Calcifediol (DB00146): A Comprehensive Pharmacological and Clinical Monograph

Executive Summary

Calcifediol, also known as 25-hydroxycholecalciferol or 25-hydroxyvitamin D3 (25(OH)D3​), is a pivotal prohormone and the major circulating metabolite within the vitamin D endocrine system. Synthesized in the liver via the hydroxylation of its precursor, cholecalciferol (vitamin D3), Calcifediol serves as the primary substrate for the renal and extra-renal production of calcitriol (1,25(OH)2​D3​), the biologically active form of vitamin D.[1] Its central role in mineral homeostasis forms the basis of its therapeutic applications.

The principal indication for Calcifediol, particularly in its extended-release formulation (Rayaldee®), is the treatment of secondary hyperparathyroidism (SHPT) in adult patients with Stage 3 or 4 chronic kidney disease (CKD) and vitamin D insufficiency (serum total 25(OH)D levels < 30 ng/mL).[4] This specific indication leverages its unique pharmacological profile. The mechanism of action involves its conversion to calcitriol, which subsequently binds to the vitamin D receptor (VDR), modulating gene expression to increase intestinal calcium and phosphorus absorption, enhance renal calcium reabsorption, and provide negative feedback to the parathyroid glands, thereby suppressing parathyroid hormone (PTH) synthesis and secretion.[1]

Pharmacokinetically, Calcifediol distinguishes itself from its precursor, cholecalciferol. It exhibits superior intestinal absorption and, by bypassing the requisite hepatic 25-hydroxylation step, provides a more direct and reliable means of elevating serum 25(OH)D levels. Clinical studies have demonstrated that Calcifediol is approximately two to three times more potent and acts more rapidly than cholecalciferol in correcting vitamin D deficiency.[7] This enhanced potency is particularly advantageous in patient populations with compromised liver function or intestinal malabsorption.

The safety profile of Calcifediol is well-characterized, with hypercalcemia representing the primary dose-limiting toxicity. This risk is managed through careful dose titration and regular monitoring of serum calcium, phosphorus, PTH, and 25(OH)D levels.[9] Notably, the medication does not carry a Black Box Warning from the U.S. Food and Drug Administration (FDA), indicating that its risks are considered manageable within standard clinical practice.

The global regulatory landscape for Calcifediol is diverse. While the FDA and the European Medicines Agency (EMA) have approved the specialized extended-release formulation for the niche indication of SHPT in pre-dialysis CKD, Australia's Therapeutic Goods Administration (TGA) has approved an immediate-release formulation (Vistella®) for the broader treatment of vitamin D deficiency in the general adult population.[4] Furthermore, European authorities have also authorized its use as a novel food ingredient in supplements, reflecting its dual role as both a potent therapeutic agent and a nutritional compound.[12] This monograph provides an exhaustive review of Calcifediol's chemical properties, pharmacology, clinical efficacy, safety, and regulatory status.

Drug Identification and Physicochemical Profile

Precise identification of a pharmaceutical agent is fundamental to clinical practice, research, and regulatory affairs. This section provides a comprehensive summary of the nomenclature, unique identifiers, and core physicochemical properties of Calcifediol.

Nomenclature and Identifiers

Calcifediol is recognized by a variety of names and codes across different scientific and commercial contexts.

• Generic/Common Names: The most widely used generic names are Calcifediol and Calcidiol.[1]
• Chemical Names: In scientific literature, it is frequently referred to by its chemical identity as 25-hydroxycholecalciferol, 25-hydroxyvitamin D3, or the abbreviation 25(OH)D3​.[2]
• Brand Names: Commercially, Calcifediol is available under several brand names, including Rayaldee® (an extended-release formulation), Calderol®, Vistella®, Dedrogyl®, Didrogyl®, and Hidroferol®.[14]
• Systematic (IUPAC) Name: The preferred International Union of Pure and Applied Chemistry (IUPAC) name for the compound is (1S,3Z)-3--7a-methyloctahydro-4H-inden-4-ylidene}ethylidene]-4-methylidenecyclohexan-1-ol.[2] Other systematic names, such as (3β,5Z,7E)-9,10-Secocholesta-5,7,10(19)-triene-3,25-diol, are also used.[18]
• Database Identifiers: To facilitate unambiguous data retrieval, Calcifediol is assigned unique identifiers in major chemical and pharmacological databases:
• DrugBank ID: DB00146 [1]
• CAS Number: 19356-17-3 (anhydrous form); 63283-36-3 (monohydrate form) [2]
• ChEBI: 17933 [2]
• PubChem CID: 5283731 [2]
• UNII: T0WXW8F54E [2]
• KEGG: C01561 [2]

Molecular and Chemical Properties

The therapeutic behavior of Calcifediol is dictated by its molecular structure and chemical characteristics.

• Chemical Formula: The molecular formula for the anhydrous form of Calcifediol is C27​H44​O2​.[1] The monohydrate form, which is also used in pharmaceutical preparations, has the formula C27​H46​O3​.[20]
• Molecular Weight: The average molecular weight of anhydrous Calcifediol is approximately 400.64 g/mol, with a monoisotopic mass of 400.334131 Da.[1] The monohydrate form has a molecular weight of approximately 418.65 g/mol.[20]
• Chemical Structure: Calcifediol is a secosteroid, a class of steroids in which one of the rings has been broken. It is structurally derived from its precursor, cholecalciferol (vitamin D3), through the enzymatic addition of a hydroxyl group (-OH) at the carbon-25 position of the side chain.[2]
• Physicochemical Characteristics:
• Appearance: In its pure form, Calcifediol is a solid.[24] In pharmaceutical formulations like Vistella®, it is presented as a clear, low-viscosity liquid contained within soft gelatin capsules.[11]
• Solubility: As a hydroxylated metabolite of vitamin D3, Calcifediol is more polar than its precursor.[25] It is soluble in organic solvents such as ethanol and dimethyl sulfoxide (DMSO) but is considered insoluble in water.[22]
• Stability and Storage: For laboratory use, long-term stability (up to 12 months) is best achieved by storing the compound at -20°C under desiccating conditions.[22] Commercially prepared pharmaceutical products are typically stored at room temperature, protected from moisture and heat.[16]

The following table consolidates the key identification and physicochemical data for Calcifediol.

Table 1: Key Identifiers and Physicochemical Properties of Calcifediol

Property	Value/Identifier
Generic Name	Calcifediol, Calcidiol
IUPAC Name	(1S,3Z)-3--7a-methyloctahydro-4H-inden-4-ylidene}ethylidene]-4-methylidenecyclohexan-1-ol
Common Synonyms	25-hydroxycholecalciferol, 25-hydroxyvitamin D3, 25(OH)D3
Key Brand Names	Rayaldee®, Vistella®, Calderol®
DrugBank ID	DB00146
CAS Number (Anhydrous)	19356-17-3
CAS Number (Monohydrate)	63283-36-3
Chemical Formula (Anhydrous)	C27​H44​O2​
Chemical Formula (Monohydrate)	C27​H46​O3​
Average Molecular Weight (Anhydrous)	400.64 g/mol
Molecular Weight (Monohydrate)	418.65 g/mol
Solubility	Soluble in ethanol, DMSO; Insoluble in water
Storage Conditions	-20°C (pure compound); Room temperature (pharmaceutical product)

In-Depth Pharmacology

The clinical utility of Calcifediol is rooted in its intricate pharmacology, which encompasses its role as a prohormone, its influence on gene transcription via the vitamin D receptor, and its ultimate physiological effects on mineral homeostasis. Its pharmacokinetic profile, particularly when compared to its precursor cholecalciferol, is a critical determinant of its therapeutic application in specific patient populations.

Mechanism of Action

Calcifediol's mechanism of action is best understood as a multi-step process that begins with its formation and culminates in the modulation of gene expression in target tissues.

