MAL-associated methyl nicotinate for topical PDT improvement

Highlights

• Methyl nicotinate for topical PDT improvement.

• Methyl nicotinate increases the depth of PpIX produced.

• Methyl nicotinate quantify the depth of PpIX produced

• Greater tissue damage by topical PDT using methyl nicotine.

Abstract

Photosensitization of all tissue in sufficient quantity to generate damage is one of the limiting factors for Photodynamic Therapy (PDT) efficiency. Methyl nicotinate (MN) is a thermogenic and vasodilating substance that facilitates the topical tissue penetration of some compounds. The topical MAL (methyl aminolevulinate) PDT is commonly used as a precursor of protoporphyrin IX (PpIX). This study investigates the safety of topical use in NM, as well as its ability to improve the efficiency of topical PDT. For this, we investigate the cytotoxicity of MN, as well as its actions in increasing cellular metabolism and vasodilation. Besides, its ability to optimize the formation of PpIX in the tissue when associated with MAL cream was investigated, besides assessing the severity of necrosis obtained by treatments. The cytotoxicity of MN was tested for concentrations of 0, 0.1, 0.25, 0.5, 0.75 and 1% in cell culture. For the concentration of 0.5%, the cellular metabolism was evaluated using confocal microscopy to calculate the redox rate. In the Chorioallantoic Membrane Model, vasodilation was evaluated for concentrations of 0.5 and 1% MN during 1 h of incubation. In the animal model, the healthy skin of Wistar rat was used to evaluate the production of PpIX in the tissue and the degree of necrosis obtained by Photodynamic therapy when using NM associated with methyl aminolevulinate. It was observed the non-cytotoxicity in vitro of MN in the concentration used (0.5%) and its ability to increase cellular metabolism. In a chorioallantoic model, the MN vasodilation power was demonstrated for different caliber of vessels. In vivo studies are showing that the incorporation of MN in the MAL cream increases the amount of PpIX produced in the tissue causing a higher effect on the epidermis after PDT. This improvement of the protocol may make the procedure more effective both in the destruction of tumor tissue and in the treatment of deeper cells decreasing possible recurrence, in addition to allowing improvements in the protocol, such as reducing the cream's incubation time.

Introduction

Photodynamic Therapy (PDT) is a therapeutic alternative for the treatment of malignant and potentially malignant lesions. In PDT, the interaction among photosensitizer (PS), light at a wavelength suitable for absorption and excitation of the PS, and molecular oxygen present in the tissue results in reactive oxygen species capable of causing death in target cells. [1,2]

Aminolevulinic acid (ALA) and methyl aminolevulinate (MAL) are precursors of protoporphyrin IX (PpIX), an endogenous photosensitizer present in small amounts in cells. After the precursor penetrates the cell, it interacts in the biosynthesis of the heme group and stimulates a greater formation of PpIX that preferentially accumulates in altered tissues. [3]

One of the limiting factors of PDT is the insufficient accumulation of PS in the entire tumor volume for the procedure to be efficient. In the topical treatment, the permeation of the PS or precursor (as in the case of the use of ALA or MAL) should be sufficient for the entire depth of the tissue to be photosensitized. [4,5] In the case of using ALA or MAL, the time required for the production of PpIX is also a challenge for treatment protocols. Long incubation times for the cream become unfeasible to ambulatory dynamics, usually 3 h in protocols to treat basal cell carcinoma, whereas short waiting times leads to an insufficient amount of PpIX to fully treat the tissue. [6,7] It is known that the depth of the PpIX is up to 2 mm with 3 h of incubation, however, the heterogeneities of the tissue causing a non-uniform distribution of the produced PpIX, with portions of tissue absent photosensitizer harming the treatment. [8,9]

In a study conducted by Stringasci et al. the thermogenic agents/vasodilators menthol, ginger, and methyl nicotinate (MN) incorporated into ALA and MAL creams were tested. The results showed that the incorporation of MN to the MAL cream improved the production of PpIX in the tissue. [10]

The use of a thermogenic and vasodilating substance such as MN can facilitate the skin permeation of the precursor, improving its penetration, and make it possible to obtain PpIX production in greater tissue depth. [11,12] In addition, the increase in metabolism associated with MN could accelerate and optimize the production process of PpIX. [5,11] Also, the substance's vasodilating action would increase the oxygen supply of the skin, again optimizing the effect of PDT. [13].

MN has been used in many percutaneous penetration studies, from in vitro to clinical studies. [14] However, there are no studies that investigate the MN ability to optimize topical PDT using topical MAL. The association of MN with MAL cream has the potential to facilitate the penetration of the cream and consequently increase the PpIX produced in the tissue, allowing improvements in the treatment protocols. This improvement of the production of PpIX would require shorter cream incubation times, in addition to allowing the treatment of thicker lesions and increasing the effectiveness of treatments.

Besides, an important aspect of topical PDT is the stinging pain reported by patients during illumination, pain so severe that many patients do not complete treatment or do not accept a new PDT treatment. [15,16] Studies have reported a possible analgesic effect of MN, thus, in addition to optimizing the treatment result, MN also has the potential to minimize the pain of patients during illumination. [17]

This study investigates the safety of topical use in NM, as well as its ability to improve the efficiency of topical PDT. So, we will investigate the cytotoxicity of MN, as well as its actions in increasing cellular metabolism and vasodilation. When associated with MAL cream, its ability to optimize the formation of PpIX in the tissue will be investigated: in quantity, depth, and time of production, in addition to assessing the severity of necrosis obtained by treatments.