Hemophilia A is the most common, inherited severe bleeding diathesis and is due to a deficiency of functional Factor (F) VIII. Two challenges to current FVIII replacement therapy are maintaining sustained FVIII corrections and treating the 30% population of Hemophilia A patients who develop FVIII inhibitors. Gene therapy promises to be an attractive treatment for hemophilia A as there is a wide therapeutic window for FVIII corrective plasma levels. Since platelets target to hemostatic injuries, ectopically expressed platelet (p) FVIII is an attractive alternative FVIII delivery strategy. We have already shown the effectiveness of platelet human B-domainless factor VIII (phBF8) in the presence of circulating inhibitors in FVIIInull mice. However, the clots formed are unstable and lead to increased embolization with phBF8. This dissertation describes platelet-specific gene therapy using three FVIII variants with enhanced activity: inactivation resistant FVIII (IR8), canine (c) B-domainless FVIII (cBF8), and hBF8R1645H. Each improved clotting efficacy in FVIIInull mice and decreased embolic risk though to different degrees. For example, we found that despite pcBF8 having ~30% antigenic levels of phBF8, it corrected clot instability seen in a cremaster injury model in FVIIInull mice. Since IR8 does not bind FVIII carrier protein, Von Willebrand Factor (vWF), efficiently, we tested the effectiveness of this variant in the presence of circulating FVIII inhibitors, and found that vWF binding was not needed for IR8 to be effective. To understand the cellular mechanism for decreased cBF8 relative to hBF8 expression level in megakaryocytes, we studied mRNA levels and the effect of pcBF8 on megakaryopoiesis and found that the level of cBF8 is related to increased cellular apoptosis. Finally to understand the mechanistic basis of pcBF8 increased hemostatic effectiveness, we examined whether mutation of the cleavage site for the Golgi apparatus enzyme PACE/furin in hBF8 to that of cBF8, which increases the FVIII’s activity, would also enhance hBF8R1645H pFVIII activity and affirm that this occurs. These studies provide new insights into pFVIII during megakaryopoiesis and how to optimize pFVIII hemostatic efficacy that should be useful in translating this therapeutic model to a large animal hemophilia A model and future clinical trials.