madman
Super Moderator
* The primary aim of this study was to compare serum TT levels between patients treated with the standard once-daily regimen and those using a twice-daily split-dose schedule. Given the real-world and self-initiated nature of the dosing modification, the present study should be considered exploratory and hypothesis-generating.
* (1) treatment with 2% testosterone gel (Tostrex®, Advanz Pharma Italia S.r.l., Italy) administered at a daily dose of 5 actuations (50 mg/day); (2) spontaneous transition from once-daily to twice-daily application (morning and evening) while maintaining the same total daily dose; (3) availability of serum TT measurements at both peak and nadir before and after regimen modification.
*Peak measurements were obtained 3 h after the morning application, and nadir samples immediately before the subsequent dose, according to standardized clinical practice instructions.
Several formulations are available—including injectable preparations, transdermal gels, patches, subcutaneous pellets, and oral tablets—and the choice depends on pharmacokinetic profile, cost, comorbidities, and, most importantly, patient preference and adherence [2].
Among these, transdermal testosterone gel is the most widely prescribed formulation [4, 12]. After topical application, serum testosterone typically peaks within 2–4 h, followed by a gradual decline over 4–6 h, with a low risk of supraphysiological levels [13].
In our clinical practice, we consider a TT threshold of 3.5 ng/mL as the lower acceptable limit during TRT. To assess treatment adequacy, guidelines recommend measuring TT levels 2–4 h [3] (or 2–8 h [1]) after gel application, corresponding to the expected pharmacokinetic peak.
Transdermal gel therapy is generally well tolerated. The most common adverse effects are local skin reactions - burning, tingling, dryness, erythema, or itching—which usually decrease or resolve with continued use [17, 18]. However, many patients fail to maintain stable concentrations throughout the day following application of TRT gel. This is unsurprising, since pharmacokinetic data from clinical trials rarely apply to the real-world setting, and as such several factors can affect not only the absorption rate through the skin, but also individual metabolism of the drug itself. In a recent study, in order to better evaluate overall exposure to TRT, the authors proposed measuring TT 23 h post-application, just before the next dose, in patients who achieved adequate TT levels 2 h after application [18]. This approach was also employed in a large multicentre American randomized controlled trial (TRT gel vs. placebo) involving 5,264 hypogonadal men, which found that assessing TT 24 h post-application was clinically useful for treatment adjustment [10]. We frequently apply this method in our clinical practice, particularly for older men or patients with demanding work schedules.
During routine clinical practice, we observed a patient who had independently modified his prescribed once-daily testosterone gel regimen by splitting the dose into two applications (morning and evening) to alleviate local skin reactions. Following this observation, we identified 11 additional patients who had adopted the same twice-daily administration schedule while maintaining an identical total daily dose. The primary aim of this study was to compare serum TT levels between patients treated with the standard once-daily regimen and those using a twice-daily split-dose schedule. Given the real-world and self-initiated nature of the dosing modification, the present study should be considered exploratory and hypothesis-generating.
Materials and methods
Patients
Patients were recruited by searching the database of hypogonadal subjects on TRT treatment followed up at the Andrology outpatient service of the Endocrinology Unit, ASST Spedali Civili of Brescia. The search criteria were as follows: (1) treatment with 2% testosterone gel (Tostrex®, Advanz Pharma Italia S.r.l., Italy) administered at a daily dose of 5 actuations (50 mg/day); (2) spontaneous transition from once-daily to twice-daily application (morning and evening) while maintaining the same total daily dose; (3) availability of serum TT measurements at both peak and nadir before and after regimen modification.
All patients had been receiving stable once-daily testosterone gel therapy for at least 6 months before switching regimen, ensuring steady-state conditions. Laboratory evaluations under the fractionated regimen were performed after a minimum of 6 months of stable twice-daily administration. This approach ensured that all measurements reflected steady-state conditions under both dosing regimens. Formal adherence measures were not available due to the retrospective design; however, adherence and treatment continuity were routinely assessed during outpatient visits through structured patient questioning, in accordance with good clinical practice.
Peak measurements were obtained 3 h after the morning application, and nadir samples immediately before the subsequent dose, according to standardized clinical practice instructions.
The study was approved by the Local Ethics Committee (N. 6182).
