GROWTH-INHIBITION OF T47D HUMAN BREAST-CANCER CELLS BY THE NEW ANTIAROMATASE STEROID 3-METHYLENE-1,4-ANDROSTADIEN-17-ONE

: The C19 steroid, 3-methylene-1,4-androstadien-17-one (3-methylene-1-dehydro-4-androsten-17-one, abbrev. 3MDA) was synthesized by microbial dehydrogenation with Arthrobacter simplex from the corresponding 1,2-saturated, 3-methylene-4-androsten-17-one. The new steroid induces a Type-1 optical difference spectrum with Cytochrome P450 aromatase from human placentae (K(s) 1.61 muM). Kinetic inhibition studies with placental microsomes show: (i) 3MDA inhibits aromatase irreversibly (k(inact) 0.026 min-1), (ii) the inhibition is time- and concentration-dependent, (iii) it follows pseudo first-order kinetics, and (iv) the inhibition requires the presence of the cofactor NADPH. Human T47D breast tumor cells are able to transform C-14-labeled androstenedione into radioactive estrone/estradiol, as we have established by repetitive HPLC purifications. We have thus clearly demonstrated the presence of an endogenous steroidal androgen-aromatizing enzyme in these breast tumor cells. Besides inhibiting human placental aromatase activity, 3MDA at concentrations of 5-20 muM also significantly decreases the growth of cultured T47D cells as measured by the incorporation of [H-3]thymidine into DNA, but without causing cell death due to cytotoxicity. These findings prompt us to propose 3MDA as a new, mechanism-based suicide substrate of the human aromatase enzyme with potential utility as a 'second generation' anti-breast cancer agent.