TY - JOUR
T1 - AP301, a synthetic peptide mimicking the lectin-like domain of TNF, enhances amiloride-sensitive Na+ current in primary dog, pig and rat alveolar type II cells
AU - Tzotzos, Susan
AU - Fischer, Bernhard
AU - Fischer, Hendrik
AU - Pietschmann, Helmut
AU - Lucas, Rudolf
AU - Dupré, Gilles
AU - Lemmens-Gruber, Rosa
AU - Hazemi, Parastoo
AU - Prymaka, Victoria
AU - Shabbir, Waheed
N1 - Funding Information:
W.S. and P.H. received financial support from the Austrian Research Promotion Agency (FFG) . This work was partially supported by a research grant from NHBLI , NIH (Grant RO1HL094609 to R.L.).
PY - 2013/6
Y1 - 2013/6
N2 - Pulmonary permeability oedema is a frequent complication in a number of life-threatening lung conditions, such as ALI and ARDS. Apart from ventilation strategies, no specific therapy yet exists for treatment of these potentially fatal illnesses. The oedema-reducing capacity of the lectin-like domain of TNF (TIP) and of synthetic peptides, mTIP and hTIP, which mimic the TIP domain of mouse and human TNF, have been demonstrated in various studies in rodents. Cell-based electrophysiological studies have revealed that the alveolar fluid clearing capacity of TNF and the TIP peptides is due to activation of the amiloride-sensitive Na+ current in alveolar epithelial cells and that the primary site of action is on the apical side of these cells.AP301, a synthetic cyclic peptide mimicking the TIP domain of human TNF is currently undergoing clinical trials as a therapy for pulmonary permeability oedema. AP301 has been shown to improve alveolar liquid clearance and lung function in a porcine model of ALI. For non-clinical regulatory assessment, dog, pig and rat are standard animal models; accordingly, pre-clinical toxicological and pharmacological safety studies have been conducted with AP301 in dogs and rats. Hitherto, no studies have assessed the pharmacodynamic effect of AP301 on primary canine or porcine type II AEC. The current study describes the effect of AP301 on the amiloride-sensitive Na+ current in type II AEC isolated from dog, pig and rat lungs. In whole cell patch clamp experiments with dog type II AEC, an increase in the amiloride-sensitive Na+ current from 3.7 pA to 49.4 pA was observed in the presence of AP301; in pig type II AEC, an increase from 10.0 pA to 159.6 pA was observed, and in rat AEC, from 6.9 pA to 62.4 pA. In whole cell patch clamp experiments in A549 cells, AP301-induced enhancement of the amiloride-sensitive current was eliminated when Na+ in the bath solution was replaced with N-methyl-d-glucamine (NMDG), and when the cells were pre-incubated with 5-aminoimidazole-4-carboxamide-1-ß-d-ribofuranoside (AICAR), an inhibitor of ENaC, but enhancement was unaffected by addition of cyclic nucleotide-gated (CNG) channel inhibitors Zn2+ or l-cis-diltiazem prior to AP301. These results provide strong evidence that AP301 activates the amiloride-sensitive Na+ current through ENaC in type II AEC from dog, pig and rat. To our knowledge, this is the first cell-based analysis of the oedema-clearing effect of AP301 observed in the porcine model of pulmonary oedema. Furthermore, the results validate the dog and pig models in non-clinical assessment of AP301.
AB - Pulmonary permeability oedema is a frequent complication in a number of life-threatening lung conditions, such as ALI and ARDS. Apart from ventilation strategies, no specific therapy yet exists for treatment of these potentially fatal illnesses. The oedema-reducing capacity of the lectin-like domain of TNF (TIP) and of synthetic peptides, mTIP and hTIP, which mimic the TIP domain of mouse and human TNF, have been demonstrated in various studies in rodents. Cell-based electrophysiological studies have revealed that the alveolar fluid clearing capacity of TNF and the TIP peptides is due to activation of the amiloride-sensitive Na+ current in alveolar epithelial cells and that the primary site of action is on the apical side of these cells.AP301, a synthetic cyclic peptide mimicking the TIP domain of human TNF is currently undergoing clinical trials as a therapy for pulmonary permeability oedema. AP301 has been shown to improve alveolar liquid clearance and lung function in a porcine model of ALI. For non-clinical regulatory assessment, dog, pig and rat are standard animal models; accordingly, pre-clinical toxicological and pharmacological safety studies have been conducted with AP301 in dogs and rats. Hitherto, no studies have assessed the pharmacodynamic effect of AP301 on primary canine or porcine type II AEC. The current study describes the effect of AP301 on the amiloride-sensitive Na+ current in type II AEC isolated from dog, pig and rat lungs. In whole cell patch clamp experiments with dog type II AEC, an increase in the amiloride-sensitive Na+ current from 3.7 pA to 49.4 pA was observed in the presence of AP301; in pig type II AEC, an increase from 10.0 pA to 159.6 pA was observed, and in rat AEC, from 6.9 pA to 62.4 pA. In whole cell patch clamp experiments in A549 cells, AP301-induced enhancement of the amiloride-sensitive current was eliminated when Na+ in the bath solution was replaced with N-methyl-d-glucamine (NMDG), and when the cells were pre-incubated with 5-aminoimidazole-4-carboxamide-1-ß-d-ribofuranoside (AICAR), an inhibitor of ENaC, but enhancement was unaffected by addition of cyclic nucleotide-gated (CNG) channel inhibitors Zn2+ or l-cis-diltiazem prior to AP301. These results provide strong evidence that AP301 activates the amiloride-sensitive Na+ current through ENaC in type II AEC from dog, pig and rat. To our knowledge, this is the first cell-based analysis of the oedema-clearing effect of AP301 observed in the porcine model of pulmonary oedema. Furthermore, the results validate the dog and pig models in non-clinical assessment of AP301.
KW - AP301
KW - Alveolar fluid clearance
KW - ENaC
KW - Lectin-like domain
KW - Patch clamp
KW - Synthetic cyclic peptide
KW - Type II alveolar cells
UR - http://www.scopus.com/inward/record.url?scp=84875913078&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84875913078&partnerID=8YFLogxK
U2 - 10.1016/j.pupt.2012.12.011
DO - 10.1016/j.pupt.2012.12.011
M3 - Article
C2 - 23313096
AN - SCOPUS:84875913078
SN - 1094-5539
VL - 26
SP - 356
EP - 363
JO - Pulmonary Pharmacology and Therapeutics
JF - Pulmonary Pharmacology and Therapeutics
IS - 3
ER -