Validación de biomarcadores de respuesta, prevención y evolución de queratosis actínicas tratadas con terapia fotodinámica con luz de día

Résumé

Actinic keratoses (AK) are the most frequent precursor lesions of cutaneous squamous cell carcinoma (SCC). Among its field treatments is Daylight Photodynamic Therapy (DLPDT), which employs natural light to activate a photosensitizer and trigger a photodynamic reaction to treat the cancerization field and reduce the risk of developing SCC. However, there are cases of poor response to DLPDT. The aim of this study is to analyze the clinical, histological, immunohistochemical and molecular response of patients with field cancerization and AK after treatment with DLPDT, in order to advance in the knowledge of the molecular mechanisms involved in the preventive effect of DLPDT and to find biomarkers of good or bad response. For this purpose, we selected patients with AK and performed skin biopsies before and three months after treatment with methyl aminolevulinate (MAL) DLPDT. Patients were divided according to their clinical response into "good responders", "intermediate responders" and "poor responders". Histological and immunohistochemical changes in the expression of proliferation and oncogenic proteins were studied. RNA arrays were also performed in selected cases to study the molecular response. Twenty-five patients were selected of whom 22 completed the study with a 6-month follow-up. The total number of AKs decreased from 515 before treatment to 315 at 3 months (38,8% reduction, p = 0,001) and 376 at 6 months (27% reduction, p = 0,013). The AKASI index decreased from 5.85 before treatment to 4.69 at 3 months (p = 0,002) and 4.9 at 6 months (p = 0,014). There were 7 good responders, 5 intermediate responders and 9 poor responders, and one case developed SCC during follow-up despite treatment with DLPDT. Poor responders developed four times more new lesions than good responders and showed twice as many treatment-resistant lesions. Histological response was determined by two dysplasia classifications for AK: KIN and Pro, the latter showing a better correlation with clinical response. The immunohistochemical response showed a decrease in Ki-67, p53 and cyclin D1 expression in good responders, while in poor responders an increase in p53 and cyclin D1 expression and a decrease in Ki67 expression were observed. Three representative cases of good responders and three representative cases of poor responders were selected for RNA array extraction. The expression of 32 genes related to SCC pathogenesis was analyzed. Of these, only the PIK3R1 gene (phosphatidylinositol 3-kinase alpha regulatory subunit) showed post-treatment under-expression in all good responders and overexpression in all poor responders. Other genes, such as CCND1 (cyclin D1), PDK1 (phosphatidyl-3-inositoldependent protein kinase 1) and MAPK3 (MAP Kinase 3) showed underexpression in all good responders and overexpression in 2 of the 3 poor responders. The expression of these genes was validated by immunohistochemistry. Additionally, the MYC oncogene was found to be overexpressed pre-treatment in poor responders, whereas the PTEN gene was found to be under-expressed. The findings of this study suggest the existence of a molecular basis that could determine a good or bad response to DLPDT, probably related to the phosphatidylinositol 3-kinase (PI3K/Akt) pathway, in which PIK3R1 as well as PDK1, MAPK3 and Cyclin D1 are involved. The development of molecular techniques to classify patients according to the genetic alterations present is key to offer targeted treatments and thus optimize good responses.