Hyesik Kong

Sulfate-conjugated methylprednisolone as a colon-targeted methylprednisolone prodrug with improved therapeutic properties against rat colitis.

Methylprednisolone (MP) is one of the most widely used corticosteroids for the treatment of inflammatory bowel disease (IBD). However, systemic adverse effects of MP limit its availability for the disease. In present study, sulfate-conjugated methylprednisolone (MPS) was evaluated in vivo as a colon-targeted prodrug of MP and its therapeutic properties against 2,4,6-trinitrobenzenesulfonic acid–induced rat colitis were investigated. Upon oral administration, a large fraction of MPS reached the large intestine, where MPS was converted to MP implying that MPS would deliver MP effectively to the large intestine. The fecal recovery of MP (after MPS administration) was much greater than that after MP administration and the urinary recovery of MP (after MPS administration) was much less than that after MP administration, suggesting that MPS should exhibit enhanced therapeutic activity and reduced systemic adverse effects. Consistent with this notion, MPS was more effective than MP in ameliorating rat colitis. Moreover, the adverse effects of MPS on adrenal function and thymus were much lower than those of MP. Taken together, MPS may be therapeutically superior to MP in IBD treatment.

Synthesis and in vitro evaluation of N-nicotinoylglycyl-2-(5-fluorouracil-1-yl)-D,L-glycine as a colon-specific prodrug of 5-fluorouracil.

N-Nicotinoylglycyl-2-(5-fluorouracil-1-yl)-d,l-glycine (NGFG) was synthesized as a colon-specific prodrug of 5-fluorouracil (5-FU) expecting that hydrolysis of nicotinoyl and glycyl moieties by microbial enzymes in the colon will give 2-(5-fluorouracil-1-yl)-d,l-glycine, which releases 5-FU spontaneously. To in vitro-evaluate colon targetability of NGFG, apparent partition coefficient and chemical/biochemical stability of NGFG in the contents or/and tissue of the various segments of the gastrointestinal tract were determined. Low partition coefficient and stability of NGFG in the upper intestinal condition suggested its delivery to the colon in intact form after oral administration. Incubation with rat cecal contents produced 5-FU and its metabolite about 16%. Structural modification to enhance amide hydrolysis, the rate determining step in NGFG bioactivation, is suggested.

Emetine Promotes von Hippel-Lindau-Independent Degradation of Hypoxia-Inducible Factor-2α in Clear Cell Renal Carcinoma

Inactivating mutations of the von Hippel-Lindau (VHL) tumor suppressor gene are associated with inherited VHL syndrome, which is characterized by susceptibility to a variety of neoplasms, including central nervous system hemangioblastoma and clear cell renal cell carcinoma (CCRCC). Mutations in the VHL gene are also found in the majority of sporadic clear cell renal carcinoma, the most common malignant neoplasm of the human kidney. Inactivation of VHL ubiquitin ligase is associated with normoxic stabilization of hypoxia-inducible factor-1α and 2-α (HIF-1α and HIF-2α), transcriptional regulators of tumor angiogenesis, invasion, survival, and glucose utilization. HIF-2α has been particularly implicated in the development of CCRCC. Although several inhibitors of HIF-1α have been described, these drugs typically have a minimal affect on HIF-2α. 786-O is a VHL-deficient CCRCC cell line that constitutively expresses only HIF-2α and is therefore suitable for the screening of novel HIF-2α inhibitors. Using this cell line, we have identified emetine as a specific inhibitor of HIF-2α protein stability and transcriptional activity. Without altering HIF-2α mRNA level, emetine rapidly and dramatically down-regulated HIF-2α protein expression in 786-O cells. HIF-2α down-regulation was accompanied by HIF-2α ubiquitination and was reversed by proteasome inhibition. Emetine-induced HIF-2α down-regulation was confirmed in three additional VHL-renal cancer cell lines, was insensitive to the prolyl hydroxylase inhibitor dimethyloxaloyl glycine, and did not require neural precursor cell expressed developmentally down-regulated-8, suggesting that emetine accesses a previously undescribed cullin-independent proteasome degradation pathway for HIF-2α. These data support the use of emetine or structurally related compounds as useful leads for the identification of novel HIF-2α inhibitors.

Sulfate-conjugated methylprednisolone: Evaluation as a colon-specific methylprednisolone prodrug and comparison with sulfate-conjugated prednisolone and dexamethasone

Methylprednisolone 21-sulfate sodium (MPS) was prepared and evaluated as a colon-specific methylprednisolone prodrug and its colon-specific property was compared with prednisolone 21-sulfate sodium (PDS) and dexamethasone 21-sulfate sodium (DS), reported previously as colon-specific prodrugs of the glucocorticoids. The synthetic process and yield of MPS was simple and high. The apparent partition coefficient of methylprednisolone (MP) was greatly reduced by sulfate conjugation. The sulfate conjugates MPS, PDS, and DS were (bio)chemically stable in the homogenates of the upper intestine. In marked contrast, the sulfate conjugates were deconjugated to liberate the corresponding glucocorticoids in the cecal contents. Although the rates of deconjugation were not significantly different, the corresponding glucocorticoids were accumulated with distinct profiles depending on the metabolic susceptibility of the unconjugated glucocorticoids to microbial reductase(s). Upon oral administration of the sulfate conjugates to rats, the plasma concentrations of the conjugates were extremely low and the urinary recoveries were less than 5% of the doses. These results suggest that, like PDS and DS, MPS administered orally is delivered efficiently to the large intestine followed by deconjugation to liberate MP and the metabolic susceptibility of the unconjugated glucocorticoids may affect therapeutic availability of the sulfate conjugates.

