154-OR: Neprilysin Inhibition Increases Glucagon Levels with Possible Implications for Hepatic Amino Acid Metabolism
Authors: KJELDSEN SASHA , ZRAIKA SAKENEH , MONGOVIN STEVE M. , HANSEN LASSE H. , TERZIC DIJANA , MARK PETER D. , PLOMGAARD PETER , GØTZE JENS P. , WINTHER-SOERENSEN MARIE , HUNT JENNA , GALSGAARD KATRINE D. , ROSENKILDE METTE M. , GOOSSENS GIJS H. , BLAAK ELLEN E. , HOLST JENS J. , WEWER ALBRECHTSEN NICOLAI J. 
Published: June 2020

Glucagon (gcg) regulates hepatic glucose and amino acid (AA) metabolism, and increased gcg levels (hyperglucagonemia) contribute to hyperglycemia in diabetes. We hypothesized that the enzyme neprilysin (NEP) contributes to gcg degradation. We measured plasma gcg levels during a mixed meal in nine healthy men after a single dose of the NEP-inhibitor/angiotensin II receptor blocker (194 mg sacubitril/206 mg valsartan, 30 min prior to meal), a DPP-4 inhibitor (sitagliptin, 2x100mg), these combined, or the meal alone. Postprandial gcg levels were 2.7-fold higher in sacubitril/valsartan treated individuals compared to controls (P=0.005) and this was not significantly altered by the addition of sitagliptin (P=0.28). Sacubitril/valsartan also lowered postprandial plasma AA levels (P=0.01), but glucose levels were unaffected (P=0.76). In obese individuals (n=7), eight weeks sacubitril/valsartan treatment increased fasting gcg levels (P=0.02). To test whether NEP degrades gcg and diminishes its signaling, we performed mass-spectrometry and assessed cells transfected with the gcg receptor (gcgr). We found that NEP cleaves gcg and that the gcg fragments produced were unable to activate the gcgr. In non-sedated C57BL/6JRj female mice (n=8) NEP inhibition (sacubitril, 0.7 nmol/g) increased gcg levels (P=0.02) and tended to increase AA disappearance (P=0.076) and urea formation (P=0.08) during an AA challenge. A gcgr antagonist (Novo Nordisk; 25-2648, 100 mg/kg) abolished the increase in urea formation observed with sacubitril alone. Gcg levels were increased 1.9-fold (P&lt;0.002) with sacubitril compared to without, after a single injection of gcg (96 ng/g). In mice with genetic ablation of NEP (n=10), fasting plasma urea (P=0.003) but surprisingly not gcg levels (P=0.57) were increased compared to controls. NEP degrades gcg and thus inhibitors of NEP may result in hyperglucagonemia with potential metabolic perturbations on hepatic AA metabolism.

Disclosure
S. Kjeldsen: None. S. Zraika: Research Support; Self; Novartis Pharmaceuticals Corporation. S.M. Mongovin: None. L.H. Hansen: None. D. Terzic: None. P.D. Mark: None. P. Plomgaard: None. J.P. Gøtze: None. M. Winther-Soerensen: None. J. Hunt: None. K.D. Galsgaard: None. M.M. Rosenkilde: Board Member; Self; Synklino. Consultant; Self; Antag Therapeutics, Bainan Biotech. G.H. Goossens: None. E.E. Blaak: None. J.J. Holst: Advisory Panel; Self; AstraZeneca, Merck Sharp & Dohme Corp., Novo Nordisk A/S, Zealand Pharma A/S. Other Relationship; Spouse/Partner; Antag Therapeutics. N.J. Wewer Albrechtsen: Research Support; Self; Mercodia, Novo Nordisk A/S, Novo Nordisk Foundation. Speaker’s Bureau; Self; Merck Sharp & Dohme Corp.

Funding
Novo Nordisk Foundation (NNF19OC0055001)