Prohormone Activation Pathway

Calcifediol is not the final active hormone but rather a crucial intermediate. The process begins with vitamin D3 (cholecalciferol), which is either synthesized in the skin upon exposure to ultraviolet light or obtained from dietary sources. This inactive form is transported to the liver, where it undergoes a critical hydroxylation reaction at the 25-carbon position, catalyzed predominantly by the microsomal enzyme vitamin D 25-hydroxylase (CYP2R1).[2] The product of this reaction is Calcifediol (

25(OH)D3​), the major circulating form of vitamin D in the bloodstream.[1]

Calcifediol then serves as the primary substrate for a second hydroxylation step. This reaction is catalyzed by the enzyme 25-hydroxyvitamin D3-1-alpha-hydroxylase (CYP27B1), which is located primarily in the mitochondria of the proximal tubules of the kidneys.[1] This conversion yields calcitriol (

1,25(OH)2​D3​), the most biologically potent and active form of vitamin D.[1] The activity of CYP27B1 is tightly regulated by factors such as PTH, serum calcium, and fibroblast growth factor 23 (FGF23), ensuring precise control over active hormone production.[2]

Genomic Action via Vitamin D Receptor (VDR)

The physiological effects of the vitamin D system are mediated by calcitriol's interaction with the vitamin D receptor (VDR), a member of the nuclear receptor superfamily.[1] Calcitriol binds with high affinity to the VDR located within the cytoplasm of target cells, which are found in tissues such as the intestine, bone, kidneys, and parathyroid glands.[1]

Upon binding, the VDR undergoes a conformational change and forms a heterodimeric complex with another nuclear receptor, the retinoid-X receptor (RXR).[6] This activated VDR-RXR complex translocates into the cell nucleus. There, it functions as a transcription factor, binding to specific DNA sequences known as Vitamin D Response Elements (VDREs), which are located in the promoter regions of hundreds of target genes.[6] This binding event typically initiates or enhances the transcription of these genes, although instances of transcriptional suppression are also known.[1] The resulting changes in protein synthesis are responsible for the downstream physiological effects of vitamin D.

Direct and Non-Genomic Actions

While the classical pathway proceeds through calcitriol, emerging research indicates that Calcifediol itself is not entirely inert. Studies have shown that Calcifediol can bind directly to the VDR, albeit with a lower affinity than calcitriol, and can exert some gene-regulatory effects.[29] Furthermore, evidence suggests the existence of rapid, non-genomic actions that do not involve changes in gene transcription. These effects are thought to be mediated by the interaction of vitamin D metabolites, including Calcifediol, with specific membrane-associated VDRs (mVDR) and other cell surface proteins such as protein disulfide isomerase family A member 3 (Pdia3).[29] These pathways may contribute to the broader spectrum of vitamin D's biological activities.

Pharmacodynamics

The pharmacodynamic effects of Calcifediol administration are the direct physiological consequences of increased calcitriol production and subsequent VDR activation. These effects are centered on the regulation of mineral homeostasis and PTH secretion.

Regulation of Mineral Homeostasis

The primary pharmacodynamic outcome is the maintenance of serum calcium and phosphorus concentrations within a narrow physiological range. This is accomplished through a coordinated set of actions on multiple organ systems [1]:

1. Increased Intestinal Absorption: VDR activation in the enterocytes of the small intestine upregulates the transcription of genes coding for proteins essential for mineral transport. This includes the apical calcium channel TRPV6 and the intracellular calcium-binding protein calbindin-D9k, which together facilitate the efficient absorption of dietary calcium and phosphorus from the intestinal lumen into the bloodstream.[1]
2. Increased Renal Reabsorption: In the kidneys, calcitriol enhances the reabsorption of calcium from the glomerular filtrate, primarily in the distal convoluted tubules. This action reduces urinary calcium loss and helps to conserve the body's calcium stores.[1]
3. Bone Mineralization and Mobilization: Vitamin D is essential for the normal mineralization of bone by ensuring an adequate supply of calcium and phosphorus.[1] In states of calcium deficiency, calcitriol works synergistically with PTH to stimulate the differentiation and activity of osteoclasts. This leads to the resorption of bone matrix, which mobilizes calcium and phosphorus from the skeletal reservoir into the plasma to restore normal serum levels.[1]

Suppression of Parathyroid Hormone (PTH)

A key therapeutic effect of Calcifediol in the context of CKD is the suppression of PTH. The parathyroid glands are a primary target of calcitriol. By raising serum calcium levels, calcitriol exerts a powerful negative feedback effect on the parathyroid glands, leading to a decrease in the transcription of the PTH gene and a reduction in PTH synthesis and secretion.[3] This action directly counteracts the pathophysiology of secondary hyperparathyroidism, which is characterized by excessive PTH production.

Other Biological Effects

Beyond its classical role in mineral metabolism, the VDR is expressed in a wide variety of tissues, suggesting broader physiological functions. VDR activation is known to play a role in regulating cellular proliferation and differentiation.[28] Additionally, preclinical data suggest that Calcifediol may possess anti-inflammatory properties, potentially through the downregulation of signaling pathways such as Toll-like receptor 4 (TLR4) and JAK1/STAT3.[24]

Pharmacokinetics

The pharmacokinetic profile of Calcifediol is distinct from that of cholecalciferol and is central to its therapeutic rationale, especially in specific clinical scenarios.

• Absorption and Bioavailability: Calcifediol is readily absorbed from the gastrointestinal tract. Its absorption is considered more efficient and reliable than that of cholecalciferol, a difference attributed to its greater polarity, which makes it less dependent on the presence of bile acids for absorption.[2] This superior bioavailability is a significant advantage in patients with conditions that impair fat absorption, such as cholestatic liver disease or malabsorption syndromes.[8]
• Distribution: Following absorption, Calcifediol is extensively bound to the vitamin D-binding protein (DBP) in the plasma.[2] It is the main circulating metabolite of vitamin D, and its serum concentration is the most widely accepted biomarker for assessing a person's overall vitamin D status.[1] Compared to the highly lipophilic cholecalciferol, Calcifediol is less readily sequestered in adipose tissue. This lower volume of distribution means that in obese individuals, a greater proportion of an administered dose remains in circulation, leading to a more predictable and robust increase in serum 25(OH)D levels.[25]
• Metabolism: As a prohormone, Calcifediol's primary metabolic fate is its conversion to active calcitriol in the kidney via the CYP27B1 enzyme.[3] A key therapeutic advantage is that administration of Calcifediol bypasses the need for the initial 25-hydroxylation step in the liver. This makes it an effective treatment option for patients with severe liver disease who may have impaired capacity to convert cholecalciferol to Calcifediol.[17]
• Elimination: The elimination half-life of circulating Calcifediol is relatively long, estimated to be between 15 and 30 days [2], or 2 to 3 weeks.[25] This long half-life contributes to its stability as a marker of vitamin D stores. In contrast, the active hormone, calcitriol, has a very short half-life of only about 4 to 6 hours, reflecting its role as a tightly regulated, potent hormone.[25]
• Comparative Potency and Onset of Action: Clinical pharmacokinetic studies have consistently shown that Calcifediol is more potent and acts more rapidly than cholecalciferol in raising serum 25(OH)D levels. Studies suggest it is approximately two to three times more potent on a microgram-for-microgram basis.[7] One comparative study found that all participants receiving daily or weekly Calcifediol achieved target serum 25(OH)D concentrations of >30 ng/mL within a mean of 16.8 days. In contrast, only 70% of participants receiving equivalent doses of cholecalciferol reached this target, and it took them a mean of 68.4 days to do so.[7] This faster onset of action is a clinically relevant advantage when rapid correction of vitamin D deficiency is desired.[8]

The unique pharmacokinetic properties of Calcifediol are not merely academic observations; they form the very foundation of its targeted therapeutic strategy and the development of specialized formulations like Rayaldee®. The drug's superior potency and more rapid onset of action compared to cholecalciferol present both a clinical opportunity and a pharmacological challenge.[7] The opportunity lies in its ability to effectively treat vitamin D deficiency and its sequelae in populations where cholecalciferol may be less effective, such as those with liver disease or obesity.[17] The challenge, however, is that this same potency increases the risk of inducing hypercalcemia, the primary dose-limiting toxicity, if not administered with precision.[34]

The development of an extended-release (ER) formulation is a direct and sophisticated pharmaceutical response to this challenge. The ER technology used in Rayaldee® is specifically engineered to harness the potency of Calcifediol while mitigating the risk of acute toxicity. By ensuring a slow, gradual release of the drug, the formulation is designed to raise serum 25(OH)D levels in a controlled manner, avoiding the sharp pharmacological surges that can occur with immediate-release preparations.[35] This controlled elevation allows for a sustained and steady supply of substrate for calcitriol production, leading to effective, long-term suppression of PTH. Simultaneously, it minimizes the risk of abrupt increases in serum calcium, enhancing the safety margin. This makes the ER formulation particularly suitable for the high-risk population of Stage 3-4 CKD patients, in whom calcium and phosphorus homeostasis is already fragile and precise hormonal modulation is paramount.[4] This illustrates a clear and deliberate pathway from understanding basic pharmacokinetic principles to designing an advanced drug delivery system for a specific, targeted clinical indication.