Laboratory assessments
Laboratory tests were performed in all patients according to standard clinical practice. All blood samples were collected in the morning after an overnight fast. Blood samples were collected to evaluate total testosterone (TT: normal range: 2.49–8.36 ng/mL; limit of quantitation 0.12 ng/mL), luteinizing hormone (LH; normal range: 1.7–8.6 IU/L; limit of quantitation 1 mIU/mL), haematocrit (HCT; normal range: 42–52%), haemoglobin (Hb; normal range for male patients: 14–18 g/dL), and prostate-specific antigen (PSA; normal range: <1.4 mcg/L) were performed. Circulating free testosterone (cFT) was calculated based on total testosterone, sex hormone-binding globulin (SHBG; normal range: 18.8–54.1 nmol/L; limit of detection 0.35 nmol/L), and albumin (normal range: 3.1–5.2 g/dL) levels according to the Vermeulen formula [19, 20]. All measurements were performed after at least 6 months of stable treatment under each dosing regimen. Serum TT levels were collected at both peak (3 h after gel application) and nadir (immediately before the subsequent application). Patients were classified as achieving therapeutic targets when peak TT values fell within the 3.5–9 ng/mL range [4]. This interval was used as a pragmatic classification tool to distinguish under-,within-, and over-treatment categories. In particular, the upper threshold (9 ng/mL) was not intended as a therapeutic target but rather to identify values approaching the upper limit of the physiological reference range, thus indicating clearly supraphysiological exposure at peak
The present study has several important limitations. First, its retrospective design and the self-initiated dosing modification introduce potential selection and behavioural bias. Patients who independently adopted fractionated dosing may represent a more motivated or treatment-aware subgroup, possibly influencing adherence and monitoring frequency. Importantly, the self-selected adoption of fractionated dosing may identify a subgroup of patients with higher motivation, treatment awareness, or intolerance to standard regimens, thus limiting the generalizability of the findings. Additionally, adherence was not formally assessed, which may represent a further source of variability in real world treatment exposure. Second, the timing of laboratory assessments, although performed according to routine clinical practice, was not prospectively standardized in relation to exact application timing under controlled conditions. This limitation should be considered central when interpreting the pharmacokinetic implications of our findings. Third, the small sample size limits statistical power and precludes adjustment for potential confounders. The cohort was heterogeneous in terms of underlying aetiology of hypogonadism, including both primary and secondary forms. This heterogeneity further limits the generalizability of the findings. Therefore, the present findings should be considered hypothesis-generating.
In conclusion, even if the present data require confirmation in larger, controlled studies, they provide a hypothesis-generating observation suggesting that dose fractionation may modify the peak-to-pre-dose biochemical pattern observed under once-daily transdermal testosterone administration. Prospective pharmacokinetic studies with standardized full 24-hour sampling are required to clarify whether these observations translate into differences in overall daily testosterone exposure.
* (1) treatment with 2% testosterone gel (Tostrex®, Advanz Pharma Italia S.r.l., Italy) administered at a daily dose of 5 actuations (50 mg/day); (2) spontaneous transition from once-daily to twice-daily application (morning and evening) while maintaining the same total daily dose; (3) availability of serum TT measurements at both peak and nadir before and after regimen modification.
*Peak measurements were obtained 3 h after the morning application, and nadir samples immediately before the subsequent dose, according to standardized clinical practice instructions.
Several formulations are available—including injectable preparations, transdermal gels, patches, subcutaneous pellets, and oral tablets—and the choice depends on pharmacokinetic profile, cost, comorbidities, and, most importantly, patient preference and adherence [2].
Among these, transdermal testosterone gel is the most widely prescribed formulation [4, 12]. After topical application, serum testosterone typically peaks within 2–4 h, followed by a gradual decline over 4–6 h, with a low risk of supraphysiological levels [13].
In our clinical practice, we consider a TT threshold of 3.5 ng/mL as the lower acceptable limit during TRT. To assess treatment adequacy, guidelines recommend measuring TT levels 2–4 h [3] (or 2–8 h [1]) after gel application, corresponding to the expected pharmacokinetic peak.
Transdermal gel therapy is generally well tolerated. The most common adverse effects are local skin reactions - burning, tingling, dryness, erythema, or itching—which usually decrease or resolve with continued use [17, 18]. However, many patients fail to maintain stable concentrations throughout the day following application of TRT gel. This is unsurprising, since pharmacokinetic data from clinical trials rarely apply to the real-world setting, and as such several factors can affect not only the absorption rate through the skin, but also individual metabolism of the drug itself. In a recent study, in order to better evaluate overall exposure to TRT, the authors proposed measuring TT 23 h post-application, just before the next dose, in patients who achieved adequate TT levels 2 h after application [18]. This approach was also employed in a large multicentre American randomized controlled trial (TRT gel vs. placebo) involving 5,264 hypogonadal men, which found that assessing TT 24 h post-application was clinically useful for treatment adjustment [10]. We frequently apply this method in our clinical practice, particularly for older men or patients with demanding work schedules.
