Discovery and Structural Analysis of Metabolites of Vitamin D Lactones.

25-Hydroxyvitamin D-23,26-lactone (1) and 1α,25-dihydroxyvitamin D-23,26-lactone (2) have long been considered as among the end metabolites of vitamin D. Recently, however, we found that these lactones exhibit biological activity related to the β-oxidation of fatty acids. We hypothesized that a metabolic pathway might exist to inactivate their physiological activity.

Complications including dysphagia following transoral non-robotic surgery for pharyngeal and laryngeal squamous cell carcinoma: A retrospective multicenter study.

Objective: Transoral surgery is a minimally invasive treatment but may cause severe dysphagia at a lower rate than chemoradiotherapy.

Methods: We compared clinical information, surgical complications, and swallowing function in patients who underwent transoral nonrobotic surgery for laryngo-pharyngeal squamous cell carcinoma between 2015 and 2021 in a multicenter retrospective study.

Results: Six hundred and forty patients were included.

A tumor endothelial cell-specific microRNA replacement therapy for hepatocellular carcinoma.

Current approved anti-angiogenic drugs (AAD) for hepatocellular carcinoma (HCC) inhibit tumor angiogenesis, but affect the hepatic vasculature resulting in adverse effects. Tumor endothelial cells (TECs) differ from normal endothelial cells. In this study, we aimed to detect TEC-specific miRNAs and develop an anti-angiogenic treatment specific for TECs.

Impact of cervical lymph node metastasis on transoral surgery for hypopharyngeal squamous cell carcinoma: A retrospective multicenter study.

Background: Hypopharyngeal carcinoma is likely to spread to the lymph nodes, but there is no established strategy for management in transoral surgery.

Methods: We compared oncologic and functional outcomes in a retrospective multicenter study of patients who underwent transoral surgery for hypopharyngeal carcinoma between 2015 and 2021.

Results: Two-hundred and thirty-two patients were included.

Fabrication Technology of Self-Dissolving Sodium Hyaluronate Gels Ultrafine Microneedles for Medical Applications with UV-Curing Gas-Permeable Mold.

Microneedles are of great interest in diverse fields, including cosmetics, drug delivery systems, chromatography, and biological sensing for disease diagnosis. Self-dissolving ultrafine microneedles of pure sodium hyaluronate hydrogels were fabricated using a UV-curing TiO-SiO gas-permeable mold polymerized by sol-gel hydrolysis reactions in nanoimprint lithography processes under refrigeration at 5 °C, where thermal decomposition of microneedle components can be avoided. The moldability, strength, and dissolution behavior of sodium hyaluronate hydrogels with different molecular weights were compared to evaluate the suitability of ultrafine microneedles with a bottom diameter of 40 μm and a height of 80 μm.

Analysis of vitamin D receptor binding affinities of enzymatically synthesized triterpenes including ambrein and unnatural onoceroids.

Onoceroids are a rare family of triterpenes. One representative onoceroid is ambrein, which is the main component of ambergris used as a traditional medicine. We have previously identified the onoceroid synthase, BmeTC, in Bacillus megaterium and succeeded in creating ambrein synthase by introducing mutations into BmeTC.

Characterization of Rickets Type II Model Rats to Reveal Functions of Vitamin D and Vitamin D Receptor.

Vitamin D has been known to exert a wide range of physiological effects, including calcemic, osteogenic, anticancer, and immune responses. We previously generated genetically modified (GM) rats and performed a comparative analysis of their physiological properties to elucidate the roles of vitamin D and vitamin D receptor (VDR). In this study, our primary goal was to investigate the manifestations of type II rickets in rats with the VDR(H301Q) mutation, analogous to the human VDR(H305Q).

Gene therapy for alopecia in type II rickets model rats using vitamin D receptor-expressing adenovirus vector.

Type II rickets is a hereditary disease caused by a mutation in the vitamin D receptor (VDR) gene. The main symptoms of this disease are bone dysplasia and alopecia. Bone dysplasia can be ameliorated by high calcium intake; however, there is no suitable treatment for alopecia.

Local recurrence and metachronous multiple cancers after transoral nonrobotic surgery for pharyngeal and laryngeal squamous cell carcinoma: A retrospective multicenter study.

Background: Late laryngopharyngeal cancers after transoral surgery include not only local recurrences but also metachronous multiple cancers.

Methods: We compared clinical information, surgical outcomes, and late laryngopharyngeal cancers in patients who underwent transoral nonrobotic surgery for laryngopharyngeal squamous cell carcinoma without lymph node metastases between 2015 and 2021 in a multicenter retrospective study.

