Stabilization of Interferon alpha-2b in a Topical Cream - Pharmaceutical Technology

Latest Issue
PharmTech

Latest Issue
PharmTech Europe

Stabilization of Interferon alpha-2b in a Topical Cream
The authors describe a proprietary process for producing a stable, topical interferon alpha-2b formulation that can deliver large drug molecules into the skin or mucosa.


Pharmaceutical Technology
Volume 33, Issue 7, pp. 80-86

Identification of Component A in the formulation

The prevention of IFNα-2b oxidation was the most significant challenge in developing a topical IFNα-2b formulation. IFNα-2b contains five methionine residues (Met16, Met21, Met59, Met111, and Met148), which are susceptible to oxidation in solution (6). Giltlin et al. demonstrated that the oxidation of the Met111 residue produced a methioninesulfoxide derivative that has similar biological activity to the native IFNα-2b (7). This derivative is potentially a major oxidative product in Interferon alpha-2b Cream and is generally known as Component A. In the authors' research, activity of IFNα-2b was measured using a cell-based antiviral assay (AVA) that was shown to be nonspecific for the active pharmaceutical ingredient (API) versus Component A. A sensitive reversed-phase high-performance liquid chromatography (RP-HPLC) assay using fluorescence detection was developed to monitor the formation of the oxidative degradation product (Component A) and to quantify the amount of IFNα-2b.


Figure 1: Reverse-phase high-performance liquid chromatography assays using fluorescence detection as follows: (a) unoxidized interferon alpha-2b (IFNα-2b) in solution 80 g/g; (b) oxidized IFNα-2b in solution 80 g/g after 5 h; (c) spiked-placebo formulation with stressed active pharmaceutical ingredient and recovery of Component A; (d) active formulation after 5 months at real-time storage; (e) forced degradation of active formulation after 5-h exposure to 0.25% hydrogen peroxide (H2O2); and (f) forced degradation of active formulation after 5-h exposure to 3% H2O2. EU is emission unit; IFN = IFNα-2b.
When analyzing concentrated solutions containing 80g/g IFNα-2b, Component A was identified as eluting before the principal IFNα-2b peak at approximately 1% of the principal peak area (see Figure 1a). To confirm the formation of Component A, the forced degradation of IFNα-2b in solution showed that while the principal IFNα-2b peak decreased by 16%, a corresponding increase was observed in the Component A peak. The forced degradation of IFNα-2b in solution was conducted in the presence of an oxidizing agent (0.25% hydrogen peroxide [H2O2]) for 5 h, and the reaction was stopped by the addition of methionine (see Figure 1b) (6).

The AVA and the enzyme-linked immunosorbent assay detected no change in biological activity and content, respectively, as a result of oxidative stress. Formation of aggregates was not observed upon oxidative stress of IFNα-2b, as suggested by ultraviolet–visible spectroscopic data and sodium dodecyl sulfate polyacrylamide gel electrophoresis studies (Data not shown).

In the chromatogram of stressed IFNα-2b in solution (see Figure 1b), Component A was positively identified as eluting prior to the principal IFNα-2b peak, and the peak area was used to calculate its theoretical concentration (assuming Component A was equivalent to IFN-a2b). The stressed API solution was used to spike placebo formulation at a target concentration of 125 ng/g Component A (see Figure 1c). The percent recovery of Component A from spiked placebo formulation (see Table II) confirmed that if IFNα-2b had been oxidized in the formulation, the main oxidative product would have been extracted and detected using the RP-HPLC method.


ADVERTISEMENT

blog comments powered by Disqus
LCGC E-mail Newsletters

Subscribe: Click to learn more about the newsletter
| Weekly
| Monthly
|Monthly
| Weekly

Survey
FDASIA was signed into law two years ago. Where has the most progress been made in implementation?
Reducing drug shortages
Breakthrough designations
Protecting the supply chain
Expedited reviews of drug submissions
More stakeholder involvement
Reducing drug shortages
70%
Breakthrough designations
4%
Protecting the supply chain
17%
Expedited reviews of drug submissions
2%
More stakeholder involvement
7%
View Results
Eric Langerr Outsourcing Outlook Eric LangerRelationship-building at Top of Mind for Clients
Cynthia Challener, PhD Ingredients Insider Cynthia ChallenerRisk Reduction Top Driver for Biopharmaceutical Raw Material Development
Jill Wechsler Regulatory Watch Jill Wechsler Changes and Challenges for Generic Drugs
Faiz Kermaini Industry Insider Faiz KermainiNo Signs of a Slowdown in Mergers
CMOs and the Track-and-Trace Race: Are You Engaged Yet?
Ebola Outbreak Raises Ethical Issues
Better Comms Means a Fitter Future for Pharma, Part 2: Realizing the Benefits of Unified Communications
Better Comms Means a Fitter Future for Pharma, Part 1: Challenges and Changes
Sandoz Wins Biosimilar Filing Race
Source: Pharmaceutical Technology,
Click here