Angiotensin-converting enzyme 2 (ACE2) is critically involved in cardiovascular physiology and pathology, and is currently clinically evaluated to treat acute lung failure. Here we show that the B38-CAP, a carboxypeptidase derived from Paenibacillus sp. B38, is an ACE2-like enzyme to decrease angiotensin II levels in mice. In protein 3D structure analysis, B38-CAP homolog shares structural similarity to mammalian ACE2 with low sequence identity. In vitro, recombinant B38-CAP protein catalyzed the conversion of angiotensin II to angiotensin 1-7, as well as other known ACE2 target peptides. Treatment with B38-CAP suppressed angiotensin II-induced hypertension, cardiac hypertrophy, and fibrosis in mice. Moreover, B38-CAP inhibited pressure overload-induced pathological hypertrophy, myocardial fibrosis, and cardiac dysfunction in mice. Our data identify the bacterial B38-CAP as an ACE2-like carboxypeptidase, indicating that evolution has shaped a bacterial carboxypeptidase to a human ACE2-like enzyme. Bacterial engineering could be utilized to design improved protein drugs for hypertension and heart failure.

Esophageal squamous cell carcinoma (ESCC) is one of the most fatal types of malignant tumors worldwide. Epitranscriptome, such as N6‐methyladenosine (m6A) of mRNA, is an abundant post‐transcriptional mRNA modification and has been recently implicated to play roles in several cancers, whereas the significance of m6A modifications is virtually unknown in ESCC. Analysis of tissue microarray of the tumors in 177 ESCC patients showed that higher expression of m6A demethylase ALKBH5 correlated with poor prognosis and that ALKBH5 was an independent prognostic factor of the survival of patients. There was no correlation between the other demethylase FTO and prognosis. siRNA knockdown of ALKBH5 but not FTO significantly suppressed proliferation and migration of human ESCC cells. ALKBH5 knockdown delayed progression of cell cycle and accumulated the cells to G0/G1 phase. Mechanistically, expression of CDKN1A (p21) was significantly up‐regulated in ALKBH5‐depleted cells, and m6A modification and stability of CDKN1A mRNA were increased by ALKBH5 knockdown. Furthermore, depletion of ALKBH5 substantially suppressed tumor growth of ESCC cells subcutaneously transplanted in BALB/c nude mice. Collectively, we identify ALKBH5 as the first m6A demethylase that accelerates cell cycle progression and promotes cell proliferation of ESCC cells, which is associated with poor prognosis of ESCC patients.

Angiotensin-converting enzyme 2 (ACE2) is a receptor for cell entry of SARS-CoV-2, and recombinant soluble ACE2 protein inhibits SARS-CoV-2 infection as a decoy. ACE2 is a carboxypeptidase that degrades angiotensin II, thereby improving the pathologies of cardiovascular disease or acute lung injury. Here we show that B38-CAP, an ACE2-like enzyme, is protective against SARS-CoV-2-induced lung injury. Endogenous ACE2 expression is downregulated in the lungs of SARS-CoV-2-infected hamsters, leading to elevation of angiotensin II levels. Recombinant Spike also downregulates ACE2 expression and worsens the symptoms of acid-induced lung injury. B38-CAP does not neutralize cell entry of SARS-CoV-2. However, B38-CAP treatment improves the pathologies of Spike-augmented acid-induced lung injury. In SARS-CoV-2-infected hamsters or human ACE2 transgenic mice, B38-CAP significantly improves lung edema and pathologies of lung injury. These results provide the first in vivo evidence that increasing ACE2-like enzymatic activity is a potential therapeutic strategy to alleviate lung pathologies in COVID-19 patients.

