Effect of a botulinum neurotoxin from an autochthonous strain of C.botulinum on breast cancer cells

CHAPANA A; GUARNIOLO D; SOSA PAREDES E; RA FERNANDEZ; SOSA MA; CARVELLI L; PATRICIA CABALLERO

Congreso; XXXVII Reunión Anual de la Sociedad de Biología de Cuyo; 2021

Botulism is a neuroparalytic disease caused by botulinum neurotoxins (BoNT, serotypes A-G) produced by Clostridium botulinum. BoNT A is currently widely used in the treatment of neuromuscular transmission disorders. Genomic and phenotypic studies showed that native strains from Cuyo region (Argentina) and their BoNTs correspond to subtype A2, and would have higher activity than A1 (Botox®), being a potential therapeutic agent. The native BoNTSu 1935obtained from soil is known to degrade actin from rat brain homogenates, suggesting that this protein would be a target for this toxin. In this study, the action of BoNTSu 1935 on the actin and tubulin cytoskeleton was evaluated in mammary tumor cells. Both, the BoNTSu 1935 and the BoNT from the A Hall strain (subtype A1) were purified by 60% saline precipitation. MCF7 and MDA-MB-231 cells were treated with 250 LD50 of the BoNTs for 5, 10, 25, 45 and 90 min. The cells were then processed for immunoblot or immunofluorescence in order to evaluate the expression and distribution of actin and tubulin. In addition, the integrity of the Golgi complex was evaluated. In MCF-7 cells, BoNTSu 1935 induced actin degradation at higher extent than BoNT A Hall. In turn, this protein was redistributed to plasma membrane location, in a time-dependent manner. Although tubulin was affect in a similar way, this protein was redistributed to perinulear patches. Meanwhile, the Golgi complex appears disorganized after treatment with the toxin. Interestingly, no changes were observed in either the distribution, or in the expression of actin and tubulin in MDA-MB-231 cells. Regarding the cytotoxicity of BoNTSu 1935, we observed ~90% mortality of MCF-7 cells at 90 min of incubation with the toxin, while that index was reached in the MDA-MB-231 cells, as early as 25 min incubation. In contrast, BoNT A Hall only induced 50% cytotoxicity in MDA-MB-231 cells by 90 min incubation. These results would indicate a higher cytotoxicity of BoNTSu 1935, whose action mechanisms would be selective for tumour cell types. This could be related to levels of malignancy of the tumor cells.