During routine clinical practice, we observed a patient who had independently modified his prescribed once-daily testosterone gel regimen by splitting the dose into two applications (morning and evening) to alleviate local skin reactions. Following this observation, we identified 11 additional patients who had adopted the same twice-daily administration schedule while maintaining an identical total daily dose. The primary aim of this study was to compare serum TT levels between patients treated with the standard once-daily regimen and those using a twice-daily split-dose schedule. Given the real-world and self-initiated nature of the dosing modification, the present study should be considered exploratory and hypothesis-generating.
Materials and methods
Patients
Patients were recruited by searching the database of hypogonadal subjects on TRT treatment followed up at the Andrology outpatient service of the Endocrinology Unit, ASST Spedali Civili of Brescia. The search criteria were as follows: (1) treatment with 2% testosterone gel (Tostrex®, Advanz Pharma Italia S.r.l., Italy) administered at a daily dose of 5 actuations (50 mg/day); (2) spontaneous transition from once-daily to twice-daily application (morning and evening) while maintaining the same total daily dose; (3) availability of serum TT measurements at both peak and nadir before and after regimen modification.
All patients had been receiving stable once-daily testosterone gel therapy for at least 6 months before switching regimen, ensuring steady-state conditions. Laboratory evaluations under the fractionated regimen were performed after a minimum of 6 months of stable twice-daily administration. This approach ensured that all measurements reflected steady-state conditions under both dosing regimens. Formal adherence measures were not available due to the retrospective design; however, adherence and treatment continuity were routinely assessed during outpatient visits through structured patient questioning, in accordance with good clinical practice.
Peak measurements were obtained 3 h after the morning application, and nadir samples immediately before the subsequent dose, according to standardized clinical practice instructions.
The study was approved by the Local Ethics Committee (N. 6182).
Laboratory assessments
Laboratory tests were performed in all patients according to standard clinical practice. All blood samples were collected in the morning after an overnight fast. Blood samples were collected to evaluate total testosterone (TT: normal range: 2.49–8.36 ng/mL; limit of quantitation 0.12 ng/mL), luteinizing hormone (LH; normal range: 1.7–8.6 IU/L; limit of quantitation 1 mIU/mL), haematocrit (HCT; normal range: 42–52%), haemoglobin (Hb; normal range for male patients: 14–18 g/dL), and prostate-specific antigen (PSA; normal range: <1.4 mcg/L) were performed. Circulating free testosterone (cFT) was calculated based on total testosterone, sex hormone-binding globulin (SHBG; normal range: 18.8–54.1 nmol/L; limit of detection 0.35 nmol/L), and albumin (normal range: 3.1–5.2 g/dL) levels according to the Vermeulen formula [19, 20]. All measurements were performed after at least 6 months of stable treatment under each dosing regimen. Serum TT levels were collected at both peak (3 h after gel application) and nadir (immediately before the subsequent application). Patients were classified as achieving therapeutic targets when peak TT values fell within the 3.5–9 ng/mL range [4]. This interval was used as a pragmatic classification tool to distinguish under-,within-, and over-treatment categories. In particular, the upper threshold (9 ng/mL) was not intended as a therapeutic target but rather to identify values approaching the upper limit of the physiological reference range, thus indicating clearly supraphysiological exposure at peak
The present study has several important limitations. First, its retrospective design and the self-initiated dosing modification introduce potential selection and behavioural bias. Patients who independently adopted fractionated dosing may represent a more motivated or treatment-aware subgroup, possibly influencing adherence and monitoring frequency. Importantly, the self-selected adoption of fractionated dosing may identify a subgroup of patients with higher motivation, treatment awareness, or intolerance to standard regimens, thus limiting the generalizability of the findings. Additionally, adherence was not formally assessed, which may represent a further source of variability in real world treatment exposure. Second, the timing of laboratory assessments, although performed according to routine clinical practice, was not prospectively standardized in relation to exact application timing under controlled conditions. This limitation should be considered central when interpreting the pharmacokinetic implications of our findings. Third, the small sample size limits statistical power and precludes adjustment for potential confounders. The cohort was heterogeneous in terms of underlying aetiology of hypogonadism, including both primary and secondary forms. This heterogeneity further limits the generalizability of the findings. Therefore, the present findings should be considered hypothesis-generating.
In conclusion, even if the present data require confirmation in larger, controlled studies, they provide a hypothesis-generating observation suggesting that dose fractionation may modify the peak-to-pre-dose biochemical pattern observed under once-daily transdermal testosterone administration. Prospective pharmacokinetic studies with standardized full 24-hour sampling are required to clarify whether these observations translate into differences in overall daily testosterone exposure.
