Bioequivalence Vasoconstriction Assay Studies for Topical Corticosteroids – BioPharma Services

Bioequivalence Vasoconstriction Assay Studies for Topical Corticosteroids

Understanding the Vasoconstriction Assay for Topical Corticosteroids

In general, traditional pharmacokinetic (PK) endpoint studies, comparing the rate and extent of absorption of the drug estimated from the drug concentrations in a biological fluid, are a strong surrogate to predict the drug’s safety and efficacy of new or generic products. However, this approach assumes the investigational drug has sufficient bioavailability and systemic exposure, which pose major challenges to the locally-applied, locally-acting (LALC) products, such as topical dermatologic corticosteroid formulations (TC).

In those formulations (e.g. cream, ointment, lotions, etc.), the drug molecules have to diffuse through the multiple layers of skin to reach the capillaries, the systemic exposure is often minimal, and relying solely on pharmacokinetic endpoints can be misleading due to the unreliable pharmacokinetic  profile of the drug. Thus, a pharmacodynamic (PD) endpoint study should provide a better approach to assess the topical dermatologic corticosteroid performance.

This is where the vasoconstriction assay comes into play. The TC can cause vasoconstrictions (narrowing blood vessels) in the skin which leads to skin blanching, providing a direct estimation of the drug’s local effect by skin color. Thus, this skin color change is selected as the pharmacodynamic endpoint for bioequivalence evaluation for TC.

This blog explores the use of vasoconstriction assay in clinical studies for topical corticosteroids, including some tips and tricks in the execution of a vasoconstriction assay study.

Instruction of Vasoconstriction Assay Study

A vasoconstriction assay (VCA) is a pharmacodynamic test that measures the degree of vasoconstriction induced by a topical corticosteroid. The mechanism of the skin blanching effect caused by corticosteroids was still not fully pictured. Still, the degree of vasoconstriction is proportional to the rate and extent of corticosteroid diffusion which relates to the potency and, consequently, its therapeutic efficacy. The assay is typically conducted by applying the corticosteroid formulation to the skin, usually on the forearm, and measuring the resultant blanching or whitening of the skin using a chromameter or visual scoring.6 Both FDA and EMA issued guidance outlining the requirements of VCA studies.

Overall Designs

Number of studies

Pilot and pivotal studies.

The pilot study should be conducted with the reference-listed drug (RLD) only to configure the optimum conditions for the pivotal study. Based on the results from the pilot study, the study conditions could be established such as optimum dose duration and time to reach ED50, where the pivotal study will follow those parameters.

PD Measurement

The most convenient, acceptable by agencies, and non-invasive method was measuring surface color by chromameter with three wavelength filters (450, 560, and 600 nm). The chromameter converted the detected signal into three coordinates of L* (luminosity), a* (amount of green or red), and b* (amount of yellow or blue) by only using a-scale (red-green) data to provide the greatest sensitivity.

Additionally, other methods could also assess the skin blanching such as reflectance spectroscopy, thermography, and laser Doppler velocimetry. Qualifying the instruments before the study is necessary to ensure the reproducibility of the equipment and minimize the intra- and inter-operator measurements.

Data analysis

baseline subtraction of blanching response with the mean of untreated control site on the same arm is required. Then the area under the effect curve (AUEC) will be computed with the trapezoidal rule. Only data from detectors will be included in the data analysis with the criteria listed in equation 2.2.

Equation 2.2 Dose duration-response criterion to define detector.

AUEC at D2 = average of AUEC at D2 (2 times of duration to reach ED50) from both arms

AUEC at D1 = average of AUEC at D1 (0.5 times of duration to reach ED50) from both arms

Dose duration response and model

Various models could correlate the drug dose and the PD effect, a simple model with the measure of effect (E) in terms of baseline effect (E0), maximal effect (Emax), and dose duration to reach ED50 is preferred unless the model is deemed unappropriated alternative model could be used with proper justification.

Equation 2.1: Simple dose duration model

Important procedures

Dosing procedure

The application of dose durations to skin sites on the forearms of each participant is being randomized. The dose durations and control sites in the pivotal study should include untreated control (UNT), D1 and D2 (the dose duration calibrated with reference standard in the pilot study), test (T), and reference (R) (duration to ED50). All the application sites should be in pairs on each arm. Figure 2.2.1 shows an example of a sequence with a total of 16 sites equally distributed on two arms of a participant.

Figure 2.2.1 An example of application sequence for a participant

Elbow
Wrist

Participants screening:

The thickness and structure of the stratum corneum may have an enormous variability in the “bleaching effect” between individuals. Therefore, it will be necessary to perform a skin blanching assessment. This screening assessment should be performed before and at 2 hours after the 4- to 6-hour application period, with either a chromameter or multiple unit (3 or 4) scale visual reading. The best participants with at least a unit difference will be classified as responders and eligible for the clinical study.

Study day activity:

Considering the pharmacodynamic effect related to blood vessel narrowing, there will be a list of activity restrictions that will need to be applied to the trial participants. For instance, participants cannot exercise or have strenuous activity until the study period is finished. Also, trial participants cannot take a bath or shower before the last assessment of skin blanching is taken. The goal is to minimize the alteration to measurements which affect the study outcome.

Applications of Vasoconstriction Studies

There were several clinical studies performed with various settings and topical steroids, let’s have a look at each of the examples which provided a better understanding of how to execute a clinical study with vasoconstriction assay.

Subjective vs chromameter assessment

Gome et. al. performed a pilot and pivotal studies. The pilot study was used to test for the optimal administration period. Only positive steroid responders were included in the study. Assessment of vasoconstriction was performed before treatment, 0.5, 4, 6, and 24h post-treatment. The measurements were taken with a subjective rating scale with 4 grades and an objective chromameter method. The study concluded the subjective scoring system is not comparable to the one from the chromameter unless the blanching effect is significant enough.

