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The binding of human sex hormone-binding globulin (SHBG) to a human prostatic cancer cell line (LNCaP) and the results of that binding were examined. Membranes derived from LNCaP cells bound unliganded SHBG on two sets of sites whose affinities were: Ka1 = 3.1 +/- 1.6 x 10(10) M-1 and Ka2 = 8.7 +/- 4.3 x 10(6) M-1. Intact cells also bound SHBG, but even after 6 h, less than 10% of specifically bound SHBG was internalized. This observation speaks against a role for the membrane binding of SHBG in steroid transport across cell membranes. When LNCaP cells were prebound with SHBG, addition of dihydrotestosterone or estradiol resulted in a dose-dependent increase in intracellular cAMP. SHBG in the absence of steroids or dihydrotestosterone in the absence of SHBG was without effect. 2-Methoxyestradiol, a steroid metabolite without biological activity, but which binds to SHBG more tightly than does estradiol, was also without effect. These observations demonstrate a potentially important role for SHBG as a regulator of cell function. They also demonstrate an additional mode of action of steroid hormones, one that does not require that the steroid interact with a steroid receptor.

We have studied the concentration and properties of a protein which binds cortisol in human milk in samples obtained from women during the first 100 days after delivery. A filter disk assay was developed both for the measurement of plasma corticosteroid-binding globulin (CBG) and for the cortisol-binding protein in milk. The concentration of CBG in milk, expressed as its capacity to bind cortisol, is highest on the day of delivery, ca, 0.80 mug/dl, falls over the next 10 days to ca. 0.25 mug/dl, and remains at that level thereafter. If the concentration of CBG is expressed relative to the concentration of serum albumin in milk, it increases from day 1 to day 3 and then remains constant. A detailed comparison of CBG derived from milk and plasma showed that the two proteins co-migrated on Sephadex, sucrose gradient ultracentrifugation, and polyacrylamide gel electrophoresis. The two proteins had the same affinity for cotrisol, progesterone, 17-OH-progesterone, and dexamethasone. Furthermore, the binding activity of CBG in milk was neutralized with anti-CBG antibodies raised against CBG isolated from plasma. Unlike CBG, the concentration of cortisol in milk, 0.8-3.5 mug/dl, showed no systematic variation as a function of the postpartum day on which the sample was obtained.