# New Insights into the Bioenergetic and Immunomodulatory Properties of Phospholipases A2 from Bothrops diporus Venom

## TL;DR

This study explores how snake venom phospholipases affect cancer cell energy and immune responses, revealing new therapeutic potential.

## Contribution

The first integrated analysis of Bothrops diporus PLA2s' effects on mitochondrial function and immune modulation.

## Key findings

- B. diporus PLA2s induce mitochondrial stress in breast cancer cells, reducing proliferation.
- PLA2s trigger a dynamic cytokine response in human immune cells, including early cytotoxic and late regulatory mediators.
- A specific PLA2 isoform uniquely activates IL-10 and CCL20, highlighting isoform-specific immunomodulatory effects.

## Abstract

Phospholipases A2 (PLA2s) are key mediators of the cytotoxic and inflammatory activities of snake venoms. While PLA2 isoforms from Bothrops diporus venom have been characterized and shown to possess antimetastatic and antiangiogenic properties, their impact on mitochondrial bioenergetics and immune modulation has not yet been investigated. In this study, we examined the bioenergetic and immunomodulatory effects of B. diporus PLA2s using integrated biochemical, metabolic, and multiplex cytokine analyses. In MDA-MB-231 breast cancer cells, pooled PLA2s induced a dose-dependent decrease in MTT-reducing activity, increased mitochondrial ROS, caused Δψm hyperpolarization, and decreased NADH autofluorescence, collectively indicating sustained mitochondrial stress. Real-time impedance measurements further revealed a marked inhibition of cell proliferation. In human PBMCs, pooled PLA2s elicited a dynamic cytokine program, inducing early cytotoxic (Granzyme B) and chemotactic (CCL2, CCL3, CCL4) mediators, followed by late pro-inflammatory and regulatory factors such as IL-6, TNF-β, IL-10 and IL-15. Analysis of a single purified PLA2 isoform (Fraction 6) confirmed activation of the canonical IL-6/TNF-α/IL-1β axis but uniquely induced IL-10 and CCL20, revealing isoform-specific immunomodulatory properties. Altogether, these findings provide the first integrated characterization of mitochondrial and immune perturbations induced by B. diporus PLA2s, expanding their recognized biological scope and underscoring their potential as molecular templates for novel pharmacological strategies targeting mitochondrial vulnerabilities or modulating the tumor immune microenvironment.