and qualitative changes in serum luteinizing hormone after injectable
testosterone undecanoate treatment in hypogonadal men
GU1,Zheng-Yan GE2, Gui-Yuan ZHANG1, William
Research Institute for Family Planning, Beijing 100081, China
Asian J Androl 2000 Mar; 2: 65-71
AbstractAim: To clarify the immuno-active LH (I-LH) and bioactive LH (b-LH) responses and qualitative changes in the circulating LH to testosterone undecanoate (TU) injection. Methods: Eight men with Klinefelters syndrome were recruited for the study. They received crossover injections of TU at doses of 500 and 1000 mg. Serum I-LH and b-LH levels before and at various time intervals after TU injection were measured and the serum I-LH, b-LH, b-LH/I-LH (B/I) and testosterone/sex hormone-binding globulin (T/SHBG) ratio in LH-responders and LH non-responders were compared. Results: A parallel suppression of serum I-LH and b-LH was consistent with their overall high correlation between each other (r=0.84, P<0.001). Mean serum i-FSH levels were decreased by TU injection at both doses without dose-response effects. LH-responders had lower baseline serum i-LH and b-LH, and higher E2 levels and T/SHBG ratio. There was a quantitative change in serum LH as induced by TU without qualitative change within LH-responders os LH-non-responders. Conclusion: A high loading dose (1000 mg) of TU is important for the initial suppression of LH. With the lower dose (500 mg), repeated injections will be required to attain such LH suppression for the purpose of fertility regulation. The lower baseline serum I-LH level may be an intrinsic characteristic of LH-responders.
1 IntroductionAlthough a number of testosterone (T) preparations, oral or parenteral, are available for clinical use, the rapidity of absorption, early peak release, relatively short-acting period and excessive hepatic toxicity limit their long-term application[1,2]. Recently, a new long-acting T preparation, T undecanoate (TU) has been shown to have a more favorable pharmacokinetic profile in Klinefelters syndrome patients with a significant suppression of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) after a single injection of 500 or 1000 mg, the serum LH suppression being non-uniform and varied between individuals, independent of the TU doses. It is not clear whether the LH response to exogenous TU was specific to men with Klinefelters syndrome whose serum LH was relatively resistant to T suppression. Recent studies showed that androgen administration is able to modulate the amount of LH secretion as well as qualitative alteration in LH molecules[5,6]. It is unclear whether the heterogeneous response of serum LH to T suppression is associated with a qualitative change in LH molecules. Therefore, the clarification of serum LH response to TU injection would be crucial for further understanding of TU.
2 Materials and methods
Eight Chinese men aged 16-26 years with Klinefelters syndrome, otherwise healthy, were recruited for the study. They had stopped previous hormonal therapy for their hypogonadism at least 6 weeks before recruitment. All the subjects gave written consent to participate in the study after understanding the studys purpose, benefit and possible risks.
TU (Zhejiang Xian Ju Pharmaceutical Corporation, Zhejiang, China) was available
in ampules containing 250 mg of the ester in 2 mL of tea seed oil.
patients were divided randomly into two groups. A prospective, crossover
clinical trial with injectable TU at doses of 500 and 1000 mg (equivalent
to 315 and 630 mg of T free base, respectively) was performed. The sample
size was 8 for 500 mg and 7 for 1000 mg (due to personal reason, one patient
only attended the 500 mg course). The first dose used for each subject
was randomized. The two treatment periods were separated by an intervening
washout period of 3 months. Two pre-treatment blood samples were taken
for baseline estimation of serum hormones. Blood samples were allowed
to clot for 24 hours before centrifugation. Serum samples
were stored at -70 until analyzed.
for RIA of T, estradiol (E2) and prolactin (PRL) were supplied
by the Diagnostic Products
Corporation, USA. All samples from one subject were analyzed in the same
assay. The lower detection limits of the assays were 0.14 nmol/L, 3.7
pmol/L and 20 mIU/L for T, E2 and PRL, respectively. The intra-assay
coefficient of variation for serum T, E2 and PRL was 7.6%,
5.8% and 6.2%, respectively. The mean inter-assay coefficient of variation
was less than 10% for all the three hormones. The serum sex hormone-binding
globulin (SHBG) was measured by the Nichols Institute (San Juan Capistrano,
CA) with a double antibody RIA kit supplied by the Diagnostic System Laboratories
(Webster, TX). The lower detection limit was
5.0 nmol/L. The mean intra-assay and inter-assay coefficient of variation for
serum SHBG was 4.0% and 8.8%, respectively.
I-LH and I-FSH were measured by time-resolved fluoroimmunoassay (Delfia,
Wallac Oy, Finland). The lower detection limit was 0.018 IU/L and 0.016
IU/L for LH and FSH, respectively.
