PT  - JOURNAL ARTICLE
AU  - Farnaz Khodabakhsh
AU  - Morteza Salimian
AU  - Ardavan Mehdizadeh
AU  - Mohammad Sadeq Khosravi
AU  - Alireza Vafabakhsh
AU  - Elmira Karami
AU  - Reza Ahangari Cohan
TI  - <strong>New proline, alanine, serine repeat sequence for pharmacokinetic enhancement of anti-VEGF single domain antibody</strong>
AID  - 10.1124/jpet.120.000012
DP  - 2020 Jan 01
TA  - Journal of Pharmacology and Experimental Therapeutics
PG  - JPET-AR-2020-000012
4099  - http://jpet.aspetjournals.org/content/early/2020/07/14/jpet.120.000012.short
4100  - http://jpet.aspetjournals.org/content/early/2020/07/14/jpet.120.000012.full
AB  - Therapeutic fragmented antibodies show a poor pharmacokinetic profile that leads to frequent high-dose administration. In the current study, for the first time, a novel proline, alanine, serine (PAS) repeat sequence, called PAS#208, was designed to extend the plasma half-life of a nano-sized anti-Vascular Endothelial Growth Factor-A single-domain antibody. Polyacrylamide gel electrophoresis, circular dichroism, dynamic light scattering, and electrophoretic light scattering were used to assess the physicochemical properties of the newly designed PAS sequence. The effect of PAS#208 on the biological activity of a single-domain antibody was studied using an in vitro proliferation assay. The pharmacokinetic parameters including terminal half-life, the volume of distribution, elimination rate constant, and clearance were determined in mice model and compared to the native protein and PAS#1(200) sequence. The novel PAS repeat sequence showed comparable physicochemical, biological, and pharmacokinetic features to the previously reported PAS#1(200) sequence. The PAS#208 increased the hydrodynamic radius and decreased significantly the electrophoretic mobility of the native protein without any change in zeta potential. Surprisingly, the fusion of PAS#208 to the single-domain antibody increased the binding potency. In addition, it did not alter the biological activity and did not show any cytotoxicity on the normal cells. The PAS#208 sequence improved the terminal half-life (14-fold) as well as other pharmacokinetic parameters significantly. The simplicity as well as superior effects on half-life extension, make PAS#208 sequence as a novel sequence for in vivo pharmacokinetic enhancement of therapeutic fragmented antibodies. Significance Statement •In the current study, a new PAS sequence was developed that showed comparable physicochemical, biological, and pharmacokinetic features to the previously reported PAS#1(200) sequence. •The simplicity as well as superior effects on half-life extension, make PAS#208 sequence as a novel sequence for in vivo pharmacokinetic enhancement of recombinant small proteins.