studies suggest that the superior labrum-biceps complex plays a role in glenohumeral stability. Superior labrum anterior to posterior (SLAP) tears can be present in acute and recurrent glenohumeral dislocations and contribute to glenohumeral instability. Isolated SLAP tears can cause instability, especially in throwing athletes. Diagnosing a SLAP tear on the basis of the clinical examination alone is difficult because of nonspecific history and physical examination findings and the presence of coexisting intra-articular lesions. Magnetic resonance arthrography is the imaging study of choice for diagnosing SLAP tears; however, arthroscopy remains the gold standard for diagnosis. Arthroscopy is the preferred technique for the repair of a type II SLAP tear and its variant types (V through X) in acute glenohumeral dislocations and instability in younger populations. Clinical outcome studies report a low recurrence of glenohumeral instability after the arthroscopic repair of a SLAP tear in addition to a Bankart repair. Long-term follow-up studies and further advances in arthroscopic fixation techniques will allow a better understanding and improvement in outcomes in patients with SLAP tears associated with glenohumeral instability.

Bone tissue has an exceptional quality to regenerate to native tissue in response to injury. However, the fracture repair process requires mechanical stability or a viable biological microenvironment or both to ensure successful healing to native tissue. An improved understanding of the molecular and cellular events that occur during bone repair and remodeling has led to the development of biologic agents that can augment the biological microenvironment and enhance bone repair. Orthobiologics, including stem cells, osteoinductive growth factors, osteoconductive matrices, and anabolic agents, are available clinically for accelerating fracture repair and treatment of compromised bone repair situations like delayed unions and nonunions. Preclinical and clinical studies using biologic agents like recombinant bone morphogenetic proteins have demonstrated an efficacy similar or better than that of autologous bone graft in acute fracture healing. A lack of standardized outcome measures for comparison of biologic agents in clinical fracture repair trials, frequent off-label use, and a limited understanding of the biological activity of these agents at the bone repair site have limited their efficacy in clinical applications.

Ex-vivo regional gene therapy with bone marrow cells (BMCs) overexpressing bone morphogenetic protein-2 (BMP-2) has demonstrated efficacy in healing critical sized bone defects in preclinical studies. The purpose of this preclinical study was to compare the osteoinductive potential of a novel "same day" ex-vivo regional gene therapy versus a traditional two-step approach, which involves culture expansion of the donor cells before implantation. In the "same day" strategy buffy coat cells were harvested from the rat bone marrow, transduced with a lentiviral vector-expressing BMP-2 for 1 hour and implanted into a rat femoral defect in the same sitting. There was no significant difference (P = 0.22) with respect to the radiographic healing rates between the femoral defects treated with the "same day" strategy (13/13; 100%) versus the traditional two-step approach (11/14; 78%). However, the femoral defects treated with the "same day" strategy induced earlier radiographic bone healing (P = 0.004) and higher bone volume (BV) [micro-computed tomography (micro-CT); P < 0.001]. The "same day" strategy represents a significant advance in the field of ex-vivo regional gene therapy because it offers a solution to limitations associated with the culture expansion process required in the traditional ex vivo approach. This strategy should be cost-effective when adapted for human use.

The purpose of this study was to investigate the influence of combined inhibition of receptor activator of nuclear factor kappa-B ligand (RANKL) and bone morphogenetic protein (BMP) activity in a mixed lytic/blastic prostate cancer lesion in bone. Human prostate cancer cells (C4 2b) were injected into immunocompromised mice using an intratibial injection model to create mixed lytic/blastic lesions. RANK-Fc, a recombinant RANKL antagonist, was injected subcutaneously three times a week (10mg/kg) to inhibit RANKL and subsequent formation, function and survival of osteoclasts. Inhibition of BMP activity was achieved by transducing prostate cancer cells ex vivo with a retroviral vector expressing noggin (retronoggin; RN). There were three treatment groups (RANK-Fc treatment, RN treatment and combined RN and RANK-Fc treatment) and two control groups (untreated control and empty vector control for the RN treatment group). The progression of bone lesion and tumor growth was evaluated using plain radiographs, hindlimb tumor size, (18)F-Fluorodeoxyglucose and (18)F-fluoride micro PET-CT, histology and histomorphometry. Treatment with RANK-Fc alone inhibited osteolysis and transformed a mixed lytic/blastic lesion into an osteoblastic phenotype. Treatment with RN alone inhibited the osteoblastic component in a mixed lytic/blastic lesion and resulted in formation of smaller osteolytic bone lesion with smaller soft tissue size. The animals treated with both RN and RANK-Fc demonstrated delayed development of bone lesions, inhibition of osteolysis, small soft tissue tumors and preservation of bone architecture with less tumor induced new bone formation. This study suggests that combined inhibition of the RANKL and the BMP pathway may be an effective biologic therapy to inhibit the progression of established mixed lytic/blastic prostate cancer lesions in bone.

