The intrarenal renin-angiotensin system (RAS) plays a key role in development of diabetic nephropathy. Recently we showed that combination therapy with an AT1 receptor blocker (ARB) and an activated vitamin D analog produced excellent synergistic effects against diabetic nephropathy, as a result of blockade of the ARB-induced compensatory renin increase. Given the diversity of vitamin D analogs, here we used a prodrug vitamin D analog doxercalciferol (1-hydroxyvitamin D₂) to further test the efficacy of the combination strategy in long-term treatment. Streptozotocin-induced diabetic DBA/2J mice were treated with vehicle, losartan, doxercalciferol (0.4 and 0.6 µg/kg) or losartan and doxercalciferol combination for 20 weeks. Vehicle-treated diabetic mice developed progressive albuminuria and glomerulosclerosis. Losartan alone moderately ameliorated kidney injury, with renin being drastically up-regulated. A similar therapeutic effect was seen with doxercalciferol alone, which markedly suppressed renin and angiotensinogen expression. Losartan and doxercalciferol combination most effectively prevented albuminuria, restored glomerular filtration barrier structure and dramatically reduced glomerulosclerosis in a dose-dependent manner. These effects were accompanied by blockade of intrarenal renin up-regulation and Ang II accumulation. These data demonstrate an excellent therapeutic potential for doxercalciferol in diabetic renal disease and confirm the concept that blockade of compensatory renin increase enhances the efficacy of RAS inhibition and produces synergistic therapeutic effects in combination therapy.