Analysis of the proton NMR spectra of 3-ethyl-3-(4-pyridyl)piperidine-2,6-dione (rogletimide, 1) shows that it exists in solution with the aromatic ring in an axial position; the same conformation was found for aminoglutethimide. Excess lithium diisopropylamide treatment of 1 formed a dianion which methylated at C-5. The major product with the methyl group trans to the pyridyl ring retained this ring in an axial position and had higher aromatase inhibitory potency than 1. The minor diastereoisomer with an equatorial pyridyl ring had low potency. Upon elongating the alkyl chain, particularly high inhibitory activity was found for the major product isomer having a C-5 octyl, coinciding with the high activity in C-3 and N-1 octyl derivatives of 1, but there was only a small difference in the activity between the enantiomers of 5-octyl-1 and activity was reduced rather than increased when octyl also replaced ethyl at C-3. The results partially support a previously described model comparing binding of androstenedione to aromatase in as much as an axial pyridyl ring is needed to mimic the axial C-19 methyl group of the steroid and bind to the heme component of the enzyme, but for the derivatives bearing a C-5 octyl, the function of the glutarimide ring seems to be simply as a spacer between the hydrophobic chain and the pyridyl ring.