Review
25-Hydroxyvitamin D-24-hydroxylase (CYP24A1): Its important role in the degradation of vitamin D

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Abstract

CYP24A1 is the cytochrome P450 component of the 25-hydroxyvitamin D3-24-hydroxylase enzyme that catalyzes the conversion of 25-hydroxyvitamin D3 (25-OH-D3) and 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) into 24-hydroxylated products, which constitute the degradation of the vitamin D molecule. This review focuses on recent data in the CYP24A1 field, including biochemical, physiological and clinical developments. Notable among these are: the first crystal structure for rat CYP24A1; mutagenesis studies which change the regioselectivity of the enzyme; and the finding that natural inactivating mutations of CYP24A1 cause the genetic disease idiopathic infantile hypercalcemia (IIH). The review also discusses the emerging correlation between rising serum phosphate/FGF-23 levels and increased CYP24A1 expression in chronic kidney disease, which in turn underlies accelerated degradation of both serum 25-OH-D3 and 1,25-(OH)2D3 in this condition. This review concludes by evaluating the potential clinical utility of blocking this enzyme with CYP24A1 inhibitors in various disease states.

Highlights

► Synopsis of anabolic and catabolic pathways of vitamin D stressing role of CYP24A1. ► CYP24A1 structure based upon rat crystal structure, homology models and mutagenesis. ► Natural mutations of human CYP24A1 cause idiopathic infantile hypercalcemia. ► Physiological role of CYP24A1 and regulation by 1,25-(OH)2D3, PTH and FGF-23. ► Upregulated CYP24A1 in chronic kidney disease and use of CYP24A1 inhibitors.

Introduction

The mitochondrial enzyme, 25-hydroxyvitamin D3-24-hydroxylase was first described in the early 1970s and initially believed to be involved solely in the renal 24-hydroxylation of 25-OH-D3 [1]. Work performed over the last 35 years has shown that the enzyme activity is the result of a combination of three components: ferredoxin, ferredoxin reductase and cytochrome P450, only the latter component, referred to as CYP24A1, being specific for this reaction [2], [3]. It is now known that CYP24A1 catalyzes the conversion of both 25-OH-D3 and 1,25-(OH)2D3 into 24-hydroxylated products targeted for excretion along well-established pathways. We review current knowledge of the structure and function of this protein as well as providing an update on the latest information regarding the physiological and clinical importance of CYP24A1.

Section snippets

CYP24A1: biochemistry and catalytic properties

Though, CYP24A1 was initially referred to as the 25-hydroxyvitamin D3-24-hydroxylase, work with the recombinant enzyme has shown that it is able to catalyze multiple hydroxylation reactions at carbons C-24 and C-23 of the side chain of both 25-OH-D3 and its hormonal form, 1,25-(OH)2D3 [2], [3]. Indeed, our view of the role of CYP24A1 has expanded greatly to suggest that this single P450, alone, is responsible for the 5-step, 24-oxidation pathway from 1,25-(OH)2D3 to produce calcitroic acid, a

CYP24A1: crystal structure, homology modeling and mutagenesis studies

In 2010, the crystal structure of the rat CYP24A1 was elucidated but in the presence of the detergents Cymal and CHAPS [11]. Although the active site of rat CYP24A1 did not contain its natural substrate, for the most part the crystal structure did confirm the predicted tertiary structure of the protein, as well as the putative active-site residues from previous homology models and mutagenesis studies [10], [12], [13], [14], [15]. The crystal structure of rat CYP24A1 reveals a canonical

CYP24A1: physiological role

During the same period of time in which the role of CYP24A1 in multi-step hydroxylation of the side-chain of vitamin D was being elucidated, it was also shown that the enzyme is expressed in many, if not all, target cells containing the vitamin D receptor (VDR),1

CYP24A1: regulation by 1,25-(OH)2D3, PTH and FGF-23

Vitamin D signaling plays a critical role in regulating bone and mineral homeostasis and consequently, enzymes such as CYP24A1 which control vitamin D levels are regulated by hormones which are integral to mineral metabolism [2] (see Fig. 6). This is an important consideration in diseases such as chronic kidney disease where key factors such as PTH and FGF-23 increase in a maladaptive response to loss of kidney function. Both PTH and FGF-23 directly affect vitamin D metabolism; overproduction

