Skin exposure to ultraviolet radiation (UVR) promotes photoaging and is the main risk factor for the development of actinic keratoses (AK).1,2 The UVA spectrum promotes the formation of reactive oxygen species (ROS), leading to oxidative stress and damage to keratinocyte DNA.2–4 The UVB spectrum is responsible for the formation of pyrimidine dimers in the keratinocyte DNA, the main ones being cyclobutane pyrimidine (CBP) and pyrimidine-pyrimidone photoproducts (6-4PP).3

AK diagnosis and treatment is important because it presents a risk of progression to squamous cell carcinoma (SCC).5,6 Generally, multiple AKs occur in the same area of photodamaged skin, which accumulates changes in the keratinocyte genetic material, which defines cutaneous field cancerisation (CFC).7

Recently, photolyase, a monomeric enzyme not found in the cells of placental mammals, has emerged as an alternative for the treatment of AK and CFC.8–10 The enzyme has the ability to break down the dimers of CBP and 6-4PP by means of visible light, a process called photoreactivation.11 Puig et al. published a review article that concluded that there is evidence to support the beneficial effects in the treatment of AK and CFC with topical photolyase.2

Considering the indirect damage caused by UVR, through the formation of ROS, studies with antioxidant agents (AOx) have started. Vitamin C (ascorbic acid) is the most prevalent water-soluble AOx in the epidermis.12 It neutralizes free radicals and ROS in the aqueous compartments of the skin and plays an important role in the regeneration of vitamin E (tocopherol).13 Vitamin E is a fat-soluble AOx present in the horny layer, produced from the sebaceous glands, characterized by the ability to protect the cell membrane from oxidative stress.12 Murray et al. demonstrated that the combination of three AOx (L-ascorbic acid 15%, alpha-tocopherol 1%, and ferulic acid 0.5%), when compared to placebo (vehicle), showed a photoprotective effect by reducing the erythema induced by UVR, significantly reducing the formation of ‘sunburn cells’, as well as reducing the expression of p53 and the formation of thymine dimers.14

Most studies using antioxidants formulas focus on its photoprotective properties. Although, no studies have evaluated its efficacy in the treatment of AK. Also, the combination of antioxidants and photolyase has not been described in the literature up to now.

The aim of this study was to evaluate the efficacy of using sunscreen with photolyase compared to regular sunscreen as well as to compare the combination of a topical formulation of antioxidants versus placebo in the treatment of advanced photodamage, characterized by the presence of AK and CFC of the forearms, after an interval of 8 weeks.

Between January 2019 and January 2020, 40 participants were eligible and randomized for the study. Twenty forearms were part of the SC+P/AOx group and the SC+P/placebo group. Twenty forearms were part of the SC/AOx and SC/placebo groups. There were two dropouts in the SC+P group, not associated with the interventions, as patients did not return for re-evaluation on the scheduled date (Fig. 1).

The main clinical and demographic data of the 40 participants at baseline (T0) are displayed in Table 1. There was no significant difference between groups for any of the variables.

There was no significant difference in primary and secondary outcomes between the regular sunscreen and sunscreen containing photolyase groups (Table 2). Total clearance was achieved by three forearms treated with topical antioxidant (7.9%; 95% CI 2.6–13.2%) and one forearm treated with placebo (2.6%; 95% CI 0–7.9%), without a difference (p = 0.997). Partial clearance was achieved by 18 forearms treated with topical antioxidant (47.4%; 95% CI 34.2–60.5%) and nine forearms treated with placebo (23.7%; 95% CI 13.2–34.2%; p = 0.018).

There was a reduction in the AK count in all groups over time (p < 0.05). In addition, there was a reduction in the AK count in the groups that used topical antioxidants (SC/AOx and SC+P/AOx) in relation to those who used a placebo (p < 0.05) (Table 2 and Fig. 2).

Figure 2.

AK count in the four treatments groups at T0 and T60.

AK, actinic keratoses; SC, regular sunscreen SPF 99; SC+P, sunscreen containing photolyase SPF 99; AOx, cream composed of antioxidant agents.

(*p < 0.05).

