Development of a large peptoid-DOTA combinatorial library

Abstract

Conventional one-bead one-compound (OBOC) library synthesis is typically used to identify molecules with therapeutic value. The design and synthesis of OBOC libraries that contain molecules with imaging or even potentially therapeutic and diagnostic capacities (e.g. theranostic agents) has been overlooked. The development of a therapeutically active molecule with a built-in imaging component for a certain target is a daunting task, and structure-based rational design might not be the best approach. We hypothesize to develop a combinatorial library with potentially therapeutic and imaging components fused together in each molecule. Such molecules in the library can be used to screen, identify, and validate as direct theranostic candidates against targets of interest. As the first step in achieving that aim, we developed an on-bead library of 153,600 Peptoid-DOTA compounds in which the peptoids are the target-recognizing and potentially therapeutic components and the DOTA is the imaging component. We attached the DOTA scaffold to TentaGel beads using one of the four arms of DOTA, and we built a diversified 6-mer peptoid library on the remaining three arms. We evaluated both the synthesis and the mass spectrometric sequencing capacities of the test compounds and of the final library. The compounds displayed unique ionization patterns including direct breakages of the DOTA scaffold into two units, allowing clear decoding of the sequences. Our approach provides a facile synthesis method for the complete on-bead development of large peptidomimetic-DOTA libraries for screening against biological targets for the identification of potential theranostic agents in the future. © 2016 The Authors. Biopolymers Published by Wiley Periodicals, Inc. Biopolymers (Pept Sci) 106: 673-684, 2016.

Keywords: DOTA; combinatorial; imaging; peptoids; theranostics.

Figure 1

(A) Schematic representation of a theranostic unit. (B) On‐bead structure of tri‐peptoid–DOTA molecule with initial linker, DOTA scaffold, spacer, and diversified peptoid region.

Scheme 1

General synthesis strategy for peptoid–DOTA library. (A) Assembly of linker on resin following DO3A insertion, spacer addition on 3 arms of DO3A. (B) General oligomerization strategy for library construction, (C) list if amines used for the library synthesis.

Figure 2

(A) Structures of test compounds 20, 21, and 22 with n = 1, 2, and 3 AEEAc spacers respectively. (B), (C), and (D) MALDI mass spectra acquired by voyager De‐Pro for compounds 20 (expected MS 3282.85 m/z, observed MS: 3283.36 m/z), 21 (expected MS 3928.24 m/z, observed MS: 3927.62 m/z), and 22 (expected MS 4612.29 m/z, observed MS: 4613.03 m/z). (E–G) are analytical HPLC for compounds 20, 21, and 22 respectively.

Figure 3

MS/MS sequencing of the test compound 20 from crude material cleaved from a single bead.

Figure 4

MS/MS sequencing of the test compound 22 from crude material cleaved from a single bead.

Scheme 2

Peptoid–DOTA OBOC combinatorial synthesis strategy.

Figure 5

Analysis of one of the representative compounds from crude material cleaved off from a single bead randomly chosen from the library. (A) Total mass (MS) spectrum. Expected MS 3778.30 m/z and observed MS 3779.84 m/z. (B) Analytical HPLC. Compound peak is marked with an arrow. (C) MS/MS sequencing spectrum and the corresponding analysis. The R1–6 peptoid region peaks considered here are derived from M + H master ion. At the same time, each of those peaks are flanked by corresponding M + Na derived peaks as well (multiple peaks found on the expanded portion of the spectrum).