Paul L. Kaplan

We report the results of a long-term follow-up of subjects in a phase 1 study of AAV2-hAADC (adeno-associated virus type 2–human aromatic L-amino acid decarboxylase) gene therapy for the treatment of Parkinson's disease (PD). Ten patients with moderately advanced PD received bilateral putaminal infusions of either a low or a high dose of AAV2-hAADC vector. An annual positron emission tomography (PET) imaging with [18F]fluoro-L-m-tyrosine tracer was used for evaluation of AADC expression, and a standard clinical rating scale [Unified Parkinson's Disease Rating Scale (UPDRS)] was used to assess effect. Our previous analysis of the 6-month data suggested that this treatment was acutely safe and well tolerated. We found that the elevated PET signal observed in the first 12 months persisted over 4 years in both dose groups. A significantly increased PET value compared with the presurgery baseline was maintained over the 4-year monitoring period. The UPDRS in all patients off medication for 12 hr improved in the first 12 months, but displayed a slow deterioration in subsequent years. This analysis demonstrates that apparent efficacy continues through later years with an acceptable safety profile. These data indicate stable transgene expression over 4 years after vector delivery and continued safety, but emphasize the need for a controlled efficacy trial and the use of a higher vector dose. Mittermeyer and colleagues report results from a phase 1 study of AAV2-hAADC (human aromatic l-amino acid decarboxylase) gene therapy for the treatment of Parkinson's disease. The study shows that AAV2-hAADC treatment was safe and that AADC gene expression was maintained at least 4 years after administration of therapy. Positron emission tomography analysis showed that a similar number of neurons continued expressing the transgene throughout the study.

We have examined culture fluids from a variety of Kirsten murine sarcoma virus (KiMSV) transformed rat and mouse cells for the presence of factors which induce normal Rat-1 cells to assume the transformed phenotype. All KiMSV transformants produced transforming factor (TF). Revertants of KiMSV transformed rat or mouse failed to release TF as did normal rat or mouse cells. Cells transformed by a temperature sensitive mutant of KiMSV produced TF at the permissive temperature but not at the nonpermissive temperature. Further, cells from a spontaneous transformant of Rat-1 cells also produced TF. TF is a small polypeptide which competes for the epidermal growth factor receptor. Its effect upon normal cells is reversible and requires de novo RNA and protein synthesis. Cells treated with TF lose the actin fibers observed in normal fibroblasts, assume a transformed cell morphology, become anchorage independent for growth, grow in low concentrations of serum, grow to a high cell density, and have an increased rate of hexose uptake.

Osteogenic protein-1 (OP-1) is a member of the transforming growth factor beta superfamily and is a potent modulator of osteogenesis and bone cell differentiation. This preclinical study in dogs sought to assess the effects of OP-1 on periodontal wound healing in surgically created critical size Class III furcation defects. Eighteen male beagle dogs were subjected to the creation of bilateral mandibular 5 mm osseous defects. A split-mouth design was utilized which randomly assigned opposing quadrants to control therapy (surgery alone or collagen vehicle) or 1 of 3 ascending concentrations of OP-1 in a collagen vehicle (0.75 mg OP-1/g collagen, 2.5 mg/g, or 7.5 mg/g). Thus, 9 quadrants per test group received OP-1, 9 quadrants per control group received surgery alone, and 9 quadrants received collagen vehicle alone. Test articles were delivered by a surgeon masked to the treatment, and fluorogenic bone labels were injected at specified intervals post-treatment. Eight weeks after defect creation and OP-1 delivery, tissue blocks of the mandibulae were taken for masked histomorphometric analysis to assess parameters of periodontal regeneration (e.g., bone height, bone area, new attachment formation, and percent of defect filled with new bone). Histomorphometry revealed limited evidence of osteogenesis, cementogenesis, and new attachment formation in either vehicle or surgery-alone sites. In contrast, sites treated with all 3 concentrations of OP-1 showed pronounced stimulation of osteogenesis, regenerative cementum, and new attachment formation. Lesions treated with 7.5 mg/g of OP-1 in collagen regenerated 3.9+/-1.7 mm and 6.1+/-3.4 mm2 (mean +/-S.D.) of linear bone height and bone area, respectively. Furthermore, these differences were statistically different from both control therapies for all wound healing parameters (P < 0.0001). No significant increase in tooth root ankylosis was found among the treatment groups when compared to the surgery-alone group. We conclude that OP-1 offers promise as an attractive candidate for treating severe periodontal lesions.

Kirsten murine sarcoma virus (KiMSV)-transformed rat-1, normal rat kidney (NRK), and BALB/c 3T3 cells are capable of continual growth in a serum-free medium supplemented with transferrin and insulin but with no exogenous mitogenic growth factors. Cells transformed by a mutant of KiMSV that is temperature sensitive for the maintenance of transformation grow in this medium at the permissive temperature only. At the nonpermissive temperature, growth is dependent upon the presence of serum-free conditioned medium from the transformed cells. Normal rat-1 cells are also dependent upon factors from the transformed cells for growth in this serum-free/mitogen-free medium. The serum-derived growth factors, epidermal growth factor, and fibroblast growth factor have no effect on the transformed cells, although epidermal growth factor can replace transforming growth factors produced by KiMSV-transformed cells for the growth of rat-1 cells. Growth of the transformed cells in serum-free medium at clonal densities is dependent upon the presence of conditioned medium collected from the same cells grown to high densities. These results show that (i) growth in serum-free/mitogen-free medium is a general property of KiMSV-transformed cells and (ii) growth of the transformed cells in this medium is dependent upon the presence of growth factors known to be produced by the cells, and they provide support for the hypothesis that serum-free growth of KiMSV-transformed cells is dependent upon ectopically produced growth factors.