The following table provides a direct comparison of the key pharmacokinetic parameters of Calcifediol and cholecalciferol, highlighting the clinical implications of their differences.

Table 2: Comparative Pharmacokinetic Profile: Calcifediol vs. Cholecalciferol

Key Pharmacokinetic Parameter	Calcifediol (25-hydroxycholecalciferol)	Cholecalciferol (Vitamin D3)	Clinical Implication
Intestinal Absorption	Readily absorbed; more polar	Requires bile for absorption; less efficient	Calcifediol is more reliable, especially in patients with malabsorption syndromes or liver disease.
Hepatic Metabolism Step	Bypasses 25-hydroxylation	Requires hepatic 25-hydroxylation to become Calcifediol	Calcifediol is effective in patients with severe liver disease who cannot perform this conversion.
Affinity for Adipose Tissue	Lower; less sequestration in fat	Higher; readily sequestered in fat	Calcifediol provides a more predictable dose-response in obese individuals.
Time to Reach Target 25(OH)D	Faster (e.g., ~17 days)	Slower (e.g., ~68 days)	Calcifediol allows for more rapid correction of vitamin D deficiency.
Relative Potency	2-3 times more potent than cholecalciferol	Standard baseline	Smaller doses of Calcifediol are required to achieve the same effect, necessitating careful dosing.

Clinical Efficacy and Therapeutic Indications

The clinical application of Calcifediol has evolved from a general vitamin D supplement to a highly targeted therapy for specific disease states. Its approved indications vary significantly across different global regulatory jurisdictions, reflecting the diverse clinical evidence submitted by sponsors for different formulations and the corresponding therapeutic strategies.

Global Approved Indications

A review of the major regulatory agencies reveals distinct and important differences in the approved uses of Calcifediol.

• U.S. Food and Drug Administration (FDA): In the United States, the extended-release formulation of Calcifediol, marketed as Rayaldee®, has a highly specific indication. It is approved for the treatment of secondary hyperparathyroidism (SHPT) in adult patients who meet two concurrent criteria: they must have Stage 3 or 4 chronic kidney disease (CKD), and their serum total 25-hydroxyvitamin D levels must be less than 30 ng/mL.[4] The FDA label includes a critical "Limitation of Use," stating that Rayaldee® is not indicated for patients with Stage 5 CKD or for those with end-stage renal disease on dialysis.[4]
• European Medicines Agency (EMA): The approved indication for Rayaldee® in the European Union is nearly identical to that in the US. It is authorized for the treatment of SHPT in adults with CKD Stage 3 or 4 who also have vitamin D insufficiency or deficiency.[5]
• Australian Therapeutic Goods Administration (TGA): In contrast to the US and EU, the approved indication in Australia for the immediate-release formulation, Vistella® (calcifediol monohydrate), is much broader. It is indicated for the treatment of vitamin D deficiency in adults, as well as for maintenance treatment as required.[11] This positions the drug as a first-line or alternative option for the general management of hypovitaminosis D, not restricted to the CKD population.
• Other Regulatory Developments: The regulatory status of Calcifediol continues to evolve. In Europe, the European Food Safety Authority (EFSA) issued a positive opinion on its use as a novel food ingredient for food supplements, establishing a conversion factor of 2.5 for labeling purposes to equate its potency to vitamin D3.[12] This dual status as both a prescription medicine and a regulated food supplement is unique. In Australia, regulatory bodies have formally moved Calcifediol to Schedule 4 (Prescription Only Medicine), distinguishing it from lower-dose, over-the-counter vitamin D preparations.[41]

The following table summarizes the approved indications for Calcifediol across these key regulatory agencies.

Table 3: Summary of Approved Indications for Calcifediol by Regulatory Agency

Regulatory Agency	Brand Name(s)	Approved Indication(s)	Key Population/Limitations
FDA (USA)	Rayaldee®	Treatment of secondary hyperparathyroidism (SHPT)	Adults with Stage 3 or 4 CKD and serum 25(OH)D < 30 ng/mL. Not for Stage 5 CKD/dialysis.
EMA (Europe)	Rayaldee®	Treatment of SHPT	Adults with Stage 3 or 4 CKD and vitamin D insufficiency or deficiency.
TGA (Australia)	Vistella®	Treatment of vitamin D deficiency, and maintenance treatment as required	General adult population.

Analysis of Key Clinical Trials

The approved indications for Calcifediol are supported by a robust body of clinical evidence from randomized controlled trials.

Secondary Hyperparathyroidism (SHPT) in CKD

The approval of Rayaldee® for SHPT was based on two identical, pivotal Phase 3, multicenter, randomized, double-blind, placebo-controlled trials. These studies enrolled a total of 429 subjects with Stage 3 or 4 CKD, SHPT (elevated iPTH), and vitamin D insufficiency.[38]

• Efficacy: The primary efficacy endpoint in these trials was the proportion of patients achieving a clinically meaningful reduction in plasma intact PTH (iPTH) of at least 30% from baseline after 26 weeks of treatment. The results demonstrated the superiority of Rayaldee® over placebo, with a significantly greater percentage of patients in the treatment arm meeting this endpoint.[38] A key secondary outcome was the correction of vitamin D insufficiency. Over 80% to 95% of patients treated with Rayaldee® achieved serum 25(OH)D levels of ≥ 30 ng/mL, compared to less than 7% of subjects receiving placebo.[36] Data from a subsequent open-label extension study showed that the PTH-lowering effect was durable and progressive, with 50% of patients achieving a ≥ 30% reduction in iPTH after 52 weeks of continuous treatment.[42]
• Safety: A crucial finding from these pivotal trials was the favorable safety profile with respect to mineral metabolism. Treatment with Rayaldee® was associated with only minor, clinically insignificant changes in mean serum calcium and phosphorus levels when compared to placebo, addressing a major concern with active vitamin D therapies.[35]
• Comparative Efficacy: A network meta-analysis was conducted to indirectly compare the efficacy and safety of ER calcifediol with paricalcitol, an active vitamin D analog also used to treat SHPT. The analysis, which included nine RCTs with over 1400 patients, found that ER calcifediol and paricalcitol had comparable efficacy in reducing PTH levels. However, treatment with paricalcitol was associated with a statistically significant greater increase in serum calcium levels, suggesting that ER calcifediol may offer a superior safety profile regarding the risk of hypercalcemia.[44]

Vitamin D Deficiency

The broader indication for treating general vitamin D deficiency is supported by trials directly comparing Calcifediol to cholecalciferol.

• Efficacy and Onset of Action: A large Phase III-IV trial in postmenopausal women with vitamin D deficiency (serum 25(OH)D < 20 ng/mL) compared the efficacy of monthly oral Calcifediol (0.266 mg) to a standard high dose of monthly cholecalciferol (25,000 IU).[33] The study's primary endpoint was the percentage of patients achieving serum 25(OH)D levels above 30 ng/mL after four months. Calcifediol was found to be significantly superior, with 35.0% of patients in the Calcifediol group reaching the target compared to only 8.2% in the cholecalciferol group. The study also confirmed Calcifediol's faster onset of action, demonstrating a significantly greater increase in serum 25(OH)D levels after just the first month of treatment.[33]
• Long-Term Maintenance and Safety: The same study followed patients for a full year and confirmed that long-term monthly treatment with Calcifediol is both effective and safe for maintaining adequate vitamin D status. However, it also highlighted that the therapeutic effect is dependent on continued administration; in a group where Calcifediol was withdrawn after four months, serum 25(OH)D levels rapidly declined back to baseline deficiency levels by the end of the study.[45] Other clinical trials have corroborated the safety and efficacy of Calcifediol for long-term use in diverse populations, including young adults and patients with severe deficiency.[46]

Off-Label and Investigational Uses

Beyond its approved indications, Calcifediol is used in other clinical contexts based on its pharmacological properties.

