A Chemoenzymatic Strategy for the Synthesis of Steroid Drugs Enabled by P450 Monooxygenase-Mediated Steroidal Core Modification

The
synthesis of steroid drugs by multistage modifications of the
steroidal core is challenging since site-specific and selective modification
is essentially required, which is often difficult or complicated for
chemocatalysis. For example, the synthesis of Trenbolone (3), a versatile anabolic–androgenic steroid, relies on a four-step
chemical procedure on its core modifications of estra-4,9-diene-3,17-dione
(1). Here, we have designed a two-step chemoenzymatic
strategy that includes a biocatalytic one-pot C11-hydroxylation/17β-ketoreduction
of 1 with a computationally designed P450 monooxygenase
and an appropriate 17-ketosteroid reductase to generate 11α-OH-9(10)-dehydronandrolone
(2a) as an intermediate followed by chemical dehydration
to introduce the double bond at carbons 11 and 12 with the formation
of Trenbolone (3). To obtain a highly active and C11-selective
enzyme, molecular dynamics simulations were performed, uncovering
a crucial role of water molecules for substrate recognition and targeted
hydroxylation of steroids. Moreover, Trenbolone is further subjected
to esterification to produce Trenbolone acetate (9) that
has been widely used in veterinary medicine. Finally, our approach
enables the regio- and stereoselective synthesis of both steroid drugs 3 and 9 on a (nearly) gram scale with 83–91%
isolated yields, showing great potential for industrial applications.