Engineered hypoxia-responding Escherichia coli carrying cardiac peptide genes, suppresses tumor growth, angiogenesis and metastasis in vivo

Mitra Samadi,Keivan Majidzadeh-A,Ahad Muhammadnejad,Neda Jalili,Shima Moradi-Kalbolandi,Leila Farahmand,Azadeh Sharif Khatibi,Zeinab Noorinejad,Marjan Mosayebzadeh,Malihe Salehi

doi:10.1186/s13036-021-00269-2

Abstract

Development of engineered non-pathogenic bacteria, capable of expressing anti-cancer proteins under tumor-specific conditions, is an ideal approach for selectively eradicating proliferating cancer cells. Herein, using an engineered hypoxia responding nirB promoter, we developed an engineered Escherichia coli BW25133 strain capable of expressing cardiac peptides and GFP signaling protein under hypoxic condition for spatiotemporal targeting of mice mammary tumors. Following determination of the in vitro cytotoxicity profile of the engineered bacteria, selective accumulation of bacteria in tumor microenvironment was studied 48 h after tail vein injection of 108 cfu bacteria in animals. For in vivo evaluation of antitumoral activities, mice with establishment mammary tumors received 3 consecutive intravenous injections of transformed bacteria with 4-day intervals and alterations in expression of tumor growth, invasion and angiogenesis specific biomarkers (Ki-67, VEGFR, CD31and MMP9 respectively), as well as fold changes in concentration of proinflammatory cytokines were examined at the end of the 24-day study period. Intravenously injected bacteria could selectively accumulate in tumor site and temporally express GFP and cardiac peptides in response to hypoxia, enhancing survival rate of tumor bearing mice, suppressing tumor growth rate and expression of MMP-9, VEGFR2, CD31 and Ki67 biomarkers. Applied engineered bacteria could also significantly reduce concentrations of IL-1β, IL-6, GC-SF, IL-12 and TNF-α proinflammatory cytokines while increasing those of IL-10, IL-17A and INF-γ. Overall, administration of hypoxia-responding E. coli bacteria, carrying cardiac peptide expression construct could effectively suppress tumor growth, angiogenesis, invasion and metastasis and enhance overall survival of mice bearing mammary tumors.

Highlights

Despite the efficacy of conventional chemotherapeutic agents in eradicating cancer cells, unacceptable adverse effects of these compounds, resulted from their nonselective toxicity toward normal cells has mostly limited their broad application in clinic and has directed oncologist toward development of novel interventions with selective toxicity toward cancer cells
In present study, we reported successful development of an engineered Hypoxia-responding E. coli BW25113 strain carrying cardiac peptides (LANP, vessel dilator (VD) and KP) and green fluorescent protein (GFP) genes which could spatiotemporally target breast tumors, that is to say, preferentially delivering therapeutic cargo to the tumor site and expressing it only after receiving the stimulatory signal at tumor site
This Selective process of bacterial colonization in tumor microenvironment has been in large part attributed to Despite the existing evidence on antitumoral activities of bacteria, their localization and proliferation alone in tumor microenvironment is not sufficient for complete attenuation of the tumor growth

Summary

Introduction

Despite the efficacy of conventional chemotherapeutic agents in eradicating cancer cells, unacceptable adverse effects of these compounds, resulted from their nonselective toxicity toward normal cells has mostly limited their broad application in clinic and has directed oncologist toward development of novel interventions with selective toxicity toward cancer cells. Pursuing this goal has resulted in development of two new approaches, namely, molecular targeted therapies and immunotherapies. Most of the tumors do not express currently actionable genetic alterations [12]

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