Results: Four hundred and fifty-seven patients were included.

Prenatal vitamin D deficiency alters immune cell proportions of young adult offspring through alteration of long-term stem cell fates.

Vitamin D deficiency is a common deficiency worldwide, particularly among women of reproductive age. During pregnancy, it increases the risk of immune-related diseases in offspring later in life. However, exactly how the body remembers exposure to an adverse environment during development is poorly understood.

Synthesis of (22)-, (22)-22-Fluoro-, and 22,22-Difluoro-25-hydroxyvitamin D and Effects of Side-Chain Fluorination on Biological Activity and CYP24A1-Dependent Metabolism.

Three novel analogues of C22-fluoro-25-hydroxyvitamin D (-) were synthesized and evaluated to investigate the effects of side-chain fluorination on biological activity and metabolism of vitamin D. These novel analogues were constructed by convergent synthesis applying the Wittig-Horner coupling reaction between CD-ring ketones (,,) and A-ring phosphine oxide (). The introduction of C22-fluoro units was achieved by stereoselective deoxy-fluorination for synthesizing and or two-step cationic fluorination for .

Analysis of vitamin D metabolites in biological samples using a nanoluc-based vitamin D receptor ligand sensing system: NLucVDR.

Many assays are currently being developed to measure the levels of vitamin D metabolites in various samples (such as blood, urine, and saliva). This study focused on the measurement of vitamin D metabolites in serum and urine using the NLucVDR assay system, which consists of a split-type nanoluciferase and ligand-binding domain (LBD) of the human vitamin D receptor. Blood and urine samples were collected from 23 participants to validate the NLucVDR assay.

Differential Metabolic Stability of 4α,25- and 4β,25-Dihydroxyvitamin D and Identification of Their Metabolites.

Vitamin D () is metabolized by various cytochrome P450 (CYP) enzymes, resulting in the formation of diverse metabolites. Among them, 4α,25-dihydroxyvitamin D () and 4β,25-dihydroxyvitamin D () are both produced from 25-hydroxyvitamin D () by CYP3A4. However, is detectable in serum, whereas is not.

Synthesis of New 26,27-Difluoro- and 26,26,27,27-Tetrafluoro-25-hydroxyvitamin D: Effects of Terminal Fluorine Atoms on Biological Activity and Half-life.

As an extension of our research on providing a chemical library of side-chain fluorinated vitamin D analogues, we newly designed and synthesized 26,27-difluoro-25-hydroxyvitamin D (1) and 26,26,27,27-tetrafluoro-25-hydroxyvitamin D (2) using a convergent method applying the Wittig-Horner coupling reaction between CD-ring ketones (13, 14) and A-ring phosphine oxide (5). The basic biological activities of analogues, 1, 2, and 26,26,26,27,27,27-hexafluoro-25-hydroxyvitamin D [HF-25(OH)D] were examined. Although the tetrafluorinated new compound 2 exhibited higher binding affinity for vitamin D receptor (VDR) and resistance to CYP24A1-dependent metabolism compared with the difluorinated 1 and its non-fluorinated counterpart 25-hydroxyvitamin D [25(OH)D], HF-25(OH)D showed the highest activity among these compounds.

Expression of Rat Cyp27b1 in HepG2 Cells Using Adenovirus Vector and Its Application to Evaluation of Self-Made and Commercially Available Anti-Cyp27b1 Antibodies.

Rat Cyp27b1 was successfully expressed in HepG2 cells using an adenovirus vector. High vitamin D 1α-hydroxylation activity was detected in them, whereas no activity was observed in non-infected cells. Similarly, vitamin D 1α-hydroxylation activity was also observed in HepG2 cells expressing Cyp27b1-Flag, which is tagged with a Flag at the C-terminus of Cyp27b1.

Functional analysis of vitamin D receptor (VDR) using adenovirus vector.

Recently, we generated type II rickets model rats, including Vdr(R270L), Vdr(H301Q), Vdr(R270L/H301Q), and Vdr-knockout (KO), by genome editing. All generated animals showed symptoms of rickets, including growth retardation and abnormal bone formation. Among these, only Vdr-KO rats exhibited abnormal skin formation and alopecia.

Fabrication of High-Resolution Fine Microneedles Derived from Hydrolyzed Hyaluronic Acid Gels in Vacuum Environment Imprinting Using Water Permeable Mold.

