Activity-based chemical proteomics approaches used for
identifying
cellular targets of drugs are mainly dependent on the availability
of probes derived from drugs. However, all chemical probes are structurally
different from the drugs themselves and cannot fully mimic the real
actions of drugs in cells. Here we present a concise and unbiased
immunoaffinity-based strategy for identifying covalent drug targets
in vivo. By using the specific antibody, we not only confirm the well-known
ibrutinib-binding target BTK, but also identify some previously undescribed
strongly binding proteins, such as CKAP4 in human cell lines and TAP1
in mouse organs. The observed target profiles between species may
partially explain why certain drug candidates are very effective in
mice but not in humans. This approach avoids the chemical modification
of drugs, eliminates the nonspecific bindings of chemical probes,
and allows to unbiasedly decode the underlying mechanisms of action
of covalent drugs.