Current Research in Cosmeceuticals R. Randall Wickett, Ph.D. James L. Winkle College of Pharmacy randy.wickett@uc.edu

What is a Cosmeceutical? Everyone knows that the term Cosmeceutical was coined by Albert Kligman about 25 years ago. Like so many things that “everyone knows” this is incorrect. The first known public use of “Cosmeceutical” that I can document was in Raymond Reed’s SCC Medal Award Speech in December of 1961, published in the Journal of the Society of Cosmetic Chemists in January of 1962. He stated that they had been using the term in his company for many years. Kligman began to use the term widely and popularized it about 25 years ago. Kligman AM. Why cosmeceuticals? Cosmet Toilet 1993;108:37-8. Albert Kligman

What is a Cosmeceutical? According the US Food and Drug Administration Office of Cosmetics and Colors “The FD&C Act does not recognize and such category as ‘cosmeceuticals’. A product can be a drug, a cosmetic, or a combination of both, but the term cosmeceutical has no meaning under the law.” Since the term has no meaning under the law (at least in the US) we are left to our own devices to define it. It has come to mean “active cosmetics” that have positive effects on the skin that may go beyond cleansing or moisturization. Most commonly it has come to refer to “actives” that have so called “anti-aging” effects by whatever mechanism.

Some Cosmeceuticals Gau et. Al. (2008) Clinics in Dermatology 26 367-374

RETINOIDS “Gold Standard” against which all other for photo-aging could be compared. Metabolites of retinol Best known is all trans retinoic acid or tretinoin Induces smoothing of skin by affecting dermis Clinically proven to reduce wrinkles It the US tretinoin is drug. Thus it is not a cosmeceutical, it is a pharmaceutical – however study of the mechanisms of tretinoin action may lead to improved “cosmeceutical approaches” to anti-aging

Retinol and metabolites

RETINOIDS Olsen et.al showed the potential effectiveness of topical tretinoin emollient cream (TEC) for treating photo-damaged skin. 320 healthy, white persons with mild to moderate facial photo-damage were selected. Subjects were equally randomized into 4 treatment groups, TEC 0.05%, TEC 0.01%, TEC 0.001% Vehicle. Olsen et al(1992) J. Amer. Acad. Dermatol 26: 215-224

Duration of treatment was 24 weeks. RETINOIDS Subjects were instructed to apply the assigned test cream to the entire face. Duration of treatment was 24 weeks. 2mm punch biopsy specimens were obtained before therapy and after 24 weeks of treatment from adjacent areas. Olsen et al(1992) J. Amer. Acad. Dermatol 26: 215-224

RETINOIDS (A) Pretreatment appearance (A) Pretreatment appearance (B) Appearance at week 24 of TEC (A) Pretreatment appearance (B) Appearance at week 24 of TEC

RETINOIDS TEC 0.05% significant difference from the vehicle in reducing the overall severity of photo- damage from baseline to the end of the therapy.

RETINOIDS Mottled pigmentation, fine wrinkling and roughness decreased to a greater extent after TEC 0.05% therapy compared to vehicle.

RETINOIDS There was increase in epidermal thickness and granular layer thickness. The melanin content was reduced by 56% in TEC 0.05% group and 57% in TEC 0.01% group.

RETINOIDS CONCLUSIONS Many visible signs such as wrinkling and mottled pigmentation are a result primarily from cumulative sun exposure. Most notable clinical changes on treating with TEC are reduction in fine wrinkling, roughness and mottled pigmentation. Thus, TEC appears to be effective in the treatment of photo-damaged skin.