All samples from one individual were analyzed in the same assay. The mean
intra- and inter-assay coefficients of variation were 4.4% and 9.3% for
LH and 5.9% and 8.7% for FSH, respectively.
in vitro bioassay of LH was a modification[7-9] of the procedure
described by van Damme et al and Dufau et al.
This assay is based on the measurement of T production by the dispersed
Leydig cells isolated from immature male Swiss Webster mice (34-45 day-old).
T was measured by RIA using reagents supplied by
the Diagnostic System Laboratories (Webster, TX). LER 907 was used as
the reference standard. Serial dilutions of serum samples containing high
LH levels were shown to be parallel to the standard curve. All the serum
samples were run in duplicate.
The minimal detection limit of b-LH was 0.04 mIU/tube and the intra-assay
and inter-assay coefficients of variation were 8.4% and 24%, respectively.
Data analysis and statistics
Serum I-LH, I-FSH, T, E2 and PRL levels
the 4 LH responders, TU suppressed the nadir of serum I-LH levels to the
upper normal threshold of eugonadal men (Figure
1). Serum I-LH levels at the baseline were significantly lower in
LH responders than in non-responders. There were marked differences between
those who response with a decline of I-LH following TU administration
(Figure 2). Mean serum I-FSH levels
were decreased by TU injection at both doses without dose response effects.
There was no apparent difference in serum T (Figure
3) and FSH levels between the two groups. However, serum E2
levels were slightly higher (P>0.05) in LH responders (Figure
4). Serum PRL levels remained unchanged in both groups throughout
the study period.
1. Serum I-LH levels before and after crossover injection of TU at
doses of 500 mg and 1000 mg
to 8 men with Klinefelters syndrome. Solid symbols (LH responders),
open symbols (LH non-responders). ......Upper normal limit.
SHBG and T/SHBG ratio
comparison of serum SHBG levels and T/SHBG ratio between the two groups
is shown in Table 1. Serum SHBG level was relatively higher at baseline
in both groups than that after
TU administration. It decreased after TU injection, especially in the
LH responders and remained within the normal range throughout the study
period. The T/SHBG ratio was significantly higher in LH responders, although
serum T levels were almost the same between
the two groups. No significant differences in serum SHBG levels were found
at different time points between the two groups as well as between the
baseline and various time points in either groups. A weak negative correlation
(r=-0.36) between T/SHBG and LH levels was observed.
1. Serum SHBG level and T/SHBG
ratio between LH responders and non-responders. meanSEM. bP<0.05,
between the two groups.
range of serum SHBG in adult men is 6-44 nmol/L.
parallel suppression of serum b-LH and I-LH was consistent with an overall
high correlation (r=0.84, P<0.001) between them (Figure
5). b-LH levels at the
baseline of the two groups
were almost at the same level, whereas i-LH levels at baseline were different.
After TU injection, b-LH significantly decreased in the LH responders
and gradually returned to baseline levels 8 weeks after treatment (Figure
6). The B/I ratio, an index of relative LH biopotency, fluctuated
slightly within each group.
In LH responders, the B/I ratio maintained a higher level throughout the
observation period except at week 7 (Figure
7). There was no correlation (r=-0.14) between serum T levels and
B/I ratio, and between T and serum i-LH or
b-LH levels. A weak negative correlation (r=-0.43) between T/SHBG
5. Correlation analysis between serum I-LH and b-LH. y=4.54+0.18x.
of supraphysiological dose of T causes profound suppression of spermatogenesis
by the depletion of gonadotropins and endogenous T[12,13]. A
decrease in the level of gonadotropins, especially LH, would be a criterion
for evaluating the potency of male hormonal contraceptives. The results
derived from pooled LH responders and non-responders showed obvious heterogeneity
in the two groups. Of considerable interest is the significant difference
in serum i-LH baseline levels between the two groups. LH responders who
had lower i-LH levels before the
first injection also had lower levels before the second injection, irrespective
of the dose used. These findings indicate that a lower baseline i-LH might be
an intrinsic characteristic of LH responders. Since these patients already
had lower baseline i-LH levels, serum LH could be easily suppressed by
TU. As LH plays an important role in male reproduction[8,9,14,15],
we propose that the present study, focusing on the changes in LH after
TU injection, will provide a baseline data for the future development
of male hormonal contraceptives. In this study, the lower baseline and
nadir levels of serum I-LH before the second injection suggests that in
order to produce profound suppression of the serum i-LH, TU injection
should be given every 4-6 weeks before the serum LH returns to the baseline.
