Fine-tuning Bacterial Cyclic di-AMP Production for Durable Antitumor Effects Through the Activation of the STING Pathway

The stimulator of interferon genes (STING) protein is an important and promising innate immune target for tumor therapy. However, the instability of the agonists of STING and their tendency to cause systemic immune activation is a hurdle. The STING activator, cyclic di-adenosine monophosphate (CDA), produced by the modified Escherichia coli Nissle 1917, shows high antitumor activity and effectively reduces the systemic effects of the off-target caused by the activation of the STING pathway. In this study, we used synthetic biological approaches to optimize the translation levels of the diadenylate cyclase that catalyzes CDA synthesis in vitro. We developed 2 engineered strains, CIBT4523 and CIBT4712, for producing high levels of CDA while keeping their concentrations within a range that did not compromise the growth. Although CIBT4712 exhibited stronger induction of the STING pathway corresponding to in vitro CDA levels, it had lower antitumor activity than CIBT4523 in an allograft tumor model, which might be related to the stability of the surviving bacteria in the tumor tissue. CIBT4523 exhibited complete tumor regression, prolonged survival of mice, and rejection of rechallenged tumors, thus, offering new possibilities for more effective tumor therapy. We showed that the appropriate production of CDA in engineered bacterial strains is essential for balancing antitumor efficacy and self-toxicity.

Automated summary

This study optimized the translation levels of diadenylate cyclase for the production of cyclic di-adenosine monophosphate (CDA) in two engineered bacterial strains. Although one strain showed stronger induction of the STING pathway, it had lower antitumor activity in an allograft tumor model, possibly due to the stability of the bacteria in the tumor tissue. The other strain exhibited complete tumor regression, prolonged survival of mice, and rejection of rechallenged tumors, suggesting new possibilities for effective tumor therapy.