Retinol a cosmetic anti-aging active or “cosmeceutical”

Effects of retinol and retinyl propionate on wrinkling and hyperpigmentation Reduction in wrinkling and hyperpigmentation caused by retinol (ROH) and retinyl propionate (RP). Expert graders (0-4 scale) evaluated reduction vs baseline in wrinkling and hyperpigmentation at 4, 8, and 12 weeks (average data presented.) The low irritation of RP vs ROH permits use of higher levels to achieve greater effects without significant negative aesthetic issues. Bissett, Clinics in Dermatology (2009) 27, 435–445

Scott reported that retinol and AHA + retinol were effective against photodamged skin on forearms Scott, proceedings of the 22nd IFSCC conference, Edinburgh 2002

In a face study retinoic acid was clearly superior to AHA+Retinol Scott, proceedings of the 22nd IFSCC conference, Edinburgh 2002

Niacinamide Water soluble form derivative of B Vitamin. When added to a sunscreen it lightened skin tone. Promoted as an ingredient the smooths surface texture Olay total effects L’Oreal Plentitude May interfere with transfer of pigment granules from pigment producing cells(melanocytes) to keratinocytes.

Niacinamide (Nicotinamide)

5% Niacinamide reduced hyperpigmentation in human clinical trial. Hakozaki et al.(2002) Brit. J. Derm. 47 20-31

5% Niacinamide over 8 weeks of treatment Greatens et al Experimental Dermatol 14 498-508 (2005)

Niacinamide does not lighten human melanocytes in culture Greatens, Wickett and Boissy, unpublished data

Flow Cytometry Analysis assay used to determine mean fluorescence two dyes used- CFDA and PE outcome indicates the % inhibition of melanosome transfer

Niacinamide inhibited melansome transfer in vitro. To demonstrate melanosome-transfer inhibition, melanocytes were labeled with a succinimidyl ester of CFDA and cocultured with keratinocytes. These cocultures were assessed by confocal microscopy for CFDA transfer in the presence or absence of 10 mM niacinamide after 6 days of treatment. Control cultures demonstrated brightly fluorescing keratinocytes (arrowheads) within a colony of keratinocytes (star) that also contain CFDA-positive melanocytes (arrows) The corresponding differential contrast image demonstrates that the melanocytes were darkly pigmented (arrows). In contrast, treatment with niacinamide resulted in only weaklyfluorescing keratinocytes within a colony (star) that also contained CFDA-positive melanocytes of equal brightness as observed in the control cultures (arrows). The corresponding differential contrast image demonstratedthat the melanocytes were darkly pigmented (arrows). Greatens et al Experimental Dermatol 14 498-508 (2005)

The effect of niacinamide on pigmentation in vivo is reversible Greatens et al Experimental Dermatol 14 498-508 (2005)

12 weeks of treatment with niacinamide reduced facial wrinkles measured by image analysis more than placebo Bissett et al (2004) Int. J. Cos. Sci. 26 231-238

Kinetin Kinetin is a plant growth hormone. It is reported to have “anti-aging” and “anti-oxidant effects on cells. Chiu et. Al. reported on the effects of combining kinetin and niacinamide in a human clinical trial. Chiu et. Al. (2007) J. Cosmet. Dermatol. 6 243-249 The authors found the comination to be effective in reducing facial wrinkles.

Effects of Niacinamide and Kinetin on facial wrinkles

ANTIOXIDANTS Free radical scavengers or chemicals that intercept and neutralize free radicals. Skin - Target organ of environmental photo- oxidative stress. Reactive oxygen species result in structural and functional alterations of skin (i.e. breakdown of type III collagen Thus, the use of anti-oxidants that attenuate photo- oxidative toxicity is believed to be an important strategy in modulating photo-aging.

Vitamin C

Topical vitamin C has been reported to improve photoaging Fitzpatric and Rostan(2002) Dermatol Surg 28:231-236

Improvement in wrinkle scores C was significant over baseline but NOT over vehicle 10% Ascorbic Acid, 7% tetrahexyldecyl ascorbate Treatment for 12 weeks, double blind split face. Fitzpatric and Rostan(2002) Dermatol Surg 28:231-236

Vitamin C and photoaging Raschke et. Al. reported that ascorbic acid in an oil-in-water emulsion was superior to its vehicle in a 12 week split face test. Raschke et. Al. (2004) Skin Parmacol Physiol 17: 200-206 The authors reported reduced oxidative stress in the skin and significant improvement in wrinkles (roughness values by PRIMOS) compared to vehicle

Decrease in rq (root mean square roughness) with treatment Data replotted from Raschke et. Al. (2004) Vertical axis is reduction in rq as measured by PRIMOS result at 12 weeks is statistically significant.

