1. Effects of Repeated GnRH Agonist Administration in Normal Men Carrie Zimmer, M.D. PhD, Robert Rosenfield, M.D. University of Chicago-Pediatric Endocrinology Abstract Background MethodsResultsSummary Results References Burris A, Clark R, Vantman D, Sherins R. A low sperm concentration dos not preclude fertility in men with isolated hypogonadotropic hypogonadism after gonadotropin therapy. Fertil Steril 1988;50:343- 347. Hoffman A, Crowley WJ. Induction of puberty in men by long-term pulsative administration of low- dose gonadotropin-releasing hormone. N Engl J Med 1982;307:1237-1241 Rosenfield R, Perovic N, Ehrmann D, Barnes R. Acute Hormonal Responses to the Gonadotropin Releasing Hormone Agonist Leuprolide: Dose-Response Studies and Comparision to Nafarelin- A Clinical Research Center Study. J Clin Endo Met 1996;81:3408-3411. Leuprolide acetate (Lupron®) injection was given as single subcutaneous injection at a dose of 1.0 or 10 mcg/kg. Blood samples began at 0700-0800 hr. Blood samples were obtained at -60 to 0 minutes in 20 minute intervals to obtain baseline levels. Samples were then obtained at 0.5, 1, 2, 3 and 4 hours following Lupron injection. -Volunteers then returned at 24, 48 and 72 hours for blood sampling. On day 5 and day 9, Lupron injection and blood sampling were repeated as on day 1. LH and FSH were measured by monoclonal sandiwich immunoassays (Delfia kit, Wallac). Plasma testosterone was measured by nonchromatographic method (Diagnostics Products kit). Data analysis: Baseline values were completed by averaging 20 minute samples from -60 minutes to injection. Peak response following injection was then compared to baseline using anova. In addition, time 0.5-4 hour was used for area under the curve comparative cumulative response to Lupron. Gonadotropin therapy is currently used to establish or restore fertility in patients with gonadotropin deficiency. Multiple agents have been used with multiple difficulties. GnRH has been used to treat gonadotropin deficiency. However, sustained administration of LHRH or its analogues results in inhibition of LH and FSH release, due to desentization (Hoffman). Therefore, it is necessary to administer frequent boluses (every 2 hours) which requires use of an infusion pump. Due to the disadvantages of previous therapy, the goal of this study was to determine if the GnRH agonist, Lupron, given in optimally spaced doses would avoid down-regulation. Initial studies were completed in normal men to determine the appropriate timing and dosage of Lupron without a diminishing effect on LH, FSH and testosterone. Figure 1. LH and FSH Response to 10mcg/kg Lupron Injection. Lupron 10 mcg/kg was administered to 5 normal men at Day 1 and Day 5 (arrows). Following Lupron injection 1 there was a significant increase in LH peak (p=.04) with maximum response at 1hr. This response returned to baseline levels by Day 3. On Day 5, Lupron administration was repeated. However, there was no significant response of LH. FSH levels did not significantly increase from baseline following either Lupron administration. Figure 2. Testosterone Response to 10mcg/kg Lupron Injection. Lupron. 10 mcg/kg was administered to 5 normal men at Day 1 and Day 5. Following Lupron injection on Day, there was a significant increase in testosterone on Day 2 & 3. By Day 4, testosterone levels had returned to baseline. Figure 3. Response to Repeated 1 mcg/kg Lupron Administration. Lupron was administered to 6 normal men at 4 day intervals (arrows). Figure 3A. LH and FSH response to Lupron administration. At 1 hr post Lupron administration, LH obtained peak response. Following each Lupron administration, LH peak had a significant increase from baseline (*p<.05). The peak LH response was significantly different between the initial injection and subsequent injections (# p<.05 ). However, the area under the curve for 0.5-4 hr was not significantly different between cycles. LH levels did return to baseline levels 48 hr following Lupron injection. FSH did not show a significant response to Lupron. Figure 3B. Testosterone and Estradiol levels following Lupron Administration. Following Lupron Injection (Day 1, 5, and 9) there was a significant increase in testosterone 24 hours later (* p<.05). By 48 hours later, testosterone levels were not significantly different than baseline. Peak testosterone levels following Lupron administration were not significantly different between cycles. Estradiol levels were not significantly elevated in response to Lupron injection. Conclusion Gonadotropin therapy is currently used to establish or restore fertility in patients with gonadotropin deficiency. However, current therapy is complicated by frequent dosing and extended therapy length to result in fertility. This current study in normal male subjects demonstrates that intermittent low dose Lupron administration can result in elevated levels of testosterone. The results of this study will be expanded to examine the ability of Lupron administration in gonadotropin deficient patients to produce fertility. As initially reported (Rosenfield), Lupron doses of 1.0-10 mcg/kg were shown to produce an acute response of LH. In this study, high dose Lupron was again able to produce a significant LH response, however subsequent Lupron dose was unable to produce an LH elevation. It is possible that at this dose Lupron results in release of a majority of stored LH and an adequate time between administration is not allowed to produce significant LH stores. This would be supported by the previous finding that Lupron 20 mcg/kg was unable to produced an elevated LH response as compared to 10 mcg/kg. In contrast, the study group that received a lower dose produced a significantly lower LH response (60 vs 30 IU/L). This level of response may allow a percentage of stored LH to remain while further production is completed. Importantly, this level of LH was able to produce similar levels of testosterone as compared to high dose. A significant finding of this study was that although the peak LH response may demonstrate a degree of down-regulation, the ability to repeatedly stimulate testosterone was not changed. * * * * LH or FSH (IU/L) * * # # * * * Testosterone (ng/dL) The acute administration of GnRH agonists has previously been shown to result in a significant increase in gonadotropins (LH, FSH) and testosterone. While chronic administration of GnRH agonists results in a decreased LH response.In patients with gonadotropin deficiency, it is necessary to maintain elevated testosterone and LH for an extended period of time to induce fertility. This study examined the effects of intermittent Lupron administration at 4-5 day intervals in normal men. The findings demonstrate that an initial high dose Lupron injection was able to produce a significant elevation of LH and testosterone, but subsequent Lupron administration was unable to replicate this elevation. When low dose Lupron was administered, the initial dose was again able to produce an elevation of LH and testosterone. In contrast to high dose, subsequent administration of low dose Lupron was able to cause a significant elevation of testosterone. In addition, the LH peak was blunted with repeated administration, however the area under the curve for LH (hr 0 – 4) was not different between cycles. The preliminary findings of this study, can now be expanded to examine patients with gonadotropin deficiency. 0 10 20 30 40 50 60 Day 1PeakDay 2Day 3Day 4Day 5Day 6 LH FSH 0 5 10 15 20 25 30 35 Day 1 Peak Day 2Day 3Day 4Day 5 Peak Day 6Day 7Day 8Day 9 Peak Day 10Day 11Day 12 LH FSH 0 100 200 300 400 500 600 700 800 900 Day 1Day 2Day 3Day 4Day 5 Testosterone (ng/dL) AB