CYP24A1: pharmacological role

In addition to its involvement in the catabolism of 1,25-(OH)2D3, CYP24A1 plays a crucial role in the clearance of various vitamin D analogs, especially those used in the treatment of CKD around the world: 19-nor-1,25-(OH)2D2 (paricalcitol or Zemplar), 1α-OH-D2 (doxercalciferol or Hectorol) and 22-oxa-1,25-(OH)2D2 (OCT or Maxacalcitol) [48], [8], [49]. All of these vitamin D analogs are vulnerable to metabolism at C-24 and/or C-23 and are thus inactivated in the target cells by CYP24A1. This is

CYP24A1: human polymorphisms and genome-wide linkage studies

Mining of several genomic databases reveals that a number of polymorphisms of CYP24A1 have been identified in recent years and the list is growing rapidly (Fig. 7). Though little is known of the effects of these polymorphisms on CYP24A1 enzyme activity, inactivating mutations would be expected to give rise to a hypercalcemic phenotype, and since these had not been reported, it was presumed that these polymorphisms must be innocuous. However, the recent reports of human inactivating mutations

CYP24A1: pathological role and implications in disease

The attenuation of vitamin D signaling by CYP24A1 has been implicated in a number of diseases including metabolic bone disease, chronic kidney disease and several types of cancer [59], [60].

CYP24A1 and genetically-linked idiopathic infantile hypercalcemia

Hypercalcemic conditions are not uncommon in the pediatric literature but they appear to be a heterogeneous group of diseases including: Williams–Beuren syndrome and idiopathic infantile hypercalcemia (IIH); all characterized by transient hypercalcemia and other features. Of these, only IIH has unknown etiology and until recently, had no gene locus assigned to it [62], [63]. Fifty years ago, a group headed by the famous US physician Harold Harrison [64] proposed that:

“….IIH is a metabolic

CYP24A1 and genetically-linked hypophosphatemia

The link between vitamin D metabolism and hypophosphatemic conditions was first established in the hypophosphatemic mouse model (Hyp), a murine homologue of X-linked hypophosphatemic rickets (XLH) in humans, initially characterized by Eicher et al. [67]. More recently, it has been shown that deletion in the 3′ region of the Phex gene [68], [69] is associated with hypophosphatemia and impaired bone metabolism arising, in part, from defective renal reabsorption of inorganic phosphate at the brush

CYP24A1: involvement in chronic kidney disease

This subject has been extensively reviewed by Petkovich and colleagues [59], [60]. Vitamin D deficiency is commonly observed in patients with CKD and is causally related to secondary hyperparathyroidism, a disorder characterized by elevated serum intact PTH levels, parathyroid gland hyperplasia and imbalances in bone and mineral metabolism [86], [87]. Declining renal mass and concomitant loss of renal CYP27B1 capacity in CKD is commonly associated with reductions in circulating levels of both

CYP24A1: involvement in pathogenesis and treatment of hyperproliferative disorders

The initial demonstration that 1α,25-(OH)2D3 is an anti-proliferative, pro-differentiating agent for certain cell types in vivo and many cell lines in vitro [95], coupled with the fact that cancer cell studies have showed decreased CYP27B1 and increased CYP24A1 expression in prostatic, colonic and breast cell lines as they progress towards a more tumorigenic phenotype [96], [97], [98], [99], [100] has caused some researchers to speculate that cancer progression involves dysfunctional vitamin D

CYP24A1 inhibitors

The possibility that increased CYP24A1 expression may be an underlying cause of vitamin D deficiency and progression of disease states suggests that this enzyme might be a potential therapeutic target. Over the past decade, a number of inhibitors have been developed in the treatment of diseases associated with elevation of vitamin D catabolism including

  • (a)

    General azole-based CYP24 inhibitors, such as ketoconazole and liarazole, which bind heme at the catalytic core of the protein [57], [91].

  • (b)

Perspectives

This is an exciting time for research on CYP24A1. The elucidation of the crystal structure of the rat enzyme [11] opens the door to determining the precise positioning of vitamin D substrates in the substrate-binding pocket. This in turn will allow us to better understand the mechanism of multiple hydroxylations and will facilitate the development of a second generation of CYP24A1 inhibitors using rational drug design [54].

Some of the first generation of CYP24A1 inhibitors [112] have reached an

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