The clinical study using DCA clearly demonstrated an efficacy toward acne, albeit that the delivery of the active ingredient could still be improved. However, the reduction in P. acnes in the second study was minimal, and this is typically con- sidered to be essential for any active ingredient to exert an anti-acne activity. Although an antimicrobial activity of DCA could not be denied (Table 1), this alone was not enough to explain its activity in patients suffering from mild-to- moderate acne. In finding an explanation for this contradiction, the data in Table 3 turned out to be of crucial importance. Whereas the vehicle alone resulted in scaling in 60% of patients, the addition of DCA reduced this percentage considerably to 13% and 20%, suggesting an anti-inflammatory side effect of DCA. A separate study did indeed suggest a slight anti-inflammatory activity, albeit statistically insignificant (data not shown). As inflammation in the human skin is regulated via the PPAR, we investigated whether DCA, an unsaturated fatty acid, did indeed bind to this receptor. After all, fatty acids are the typical ligands for these receptors.
A reporter gene assay using HeLa cells transfected with chimeric receptor genes fused to a PPAR ligand-binding domain and a reporter gene with the luciferase/luciferine system was performed. Separate cell lines were developed for PPARa, d, and g. A total of 2.5 104 cells were plated into 96-well microtiter plates with phenol red-free Dulbecco’s Modified Eagles Medium (DMEM) culture media containing 6% dextran-coated, charcoal-treated fetal calf serum (DCC FCS). Cells were incubated with pharmaceutical PPAR agonists (10211 2 1025 M), DCA (1027 –1025 M), or pioglitazone [1028 –1025 M; a positive control for PPARg binding (22)] for 16 hours, and at the end of the incubation, the medium was removed and replaced with culture medium containing luciferine and a luminescent signal measured after five minutes using a Microbeta lumin- ometer (Wallac, Turku, Finland). All experiments were performed in quadruplicate.
These experiments, the results of which are illustrated in Figure 9, showed that DCA is a pan-PPAR agonist with a greater specificity for PPARg. As a specific PPARg agonist, pioglitazone naturally only binds to PPARg. Other controls for PPARa and PPARd were included but not shown here (13).
FIGURE 9 Induction of luciferase expression by octadecenedioic acid (A) and pioglitazone (B) in three separate HeLa cell lines expressing peroxisome proliferator-activated receptor a, d, and g separately. Abbreviation: PPAR, peroxisome proliferator-activated receptor.
RATIONALE FOR OCTADECENEDIOIC ACID AS A NOVEL TOPICAL ACNE TREATMENT
Acne is a common disease affecting, according to surveys, nearly one out of three teenagers, although the disease is not limited to preadolescents only. It markedly influences the quality of life and constitutes a socio-economic problem. Worldwide costs for systemic and topical acne treatment were calculated to represent 12.6% of the annual overall costs for the treatment of skin diseases (23). Its pathogenesis is multifactorial, with abnormal follicular differentiation and increased cornification, enhanced sebaceous gland activity and hyperseborrhea, bacterial hypercoloniza- tion as well as inflammation, and immunological host reactions being the major contributors (23). It is characterized by a variety of expressions and degrees depending on the distribution, type of lesions, tendency to and manifestation of scarring, start of disease in puberty, or persistence after time of physiological
regression, each form requiring a different active ingredient or drug or combination thereof (24).
There are basically four modes of action for treating acne in line with the pathogenesis described earlier: most anti-acne agents have an antimicrobial activity to counteract the hypercolonization (in particular, e.g., BPO), some agents interact with the hyperkeratinization aspect of acne to fight the abnormal follicular differen- tiation and increased cornification (in particular, the retinoids such as tretinoin, iso- tretinoin, and adapalene), a few have only a moderate anti-inflammatory activity (e.g., adapalene, AZA, and BPO), whereas none have a strong sebo-suppressive effect to fight the hyperseborrhea (24). Because of the complexity of the disease, it is very difficult to differentiate between cause and effect, and as a consequence, relapses often occur because the fundamental underlying cause may not have been dealt with. As a consequence, combination therapies are popular, involving both the topical and the oral route of administration. Mammone et al. (25), for instance, proposed a combination of chemicals with antimicrobial, anti-inflammatory, and antiandrogen properties and a desquamation enhancer, resulting in a statistically significant higher reduction in noninflamed lesions relative to a 5% BPO gel, whereas there was no statistically significant additional benefit toward inflamed lesions. The downside of combination therapy, however, is that this increases the chance of side effects, as most of these products have some. AZA, for instance, causes a very strong burning and BPO results in strong scaling and erythema (24), which was con- firmed in our studies (Table 3).
Here, we report the use of DCA as a novel topical treatment of acne that involves all four modes of action within a single molecule. Similar to most other anti-acne agents, DCA has an antimicrobial activity as indicated in Table 1. All three other modes of action may be produced via its binding to PPAR, as shown in Figure 9, which illustrates its capability to act as a pan-PPAR agonist. Binding to PPARg is most pronounced, followed by that to PPARa and PPARd. As shown by Downie et al. (5), activators of PPARa and PPARg inhibit the rate of sebaceous lipogenesis and reduce the synthesis of the sebum-specific lipids squalane and tria- cylglycerol in human sebaceous glands. DCA, as a PPARg and PPARa agonist, is therefore likely to do the same, provided it is delivered well enough. The import- ance of PPAR in the regulation of lipid synthesis and metabolism in human sebo- cytes is recognized, albeit far from clear yet. Makrantonaki and Zouboulis (26), for instance, described the stimulatory interactions between androgens and PPAR ligands, similar to the natural PPARd/g ligand linoleic acid, on neutral and polar lipid synthesis. Although linoleic acid is stimulating lipid synthesis in vitro in this communication (26), topical administration of 2.5% linoleic acid in a Carbopol gel formulation for one month resulted in a significant reduction (25%) in the overall size of follicular casts and microcomedones, whereas no change was found at placebo-treated sites (27). Further research will be necessary to elucidate the interrelationships between all factors involved.
DCA, however, is one of the very few molecules that has the potential to interact at all levels with acne: enhanced differentiation to prevent the increased cornification of the sebaceous duct (via PPARd), reduction in sebum production to counteract hyperseborrhea (via PPARg), and reduction in inflammation (PPARa). These PPAR-related effects combined with its demonstrated antimicro- bial properties to overcome the hypercolonization with microbes such as P. acnes and S. aureus, make DCA a viable nonantibiotic alternative for the treatment of acne.