(0.2MB).

The FPS decreased for all groups over time (p < 0.05), regardless of the treatment group (Table 2). There was no statistically significant difference between the other interventions for this outcome.

Likewise, there was a reduction in the AKSS for all groups over time (p < 0.05), without superiority for any treatment (Table 2).

The effect sizes for the outcomes are shown in Table 3. The difference in the AKSS means and partial clearance was higher in the groups that used sunscreen with photolyase and topical antioxidant (SC+P/AOx) compared with the groups that used the placebo.

The authors of the present study found no difference in the treatment of advanced photodamage and CFC when comparing the use of sunscreen with photolyase and regular sunscreen without photolyase. However, the use of an antioxidant formula gave a reduction in the AK count, which reflects a reduction in CFC activity. Although topical antioxidant is frequently used as an anti-aging product, the present study showed improvement in AK count without a significant change in other aspects of the forearm photoaging scale.

When interventions were analyzed over time, all parameters showed statistically significant improvements. The reduction or resolution of AKs can occur in up to 21% of patients within one year.18 In the present study’s clinical trial, all participants used at least regular sunscreen, with a reduction or resolution of the AKs in the forearms of 38.7%. In addition, two articles found that regular use of sunscreen reduced the AK count by up to 25% in immunocompetent patients and 50% in patients with transplanted solid organs.19,20

Puig et al. analyzed the evidence available in the previous ten years of using sunscreen containing photolyase. The authors analyzed 11 studies, totaling 228 participants, only three of which were RCTs, and concluded that there was evidence to support the use of photolyase for the treatment of AK and CFC.2

One of the clinical trials was that of Moscarela et al., who compared the use of sunscreen with photolyase and regular sunscreen SPF 50 in patients with four or more AKs on the face and scalp. The study included 50 participants but had 14 dropouts, resulting in 17 participants who used sunscreen with photolyase and 19 regular sunscreen SPF 50. After six months of follow-up, there was a reduction in the AK count in both groups, which did not reach statistical significance. When evaluating the subgroup of participants with up to ten AKs (n = 20), the sunscreen group with photolyase proved to be superior in reducing the AK count. In this subgroup, 14% of participants who used sunscreen with photolyase had new lesions versus 54% in those who used regular sunscreen SPF 50.21 In the present study, only one patient in the sunscreen with photolyase group presented with new AKs versus none in the regular sunscreen group.

Eibenschutz et al. conducted a clinical trial with 30 participants to evaluate the effectiveness of sunscreen with photolyase versus regular sunscreen in the treatment of AK and CFC of the face and scalp after one session of MAL-PDT. An increase in the number of AKs was observed after nine months of treatment in the participants who used regular sunscreen, while there was a reduction in the AKs in those who used sunscreen with photolyase (p = 0.001).22

Puig et al. developed a prospective, controlled, non-randomized study with 13 participants with AKs in exposed areas to evaluate the effectiveness of sunscreen with photolyase versus regular sunscreen for 4 weeks in the treatment of AKs and CFC. The groups were formed by nine participants in the sunscreen with photolyase and three in the regular sunscreen. The study analyzed the following outcomes: clinical improvement, dermatoscopy, immunohistochemistry, and appearance on confocal microscopy. There was a statistically significant improvement in all the outcomes studied in the group that used sunscreen with photolyase, but there was no improvement in the group that used regular sunscreen. However, this was a non-randomized study with a low number of participants, and the AK count was not analyzed as an outcome, in addition to presenting a disproportionate distribution between the groups, which compromises the data validity.11

The RCT did not show superior efficacy of sunscreen containing photolyase in the treatment of AKs of the forearms of immunocompetent patients, which contradicts the results of previous studies.

