Abstract A63: Combination of nonlethal photosensitization with a new imidazacridine derivate reduces UACC-62 cell viability
Authors: Santos Renata V. C. , Barbosa Artur F. S. , Chagas Mardonny B. O. , Lima Maria C. A. , Pereira Michelly C. , Pitta Ivan R. , Pitta Maira G. R. , Rego Moacyr J. B. M. 
Published: May 2019

Abstract
Melanoma is the skin cancer with the lowest incidence, but the highest aggressiveness and mortality rate. It is known that the actual common resistance to chemotherapy in clinical settings reduces the efficiency of treatments, making urgent the search for new molecules and approaches. The 5-aminolevulinic acid-mediated nonlethal photosensitization (ALA-PDT) is currently one of the most promising forms of cancer treatment, because of its benefits such as noninvasiveness and minimum side effects, usually associated with a transient sensitivity when exposed to the light. In the same context, previous work of our group has shown that the acridine nucleus, present in the imidazacridines compounds, has an essential antiproliferative role and inhibits topoisomerases I and II. Based on that, this in vitro study proposed to evaluate the antineoplastic activity of AC05 (C7H11N3OS), an imidazacridine derivate, in combination with ALA-PDT, against UACC-62, SKMEL28, and A375 melanoma lineages. The cytotoxicity of AC05 and ALA-PDT was analyzed in peripheral blood mononuclear cells (PBMC) from healthy volunteers through an MTT assay, aiming to evaluate the selectivity of therapeutic strategies. Then, using the same method, 1x104 cells of each melanoma lineage were plated in 96-well dark plates and maintained in complete medium, at 37°C in a humidified environment with 5% CO2. After 24 hours, each condition was incubated in triplicate as follow: (a) cells; (b) vehicle control (0.1% dimethyl sulfoxide—DMSO); (c) 1µM doxorubicin (a standard drug commonly used at clinical treatment); (d) 1, 25 and 50µM AC05 dissolved in DMSO and complete medium; (e) 250, 1000 and 2000 µM ALA dissolved in complete medium; (f) laser irradiation - 1, 2 and 3 J/cm2; (g) 250µM ALA + 2J/cm2 laser; (h) 1, 25 and 50 µM AC05 + 2J/cm2 laser; (i) 1, 25 and 50 µM AC05 + 250µM ALA + 2J/cm2 laser. Then, 72 hours later, the cells were incubated with 20 µl of 0.5 mg/ml MTT 3-(4,5-dimethylthiazol-2)-2,5-diphenyl tetrazolium bromide salt by well, during 3 hours, when the reaction was stopped by the incubation of 130 µl of 20% sodium dodecyl sulfate by well. The cell survival was determined after 24 hours, through absorbance measurement at 570 nm by a multiple reader. Results were calculated based on three independent experiments from each lineage. AC05 compound and ALA-PDT did not modify the viability (≤50%) of the PBMC (IC50AC05 ≥ 100 µM and ALA-PDT ≥ 2000 µM). Concerning the melanoma lineages, the SKMEL-28 was the most resistant lineage, since even the approaches in combination (i) were not capable of reduce the viability to less than 50%, except the 2000 µM ALA-PDT (e) that was effective alone. In the A375 melanoma, the single approaches (d and e) did not change the cellular survival ((d)IC50AC05 ≥ 50 µM and (e) ALA-PDT ≥ 2000 µM), although together (i) they reduced this parameter to less than 50% of cell viability. Regarding the UACC-62, either the imidazacridine derivate (d) or the ALA-PDT (e), separately, reduced the cellular viability ((d)IC50AC05 = 28.29 ± 0.6 µM and (e)IC50ALA = 1669,. ± 93.57 µM). However, when in combination they demonstrated a 31% of decreasing in the IC50AC05 capable to reduce the cell viability to less than 50%, when compared to their single effects ((i) IC50AC05 = 20.41 ± 4.37 µM and ALA-PDT = 250 µM). As expected, the experimental conditions with the lowest concentration of ALA (g) or the three concentrations of AC05 (h) combined with the photosensitization have not been able to reduce the cell viability to less than 50%. Altogether, the results showed that the association of the chemotherapy with the photosensitization enhanced the effectiveness of the treatment with AC05. Beyond, the work suggests a new therapeutic potential approach to be explored against UACC-62 and A375 melanoma lineages.
Citation Format: Renata V. C. Santos, Artur F. S. Barbosa, Mardonny B. O. Chagas, Maria C. A. Lima, Michelly C. Pereira, Ivan R. Pitta, Maira G. R. Pitta, Moacyr J. B. M. Rego. Combination of nonlethal photosensitization with a new imidazacridine derivate reduces UACC-62 cell viability [abstract]. In: Proceedings of the AACR International Conference held in cooperation with the Latin American Cooperative Oncology Group (LACOG) on Translational Cancer Medicine; May 4-6, 2017; São Paulo, Brazil. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(1_Suppl):Abstract nr A63.