Inhibition of prolyl 4-hydroxylase in vitro and in vivo by members of a novel series of phenanthrolinones

Abstract

Examples of a novel series of phenanthrolinones are shown to be potent competitive inhibitors of avian prolyl 4-hydroxylase, and of collagen hydroxylation, in embryonic chick tendon cells and human foreskin fibroblasts in vitro and in the oestradiol-stimulated rat uterus in vivo. Two compounds, Compound 1 (1,4-dihydrophenanthrolin-4-one-3-carboxylic acid) and Compound 5 [8-(N-butyl-N-ethylcarbamoyl)-1,4-dihydrophenathrolin-4-one-3-carboxylic acid], with comparable potencies in vivo, were chosen to investigate the effect of the inhibition of the hydroxylation of newly synthesized uterine collagen on the turnover of this protein in vivo. Inhibition of hydroxylation by more than 50% for approx. 8 h following single oral doses of the compounds was associated with significant losses of radiolabelled proline and 4-hydroxyproline from collagen during this period. Progressive hydroxylation of collagen over 48 h, as the inhibitory action of the compounds declined, was accompanied by a decreased loss of radiolabel from the uterine collagen. Earlier reports indicated that underhydroxylated collagen, accumulating within the endoplasmic reticulum in cells where prolyl 4-hydroxylase is inactivated, is slowly degraded, but is then rapidly hydroxylated and secreted when the activity of prolyl 4-hydroxylase is restored. Taken with the present results, this suggests that the potential use of inhibitors of prolyl 4-hydroxylase to control excessive collagen deposition in pathological fibrosis may be limited by the need to maintain continuous inhibition of collagen hydroxylation so as to facilitate intracellular degradation of the accumulated protein.

The HAG Mechanism: A Molecular Rationale For The Therapeutic Application Of Iron Chelators In Human Diseases Involving the 2-Oxoacid Utilizing Dioxygenases

“…the anthracyclines, label both the catalytic and the noncatalytic subunits [24]. The binding of other polycyclic inhibitors, such as the hydroxyanthraquinones [66] or the phenanthrolinones [35], may also involve additional interactions outside of the pocket itself.…”

“…1C). The HAG mechanism was also employed to guide the de novo synthesis of prolyl 4-hydroxylase inhibitors [5,[28][29][30][31][32][33][34][35]. In fact, major reviews on the development of antifibrotic drugs have routinely included the reproduction of some version of Fig.…”

Oxygen, Metabolism, and Regeneration: Lessons from Mice

“…For instance, the compound 1,4-dihydrophenonthrolin-4-one-3-carboxylic acid (1,4-DPCA) was identified as a blocker of P4H collagen prolyl hydroxylation in rats, with the potential of reducing collagen crosslinking and scar formation, both inhibiting regeneration (90) and inhibiting human MDA-MB-231 breast cancer metastasis in in vitro 3D cultures (91). However, it was also shown to act on PHDs and to modulate HIF-1α levels (6).…”

“…Given the results that HIF-1α protein expression appeared to be key in the regenerative response of MRL mice (6), 1,4-DPCA was used in combination with a hydrogel delivery system to slowly release low concentrations of the drug over a 5 day period in mice (92). The amount of drug used was 75-fold less than the dose for soluble drug given orally in rats for prolyl hydroxylation studies in vivo (6,90). Mimicking the kinetics of the MRL- HIF-1α expression response, 1,4-DPCA/hydrogel was injected subcutaneously at the base of the neck into non-regenerative mice and regenerative ear hole responses assessed (92).…”

Selective Inhibition of Collagen Prolyl 4-Hydroxylase in Human Cells

“…These compounds suffer from low potency in cellular assays, insufficient selectivity for CP4H, and intolerable cytotoxicity. 21,22 Still, the ethyl ester of DHB (that is, EDHB) is often used as a cellular “P4H” inhibitor, 23,24 even though DHB is not selective for CP4H compared to other FAKGDs, requires high dosing, and leads to an iron-deficient phenotype. 24 …”