Hydrolyzed hyaluronic acid high-resolution fine microneedles of 13 µm in diameter and 24 µm in height were fabricated from hydrolyzed hyaluronic acid gels made in mixtures of water using vacuum environment imprint lithography processes with a water permeable mold. The gas traps of water and volatile solvents in the imprint materials cause transfer failure in the conventional water impermeable molds of quartz and metal. However, the water permeable mold allows the use of 67 wt% dilution water with high solubility to increase the fluidity of the hydrolyzed hyaluronic acid during the patterning of high-resolution fine microneedles for cosmetics and pharmaceuticals.

Development of nanoluciferase-based sensing system that can specifically detect 1α,25-dihydroxyvitamin D in living cells.

Previously, we reported a FLucN-LXXLL+LBD-FLucC system that detects VDR ligands using split firefly luciferase techniques, ligand binding domain (LBD) of VDR, and LXXLL sequences that interact with LBD after VDR ligand binding. In vivo, 25-hydroxyvitamin D (25(OH)D) and 1α,25-dihydroxyvitamin D (1α,25(OH)D) act as VDR ligands that bind to VDR, and regulate bone-related gene expression. Therefore, the amount of 25(OH)D and 1α,25(OH)D are indicators of bone-related diseases such as rickets and osteoporosis.

The First Convergent Synthesis of 23,23-Difluoro-25-hydroxyvitamin D and Its 24-Hydroxy Derivatives: Preliminary Assessment of Biological Activities.

In this paper, we report an efficient synthetic route for the 23,23-difluoro-25-hydroxyvitamin D () and its 24-hydroxylated analogues (,), which are candidates for the CYP24A1 main metabolites of . The key fragments, 23,23-difluoro-CD-ring precursors (-), were synthesized starting from Inhoffen-Lythgoe diol (), and introduction of the C23 difluoro unit to α-ketoester () was achieved using ,-diethylaminosulfur trifluoride (DAST). Preliminary biological evaluation revealed that 23,23-F-25(OH)D () showed approximately eight times higher resistance to CYP24A1 metabolism and 12 times lower VDR-binding affinity than its nonfluorinated counterpart 25(OH)D ().

In Silico Prediction of the Metabolic Resistance of Vitamin D Analogs against CYP3A4 Metabolizing Enzyme.

The microsomal cytochrome P450 3A4 (CYP3A4) and mitochondrial cytochrome P450 24A1 (CYP24A1) hydroxylating enzymes both metabolize vitamin D and its analogs. The three-dimensional (3D) structure of the full-length native human CYP3A4 has been solved, but the respective structure of the main vitamin D hydroxylating CYP24A1 enzyme is unknown. The structures of recombinant CYP24A1 enzymes have been solved; however, from studies of the vitamin D receptor, the use of a truncated protein for docking studies of ligands led to incorrect results.

Robust osteogenic efficacy of 2α-heteroarylalkyl vitamin D analogue AH-1 in VDR (R270L) hereditary vitamin D-dependent rickets model rats.

Active vitamin D form 1α,25-dihydroxtvitamin D (1,25(OH)D) plays pivotal roles in calcium homeostasis and osteogenesis via its transcription regulation effect via binding to vitamin D receptor (VDR). Mutated VDR often causes hereditary vitamin D-dependent rickets (VDDR) type II, and patients with VDDR-II are hardly responsive to physiological doses of 1,25(OH)D. Current therapeutic approaches, including high doses of oral calcium and supraphysiologic doses of 1,25(OH)D have limited success and fail to improve the quality of life of affected patients.

Metabolism of non-steroidal anti-inflammatory drugs (NSAIDs) by Streptomyces griseolus CYP105A1 and its variants.

In the field of drug development, technology for producing human metabolites at a low cost is required. In this study, we explored the possibility of using prokaryotic water-soluble cytochrome P450 (CYP) to produce human metabolites. Streptomyces griseolus CYP105A1 metabolizes various non-steroidal anti-inflammatory drugs (NSAIDs), including diclofenac, mefenamic acid, flufenamic acid, tolfenamic acid, meclofenamic acid, and ibuprofen.

Novel DNA Aptamer for CYP24A1 Inhibition with Enhanced Antiproliferative Activity in Cancer Cells.

Overexpression of the vitamin D3-inactivating enzyme CYP24A1 (cytochrome P450 family 24 subfamily and hereafter referred to as CYP24) can cause chronic kidney diseases, osteoporosis, and several types of cancers. Therefore, CYP24 inhibition has been considered a potential therapeutic approach. Vitamin D3 mimetics and small molecule inhibitors have been shown to be effective, but nonspecific binding, drug resistance, and potential toxicity limit their effectiveness.