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Angiotensin-converting enzyme 2 (ACE2) is a receptor for cell entry of SARS-CoV-2, and recombinant soluble ACE2 protein inhibits SARS-CoV-2 infection as a decoy. ACE2 is the carboxypeptidase to degrade angiotensin II (Ang II) to angiotensin 1-7 and improves the pathologies of cardiovascular disease and acute lung injury. To address whether the carboxypeptidase enzymatic activity of ACE2 is protective against COVID-19, we investigated the effects of B38-CAP, an ACE2-like enzyme, on SARS-CoV-2-induced lung injury. Expression of ACE2 protein was significantly downregulated in the lungs of SARS-CoV-2-infected hamsters. Recombinant S1 domain or receptor-binding domain (RBD) of the SARS-CoV-2 Spike protein also directly downregulated ACE2 expression and elevated Ang II levels and considerably worsened acid-induced lung injury in hamsters. Treatment with B38-CAP downregulated Spike RBD-induced high Ang II levels, severe inflammation and pulmonary edema through its ACE2-like enzymatic activity. Consistently, elevated cytokine mRNA levels and impaired lung functions were improved by B38-CAP treatment. Moreover, in SARS-CoV-2-infected humanized ACE2 transgenic mice, B38-CAP significantly improved the pathologies of lung injury, alleviated the cytokine storms and downregulated viral RNA levels. These results provide the first experimental in vivo evidence that increasing ACE2-like enzymatic activity is a potential and powerful therapeutic strategy for lung pathologies in COVID-19.

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Angiotensin-converting enzyme 2 (ACE2) is the carboxypeptidase to degrade angiotensin II (Ang II) to angiotensin 1–7 (Ang 1–7) and improves the pathologies of cardiovascular disease and acute respiratory distress syndrome (ARDS)/acute lung injury. B38-CAP is a bacteria-derived ACE2-like carboxypeptidase as potent as human ACE2 and ameliorates hypertension, heart failure and SARS-CoV-2-induced lung injury in mice. Recombinant B38-CAP is prepared with E. coli protein expression system more efficiently than recombinant soluble human ACE2. Here we show therapeutic effects of B38-CAP on abdominal sepsis- or acid aspiration-induced acute lung injury. ACE2 expression was downregulated in the lungs of mice with cecal ligation puncture (CLP)-induced sepsis or acid-induced lung injury thereby leading to upregulation of Ang II levels. Intraperitoneal injection of B38-CAP significantly decreased Ang II levels while upregulated angiotensin 1–7 levels. B38-CAP improved survival rate of the mice under sepsis. B38-CAP suppressed the pathologies of lung inflammation, improved lung dysfunction and downregulated elevated cytokine mRNA levels in the mice with acute lung injury. Thus, systemic treatment with an ACE2-like enzyme might be a potential therapeutic strategy for the patients with severe sepsis or ARDS.

Angiotensin-converting enzyme 2 (ACE2) is a receptor for cell entry of SARS-CoV-2, and recombinant soluble ACE2 protein inhibits SARS-CoV-2 infection as a decoy. ACE2 is the carboxypeptidase to degrade angiotensin II (Ang II) to angiotensin 1-7 and improves the pathologies of cardiovascular disease and acute lung injury. To address whether the carboxypeptidase enzymatic activity of ACE2 is protective against COVID-19, we investigated the effects of B38-CAP, an ACE2-like enzyme, on SARS-CoV-2-induced lung injury. Expression of ACE2 protein was significantly downregulated in the lungs of SARS-CoV-2-infected hamsters. Recombinant S1 domain or receptor-binding domain (RBD) of the SARS-CoV-2 Spike protein also directly downregulated ACE2 expression and elevated Ang II levels and considerably worsened acid-induced lung injury in hamsters. Treatment with B38-CAP downregulated Spike RBD-induced high Ang II levels, severe inflammation and pulmonary edema through its ACE2-like enzymatic activity. Consistently, elevated cytokine mRNA levels and impaired lung functions were improved by B38-CAP treatment. Moreover, in SARS-CoV-2-infected humanized ACE2 transgenic mice, B38-CAP significantly improved the pathologies of lung injury, alleviated the cytokine storms and downregulated viral RNA levels. These results provide the first experimental in vivo evidence that increasing ACE2-like enzymatic activity is a potential and powerful therapeutic strategy for lung pathologies in COVID-19.