• Documented Off-Label Use: A logical off-label application of Calcifediol is for the treatment of metabolic bone diseases, such as osteomalacia, that arise secondary to severe liver disease.[17] Because Calcifediol administration bypasses the hepatic 25-hydroxylation step, it can effectively replete the body's vitamin D stores even when the liver's metabolic capacity is compromised.
• Investigational Use in Dialysis (Stage 5 CKD): The Rayaldee® label explicitly excludes patients on dialysis. However, this limitation has been challenged by recent investigational studies. Pilot randomized controlled trials have explored the use of high-dose ER calcifediol in patients with end-stage kidney disease requiring hemodialysis.[49] These studies have shown that providing high levels of the Calcifediol substrate can safely raise serum 25(OH)D to very high levels, which in turn drives a significant increase in serum 1,25(OH)D (calcitriol) and effectively stabilizes or reduces elevated iPTH levels.

The positive results from these dialysis trials provide compelling clinical evidence for an important physiological concept: the capacity for extra-renal 1-alpha-hydroxylation to become a therapeutically relevant pathway. In advanced CKD, the kidneys progressively lose their ability to express the CYP27B1 enzyme, leading to a failure of calcitriol production.[28] The standard therapeutic approach has been to circumvent this problem by administering exogenous active vitamin D analogs like calcitriol or paricalcitol. However, these potent agents carry a significant risk of causing hypercalcemia and hyperphosphatemia.[44]

The dialysis studies testing high-dose ER calcifediol were designed to evaluate an alternative hypothesis: that providing a sufficiently high concentration of the precursor, Calcifediol, could drive substrate-dependent conversion to calcitriol in other tissues that are known to express the CYP27B1 enzyme, such as immune cells, endothelial cells, and the parathyroid gland itself.[2] The findings that high-dose ER calcifediol successfully normalized serum calcitriol levels and controlled SHPT in anuric patients strongly support this hypothesis.[50] This represents a potential paradigm shift in the management of SHPT in dialysis patients. It suggests that prohormone repletion, which may allow for more physiological, tissue-level regulation of active hormone production, could be a safer and effective alternative to systemic administration of potent, active hormone replacements. These findings directly challenge the current "Limitation of Use" on the Rayaldee® label and identify a critical area for future large-scale clinical trials and potential label expansion.

Dosage, Formulations, and Administration

The safe and effective use of Calcifediol requires a thorough understanding of its available formulations, specific dosing protocols for each indication, and the essential monitoring parameters that guide therapy.

Available Formulations

Calcifediol is marketed in several different formulations and strengths, tailored to different therapeutic strategies.

• Extended-Release (ER) / Prolonged-Release Capsules: This is a specialized formulation marketed under the brand name Rayaldee®. Each capsule contains 30 mcg of Calcifediol and is designed for a gradual release over several hours. This design aims to provide a controlled increase in serum 25(OH)D levels, which is critical for optimizing efficacy while minimizing the risk of hypercalcemia in the sensitive CKD patient population.[5]
• Soft Gelatin Capsules (Immediate-Release): Various immediate-release soft gelatin capsules are available globally. In Australia, Vistella® is supplied as a 255 mcg calcifediol monohydrate capsule intended for intermittent (monthly) dosing.[11] Other strengths, such as 20 mcg and 50 mcg, are available for daily or every-other-day dosing for conditions like hypocalcemia.[53] Clinical trials have also utilized a range of strengths for weekly dosing regimens, including 75 mcg, 100 mcg, and 125 mcg capsules.[54]

Dosing and Titration Protocols

The dosing regimen for Calcifediol is highly dependent on the specific indication and formulation being used.

For SHPT in Stage 3-4 CKD (Rayaldee®)

The dosing protocol for Rayaldee® is carefully structured to ensure a gradual and safe response.

• Prerequisites for Initiation: Before starting therapy, it is mandatory to ensure the patient's serum calcium level is below 9.8 mg/dL (2.45 mmol/L) to minimize the risk of exacerbating or causing hypercalcemia.[4]
• Initial Dose: The recommended starting dose is 30 mcg administered orally once daily, typically at bedtime.[26]
• Dose Titration: The patient's response should be evaluated after approximately three months of treatment. If the iPTH level remains above the desired therapeutic range, the dose may be increased to 60 mcg once daily.[4] This dose escalation is contingent on several safety parameters being met: serum calcium must remain below 9.8 mg/dL, serum phosphorus must be below 5.5 mg/dL, and the serum total 25(OH)D level should be below 100 ng/mL.[4]
• Dose Suspension and Adjustment: Therapy should be suspended if any of the following occur: iPTH levels become persistently and abnormally low (which increases the risk of adynamic bone disease), serum calcium is consistently above the normal range, or serum 25(OH)D is consistently above 100 ng/mL. Once these laboratory values have normalized, treatment may be restarted at a reduced dose.[4]

For Vitamin D Deficiency (e.g., Vistella®)

For the broader indication of treating vitamin D deficiency, an intermittent dosing schedule is used.

• Standard Dose: The typical dosage is one 255 mcg capsule taken once per month. It is explicitly stated that this formulation must not be administered daily.[11]
• Dose Adjustment: In cases of severe deficiency, higher doses, such as one capsule every fortnight, may be required. The dose and frequency are determined by the prescribing physician based on baseline 25(OH)D levels and the patient's clinical response.[11]

For Hypocalcemia/Renal Osteodystrophy (Immediate-Release Formulations)

Older indications for immediate-release forms follow a different dosing pattern.

• Initial Dose: Therapy is often initiated at 50 mcg once a day or 100 mcg every other day.[16]
• Maintenance Dose: Most patients respond to a maintenance dose of 50 to 100 mcg per day, though some may be managed on as little as 20 mcg every other day.[16]

Administration Guidelines

Proper administration is key to ensuring the intended pharmacokinetic profile and patient adherence.

• Rayaldee® (ER Capsules): Patients must be instructed to swallow the extended-release capsules whole. The capsules should not be crushed, chewed, broken, or opened, as this would destroy the extended-release mechanism and could lead to dose dumping and an increased risk of toxicity.[16] The recommended administration time is at bedtime.[16]
• Missed Doses: If a patient misses a dose, they should be instructed to skip that dose and resume their normal dosing schedule at the next regularly scheduled time. They should not take an extra dose to make up for the missed one.[16]

Essential Monitoring

Given the potent effects of Calcifediol on mineral metabolism, regular laboratory monitoring is a critical component of safe therapy.

• Key Parameters: The core parameters to monitor are serum calcium, serum phosphorus, serum total 25(OH)D, and intact PTH.[4]
• Monitoring Schedule: For patients on Rayaldee®, these parameters should be checked at a minimum of three months after initiating therapy or after any dose adjustment. Subsequently, monitoring should continue at least every 6 to 12 months.[4] More frequent monitoring (e.g., weekly or bi-weekly for serum calcium) may be necessary during the initial dose titration phase for other indications.[53] The goal of therapy is to maintain iPTH within the target range for the stage of CKD, serum 25(OH)D between 30 and 100 ng/mL, serum calcium within the normal range, and serum phosphorus below 5.5 mg/dL.[55]

Comprehensive Safety and Tolerability Profile

The safety profile of Calcifediol is well-defined, with the majority of adverse effects being predictable extensions of its vitamin D-related pharmacology. A proactive approach to monitoring and patient education is essential for mitigating these risks.

Adverse Effects

Adverse events associated with Calcifediol therapy are primarily related to its potent effects on calcium and phosphorus homeostasis.

Most Significant Risk - Hypercalcemia

The principal and most clinically significant adverse effect of Calcifediol is hypercalcemia (elevated serum calcium levels).[9] The risk is dose-dependent and is the primary reason for the careful titration and monitoring protocols.

