Novel Biosynthetic Pathway for Secosteroids and the Skin

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DESCRIPTION (provided by applicant): Cytochrome P450scc system can efficiently transform 7-dehydrocholesterol (7-DHC) to 7- dehydropregnenolone (7-DHP) and hydroxylate vitamin D2 and D3, and ergosterol. Thus, we uncovered novel steroido/secosteroidogenic pathways generating new biologically active products. The cutaneous pathway would elicit mostly intra-, auto- or paracrine modes of action. In SLOS, UVB light-induced conversions (exclusive for the skin) would proceed with substrates derived from the systemic circulation (5, 7-steroidal dienes) to generate vitDL compounds. These novel pathways could generate a number of molecules with structures determined by the local type and activity of steroidogenic system, as well as the access to UVB photons. To test this hypothesis the following specific aims have been designed. 1. Testing the biological activity of P450scc derived 20-hydroxyvitamin D3 and 20,22 dihydroxyvitamin D3, 17a,24- dihydroxyergosterol, its photolytic derivative, and of 20-hydroxyvitamin D2 and 17a, 20-dihydroxyvitamin D2. The above effects will be compared to the parent compounds, i.e. vitamins D3 and 2, and ergosterol.; 2. Testing the biological activity of 7-DHP, its hydroxyderivatives and the secosteroidal products of their photolytical (UVB-induced) transformation; 3. Characterization of the enzymatic conversion of 7-DHP into hydroxy-derivatives, particularly at positions 17, 20, 21 and 11. The studies will include synthesis of the standards to fully characterize the chemical structures and properties of the reaction products. 4. Testing the predicted action of UVB in inducing B-ring intramolecular rearrangements of 7-DHP and its hydroxy- derivatives or 17a,24-dihydroxyergostero resulting in the anticipated vitDL - derivatives. Tests of the structural stability of the resulting molecules will also be performed; 5. Defining a role for the P450scc in the in vivo metabolism of 5, 7- dienesterols, vitamin D and ergosterol: a) characterization of 7-DHC transformation by P450scc in vivo in cultured skin cells in comparison to an adrenal cell line. Secosteroids production after exposure of the cells to ultraviolet radiation will also be investigated.; b) characterization of vitamin D and ergosterol transformation by P450scc in cultured skin cells. 6. Defining the effect of 7-DHP, its hydroxy and seco-derivatives and of vitamin D (modified or unmodified) on P450scc mediated pregnenolone production from cholesterol.