Novel Biomaterials To Control Igf-I Binding And Enhance Chondrocyte Gene Therapy
Articular cartilage lesions could lead to Osteoarthritis (OA) due to the inability of cartilage to heal. Gene therapy could be used to genetically engineer chondrocytes to repair cartilage lesions. A current challenge in gene therapy is the transient exposure of the gene product and growth factor to the cells. This transient exposure of the growth factor is not sufficient to promote tissue generation. This thesis reports the development of a new material engineered to have high affinity for a therapeutic gene product. The growth factor studied in this thesis is insulin-like growth factor-I (IGF-I) for its highly anabolic effects in cartilage. One of the ways that cartilage stores IGF-I is through a group of insulin-like growth factor binding proteins (IGFBPs), such as IGFBP-5. IGF-I binding peptide sequence from IGFBP-5 was grafted onto alginate in order to retain the endogenous IGF-I produced by transfected chondrocytes. The chosen peptide sequence was KPLHALL due to its proximity to IGF-I in vivo. However, the reaction efficiency of KPLHALL is affected by the electrically charged side chain of Lysine, Ɛ amino group NH3+. In order to perform the carbodiimide chemistry, the Ɛ amino group NH3+ had to be protected with a protecting group. After attaching the amino acid sequence, the protecting group had to be removed with a base or acid. Such treatments can reduce the reaction efficiency by possibly reacting with the alginate or with other peptide sequences in its vicinity. The deprotection step could be avoided by using peptide sequences that avoid Lysine, but such modifications may affect the affinity of the peptide to IGF-I. Therefore, new materials were developed to test the relationship between the peptide lengths, the efficiency of grafting, and their binding affinity for IGF-I. KPLHALL, PLHALL and ALL sequences were compared by assessing the efficiency of grafting to alginate, the ability of modified polymers to bind IGF-I, and the ability to enhance matrix production by transfecting chondrocytes to produce IGF-I. This study reinforced the importance of IGF-I and the utility of controlling both growth factor production and binding, yielding an increase in matrix production.
Osteoarthritis; Growth Factor, IGF-I; Binding Peptide
Putnam,David A.; Fortier,Lisa Ann
Ph.D. of Biomedical Engineering
Doctor of Philosophy
dissertation or thesis