Synthesis and evaluation of sulfate conjugated metronidazole as a colon-specific prodrug of metronidazole.

For an effort to improve therapeutic property of metronidazole (MTZ) which is a drug of choice for protozoal infections such as luminal amoebiasis, sulfate conjugated metronidazole (MTZS) was prepared and evaluated as a colon-specific prodrug of MTZ. The apparent partition coefficient of MTZ was greatly reduced by the sulfate conjugation. While (bio)chemically stable in the contents of the upper intestine, MTZS was rapidly cleaved to liberate MTZ on incubation with the cecal contents of rats. MTZ liberated from MTZS metabolized quickly at least partly by a microbial nitroreductase, suggesting the relevance of the metabolism to bioactivation of MTZ for antimicrobial action. Consistent with the hypothesis, MTZS elicited antibacterial activity in the cecal contents, which was as potent as free MTZ. The systemic absorption of MTZS was very low after oral administration of MTZS. In parallel with this, whereas MTZ disappeared mostly during the transit of the proximal small intestine, a substantial amount of MTZS remained in the small intestine, moving down to the large intestine where it metabolized rapidly. In addition to the efficient colonic delivery of MTZS, MTZS markedly reduced the systemic absorption of MTZ. Taken together, MTZS may be a potential colon-specific prodrug of MTZ which possesses improved therapeutic and toxicological properties.

Structural effects of N-aromatic acyl-amino acid conjugates on their deconjugation in the cecal contents of rats: implication in design of a colon-specific prodrug with controlled conversion rate at the target site

N‐aromatic acyl‐amino acid conjugates possess a colon‐targeted property, implying that such conjugates are stable and are not absorbable until reaching the large intestine in which they are microbially converted (hydrolysed) to the parent drugs that are therapeutically active. To investigate the structural effect of N‐aromatic acyl‐amino acid conjugates on the large intestinal deconjugation, the hydrolysis of various N‐aromatic acyl‐amino acid conjugates was examined in the cecal contents. On incubation of conjugates with glycine,d or/andl forms of alanine or phenylalanine in the cecal contents, the conjugates withd amino acids were not hydrolysed. The other conjugates are susceptible to the hydrolysis, the rates of which decreased as the size of the substituent on the 2‐position of the amino acids increased. The conjugates with alkyl analogs (2–4 carbons) of glycine and taurine were resistant to the hydrolysis, while taurine‐ and glycine‐conjugates were hydrolysed effectively. The hydrolysis of N‐aromatic acyl‐glycine conjugates was enhanced by para‐substitution of electron withdrawing groups on the aromatic acyl moiety and vice versa for electron‐donating groups. While a methyl, methoxy or chloro group on the ortho‐position retarded the hydrolysis, a hydroxyl group on the position accelerated it. Our data may provide useful information for the design of a colon‐specific prodrug with controlled conversion rate in the large intestine. Copyright

Synthesis and Properties of N,N′-Bis(5-Aminosalicyl)-L-Cystine as a Colon-Specific Deliverer of 5-Aminosalicylic Acid and Cystine

N,N′-bis(5-aminosalicyl)-L-cystine (5-ASA-Cys) was prepared by a simple synthetic route. 5-ASA-Cys was not degraded in pH 1.2 and 6.8 buffer solutions, and in the homogenates of the upper intestine. In marked contrast, 5-ASA-Cys was deconjugated extensively to liberate 5-ASA in the cecal contents. Upon oral administration of 5-ASA-Cys to rats, the plasma concentration of 5-ASA-Cys was extremely low and the urinary recovery of 5-ASA-Cys was ∼ 10% of the dose. These results suggest that 5-ASA-Cys administered orally is delivered efficiently to the large intestine followed by deconjugation to liberate 5-ASA and cystine.

Susceptibility of glucocorticoids to colonic metabolism and pharmacologic intervention in the metabolism: implication for therapeutic activity of colon‐specific glucocorticoid 21‐sulfate sodium at the target site

Objectives  The systemic side effects of glucocorticoids have prevented their long‐term use for treatment of inflammatory bowel disease. Colon‐specific delivery of glucocorticoids has been adopted as a strategy to circumvent the toxicological trouble. Glucocorticoids delivered to the large intestine might undergo metabolisms by colonic microflora, which should affect therapeutic availability at the target site. It was investigated whether the susceptibility of glucocorticoids to the colonic metabolisms and pharmacologic intervention in the metabolism could modulate the therapeutic availability of colon‐targeted glucocorticoids.