Background: D-004, a lipid extract of the fruit of the Cuban royal palm (Roystonea regia), has been found to reduce prostatic hyperplasia (PH) induced with testosterone (T), but not PH induced with dihydrotestosterone (DHT), in rodents, suggesting the inhibition of prostate 5α-reductase activity.
Objectives: The aims of this study were to assess whether D-004 inhibits prostate 5α-reductase activity in vitro and to examine the effects of D-004 on enzyme kinetics.
Methods: This experimental study was conducted at the Pharmacology Department, Center of Natural Products, National Center for Scientific Research, Havana, Cuba. Soluble rat prostate preparations were used as the source of 5α-reductase, and ((3)H)-DHT production was measured to determine prostate 5α-reductase activity. Cell-free rat prostate homogenates were pre-incubated with carboxymethyl cellulose 2% alone (control tubes) or D-004 (0.24-125 μg/mL) suspended in the vehicle (treated tubes) for 10 minutes prior to adding the labeled substrate ((3)H)-T Once the reaction was stopped, sterols were extracted with chloroform and aliquots were applied on silica gel plates developed in benzene-acetone (4:1, v/v). Areas containing DHT were scraped and radioactivity was counted. The median inhibitory concentration (IC50) was determined by measuring the conversion of T to DHT The apparent Michaelis-Menten constant (Km) and Vmax values before and after adding D-004 were determined in kinetic studies using labeled T (0.5-25 μmol/L).
Results: Compared with controls, D-004 significantly and dose-dependently inhibited the enzymatic reaction at doses of 1.95 to 125.0 μg/mL) (all, P < 0.05). The IC50 of D-004 required to inhibit 5a-reductase activity was 2.25 μg/mL. Enzyme inhibition was noncompetitive, since D-004 lowered the Vmax from 15.3 to 10.0 nmol DHT/min · mg(-1) protein, while the Km (4.54 μmol/L) was almost unaffected.
Conclusions: D-004 dose-dependently and noncompetitively inhibited in vitro 5α-reductase activity in soluble fractions of rat prostate. Although the extent of the maximal inhibition was high and the value of IC50 was low, the relevance of such inhibition requires further study in vivo.
Keywords: D-004; free fatty acids; prostate 5α-reductase inhibitors; prostate hyperplasia.