VCA-based bioequivalence of new reformulated mometasone cream

One pilot and one pivotal study were conducted by Krishna et. al.7, and the results of the pilot study configured the ED50 of mometasone cream. The dosing consisted of a total of 20 sites with 10 sites spread on one arm. Within the 10 sites, 8 of them were dosed with active drugs, and 2 sites were left as untreated control sites.

The vasoconstriction assay for both studies was executed with a chromameter. The chromameter’s reading was corrected for baseline reading by the average of untreated site reading. Then a negative area under the effect curve (AUEC) values, for the duration of 0–24 h post-dose removal was calculated from the final corrected (a*) values (obtained by chromameter reading) according to the trapezoidal rule. Plots were constructed and presented in Figure 2.2.1.1. The pilot study has defined the time reaching ED50 to be 90 minutes with the model in Equation 2.1, thus, the D1 and D2 were defined as 45 minutes and 180 minutes.

Figure 2.2.1.1: Overlay graph of mean a-values chromameter scores vs reading time for each dose duration in the pilot VCA study conducted by Krishna et. al.

Based on the D1, D2, and ED50 found in the pilot study, a pivotal study was conducted with 162 subjects. The dose duration was based on the optimum dose duration configured from the pilot study. Only subjects identified as “detectors” based on the definition listed in Equation 2.2 were included in the statistical analysis, thus, 105 subjects were qualified at the end. The geometric mean ratio (GMR) of the test/reference of AUEC from the study was 112.91% with a 90% confidence interval (CI) contained within 80% to 125%. Therefore, the study concludes the bioequivalence between the reformulated mometasone cream with the marketed reference product.

Potency and Emax relationship study by VCA

Clinical studies were done by Zvidzayi et. al.8 with four different topical corticosteroids, clobetasol propionate, mometasone furoate, halcinonide, and fluocinolone acetonide were chosen and a 0.1% topical solution was prepared for each molecule. Subjects were screened and those who showed acceptable blanching response were included in the study.

The dosing response duration of the study was chosen as 5, 10, 20, 40, 60, 90 and 150 minutes. Figure 2.2.2.1, illustrates how the sites were distributed for the two products and the skin blanching effect on the forearm of participants.

Figure 2.2.2.1 The blanching response after randomized application site of halcinonide and clobetasol propionate solution

The blanching response after randomized application site of halcinonide and clobetasol propionate solutio

The blanching response after randomized application site of halcinonide and clobetasol propionate solutio

All measurements were taken by chromameter, and only a-scale data was used in the statistical analysis. Like the study mentioned in section 2.2.1, the data were plotted with the Emax model listed in Equation 2.1. Before plotting the AUEC curve for each dose duration against the time of data collection, a baseline correction with a-value obtained from the untreated site was performed with the negative number multiplied by -1 to get a plot of that dose duration in proportion to skin blanching response.

The study rejected the null hypothesis where Emax values for all topical corticosteroids are equal. This study provides insight for using VCA to collect the inherent potencies of individual active pharmaceutical ingredients. Such an approach could aid the future development and optimization of topical corticosteroid products.

Limitations and Considerations when utilizing VCA for clinical study

While the VCA is powerful and easy to execute, there are still some limitations and considerations:

    • Sample size and inter-subject variability: Differences in skin thickness, vascularization, and other factors can lead to variability in response between individuals, which must be accounted for in the study design. An additional factor that affects the sample size estimation is the definition of detector mentioned in section 2.1. Data from participants not fulfilling the detector criteria will be excluded, therefore, a large sample size with buffers will be required to have sufficient power to demonstrate bioequivalence with VCA.
    • Subjectivity in Scoring: Although chromameters can provide objective measurements and are recommended by regulatory agencies, visual scoring by trained observers is still common in screening, which can introduce subjectivity in subject recruitment.
    • Limited to Corticosteroids: The assay is specifically designed for corticosteroids and may not apply to other classes of topical drugs. Corticosteroids produce an effect of vasoconstriction that causes the skin to turn pale and VCA is making use of this PD effect. However, other classes of topical drugs may not share a similar property.

Why Choose BioPharma Services for your Next Drug Development Project?

The vasoconstriction assay provided an alternative path in the bioequivalence testing of topical corticosteroids. Its ability to directly measure the pharmacodynamic effect of these drugs makes it an invaluable tool in ensuring that generic formulations meet the same standards of efficacy as their brand-name counterparts. However, like any scientific method, it requires careful execution and interpretation to account for its inherent variability and limitations. Even though the pharmaceutical industry keeps evolving, the vasoconstriction assay will remain a critical component of the bioequivalence landscape, ensuring patients can access safe, effective, and affordable medications.

Choosing a partner like BioPharma Services for your drug development process ensures that you are working with experts who are knowledgeable and experienced to evaluate, design, and execute a clinical study with vasoconstriction assay tailored to your needs. With our expertise in pharmacokinetics and a thorough understanding of regulatory requirements, we can navigate you through the obstacles during drug development, ensuring your product meets the expected safety and efficacy requirements.

Written By:

Anson Wu

Pharmacokinetics Associate

BioPharma Services, Inc., a HEALWELL AI and clinical trial services company, is a full-service Contract Clinical Research Organization (CRO) based in Toronto, Canada, specializing in Phase 1 clinical trials 1/2a, Human Abuse Liability(HAL) and Bioequivalence clinical trials for international pharmaceutical companies worldwide. BioPharma Services conducts clinical research operations from its Canadian facility, with access to healthy volunteers and special populations.

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