Researchers led by Emanuel Hanski at the Hebrew University of Jerusalem have identified a class of molecules that inhibit endoplasmic reticulum (ER) stress and could decrease mortality in mice infected with Streptococcus pyogenes, also known as the group A streptococci (GAS).

The treatment was effective even when given as late as 12 hours after infection, suggesting they could help alleviate invasive necrotizing fasciitis (NF) infections, the team reported in the August 4, 2021, issue of Science Translational Medicine.

GAS is an obligate human pathogen and the one of the most common bacterial causes of human mortality. GAS causes a vast array of human manifestations ranging from mild infections such as pharyngitis and impetigo to highly invasive and life-threatening infections such as NF and toxic shock.

NF is an infection of the deeper subcutaneous tissues and fascia that is characterized by extensive and rapidly spreading necrosis (gangrene) of the skin and underlying structure. NF kills roughly 30% of its victims and leaves the rest disfigured. Penicillin group antibiotics and surgical interventions, the standard treatments for NF, often fail, and hence there is a critical need for approaches to combat invasive soft tissue GAS infection.

Speaking to BioWorld Science, corresponding author Hanski said that "induction of endoplasmic reticulum stress and unfolded protein response (UPR) constitutes a pathogenic strategy of group A streptococcus. Our team had previously discovered that GAS induces unfolded protein response to upregulate asparagine synthetase transcription leading to production and release of asparagine. GAS senses the increased asparagine and alters gene expression profile accordingly to increase the rate of multiplication."

Hanski is a professor of microbiology and molecular genetics in the faculty of medicine at the Hebrew University, where he studies microbial toxins' mechanisms of action.

Different perturbations at the cellular level can induce accumulation of misfolded proteins within the ER lumen or change its lipid composition, leading to ER stress. To alleviate these conditions, the ER launches the UPR, allowing the cells to adapt to the environmental stresses and survive.

However, under prolonged stress conditions, when stresses remain unmitigated, UPR triggers programmed cell death.

Three ER transmembrane proteins sense ER stress: protein kinase RNA (PKR)-like ER kinase (PERK), activating transcription factor 6 (ATF6) and inositol-requiring protein 1 (IRE1). The branch of UPR involving PERK upregulates asparagine levels in host cells and helps GAS to infiltrate connective and soft tissue.

According to Hanski, "the goals of the present investigation were to understand the precise signaling pathway that GAS uses to obtain asparagine from the host and use pharmacological inhibitors of this pathway as treatments against invasive GAS infections."

Hanski's team analyzed human skin cells infected with GAS and found that asparagine release occurred through the PERK-eIF2a-ATF4 branch of the UPR. Inhibitors of PERK or the integrated stress response (ISR) blocked the formation and release of asparagine by infected mammalian cells in a dose- and time-dependent manner, and externally added asparagine overcame this inhibition.

Further, the experimental molecules ISRIB or GSK-2656157 – which inhibit the UPR directly or indirectly – lowered mortality rates and reduced bacterial counts in infected mice. The two inhibitors also accelerated wound healing and shrank bacterial lesions in mice with NF, even if given 12 hours after infection.

Neutrophils are an essential part of the host's primary innate immune defense against GAS infections. NF is characterized by spreading inflammation and subsequent necrosis of surrounding subcutaneous tissue that inhibits neutrophil infiltration at the site of infection. On days 2 and 4 after infection, GSK-2656157-treated mice displayed a substantially enhanced neutrophil infiltration into the fascial zone filled with GAS bacteria, "indicating the body's immune system mounting an inflammatory response to curb the spread of the bacteria," noted Hanski. Though the current study does not investigate the role of the immune system in GAS infection, Hanski said his team was planning to run a dual RNA sequencing analysis to understand how inflammatory molecules can affect GAS Invasion, and "hopefully strengthen those components that fight against the invasion." One of the major challenges faced by Hanski was to create an animal model of GAS, as GAS is a human specific pathogen.

Hanski believes that host-directed therapies are a new and promising approach for treating infectious diseases and can allow repurposing of drugs to improve patient outcomes in severe GAS infections. Hanski also added further work was needed to assess the validity of GSK-2656157 and ISRIB in human tissues, and his team's future studies would involve experimentation on abraded human skin tissue and NF lesions