• Symptoms of Hypercalcemia: Patients should be educated to recognize the signs and symptoms of hypercalcemia. Early or mild symptoms can be non-specific and include fatigue, confusion or difficulty thinking clearly, loss of appetite, nausea, vomiting, constipation, increased thirst (polydipsia), increased urination (polyuria), and unexplained weight loss.[16]
• Complications of Hypercalcemia: If left unmanaged, severe or chronic hypercalcemia can lead to serious complications. Acute severe hypercalcemia can precipitate life-threatening cardiac arrhythmias and seizures.[4] Chronic hypercalcemia promotes the deposition of calcium in soft tissues, leading to generalized vascular calcification, nephrocalcinosis (kidney calcification), and calcification of other tissues.[4] Additionally, hypercalcemia significantly potentiates the effects of digitalis glycosides (e.g., digoxin), increasing the risk of toxicity.[4]

Other Clinically Relevant Adverse Events

• Adynamic Bone Disease: A potential consequence of over-treatment with any agent that suppresses PTH is adynamic bone disease. If PTH levels are suppressed to persistently and abnormally low levels, normal bone turnover is inhibited. This can lead to a brittle skeleton with an increased risk of fractures.[5] This is why monitoring iPTH and suspending the dose if it is too low is a key part of the therapeutic guidelines.
• Common Adverse Reactions: Data from the pivotal clinical trials of Rayaldee® identified several adverse reactions that occurred in ≥ 3% of patients and were more frequent than in the placebo group. These included anemia, nasopharyngitis (common cold), increased blood creatinine, dyspnea (shortness of breath), cough, congestive heart failure, and constipation.[9]
• Less Common/Rare Side Effects: Other potential adverse effects include hyperphosphatemia, pancreatitis (which can be a late manifestation of severe hypercalcemia), photophobia (light sensitivity), conjunctivitis with corneal calcifications (at very high doses), and metallic taste.[10]

Contraindications and Precautions

Certain patient populations and pre-existing conditions preclude the use of Calcifediol or require special caution.

• Absolute Contraindications:
• Calcifediol is contraindicated in patients with pre-existing hypercalcemia.[59]
• It is also contraindicated in individuals with a known history of a hypersensitivity reaction to Calcifediol or any of the excipients in the formulation.[5]
• Precautions and Use with Caution:
• Pregnancy and Lactation: The safety of Calcifediol during pregnancy has not been definitively established in adequate and well-controlled studies. It should be used during pregnancy only if the potential maternal benefit justifies the potential risk to the fetus.[16] Calcifediol is excreted into breast milk, and while data are limited, infants of breastfeeding mothers should be closely monitored for signs and symptoms of hypercalcemia, such as vomiting, constipation, or seizures.[16]
• Pediatric Use: The safety and efficacy of Calcifediol have not been established in children, and it is generally not recommended for this population.[26] However, the EMA is overseeing a Paediatric Investigation Plan (PIP) to evaluate its use for SHPT in children aged 8 years and older, suggesting this may change in the future.[40]
• Renal Impairment: Although specifically indicated for patients with Stage 3-4 CKD, these patients are inherently more susceptible to developing hypercalcemia due to their impaired ability to excrete calcium. Therefore, close monitoring is essential.[11] As per the Rayaldee® label, its use is not indicated in Stage 5 CKD or dialysis patients.[4]
• Other Conditions: Caution is warranted in patients with conditions that may increase sensitivity to vitamin D. This includes patients with a history of kidney stones (nephrolithiasis), as well as those with sarcoidosis or other granulomatous diseases, in which activated macrophages can lead to unregulated extra-renal production of calcitriol, increasing the risk of hypercalcemia even at standard doses.[11]

Black Box Warning Status

A review of regulatory documents and drug safety databases indicates that Calcifediol and its branded formulations (e.g., Rayaldee®, Vistella®) do not carry a Black Box Warning from the U.S. FDA or other major regulatory agencies.[16]

The absence of this most stringent warning is a significant statement about the drug's perceived risk-benefit profile. This is particularly insightful when considered in the context of the broader therapeutic area of CKD-Mineral and Bone Disorder (CKD-MBD). For instance, another drug used in patients with bone metabolism disorders, denosumab (Prolia), recently had a Black Box Warning added by the FDA due to the increased risk of severe hypocalcemia specifically in patients with advanced CKD.[65] Calcifediol's primary risk is the opposite—

hypercalcemia. The FDA's differential action suggests that the risk associated with Calcifediol is considered to be more predictable and manageable through the standard warnings, precautions, and explicit monitoring and dosing strategies outlined in the product labeling. The risk of hypercalcemia with Calcifediol is a well-understood, dose-dependent phenomenon that can be effectively mitigated by routine laboratory monitoring and patient education. In contrast, the risk of severe hypocalcemia with denosumab in the advanced CKD population was evidently deemed by regulators to be of a severity and nature that warranted the highest level of warning. Therefore, the lack of a Black Box Warning for Calcifediol implies a regulatory consensus that its safety profile, while requiring careful management, is well-characterized and controllable within the framework of standard clinical practice.

Drug and Food Interactions

The efficacy and safety of Calcifediol can be significantly altered by concomitant medications and dietary factors.

Drug-Drug Interactions

• CYP3A4 Enzyme Modulators: Calcifediol's metabolism is influenced by the cytochrome P450 enzyme system.
• CYP3A4 Inhibitors (e.g., ketoconazole, clarithromycin, ritonavir) can inhibit the breakdown of Calcifediol, potentially increasing its serum levels and heightening the risk of hypercalcemia. Close monitoring and possible dose adjustments are necessary when co-administering these agents.[55]
• CYP3A4 Inducers (e.g., phenobarbital, phenytoin, rifampin) can accelerate the metabolism of Calcifediol, potentially reducing its serum levels and diminishing its therapeutic effect. Dose increases and careful monitoring may be required.[11]
• Agents Affecting Calcium Excretion:
• Thiazide Diuretics (e.g., hydrochlorothiazide) decrease the renal excretion of calcium. When used with Calcifediol, this can lead to an additive effect and an increased risk of developing hypercalcemia.[5]
• Agents Affecting Absorption:
• Bile Acid Sequestrants (e.g., cholestyramine, colestipol) can bind to Calcifediol in the intestine, impairing its absorption and reducing its effectiveness. It is recommended to administer these drugs at least 2 hours apart from Calcifediol.[11]
• Agents with Synergistic Toxicity:
• Cardiac Glycosides (e.g., digoxin): The toxic effects of digoxin are potentiated by hypercalcemia. If a patient on digoxin develops hypercalcemia from Calcifediol, they are at a significantly increased risk of cardiac arrhythmias. Serum calcium and signs of digoxin toxicity must be monitored closely.[4]
• Other Vitamin D Compounds: Co-administration of Calcifediol with other vitamin D analogs (e.g., calcitriol, paricalcitol) or high-dose nutritional vitamin D supplements should be avoided due to the high risk of additive effects and severe hypercalcemia.[1]

Drug-Food Interactions

While specific studies on Calcifediol and food are limited, the principles governing its active metabolite, calcitriol, are directly applicable.[71]

• Calcium, Phosphorus, and Vitamin D Intake: Patients should be advised to maintain a consistent dietary intake of calcium, phosphorus, and vitamin D. Abrupt increases in the consumption of foods rich in these nutrients—such as dairy products, fortified cereals, and certain processed foods—can precipitate hypercalcemia and hyperphosphatemia, especially in patients with impaired renal function.[59] Uncontrolled use of over-the-counter calcium or vitamin D supplements must be avoided.
• Dietary Fat: As a fat-soluble compound, the absorption of Calcifediol may be influenced by the fat content of a meal.[29] While the clinical significance of this for the ER formulation is noted as unclear, consistency in taking the medication with respect to meals may be advisable.

The following table provides a summary of the most clinically significant drug interactions.

