A Japanese study has found that the Helicobacter pylori oncoprotein, CagA (cytotoxin-associated gene A), elicited transient 'BRCAness', inducing genomic instability via DNA double-strand breaks (DSBs) and defective homologous recombination (HR). The effects may underlie the gastric carcinogenesis associated with chronic H. pylori infection.

Reported in the May 13, 2021, online edition of Cell Host & Microbe, the study also demonstrated that artificial induction of BRCAness and inactivation of Hippo signaling, which regulates cell proliferation and apoptosis, may improve gastric cancer (GC) treatment with DNA-damaging agents.

Chronic H. pylori infection, especially with cagA-positive strains, plays a key role in development of GC, the third leading cause of global cancer mortality.

Histologically, GC is classified into intestinal and diffuse types corresponding to chromosomal instability (CIN) and genomically stable (GS) types, respectively.

Intestinal-type GC predominates in GC high-risk areas, notably East Asia, and is closely associated with chronic H. pylori infection, whereas diffuse-type GC is common in low-risk areas.

The CagA protein is located in the H. pylori genome's cag pathogenicity island (cag PAI), which is contained by almost all East Asian H. pylori isolates, while approximately 40% of Western isolates are cag PAI-negative, and thus do not produce CagA.

Transgenic expression in animal models has identified various CagA molecular activities that seem to contribute to gastric carcinogenesis.

"Once established, GC is not curable by H. pylori eradication, suggesting CagA-mediated carcinogenesis occurs through a 'hit-and-run' mechanism, whereby the etiologic agent of a given cancer is no longer required or present in the established cancer," study leader Masanori Hatakeyama told BioWorld Science.

"Early GC could therefore be cured by H. pylori eradication with antibiotics, if the cancer cells are still dependent on the bacterium for their survival and expansion," said Hatakeyama, a professor in the Department of Microbiology at the University of Tokyo's Graduate School of Medicine.

Such hit-and-run factors probably cause cancer by inducing genomic instability, facilitating accumulation of mutations inducing carcinogenesis.

In vitro infection of gastric epithelial cells with H. pylori generates DNA DSBs induced by reactive oxygen/nitrogen species (ROS/RNS) production and upregulation of a DNA repair-related long noncoding RNA.

CagA potentiates DSB formation, with one study reporting that CagA-mediated partitioning-defective 1b (PAR1b) kinase inhibition generates DSBs in H. pylori-infected human gastric epithelial cells via unknown mechanisms.

However, in the new study, CagA-induced PAR1b kinase inhibition was found to prevent PAR1b-mediated phosphorylation of the BRCA1 breast/ovarian tumor suppressor on Ser-616 that is required for cytoplasmic-to-nuclear translocalization of BRCA1.

"CagA strongly inhibits PAR1b's kinase activity by directly binding to its kinase catalytic domain," explained Hatakeyama.

"BRCA1 shuttles between the cell cytoplasm and nucleus, with Par1b-mediated phosphorylation of BRCA1 in cytoplasmic BRCA1 being essential for nuclear delivery of BRCA1."

Accordingly, "CagA-mediated inhibition of PAR1b kinase activity and the subsequent failure of BRCA1 phosphorylation by PAR1b prevents nuclear translocalization of BRCA1."

The ensuing shortage of nuclear BRCA1 was then shown to elicit BRCAness, causing genome instability through error-prone DSB repair.

"BRCA1 protects against genomic DNA damage in the nucleus, but not in the cytoplasm," noted Hakeyama. "Thus, loss of nuclear BRCA1 in CagA-expressing cells elicits BRCAness, leading to loss of HR-mediated error-free DSB repair and genomic instability."

CagA/PAR1b interaction was then shown to stimulate Hippo signaling, circumventing the apoptosis of DNA-damaged cells, giving cells time to repair DSBs through error-prone mechanisms.

"Under physiological conditions, cytoplasmic PAR1b inhibits Hippo mammalian STE20-like (MST) kinase via inhibitory phosphorylation of MST through complex formation," said Hatakeyama.

"Notably, in CagA-delivered cells, CagA sequestrates PAR1b to the membrane periphery, where it inhibits PAR1b kinase activity, with CagA-PAR1b complex formation competitively inhibiting CagA-MST complex formation."

"Consequently, MST is released from constraints imposed by PAR1b, enabling activation of Hippo signaling," he said.

"We suspect that inhibition of the Hippo signaling in GC cells might improve the cytotoxic effect of DNA-damaging agents such as cisplatin and mitomycin, as Hippo signal-mediated nuclear delivery of YAP [yes-associated protein] promotes complex formation of YAP with proapoptotic nuclear protein p73, a member of the [tumor protein] p53 family."

Importantly, DSB-activated p53 was demonstrated to inhibit CagA-delivered cell proliferation, which could be overcome by p53 inactivation.

"We consider functional p53 to be a game changer in H. pylori-associated gastric diseases, as CagA-delivered cells with DSBs are enforced to cease proliferation and undergo premature cell senescence, followed by cell death, which may induce gastritis or ulceration," said Hatakeyama.

"Upon functional p53 inactivation in gastric epithelial cells, which may be potentiated by H. pylori-induced activation-induced cytidine deaminase (AID) expression, CagA-delivered cells exhibiting BRCAness-induced genome instability can survive despite severe DNA damage, prompting accumulation of mutated precancerous cells, from which CagA-independent GC cells may eventually arise."

Together, these findings show that expansion of CagA-delivered cells with BRCAness-mediated genome instability, from which CagA-independent cancer-predisposing cells arise, provides a credible 'hit-and-run' mechanism of H. pylori CagA for gastric carcinogenesis.

Treatment implications

Mechanistically, "the hit-and-run gastric carcinogenesis induced by H. pylori CagA supports the importance of early eradication of H. pylori in preventing GC," said Hatakeyama, "but also indicates the presence of a gene(s), the mutation of which makes cells independent of H. pylori eradication in GC development.

"Considering the presence of a common mechanism, namely BRCAness, in the development of GC, as well as in hereditary breast and ovarian cancers (HBOCs) and in some pancreatic and prostate cancers, it may be possible to develop new therapeutics targeting BRCAness, in addition to poly [ADP ribose] polymerase (PARP) inhibitors."