Source Name: 2021_Kelly

Literature Information

Literature Title Geometrically Diverse Lariat Peptide Scaffolds Reveal an Untapped Chemical Space of High Membrane Permeability
Doi 10.1021/jacs.0c06115
Research Group
  1. Department of Chemistry and Biochemistry, University of California Santa Cruz
  2. Department of Bioengineering and Therapeutic Sciences, University of California San Francisco
  3. Unnatural Products Inc.
  4. -
  5. -
  6. -
  7. -
  8. -
  9. -
Data Number 1519
Minimum Molecular Weight 974.3
Maximum Molecular Weight 1220.4

Assay Information 1

Assay Type PAMPA
Permeability Type logPapp
Membrane Measurement Direction (Exclude PAMPA) -
Assay Detail The analyte concentration in the PAMPA assay was 250 μM for sub-libraries (total concentration, roughly 1 μM per compound) and 1 μM for compounds assayed individually. Internal standards were included in the assay at a concentration of 1 μM.For sub-libraries, 1NMe3, synthesized according to published procedures, was used as the internal standard and for compounds assayed individually, carbamazepine was used as the standard. The assay was run for 18 h. A 96-well donor plate with 0.45 μm hydrophobic Immobilon-P membrane supports (Millipore MAIPNTR10) and a 96-well Teflon acceptor plate (Millipore MSSACCEPTOR) were used in the PAMPA permeability test. The acceptor plate was prepared by adding 300 μL of 5% DMSO in PBS to each well. Donor well solutions were prepared by diluting 50 μL DMSO stock solutions prepared above to a final volume of 1000 μL with PBS and mixed thoroughly. The frits were infused with 5 μL of dodecane containing 1% (w/v) soy lecithin (90%, Alfa Aesar). The membranes were allowed to equilibrate for 5 minutes before adding the donor well solution and placing on top of the acceptor well solution to begin the assay.
Samples were prepared for LC-MS analysis by diluting with an equal volume of ACN. The donor wells were further diluted tenfold with 1:1 ACN/H2O for approximately even analyte concentration in the donor and acceptor wells. The assay was run as described above, except that the donor well contained 0.2% (v/v) polysorbate 80 and the acceptor well contained 0.2% (w/v) TPGS-750M. Donor well samples were prepared for LC-MS analysis by diluting with an equal volume of 9:1 ACN / 2% (w/v) TPGS-750M in water. Acceptor well samples were prepared for LC-MS analysis by diluting with an equal volume of 9:1 ACN / 2% (v/v) polysorbate 80 in water.

Assay Information 2

Assay Type MDCK
Permeability Type logPapp
Membrane Measurement Direction (Exclude PAMPA) Apical to Basolateral (AB)
Assay Detail The MDCK assay was carried out according to the procedure utilized by Furukawa et al. Transcellular transport of the test compounds from apical to basal direction using MDCK II cells was investigated. For the transcellular transport assay, the culture medium on the apical side and the basal side was replaced with buffer (pH 6.5) and buffer (pH 7.4) containing BSA, respectively. The buffer on the apical side was replaced with the buffer containing 10 μM of test compounds to start the incubation. After the incubation, aliquots of the solutions were sampled from both the apical side and basal side, and the concentrations of each compound were determined by LC-MS/MS.
Apparent permeability coefficient (Papp) was calculated by the following equations. Papp = (Cb × V) / (Ca × t × A), Papp: apparent permeability (10^-6 cm/sec), Ca: test compound concentration added to the apical side (μM), Cb: test compound concentration on the basal side (μM), A: surface area of cell monolayer (cm^2), V: volume of buffer on the basal side (cm^3), t: incubation time (sec).