Table 4: Clinically Significant Drug Interactions with Calcifediol

Interacting Drug/Class	Example Drugs	Mechanism of Interaction	Clinical Consequence	Recommended Management Action
Strong CYP3A4 Inhibitors	Ketoconazole, Clarithromycin, Ritonavir	Inhibition of Calcifediol metabolism	Increased Calcifediol levels, risk of hypercalcemia	Monitor serum calcium, phosphorus, and 25(OH)D closely. Dose adjustment of Calcifediol may be required.
CYP3A4 Inducers	Phenytoin, Phenobarbital, Rifampin	Acceleration of Calcifediol metabolism	Decreased Calcifediol levels, reduced efficacy	Monitor serum 25(OH)D and iPTH. Dose increase of Calcifediol may be required.
Thiazide Diuretics	Hydrochlorothiazide, Chlorthalidone	Decreased renal excretion of calcium	Increased risk of hypercalcemia	Monitor serum calcium frequently. Avoid concomitant use if possible, or use with extreme caution.
Bile Acid Sequestrants	Cholestyramine, Colestipol	Impaired intestinal absorption of Calcifediol	Decreased efficacy	Separate administration times by at least 2 hours. Monitor therapeutic response.
Cardiac Glycosides	Digoxin, Digitoxin	Hypercalcemia potentiates digitalis toxicity	Increased risk of cardiac arrhythmias	Monitor serum calcium and for signs of digitalis toxicity, especially upon initiation or dose adjustment of Calcifediol.
Other Vitamin D Compounds	Calcitriol, Paricalcitol, Ergocalciferol	Additive pharmacodynamic effects	High risk of severe hypercalcemia	Concomitant use should be avoided.

Regulatory and Commercial Overview

The path of Calcifediol from a known metabolite to a marketed therapeutic product has been shaped by targeted clinical development and distinct regulatory strategies in different parts of the world. This has resulted in a varied commercial landscape where the drug occupies both a niche therapeutic role and a broader supplemental one.

Global Regulatory History

The approval timelines and responsible entities for key Calcifediol products highlight its modern development history.

Rayaldee® (FDA, USA)

• Approval and Sponsor: The extended-release formulation, Rayaldee®, was first approved by the U.S. FDA on June 17, 2016.[43] The New Drug Application (NDA) was submitted by OPKO Health, Inc.
• Development Pathway: The approval was granted based on the strength of two pivotal Phase 3, randomized, double-blind, placebo-controlled trials that demonstrated its efficacy and safety in the target population.[38] The path to approval was not entirely direct; OPKO received a Complete Response Letter from the FDA in March 2016, indicating that initial deficiencies needed to be addressed before the final approval was granted in June of the same year.[73]
• Commercial Launch: Following the FDA approval, OPKO Health commenced the U.S. launch of Rayaldee® in the second half of 2016, with shipments to distributors beginning in late November 2016.[38]

Rayaldee® (EMA, Europe)

• Authorization and Commercialization: Rayaldee® is now approved for use in 11 European countries.[75] The commercialization in Europe is managed by OPKO's licensee, Vifor Fresenius Medical Care Renal Pharma (VFMCRP). The European launch began with Germany in February 2022.[75]
• Pediatric Development: The EMA's Paediatric Committee (PDCO) has overseen the development of a Paediatric Investigation Plan (PIP) for Rayaldee®. This plan outlines the required studies to support a potential future indication for the treatment of SHPT in children. The PIP has undergone modifications, with the most recent acceptance of changes occurring in June 2024, focusing on studies in children aged 8 years and older.[40]

Vistella® (TGA, Australia)

• Approval and Sponsor: In Australia, the immediate-release formulation, Vistella®, was approved by the Therapeutic Goods Administration (TGA) as a new chemical entity. The decision to approve was made on July 16, 2024, with the product scheduled for entry onto the Australian Register of Therapeutic Goods (ARTG) on March 6, 2025.[77] The sponsor for this application is Aspen Pharmacare Australia Pty Ltd.[77]

Recent Regulatory Developments

The regulatory landscape for Calcifediol is not static. Recent decisions reflect a growing recognition of its distinct properties compared to other forms of vitamin D.

• Scheduling in Australia: In a significant regulatory reclassification, an interim decision was made in Australia in November 2019 to create a new, specific entry for Calcifediol in Schedule 4 of the Poisons Standard, designating it as a "Prescription Only Medicine." Previously, it had been covered under the general schedule entries for Vitamin D. This change acknowledges its higher potency and the need for medical supervision for its use.[41]
• Novel Food Status in Europe: In a parallel but distinct regulatory pathway, the European Commission officially authorized the placing of calcidiol monohydrate on the market as a novel food in April 2024. This authorization permits its use in food supplements at specified maximum daily doses (e.g., up to 10 µg/day for individuals over 11 years old).[13] This decision was preceded by a positive safety opinion from the European Food Safety Authority (EFSA) in 2021.[12] A critical component of this authorization was the scientific derivation and adoption of a conversion factor, establishing that for labeling and dosage purposes, the bioavailability of Calcifediol is 2.5 times that of vitamin D3.[13] This dual regulatory identity—as a potent prescription drug for specific diseases and as a regulated over-the-counter novel food supplement—is a defining feature of its current status and highlights its versatility.

Conclusion

Calcifediol (25(OH)D3​) occupies a unique and increasingly important position in both pharmacology and clinical medicine. As the direct precursor to the active hormone calcitriol, it serves as a cornerstone of the vitamin D endocrine system. This comprehensive monograph has detailed its multifaceted nature, from its fundamental physicochemical properties to its complex role in global health regulation.

Pharmacologically, Calcifediol's key distinction lies in its pharmacokinetic advantages over its parent compound, cholecalciferol. Its superior intestinal absorption, circumvention of the rate-limiting hepatic hydroxylation step, and reduced sequestration in adipose tissue collectively result in a more rapid, potent, and predictable elevation of serum 25(OH)D levels. These properties render it a particularly valuable therapeutic option for patients with malabsorption syndromes, liver disease, or obesity, in whom traditional vitamin D supplementation may be less effective.

The clinical development of the extended-release formulation, Rayaldee®, exemplifies a sophisticated application of these pharmacokinetic principles. By engineering a gradual release of this potent prohormone, the formulation successfully balances efficacy with safety, enabling the sustained suppression of PTH in patients with Stage 3-4 CKD while minimizing the inherent risk of hypercalcemia. The robust clinical trial data supporting its approval in the United States and Europe for this specific indication underscore its value in managing a complex metabolic complication of chronic kidney disease.

Furthermore, emerging research is expanding the potential therapeutic horizons for Calcifediol. Clinical investigations in end-stage renal disease patients on dialysis have provided compelling evidence for the viability of an extra-renal activation pathway, suggesting that high-dose prohormone repletion could represent a paradigm shift in the management of SHPT, potentially offering a safer alternative to direct administration of active vitamin D analogs.

The global regulatory landscape reflects the compound's dual identity. While specialized formulations are tightly regulated as prescription medicines for specific, high-risk conditions, its fundamental role as a vitamin D metabolite has also led to its authorization as a novel food supplement in Europe. This divergence highlights the need for clinicians and consumers to be aware of the different formulations, potencies, and intended uses available.

In summary, Calcifediol is far more than a simple vitamin supplement. It is a potent prohormone whose unique pharmacological profile has been strategically leveraged to create targeted therapies for challenging clinical conditions. Its safety profile is well-understood and manageable with appropriate clinical oversight. As research continues to elucidate its full range of biological activities and clinical applications, Calcifediol is poised to play an even more significant role in the treatment of disorders related to mineral metabolism and vitamin D deficiency.

