Targeting the sodium-dependent multivitamin transporter (SMVT) for improving the oral absorption properties of a retro-inverso Tat nonapeptide

Purpose. To investigate the potential for delivering large peptides orally by altering their absorptive transport pathways and improving intestinal pe rmeability. The absorptive transport of retro-inverso (R.I.-) K-Tat9 and R. I.-K(biotin)-Tat9, novel peptidic inhibitors of the Tat protein of HIV-I, a nd their interactions with human SMVT (hSMVT), a high affinity, low capacit y transporter, were investigated using Caco-2 and transfected CHO cells. Methods. Following synthesis on a PAL resin using Fmoc chemistry, the trans port of R.I.-K-Tat9 (0.01-25 muM) and R.I.-K(biotin)-Tat9 (0.1-25 muM) was evaluated across Caco-2 cells. The transport and kinetics of biotin, biocyt in and desthiobiotin (positive controls for SMVT) were also determined. Upt ake of R.I.-K-Tat9 and R.I.- K(biotin)-Tat9 (both 0.1-10 muM) was determine d in CHO/hSMVT and CHO/pSPORT (control) cells. Results, The absorptive transport of R.I.-K-Tat9 was passive, low (P(m)simi lar to1 x 10(-6) cm/sec) and not concentration dependent. R.I.- K(biotin)-T at9 permeability was 3.2-fold higher than R.I.-K-Tat9 demonstrating active (E-a = 9.1 kcal/mole), concentration dependent and saturable transport (K-m = 3.3 muM). R.I.-K(biotin)-Tat9 uptake in CHO/hSMVT cells (K-m = 1.0 muM) was similar to 500-fold greater than R.I.-K-Tat9 (nt 10 muM). R.I.-K(biotin )-Tat9 transport in Caco-2 and CHO/hSMVT cells was significantly inhibited by known substrates of SMVT including biotin. biocytin, and desthiobiotin. Passive uptake of R.I.-K(biotin)-Tat9 was significantly greater than R.I.-K -Tat9 uptake in CHO/pSPORT cells. Conclusions. These results demonstrate that the structural modification of R.I.-K-Tat9 to R.I.-K(biotin)-Tat9 altered its intestinal trans port pathwa y resulting in a significant improvement in its absorptive permeability by enhancing nonspecific passive and carrier-mediated uptake by means of SMVT. The specific interactions between R.I.-K(biotin)-Tat9 and SMVT suggest tha t targeting approaches utilizing transporters such as SMVT may substantiall y improve the oral delivery of large peptides.