Targeting DNA repair by coDbait enhances melanoma targeted radionuclide therapy

Abstract

Radiolabelled melanin ligands offer an interesting strategy for the treatment of disseminated pigmented melanoma. One of these molecules, ICF01012 labelled with iodine 131, induced a significant slowing of melanoma growth. Here, we have explored the combination of [131I]ICF01012 with coDbait, a DNA repair inhibitor, to overcome melanoma radioresistance and increase targeted radionuclide therapy (TRT) efficacy. In human SK-Mel 3 melanoma xenograft, the addition of coDbait had a synergistic effect on tumor growth and median survival. The anti-tumor effect was additive in murine syngeneic B16Bl6 model whereas coDbait combination with [131I]ICF01012 did not increase TRT side effects in secondary pigmented tissues (e.g. hair follicles, eyes). Our results confirm that DNA lesions induced by TRT were not enhanced with coDbait association but, the presence of micronuclei and cell cycle blockade in tumor shows that coDbait acts by interrupting or delaying DNA repair. In this study, we demonstrate for the first time, the usefulness of DNA repair traps in the context of targeted radionuclide therapy.

Keywords: DNA repair; coDbait; melanoma; targeted radionuclide therapy.

Figure 1. Effect of coDbait added to [¹³¹I]ICF01012 TRT on tumor growth and median survival in B16Bl6 model (n = 20/group)

(A) The tumor growth was calculated with the ratio of final volume (FV) minus initial volume (IV) to initial volume (IV). The doubling time (DT) was calculated with the exponential value of tumor volume curves. (B) Survival curves were established as the percentage of animals that remained in each group (n = 20/group) at indicated times (C) The number of the cells presenting micronuclei in the cytoplasm (black arrows) in non-necrotic and proliferative areas was increased in [¹³¹I]ICF01012 TRT group compared to control and coDbait, this was further enhanced by coDbait addition (n = 4 /group; 300000 cells counted). *p < 0.05.

Figure 2. Side effects associated with treatments in pigmented C57Bl6 mice

(A) Evaluation of [¹³¹I]ICF01012 + coDbait toxicity on mice retina 10 days post irradiation (n = 4 per group). The significant ratio decrease observed following [¹³¹I]ICF01012 around the optic nerve was not further increased with coDbait addition. (B) White blood cell quantification 10 days post irradiation showed a non-significant decrease in irradiated mice. (C) % variation from initial values showed no statistically significant weight loss in the four groups. (D) Evaluation of [¹³¹I]ICF01012 radiotoxicity on skin by melanocyte-specific staining of PS100 protein. Representative histological sections of mice skins from the different groups showed a similar number of melanocytes in the hair follicles. (E) Quantification of PS-100 stained cells showed no statistically variation of the hair follicle melanocytes. *p < 0.05.

Figure 3. Effect of coDbait ± [¹³¹I]ICF01012 TRT in SK-Mel 3 model

(A) The tumor growth was calculated with the ratio of final volume (FV) minus initial volume (IV) to initial volume (IV). The doubling time (DT) was calculated with the exponential value of tumor growth with six mice in each group. (B) Percentage survival of mice (n = 6 /group) receiving the different treatments. (C) HES representative histological sections and of SK-Mel 3 sampled 24 h after [¹³¹I]ICF01012 injection (25 MBq) and/or one injection of coDbait (1 mg). (D) The percentage of necrosis established by counting the cells on 3 HPF. *p < 0.05.

Figure 4. Mechanistic studies of [¹³¹I]ICF01012 ± coDbait on tumor extracts

Western blot analysis of p53 phosphorylation (S15), ATM phosphorylation (S1981), Chk2 phosphorylation (T68) p21 expression and PARP cleavage in tumors 24 h p.i. in B16Bl6 (n = 3 tumors per group) (A) and SK-Mel 3 models (n = 3 tumors per group) (B). Cell cycle modifications induced 24 h after treatment in B16Bl6 (C) and SK-Mel 3 models (D).*p < 0.05.