Works cited

1. Calcifediol: Uses, Interactions, Mechanism of Action | DrugBank Online, accessed September 28, 2025, https://go.drugbank.com/drugs/DB00146
2. Calcifediol - Wikipedia, accessed September 28, 2025, https://en.wikipedia.org/wiki/Calcifediol
3. Calcifediol, EP - Formosa Laboratories, Inc., accessed September 28, 2025, https://www.formosalab.com/calcifediol
4. This label may not be the latest approved by FDA. For current ..., accessed September 28, 2025, https://www.accessdata.fda.gov/drugsatfda_docs/label/2024/208010Orig1s013lbl.pdf
5. Prescribing Information | Rayaldee Europe, accessed September 28, 2025, https://www.rayaldee.eu/prescribing-information-global
6. What is the mechanism of Calcifediol? - Patsnap Synapse, accessed September 28, 2025, https://synapse.patsnap.com/article/what-is-the-mechanism-of-calcifediol
7. Pharmacokinetics of oral vitamin D3 and calcifediol, accessed September 28, 2025, https://www.zora.uzh.ch/id/eprint/87916/
8. Hypovitaminosis D: Is It Time to Consider the Use of Calcifediol? - MDPI, accessed September 28, 2025, https://www.mdpi.com/2072-6643/11/5/1016
9. Rayaldee ® (calcifediol), accessed September 28, 2025, https://rayaldee.com/
10. Calcifediol Side Effects: Common, Severe, Long Term - Drugs.com, accessed September 28, 2025, https://www.drugs.com/sfx/calcifediol-side-effects.html
11. AusPAR - VISTELLA - calcifediol - Aspen Pharmacare Australia Pty Ltd, accessed September 28, 2025, https://www.tga.gov.au/sites/default/files/2025-07/auspar-vistella-pi-250709.pdf
12. DSM marks milestone in authorization of ampli-D® in Europe, accessed September 28, 2025, https://www.dsm-firmenich.com/en/businesses/health-nutrition-care/news/press-releases/archive/dsm/2021/dsm-marks-milestone-in-authorization-of-ampli-d-in-europe.html
13. Commission Implementing Regulation (EU) 2024/1052 of 10 April 2024 authorising the placing on the market of calcidiol monohydra - EUR-Lex, accessed September 28, 2025, https://eur-lex.europa.eu/legal-content/EN/TXT/PDF/?uri=OJ:L_202401052
14. calcifediol | Ligand page - IUPHAR/BPS Guide to PHARMACOLOGY, accessed September 28, 2025, https://www.guidetopharmacology.org/GRAC/LigandDisplayForward?tab=biology&ligandId=6921
15. Calcifediol: CAS # 19356-17-3 Compound Information and Applications for GC (Gas Chromatography) and LC (Liquid Chromatography) Analysis - EZGC Method Translator, accessed September 28, 2025, https://ez.restek.com/compound/view/en/19356-17-3/25-Hydroxyvitamin%20D3
16. Calcifediol Uses, Side Effects & Warnings - Drugs.com, accessed September 28, 2025, https://www.drugs.com/mtm/calcifediol.html
17. CALCIFEDIOL | 19356-17-3 - ChemicalBook, accessed September 28, 2025, https://www.chemicalbook.com/ChemicalProductProperty_EN_CB5727250.htm
18. Calcifediol | C27H44O2 - ChemSpider, accessed September 28, 2025, https://www.chemspider.com/Chemical-Structure.4446820.html
19. Calcifediol - Healthdirect, accessed September 28, 2025, https://www.healthdirect.gov.au/medicines/medicinal-product/aht,50532/calcifediol
20. Calcifediol hydrate | C27H46O3 | CID 88810850 - PubChem, accessed September 28, 2025, https://pubchem.ncbi.nlm.nih.gov/compound/Calcifediol-hydrate
21. Calcifediol - 25-Hydroxyvitamin D3 monohydrate, 25-Hydroxycholecalciferol - Sigma-Aldrich, accessed September 28, 2025, https://www.sigmaaldrich.com/US/en/substance/calcifediol4186563283363
22. Calcifediol monohydrate - vitamin D3 metabolite - APExBIO, accessed September 28, 2025, https://www.apexbt.com/calcifediol-monohydrate.html
23. CAS 19356-17-3 - Sigma-Aldrich, accessed September 28, 2025, https://www.sigmaaldrich.com/US/en/search/19356-17-3?focus=products&page=1&perpage=30&sort=relevance&term=19356-17-3&type=cas_number
24. 25-Hydroxyvitamin D3 (Calcifediol), metabolite of vitamin D3 (CAS 19356-17-3) | Abcam, accessed September 28, 2025, https://www.abcam.com/en-us/products/biochemicals/25-hydroxyvitamin-d3-calcifediol-metabolite-of-vitamin-d3-ab142766
25. Pharmacokinetic profile and effect on bone markers and muscle strength of two daily dosage regimens of calcifediol in osteopenic/osteoporotic postmenopausal women - PMC, accessed September 28, 2025, https://pmc.ncbi.nlm.nih.gov/articles/PMC8429405/
26. Calcifediol (oral route) - Side effects & dosage - Mayo Clinic, accessed September 28, 2025, https://www.mayoclinic.org/drugs-supplements/calcifediol-oral-route/description/drg-20312274
27. go.drugbank.com, accessed September 28, 2025, https://go.drugbank.com/drugs/DB00146#:~:text=It%20promotes%20renal%20reabsorption%20of,active%20form%20of%20vitamin%20D3.
28. Vitamin D - Wikipedia, accessed September 28, 2025, https://en.wikipedia.org/wiki/Vitamin_D
29. Calcifediol: Mechanisms of Action - MDPI, accessed September 28, 2025, https://www.mdpi.com/2072-6643/15/20/4409
30. Calcifediol: Why, When, How Much? - PMC, accessed September 28, 2025, https://pmc.ncbi.nlm.nih.gov/articles/PMC10222038/
31. Hypovitaminosis D: Is It Time to Consider the Use of Calcifediol? - PMC, accessed September 28, 2025, https://pmc.ncbi.nlm.nih.gov/articles/PMC6566727/
32. Cholecalciferol or Calcifediol in the Management of Vitamin D Deficiency - MDPI, accessed September 28, 2025, https://www.mdpi.com/2072-6643/12/6/1617
33. Calcifediol is superior to cholecalciferol in improving vitamin D status in postmenopausal women: a randomized trial - Oxford Academic, accessed September 28, 2025, https://academic.oup.com/jbmr/article/36/10/1967/7516555
34. What are the side effects of Calcifediol? - Patsnap Synapse, accessed September 28, 2025, https://synapse.patsnap.com/article/what-are-the-side-effects-of-calcifediol
35. About Rayaldee, accessed September 28, 2025, https://rayaldee.com/hcp/about-rayaldee/
36. Modified-Release Calcifediol Effectively Controls Secondary Hyperparathyroidism Associated with Vitamin D Insufficiency in Chronic Kidney Disease, accessed September 28, 2025, https://rayaldee.com/wp-content/themes/rayaldee/pdfs/hcp/modified-release-calcifediol-effectively-controls-shpt.pdf
37. Pharmacy Medical Necessity Guidelines: - Rayaldee (calcifediol) - Point32Health, accessed September 28, 2025, https://www.point32health.org/provider/wp-content/uploads/sites/2/2024/01/Rayaldee-calcifediol-comm-pmng.pdf
38. FDA Approves New Drug Application for RAYALDEE® to Treat Secondary Hyperparathyroidism Associated with Vitamin D Insufficiency in Stage 3-4 Chronic Kidney Disease - OPKO Health, accessed September 28, 2025, https://www.opko.com/investors/news-events/press-releases/detail/268/fda-approves-new-drug-application-for-rayaldee-to-treat-secondary-hyperparathyroidism-associated-with-vitamin-d-insufficiency-in-stage-3-4-chronic-kidney-disease
39. 208010Orig1s000 - accessdata.fda.gov, accessed September 28, 2025, https://www.accessdata.fda.gov/drugsatfda_docs/nda/2016/208010Orig1s000MedR.pdf
40. European Medicines Agency decision P/0211/2024 of 14 June 2024 on the acceptance of a modification of an agreed paediatric inves, accessed September 28, 2025, https://www.ema.europa.eu/en/documents/pip-decision/p-0211-2024-ema-decision-14-june-2024-acceptance-modification-agreed-paediatric-investigation-plan-calcifediol-rayaldee-emea-002093-pip02-17-m02_en.pdf
41. 1.3. Interim decision in relation to calcifediol - Therapeutic Goods Administration (TGA), accessed September 28, 2025, https://www.tga.gov.au/resources/publication/scheduling-decisions-interim/interim-decisions-and-invitation-further-comment-substances-referred-november-2019-acmsaccs-meetings/13-interim-decision-relation-calcifediol
