The VALIDATE-D study is a unique randomized and double-blinded human intervention study that is designed to elucidate the mechanism by which vitamin D therapy may impart renoprotective benefits in diabetes. Vitamin D metabolism and regulation of the RAS are complex processes that involve multiple genes, gene-products, and environmental inputs (Figure 1); the genetic variation, dietary intake, and other environmental exposures for each individual have been shown to alter the balance of each of these hormonal systems [33–35]. In this regard, using supplementation with vitamin D3 in an intervention study to induce a clinical outcome that is mediated by the interaction between 1,25(OH)2D and the VDR (such as RAS inhibition to influence renal-vascular function) is analogous to administering a pre- or pro-drug that is dependent on multiple subsequent enzymatic steps, each regulated and modified by genetics and environment. The strengths of the VALIDATE-D study are that it focuses on overcoming this individual variability by investigating the influence of direct VDR-agonist therapy in modulating RAS activity and renal-vascular function, and employs a meticulous design to evaluate the underlying mechanism of potential findings (regulation of RAS activity).
Increasing evidence highlights a relationship between vitamin D and the RAS, with relevant implications for renal-vascular and kidney disease [1, 36]. Lowering unfavorable RAS activity has known vascular benefits in diabetes , and the use of RAS inhibitors is common practice to prevent or delay the progression of diabetic nephropathy. Animal studies have elegantly shown that activating the VDR results in lowering of the renal-vascular tissue-RAS, and that this interaction translates to reduced kidney injury, particularly in the setting of animal models of diabetes [19–21]. While human studies often focus on influencing clinical outcomes by supplementing with vitamin D3[5, 38], research studies that employ direct VDR-agonists may be most effective at invoking VDR-mediated effects such as RAS modulation [2, 39]. The advantage of using direct VDR-agonists in intervention studies is that they likely maximize any effect mediated by the VDR, eliminate inter-individual variations in vitamin D metabolism, and may influence clinical outcomes in a shorter duration of time than would take with vitamin D3 supplementation. The disadvantage of using VDR-agonists is that they are not commonly used in individuals without CKD and therefore direct clinical translation of study findings is limited; however, positive findings would support any therapy (vitamin D3 or VDR-agonist) that raises 1,25(OH)2D levels or activates the VDR as an equal surrogate.
The results of the VALIDATE-D study will shed light on: 1) the mechanism by which VDR activation influences renal-vascular function in human diabetes (via modulation of the RAS), and 2) whether VDR activation modulates renal hemodynamics and proteinuria in human diabetes. These findings would not only carry major implications for the recommendation of vitamin D supplementation in the primary prevention of diabetic nephropathy, but they could help interpret the findings of many other studies that evaluated clinical outcomes of vitamin D therapy without accounting for the mechanisms involved. In this regard, the VALIDATE-D study may explain the reasons prior negative studies failed to see outcomes and why positive studies may have succeeded.
At least three unique findings will emerge from the VALIDATE-D study. First, this intervention study will conclusively evaluate whether vitamin D therapy lowers the circulating human RAS. By using direct VDR agonists and a meticulous study design to account for confounders of the RAS (diet, posture, medications, time of day), this study will investigate whether calcitriol therapy lowers the maximally stimulated RAS on a LS diet in comparison to placebo. Second, this study will evaluate whether direct VDR activation improves renal-vascular hemodynamics by increasing renal blood flow and the renal-vascular sensitivity to angiotensin II; both of these observations are consistent with reductions in the renal-vascular tissue-RAS as would be induced by an ACE inhibitor. The study design includes a comparison of renal-vascular hemodynamics (while the circulating RAS is suppressed on a HS diet) during treatment with calcitriol vs. placebo, and then again during treatment with calcitriol + lisinopril vs. placebo + lisinopril (Figure 2). In this regard, the study design is optimized to decipher the individual role of VDR activation, and the effect of combining VDR activation with conventional RAS inhibition, since this combination therapy has been shown to provide synergistic renoprotection in animal models of diabetes [19–21]. The assessment of the renal-vascular tissue-RAS in humans is a unique and defining aspect of this study protocol. The renal vasculature is specifically sensitive to the effects of angiotensin II, and the magnitude of renal-vascular responses to angiotensin II is inversely proportional to the degree of local tissue-RAS activity [5, 11, 16]. High endogenous tissue-RAS activity results in a blunted local sensitivity to the effect of angiotensin II, while inhibiting the local renal-vascular tissue-RAS (for example with an ACE inhibitor) improves the renal-vascular sensitivity to angiotensin II  (Figure 3). Therefore, our ability to measure renal hemodynamics via para-aminohippurate clearance permits us to objectively ascertain the state of the renal-vascular tissue-RAS in response to drug intervention (calcitriol, lisinopril, placebo) and in response to provocation by angiotensin II. Lowering renal-vascular tissue-RAS activity in pathologic states such as diabetes and obesity with conventional pharmacotherapy (ACE inhibitor or angiotensin receptor blocker) has shown efficacy for the primary prevention of diabetic nephropathy [15, 40, 41]. Thirdly, the impact of adding chronic VDR-agonist therapy to ACE inhibitor therapy on proteinuria will be assessed. Since the study population is strategically selected to have diabetes without CKD, together these study findings will have implications for recommending vitamin D supplementation to lower unfavorable RAS activity for the primary prevention of diabetic nephropathy.
We have previously shown in a pilot study that high-dose vitamin D3 therapy to markedly raise 1,25(OH)2D concentrations can lower renal-vascular tissue-RAS activity in non-diabetics with normal renal function . Like an ACE inhibitor, this intervention induced a favorable state of low tissue-RAS resulting in increased renal-vascular sensitivity to angiotensin II . The VALIDATE-D study will extend these findings by employing a double-blinded randomization, employing direct VDR agonist therapy, and focusing on patients with diabetes without CKD. In prior clinical studies of secondary prevention in diabetes with pre-existing CKD, direct VDR agonists (calcitriol and paracalcitol) reduced proteinuria [7, 42–45]; whether this effect was mediated by the RAS, or could be extrapolated for the primary prevention of diabetic nephropathy, are questions that the VALIDATE-D study will clarify.
VALIDATE-D is the first human study to investigate whether vitamin D therapy to activate the VDR can lower circulating and renal-vascular tissue-RAS activity in diabetics without CKD. This study is designed to specifically focus on whether modulating the RAS represents a key mechanism by which vitamin D intervention impacts renal-vascular hemodynamics and kidney function. In this regard, VALIDATE-D will provide evidence that may influence clinical recommendations for maintaining vitamin D status for the primary prevention of diabetic nephropathy. In addition, VALIDATE-D will examine whether the action of potent VDR agonist therapy complements the known beneficial effects of ACE inhibition, as has been observed in animal models . If so, the results of VALIDATE-D could influence pharmacologic practices by favoring vitamin D3 supplementation instead of, or in addition to, ACE inhibitor therapy for patients with diabetes.