Treasure hunt for peptides with undefined chemical modifications: Proteomics identification of differential albumin adducts of 2‐nitroimidazole‐indocyanine green in hypoxic tumor

Abstract

2‐Nitroimidazole is a well‐known chemical probe targeting hypoxic environments of solid tumors, and its derivatives are widely used as imaging agents to investigate tissue and tumor hypoxia. However, the underlying chemistry for the hypoxia‐detection capability of 2‐nitroimidazole is still unclear. In this study, we deployed a biotin conjugate of 2‐nitroimidazole‐indocyanine green (2‐nitro‐ICG) for the investigation of in vivo hypoxia‐probing mechanism of 2‐nitro‐ICG compounds. By implementing mass spectrometry‐based proteomics and exhaustive data mining, we report that 2‐nitro‐ICG and its fragments modify mouse serum albumin as the primary protein target but at two structurally distinct sites and possibly via two different mechanisms. The identification of probe‐modified peptides not only contributes to the understanding of the in vivo metabolism of 2‐nitroimidazole compounds but also demonstrates a competent analytical workflow that enables the search for peptides with undefined modifications in complex proteome digests.

Measurement and utilization of the proteomic reactivity by mass spectrometry

“…The Et4‐Rhod probe (Rea 1 (Rec/Rep) 1 ) has both Rec and Rep functions in the same molecular skeleton (with Rep enabling fluorescence rather than MS detection). Biotinyl 2‐nitroimidazole‐indocyanine green (2‐nitro‐ICG, Rea 1 Rec 1 Rep 1 ), derived from 2‐nitro‐ICG for studying hypoxic environments in cancer, reveals targeting mechanisms of 2‐nitroimidazole in animals (L. Wang et al, 2020). The creation of CCPs is a challenging and individualized process, as it can vary depending on the structure of the compound being derived from (Baillie, 2016; Berdan et al, 2019; Lanning et al, 2014; P. Y. Yang et al, 2010; Zhao et al, 2017).…”