42. Use of Extended-Release Calcifediol to Treat Secondary Hyperparathyroidism in Stages 3 and 4 Chronic Kidney Disease, accessed September 28, 2025, https://rayaldee.com/wp-content/themes/rayaldee/pdfs/hcp/use-of-erc-treat-shpt-stages%203-4-ckd.pdf
43. Pharmaceutical Approval Update - PMC, accessed September 28, 2025, https://pmc.ncbi.nlm.nih.gov/articles/PMC5083074/
44. Paricalcitol and Extended-Release Calcifediol for Treatment of Secondary Hyperparathyroidism in Non-Dialysis Chronic Kidney Disease: Results From a Network Meta-Analysis - Oxford Academic, accessed September 28, 2025, https://academic.oup.com/jcem/article/108/11/e1424/7180818
45. Long‐Term Treatment and Effect of Discontinuation of Calcifediol in Postmenopausal Women with Vitamin D Deficiency: A Randomized Trial - Oxford Academic, accessed September 28, 2025, https://academic.oup.com/jbmr/article/38/4/471/7499956
46. Efficacy and Safety of Calcifediol in Young Adults with Vitamin D Deficiency: A Phase I, Multicentre, Clinical Trial—POSCAL Study - MDPI, accessed September 28, 2025, https://www.mdpi.com/2072-6643/16/2/306
47. A Randomized Phase II/III Trial Evaluating the Efficacy and Safety of 100 and 125 µg of Calcifediol Weekly Treatment of Severe Vitamin D Deficiency - PMC - PubMed Central, accessed September 28, 2025, https://pmc.ncbi.nlm.nih.gov/articles/PMC11857939/
48. Calcifediol: MedlinePlus Drug Information, accessed September 28, 2025, https://medlineplus.gov/druginfo/meds/a616041.html
49. High-dose extended-release calcifediol successfully treated advanced secondary hyperparathyroidism in an end-stage kidney disease patient: a case report - PubMed, accessed September 28, 2025, https://pubmed.ncbi.nlm.nih.gov/40775686/
50. Extended-Release Calcifediol Normalized 1,25-Dihydroxyvitamin D and Prevented Progression of Secondary Hyperparathyroidism in Hemodialysis Patients in a Pilot Randomized Clinical Trial - PubMed, accessed September 28, 2025, https://pubmed.ncbi.nlm.nih.gov/40466630/
51. Extended-Release Calcifediol Normalized 1,25-Dihydroxyvitamin D and Prevented Progression of Secondary Hyperparathyroidism in Hemodialysis Patients in a Pilot Randomized Clinical Trial - Karger Publishers, accessed September 28, 2025, https://karger.com/ajn/article/doi/10.1159/000546615/928316/Extended-Release-Calcifediol-Normalized-1-25
52. 208010Orig1s000 | FDA, accessed September 28, 2025, https://www.accessdata.fda.gov/drugsatfda_docs/nda/2016/208010Orig1s000Lbl.pdf
53. Calcifediol Dosage Guide + Max Dose, Adjustments - Drugs.com, accessed September 28, 2025, https://www.drugs.com/dosage/calcifediol.html
54. Study Details | NCT04735926 | Efficacy and Safety of Calcifediol vs Placebo in Subjects With Vitamin D Deficiency | ClinicalTrials.gov, accessed September 28, 2025, https://www.clinicaltrials.gov/study/NCT04735926
55. Rayaldee (calcifediol) dosing, indications, interactions, adverse effects, and more, accessed September 28, 2025, https://reference.medscape.com/drug/rayaldee-calcifediol-1000075
56. Getting Started - Rayaldee ® (calcifediol), accessed September 28, 2025, https://rayaldee.com/patient/getting-started/
57. Calcifediol Monograph for Professionals - Drugs.com, accessed September 28, 2025, https://www.drugs.com/monograph/calcifediol.html
58. Calcifediol: Side Effects, Uses, Dosage, Interactions, Warnings - RxList, accessed September 28, 2025, https://www.rxlist.com/calcifediol/generic-drug.htm
59. Calcifediol (Rayaldee) | Davis's Drug Guide - Nursing Central, accessed September 28, 2025, https://nursing.unboundmedicine.com/nursingcentral/view/Davis-Drug-Guide/110769/all/calcifediol?q=f
60. OPKO Announces U.S. Launch of RAYALDEE, accessed September 28, 2025, https://www.opko.com/investors/news-events/press-releases/detail/283/opko-announces-u-s-launch-of-rayaldee
61. RAYALDEE Approved for Treatment of SHPT in Pre-dialysis CKD Patients, accessed September 28, 2025, https://nephrology.wustl.edu/rayaldee-approved-treatment-shpt-pre-dialysis-ckd-patients/
62. Calcifediol Extended-Release Capsules - Cleveland Clinic, accessed September 28, 2025, https://my.clevelandclinic.org/health/drugs/19504-calcifediol-extended-release-capsules
63. Calcifediol (Rayaldee) | Davis's Drug Guide - Nursing Central, accessed September 28, 2025, https://nursing.unboundmedicine.com/nursingcentral/view/Davis-Drug-Guide/110769/all/calcifediol
64. What is Calcifediol used for? - Patsnap Synapse, accessed September 28, 2025, https://synapse.patsnap.com/article/what-is-calcifediol-used-for
65. FDA adds Boxed Warning for increased risk of severe hypocalcemia in patients with advanced chronic kidney disease taking osteoporosis medicine Prolia (denosumab), accessed September 28, 2025, https://www.fda.gov/drugs/drug-safety-and-availability/fda-adds-boxed-warning-increased-risk-severe-hypocalcemia-patients-advanced-chronic-kidney-disease
66. FDA adds Boxed Warning for increased risk of severe hypocalcemia in patients with advanced chroni... - YouTube, accessed September 28, 2025, https://www.youtube.com/watch?v=QJREmgCc7bk
67. Drug-vitamin D interactions: A systematic review of the literature - PMC, accessed September 28, 2025, https://pmc.ncbi.nlm.nih.gov/articles/PMC5623087/
68. PRODUCT MONOGRAPH INCLUDING PATIENT MEDICATION INFORMATION PrRAYALDEE® Calcifediol modified-release capsules Capsules, 30 mcg,, accessed September 28, 2025, https://pdf.hres.ca/dpd_pm/00069537.PDF
69. go.drugbank.com, accessed September 28, 2025, https://go.drugbank.com/drugs/DB00146#:~:text=Trilostane-,The%20therapeutic%20efficacy%20of%20Calcifediol%20can%20be,used%20in%20combination%20with%20Trilostane.&text=Vamorolone-,The%20therapeutic%20efficacy%20of%20Calcifediol%20can%20be,used%20in%20combination%20with%20Vamorolone.&text=Vitamin%20D-,The%20risk%20or%20severity%20of%20adverse%20effects%20can%20be%20increased,is%20combined%20with%20Vitamin%20D.
70. Drug Interaction Report: calcifediol, Vitamin D3 - Drugs.com, accessed September 28, 2025, https://www.drugs.com/interactions-check.php?drug_list=458-0,646-5790&professional=1
71. 3 foods to avoid while taking calcitriol - SingleCare, accessed September 28, 2025, https://www.singlecare.com/blog/foods-to-avoid-while-taking-calcitriol/
72. Calcitriol and Alcohol/Food Interactions - Drugs.com, accessed September 28, 2025, https://www.drugs.com/food-interactions/calcitriol.html
73. Rayaldee (calcifediol) FDA Approval History - Drugs.com, accessed September 28, 2025, https://www.drugs.com/history/rayaldee.html
74. EX-99.1 - SEC.gov, accessed September 28, 2025, https://www.sec.gov/Archives/edgar/data/944809/000129993316002648/exhibit1.htm
75. RAYALDEE® Launched in Germany by OPKO Health's Licensee, VFMCRP, accessed September 28, 2025, https://www.opko.com/investors/news-events/press-releases/detail/461/rayaldee-launched-in-germany-by-opko-healths-licensee-vfmcrp
76. EMEA-002093-PIP02-17-M02 - paediatric investigation plan - European Medicines Agency, accessed September 28, 2025, https://www.ema.europa.eu/en/medicines/human/paediatric-investigation-plans/emea-002093-pip02-17-m02
77. VISTELLA - Therapeutic Goods Administration (TGA), accessed September 28, 2025, https://www.tga.gov.au/resources/auspar/vistella
78. Australian Public Assessment Report for VISTELLA - Therapeutic Goods Administration (TGA), accessed September 28, 2025, https://www.tga.gov.au/sites/default/files/2025-07/auspar-vistella-250709.pdf