Targeting MYC Transcription with Small Peptide Derived from KSHV Transactivator (CUT&RUN)

Herpesviruses rely on host cell transcription and translation machineries for replication. Viral proteins thus function to redirect multiple cellular proteins for viral replication. In herpesvirus replicating cells, host cell gene transcription is frequently down-regulated because important transcriptional apparatuses are appropriated by viral transcription factors. Here we show that an evolutionally-shaped viral protein sequence is a great starting material for unique drug development to modulate cellular transcription. Cellular c-Myc protein (MYC) is overexpressed in over 70% of all types of cancer cells and therefore a very attractive target to control cancer cell growth. We identified a small functional peptide derived from the Kaposi's sarcoma-associated herpesvirus transactivator (K-Rta), which strongly attenuates MYC expression, reduces cell proliferation, and selectively kills cancer cells in both tissue culture and a xenograft tumor mouse model. Mechanistically, the peptide blocks promoter-enhancer interactions by preventing coactivator complex consisting of Nuclear receptor coactivator 2, p300, and SWI/SNF proteins from engaging the MYC promoter in leukemia cells. Target gene profiling with SLAM-seq suggests that the viral peptide attenuates MYC expression through a mechanism likely different from that of BET bromodomain inhibitors. Furthermore, fusing the 13 amino acids peptide with humanized anti-CD22 single chain armed the antibody drug with cell killing ability, and inhibited cell growth in soft agar. Our studies thus demonstrate the utility of the peptide sequence as a therapeutics module, which may be used to modulate MYC activity in a cell type-specific manner. Overall design: The goal of these studies was to define the genome-wide binding sites host cellular proteins and sites of histone modifications on the human and Kaposi's sarcoma-associated herpesvirus (KSHV) genome using CUT&RUN. This was performed in the context of the BCBL-1, (TREx)BCBL-1, and BC-1 cell line models, which are derived from KSHV-infected human primary effusion lymphomas (PEL) and contain latent KSHV genomes. TREx-K-Rta BCBL-1, also referred to as (TREx)BCBL-1, is an engineered subline, which contains a Tetracycline/Doxycycline (Dox)-inducible Flagx3- and HAx3-tagged K-Rta expression cassette. Viral reactivation can be stimulated by inducing K-Rta expression in individual TREx-K-Rta BCBL-1 cultures by treatment with Dox for 24 hours. For this study, replicate cell cultures were grown to 75% confluence and then harvested for CUT&RUN analyses for binding sites of SMARCC1, SMARCA5, RNA polymerase II, and sites of H3K27Ac and H3K4me1 histone modifications. Non-specific IgG was used as a control.