Multidetector Computed Tomography (MDCT) technology plays an important role in the evaluation of injured patients. At our institution pelvic X-ray (PXR) is obtained routinely on trauma patients. Many also receive MDCT of the abdomen and pelvis for other indications. We hypothesized that there would be a substantial cost savings in adopting a policy of deferring PXR in a hemodynamically normal patient who will also proceed to MDCT for other indications. We retrospectively reviewed the charts of trauma patients from February 1, 2008 to February 1, 2009. We reviewed whether a PXR was done, the result, whether an MDCT was also done, and the presence or absence of pelvic fractures. We collected billing and cost data from various hospital sources. We identified 1,330 patients with PXR between February 1, 2008 and February 1, 2009. Of those patients, 810 (61%) had MDCT after PXR. Sixty-six patients (8.0%) had pelvic fractures; 39 were correctly identified on PXR (59% of fractures). Twenty-seven were detected only by MDCT (41% of fractures); all pelvic fractures were identified on MDCT. Seven hundred and forty-four patients (92% of patients with both PXR and MDCT) had negative PXR and negative MDCT. Using three methods of cost analysis, the estimated cost savings range is from $77,011 to $331,080. MDCT of the pelvis is more sensitive and more specific than PXR. In patients who are hemodynamically normal and asymptomatic, forgoing routine PXR could result in an estimated savings from $77,011 to $331,080, depending on the method used to calculate costs.

BACKGROUND: Both adenoviral and lentiviral vectors have been successfully used to induce bone repair by over-expression of human bone morphogenetic protein 2 (BMP-2) in primary rat bone marrow stromal cells in pre-clinical models of ex vivo regional gene therapy. Despite being a very efficient means of gene delivery, there are potential safety concerns that may limit the adaptation of these viral vectors for clinical use in humans. Recombinant adeno-associated viral (rAAV) vector is a promising viral vector without known pathogenicity in humans and has the potential to be an effective gene delivery vehicle to enhance bone repair. In this study, we investigated gene transfer in rat and human bone marrow stromal cells in order to evaluate the effectiveness of the self-complementary AAV vector (scAAV) system, which has higher efficiency than the single-stranded AAV vector (ssAAV) due to its unique viral genome that bypasses the rate-limiting conversion step necessary in ssAAV. METHODS: Self-complementaryAAV2 encoding GFP and BMP-2 (scAAV2-GFP and scAAV2-BMP-2) were used to transduce human and rat bone marrow stromal cells in vitro, and subsequently the levels of GFP and BMP-2 expression were assessed 48 hours after treatment. In parallel experiments, adenoviral and lentiviral vector mediated over-expression of GFP and BMP-2 were used for comparison. RESULTS: Our results demonstrate that the scAAV2 is not capable of inducing significant transgene expression in human and rat bone marrow stromal cells, which may be associated with its unique tropism. CONCLUSIONS: In developing ex vivo gene therapy regimens, the ability of a vector to induce the appropriate level of transgene expression needs to be evaluated for each cell type and vector used.

BACKGROUND: The efficacy of prostate cancer gene therapy is limited by the inefficiency of prostate-specific promoters as compared to ubiquitous viral promoters. The purpose of this investigation was to evaluate the specificity and efficacy of a lentiviral vector driven by a PSCA promoter. METHODS: Prostate cancer (LNCap, C42-B, and LAPC-4) and non-prostate cancer (HeLa, MB231, and MCF-7) cells were transduced with a lentiviral vector expressing either the luciferase or the HSV-TK suicide gene and driven by a short PSCA promoter. Specificity and efficacy were evaluated in vitro and in vivo. RESULTS: Luciferase expression was only detected in prostate cancer cells and was comparable to the universal CMV promoter. Luciferase expression in prostate cancer cells cultured with androgen was higher than that in cells cultured without androgen. In subsequent cytotoxicity experiments in which the luciferase marker gene was replaced with the HSV-TK gene, the lentiviral vector harboring the PSCA promoter induced cytotoxicity in prostate cancer cell lines while demonstrating a minimal effect on non-prostate cells. Cellular toxicity was correlated to increasing concentrations of the prodrug ganciclovir. Androgen had a positive effect on the cytotoxicity of this lentiviral construct. Intratumoral injection of prostate cancer xenografts with the lentiviral construct induced tumor growth inhibition versus saline controls. CONCLUSION: Our results indicate that a lentiviral gene therapy vector driven by a short PSCA promoter can induce prostate-specific cellular toxicity in vivo and in vitro and may provide a strategy to selectively treat local and advanced metastatic prostate cancer. Prostate 69: 1422-1434, 2009. (c) 2009 Wiley-Liss, Inc.

BACKGROUND: Mesenchymal stem cells are multipotent and have the ability to differentiate into bone. We conducted a preclinical trial comparing the osteogenic potential of human bone marrow aspirate with that of mesenchymal stem cell-enriched bone marrow aspirate (both mixed with demineralized bone matrix) in a critical-sized rat femoral defect model. METHODS: The buffy coat was extracted from human bone marrow aspirate to obtain mesenchymal stem cell-enriched bone marrow aspirate. Fifty-nine athymic rats, each with a 6-mm femoral defect, were divided into six treatment groups: defect only (Group I), demineralized bone matrix and saline solution (Group II), demineralized bone matrix and bone marrow aspirate (Group III), demineralized bone matrix and mesenchymal stem cell-enriched bone marrow aspirate (Group IV), demineralized bone matrix and recombinant human bone morphogenetic protein-2 (rhBMP-2) (Group V [positive control]), and absorbable collagen sponge and rhBMP-2 (Group VI [positive control]). All animals were killed at twelve weeks for radiographic, micro-computed tomography, histomorphometric, and histologic analysis. RESULTS: There was wide variability in the mesenchymal stem cell concentrations obtained from the human donors. All ten defects healed in the positive control groups (Groups V and VI). Only one defect healed in each experimental group (Groups II, III, and IV) (i.e., three of forty-four defects healed). There was no significant difference among the radiographic scores of Groups II, III, and IV (p = 0.59), and the score for each of those groups was significantly higher than that for Group I (p

PMID: 19411455

ISSN: 1535-1386

CID: 1667812