ANTIOXIDANTS CONCLUSIONS Anti – oxidants inhibit the propagation of lipid peroxidation and should help to reduce skin damage associated with free radicals. Topical vitamin C treatment was reported to reduce signs of photoaging but was not significantly better than its vehicle in one study but did have a significant effect on fine wrinkles measured by PRIMOS in another study.

Lipoic acid 6-8 dithiooctanoic acid Reported to be effective for treating photoaging

Effect of 3 month treatment with 5% LA Beitner Brit. J. Dermatol. 149 841-849 (2003)

Laser profilometry indicated a significant improvement in wrinkles

Peptides Recently there has been considerable interest in the use of short peptide sequences as cosmeceutical actives Peptide sequences that can stimulate fibroblasts to produce collagen in-vitro are used. The most widely studied sequence is lysine-threonine-threonine-lysine-serine (KTTKS) found on type I procollagen. See Lupo and Cole (2007) Cosmeceutical Peptides Dermatologic Therapy 20:343-349 for a review.

Robinson et al reported that topical application of a palmitoyl derivative of KTTKS, PAL-KTTKS improved signs of photoaging. Robinson et. Al(2005) Int. J. Cosmet. Sci 27 155-160

Improvement of wrinkle grades by pal-KTTKS Robinson et. Al(2005) Int. J. Cosmet. Sci 27 155-160

Image analysis also showed directional improvement

Results Expert grading of photographs did show a significant effect versus placebo at 4 weeks (but not at 12) Subjects self assessment of “age spots” showed a significant advantage over placebo. The authors concluded that “Topical 3-ppm pal-KTTKS was shown to provide a reduction in facial lines/wrinkles in a 12 week clinical test”.

NAG Glucosamine and N-acetyl glucosamine are precursors of hyaluronic acid. These “sugar amines” have been investigated as both oral and topical ingredients for improving skin condition Bissett reports that 2% NAG applied topically has been found to have positive effects on both wrinkles and hyperpigmentation. Bissett, D. (Clinics in Dermatology (2009) 27, 435–445

NAG enhanced the ability of Niacinamide to reduce “age spots” Computer image analysis of Caucasian facial digital images for change in spot area fraction. More negative numbers indicate reduction in hyperpigmentation (improvement). N, niacinamide; NAG, N-acetyl glucosamine. P is for 4% N + 2% NAG vs 4% N. Bissett, D. (Clinics in Dermatology (2009) 27, 435–445

Gene expression and cosmeceutical research A recent trend in cosmeceutical research is investigate the effects of treatments on gene expression in the skin Studies are done either in-vitro with explants from surgery or organotypic skin cultures such as those from MatTek or in-vivo followed by biopsies. Either gene chips such as Affymetrix or rt-PCR techniques are used investigate treatment effects on gene expression

The Affymetrix gene chip

Gene expression before and after UV exposure from microarray analysis Enk et. Al. Photodermatol Photoimmunol Photomed 2004; 20: 129–137

Verification of specific genes by rt-PCR

Challenges with genetic analysis and cosmeceutical research Dozens or even hundreds or perhaps thousands of genes may be changed over the course of a treatment Some will be up regulated some down regulated How do you decide what is a positive effect? Which genes are most important to a positive interpretation?

Summary While cosmeceutical is not a term with legal standing in the US we see that there are wide variety of approaches to treating skin that may be considered “cosmeceutical”. We have only discussed a few of the major categories today. Cosmeceuticals for anti-aging treatment can reduce wrinkles and age spots and may protect the skin from oxidative damage. Hot areas of research include peptides and the use of gene analysis to investigate cosmeceutical effects on the skin.