# Chemical-Class Submixture Screening Reveals Drivers of Endocrine Disruption in Personalized Human Blood POP Mixtures

## TL;DR

This study identifies specific chemical classes in blood mixtures that drive endocrine disruption effects in human cells.

## Contribution

The study introduces a submixture screening approach to identify toxic drivers in complex chemical mixtures found in human blood.

## Key findings

- PFAS and OCP submixtures significantly affected testosterone synthesis at low and medium concentrations.
- Some submixtures altered estradiol and testosterone levels in ways not seen in full mixtures, indicating antagonistic interactions.
- High concentrations of PM#3 and PM#4 downregulated CYP11A1, suggesting a feedback mechanism suppressing testosterone synthesis.

## Abstract

Multiple studies
demonstrate mixture effects arising
from the interactive
toxicity of environmental chemicals in human blood, but identifying
the main toxic drivers remains challenging. In a recent proof-of-principle in vitro study, we showed that personalized mixtures (PMs),
reconstructed from 24 persistent organic pollutant (POPs) concentrations
measured in individual blood samples from Swedish adults, induced
unique interindividual effects on H295R cell viability and steroidogenesis.
Here, we followed up by testing submixtures of four PMs (PM#3, PM#4,
PC1-OC-Mix, and PC2-PFAS-Mix), separated by the chemical classes perfluoroalkyl
substances (PFASs), organochlorine pesticides (OCPs), polychlorinated
biphenyls (PCBs), and polybrominated diphenyl ethers (PBDEs). Submixtures
of PFAS and OCPs induced significant effects on testosterone synthesis
at low (1×) and medium (10×) concentrations, consistent
with effects observed in the corresponding whole PMs, and were therefore
likely the primary drivers of the whole-mixture effects on testosterone.
Notably, some submixtures altered estradiol and testosterone levels
in ways not observed in full PMs, suggesting antagonistic interactions
across chemical classes when combined. Potential antagonistic interaction
in more complex mixtures, independent of concentration, was also observed
within OCP submixtures, as only the less complex OCP mixtures lacking
DDE or DDT induced testosterone synthesis. For additional mechanistic
insight, we expanded the H295R assay to include oxidative stress analyses,
which revealed no effects from the PMs. RT-qPCR analysis showed downregulation
of CYP11A1 after exposure to PM#3 and PM#4 at high
concentrations (100×), suggesting a feedback mechanism contributing
to suppressed testosterone synthesis.