studies have shown that administration of exogenous T to men can result
in a dose-dependent decrease in LH[16-19]. Unexpectedly, higher
serum T levels were not seen in LH responders in our study. This finding
seemed not to be a reasonable interpretation for the results when taking
into consideration serum SHBG and
T/SHBG ratio. An increase in T/SHBG ratio seems justifiable for LH-responders
who do not have higher serum T levels, as the increase in T/SHBG may be
the result of a decreased serum SHBG that induces a rise in the MCR of
T. An inverse relationship between SHBG and MCR of T has been proved by
earlier studies. Thus, it is not difficult to explain why
higher serum T levels were not seen in LH responders. It has been reported
that a fall in SHBG leads to an increase in free T; whether
other endocrine parameters have a strong negative feedback regulation
on LH, independent of the total T concentration, is still unknown.
is converted to estradiol (E2) by aromatase. Previous studies
have demonstrated that E2
has an important role on the negative feedback regulation of serum LH[22-24].
In the present study, the E2 levels were relatively higher
in LH responders, which may indicate a higher conversion rate of T to
E2. This finding also suggests that 1) the suppressive effect
of T on serum LH might be partially mediated by E2, and 2)
a combination of supraphysiological dose of T with higher physiological
levels of E2 might have a synergistic effect on the negative
feedback regulation. This
finding raises further concern in the application of T/estrogen combination
for the enhancement of LH/FSH suppression in male contraceptive research.
results demonstrated that both I-LH and b-LH were similarly suppressed.
However, some inconsistent changes between I-LH and b-LH were also seen
(Figure 5). When I-LH value was
over 30 IU/L, relative higher b-LH activity was not seen, indicating that
these LH molecules are more immuno-active but less biologically active.
In contrast, at points where b-LH value .
contrast to the significant difference in baseline I-LH levels between
the two groups, their baseline b-LH levels were similar. Thus, B/I ratio
at the baseline was lower in LH non-responders. LH non-responders may
was over 200 IU/L, these LH molecules were found to be more bio-active
but less immuno-active. These findings suggest that there are differences
in modification of LH molecules.
a relatively higher ratio of biologically inactive LH molecules (deglycosylated
form) or LH subunits before TU treatment. In both groups the pattern of
b-LH suppression curve was similar
to that of I-LH. The B/I ratio was slightly fluctuating within each group,
which indicated that TU might be inducing a quantitative change in serum
LH without any qualitative change. This finding is not in consistence
with previous reports that T enhanced LH biopotency and increased B/I
ratio. A possible explanation for the enhancement of b-LH
activity after T therapy was attributed to the fact
that the synthesis of the
abnormal LH molecules could have been altered by T or its metabolites
within the pituitary since gonadal steroids were known to modify the process
of transcription. However, Bagatell reported that administration
of T or E2 to patients with idiopathic hypothalamic hypogonadism,
who were treated with GnRH infusion until their gonadotropins reached
the normal range, resulted in decreased levels of b-LH and i-LH in the
same ratio. In interpreting whether T treatment will induce
an increase in B/I ratio or not, the hormonal status of patients, such
as hypo- or hypergonadotropic hypogonadism, and the type and dose of androgen
administered should be considered carefully.
present study confirmed that men with Klinefelters syndrome, who had
a higher baseline level of I-LH, had a higher resistance of I-LH to T
treatment. In KlInefelters syndrome, androgen replacement therapy was
still efficacious even without normalization of serum I-LH and FSH.
Therefore, during androgen replacement therapy in Klinefelters syndrome,
the usefulness of serum i-LH assessment remains questionable. In this
situation, the efficacy of androgen replacement therapy may be estimated
mainly through improvement of symptoms and signs, clinical chemistry and
metabolic parameters. Klinefelters syndrome had a lower resistance of
I-LH to exogenous T, thus serum I-LH, T (T/SHBG) and E2 would
be appropriate parameters for assessing the adequacy of androgen replacement
summary, serum I-LH suppressive response to exogenous TU in men
with Klinefelters syndrome was heterogenous. LH responders had a lower
baseline serum I-LH, and higher T/SHBG and E2 levels during
TU treatment. These endocrine parameters may be responsible for a decrease
in serum I-LH. In both LH responders and non-responders, TU induced a
quantitative change in serum LH without any qualitative change. The authors
suggest that a higher initial dose of TU injection (1000 mg) is
important for adequate suppression of LH. Repeated injections with a lower
dose (500 mg) should be given (every 4-6 weeks) before LH returns to the
baseline in order to produce appropriate LH suppression for the purpose
of male fertility regulation.
Nieschlag E, Wang C, Handelsman DJ, Swerdloff RS, Wu FCW. Guidelines for
the use of androgens in men.
Geneva: WHO Special programme of research development and research training
in human reproduction. 1992.
to: Yi-Qun GU, M.D., Reproductive Medical Center, National Research Institute
for Family Planning (WHO Collaborating Center for Research in Human Reproduction),
Beijing 100081, China.