Biodesign of a renal-protective peptide based on alternative splicing of B-type natriuretic peptide

Abstract

Alternative RNA splicing may provide unique opportunities to identify drug targets and therapeutics. We identified an alternative spliced transcript for B-type natriuretic peptide (BNP) resulting from intronic retention. This transcript is present in failing human hearts and is reduced following mechanical unloading. The intronretained transcript would generate a unique 34 amino acid (aa) carboxyl terminus while maintaining the remaining structure of native BNP. We generated antisera to this carboxyl terminus and identified immunoreactivity in failing human heart tissue. The alternatively spliced peptide (ASBNP) was synthesized and unlike BNP, failed to stimulate cGMP in vascular cells or vasorelax preconstricted arterial rings. This suggests that ASBNP may lack the dose-limiting effects of recombinant BNP. Given structural considerations, a carboxyl-terminal truncated form of ASBNP was generated (ASBNP.1) and was determined to retain the ability of BNP to stimulate cGMP in canine glomerular isolates and cultured human mesangial cells but lacked similar effects in vascular cells. In a canine-pacing model of heart failure, systemic infusion of ASBNP.1 did not alter mean arterial pressure but increased the glomerular filtration rate (GFR), suppressed plasma renin and angiotensin, while inducing natriuresis and diuresis. Consistent with its distinct in vivo effects, the activity of ASBNP.1 may not be explained through binding and activation of NPR-A or NPR-B. Thus, the biodesigner peptide ASBNP.1 enhances GFR associated with heart failure while lacking the vasoactive properties of BNP. These findings demonstrate that peptides with unique properties may be designed based on products of alternatively splicing.vasoactive ͉ myocardial ͉ kidney G enome-wide analyses have revealed the prevalence of alternative splicing of multiexonic genes (1, 2). In fact, much of the complexity of the human proteome is accounted for by alternative splicing of messenger RNA. Identification of these altered forms may allow for unique opportunities to diagnose, understand, and treat human disease. Therefore, we hypothesized that it might be possible to identify splice variants and to design therapeutics based on their unique structure and function.As an example of the wide potential of this technology to alter disease states, we focused on a peptide with broad mechanistic, diagnostic, and therapeutic importance in cardiovascular disease, B-type natriuretic peptide (BNP). BNP is encoded by a small multiexonic gene, and although discovered in brain (3), is expressed primarily in the heart (4). BNP, like atrial natriuretic peptide and C-type natriuretic peptide, is expressed as a prepro-hormone that is processed to a mature [32-amino acid (aa)] form by extracellular proteases (5). Mature BNP contains short carboxyl and amino termini and a central 17-aa ring. BNP has important autocrine, paracrine, and endocrine actions that are mediated through the NPRA receptor and activation of cGMP in target cells (6). Infusion of a recombinant form of mat...

A Novel Atrial Natriuretic Peptide Based Therapeutic in Experimental Angiotensin II Mediated Acute Hypertension

“…38 Indeed, human BNP activates cGMP generation in isolated canine glomeruli. 39 While our results suggest comparable actions between M-ANP and human BNP our conclusions need to be cautious due to differences in human and canine BNP.…”

Can we prevent or treat renal dysfunction in chronic heart failure?

“…Another example of biodesigner peptide is alternatively spliced BNP (ASBNP). This peptide has also shown to retain renal effects while lacking vascular effects of BNP in a canine model of CHF [51]. Specifically, ASBNP did not alter mean arterial pressure and increased GFR; also, ASBNP suppressed plasma renin and angiotensin II while inducing natriuresis and diuresis compared with BNP.…”

Innovative Therapeutics

“…In vivo, intravenous administration of ANX-042 in a canine model of pacing-induced HF significantly enhanced diuresis, natriuresis, and GFR without reducing mean arterial pressure (65). Data from a Healthy Volunteer Dose Escalation Study (NCT01638104) furthermore showed that infusion with ANX-042 is safe and results in cGMP-activation (Table 1).…”