The use of antioxidant substances associated with sunscreen has become frequent since several studies have demonstrated a photoprotective effect by reducing erythema induced by UVR, reducing the expression of p53, reducing the formation of ‘sunburn cells’, and decreasing the expression of immunosuppressive cytokines.14

Vitamin C neutralizes free radicals in the aqueous compartments of the skin, in addition to playing an important role in the regeneration of vitamin E.12,13 The main function of vitamin E is to protect cell membranes from oxidative stress.12

The association of 15% L-ascorbic acid with 1% alpha-tocopherol demonstrated that they have a synergistic effect on the skin. When alpha-tocopherol neutralizes oxidative stress in lipids, its oxidized products can be regenerated by L-ascorbic acid. This interaction helps to renew antioxidant protection in tissues. The formulation containing the two products doubled the UV protection for the skin when compared to L-ascorbic acid alone.23,24 Lin et al. demonstrated that the addition of 0.5% ferulic acid increased both the stability of the formula and doubled the UV protection for the skin. It is postulated that ferulic acid protects L-ascorbic acid and alpha-tocopherol, undergoing oxidation before the others in the formula.25

There have been no studies evaluating the use of topical antioxidants for the treatment of AK and CFC. The present study demonstrated a beneficial and safe effect of the combination of 15% L-ascorbic acid, 1% alpha-tocopherol, and 0.5% ferulic acid in reducing the number of AKs, consequently controlling CFC activity.

One of the limitations of this study was that it did not directly compare the regular sunscreen and sunscreen with photolyase groups, making this comparison inferior to the topical antioxidant versus placebo comparison. In addition, the study included only patients with AK counts between three and ten, and the results may not apply to those patients with greater numbers of lesions as well as immunosuppressed patients.

The authors acknowledge that the 60-day follow-up interval might be related to the poor results of photolyase. Perhaps to achieve better results, its use should be prolonged. However, further studies must be carried out to confirm this hypothesis. In addition, as demonstrated by Eibenschutz et al.,22 it is possible that photolyase has the best indication for preventing the recurrence of AK treated by other methods, being an alternative to stabilize but not treat CFC.

The authors also believe that prolonged use of topical antioxidants might improve other signs of photoaging, since studies showed that reduction in the transcription of matrix metalloproteinase-1 inhibits the formation of thymine dimers, reducing collagen breakdown process and carcinogenesis.26

The strength of the present study was the possibility of evaluating the effects of three interventions in a factorial study design in a group of immunocompetent patients who frequently present AK in their forearms.

Sunscreen (SPF 99) containing photolyase was not superior to regular sunscreen (SPF 99) in terms of the studied outcomes after eight weeks.

The use of the topical antioxidant formula is safe, tolerable, and also had more efficacy in reducing the AK count in the forearms when associated with regular sunscreen or sunscreen with photolyase.

This study received financial support from the Sao Paulo State Dermatology Support Fund (FUNADERSP), no. 73-2018.

Bruno Augusto Alvares: Conception and design of the study; survey of data, or analysis and interpretation of data; statistical analysis; writing of the article or critical review of important intellectual content; obtaining, analysis and data interpretation; effective participation in research guidance; intellectual participation in propaedeutic and/or therapeutic conduct of studied cases; critical literature review; final approval of the final version of the manuscript.

Anna Carolina Miolla: Writing of the article or critical review of important intellectual content; obtaining, analyzing and interpreting the data; effective participation in research guidance; intellectual participation in propaedeutic and/or therapeutic conduct of studied cases.

Juliano Vilaverde Schimitt: Conception and design of the study; statistical analysis; writing of the article or critical review of important intellectual content; effective participation in research guidance; intellectual participation in propaedeutic and/or therapeutic conduct of studied cases; critical literature review.

Helio Amante Miot: Conception and design of the study; survey of data, or analysis and interpretation of data; statistical analysis; writing of the article or critical review of important intellectual content; obtaining, analysis and data interpretation; effective participation in research guidance; intellectual participation in propaedeutic and/or therapeutic conduct of studied cases; critical literature review; final approval of the final version of the manuscript.

Luciana Patricia Fernandes Abbade: Conception and design of the study; survey of data, or analysis and interpretation of data; statistical analysis; writing of the article or critical review of important intellectual content; obtaining, analysis and data interpretation; effective participation in research guidance; intellectual participation in propaedeutic and/or therapeutic conduct of studied cases; critical literature review; final approval of the final version of the manuscript.

None declared.