Biology subjects

Kokkonda, P.

Publications and source records attributed to Kokkonda, P..

4 recordsLinked to original sources

bioRxiv2025

Phosphorylation-inducing molecules for regulating dynamic cellular processes

Dynamic cellular processes often employ protein phosphorylation for rapid information transfer within and between cells. Phosphorylation-inducing chimeric small molecules (PHICS) have been developed for targeted protein phosphorylation by on-demand inducing a kinase-protein pairing. However, widespread application of PHICS has been limited as previously reported PHICS that recruited AMP-activated protein kinase (AMPK) required serum starvation and target protein overexpression, recruited only a few of the potential AMPK complexes, and exhibited poor dose- and temporal control. Herein, we report an AMPK PHICS platform that operates under physiological conditions (i.e., no serum starvation or target overexpression), recruits multiple AMPK complexes, and can induce target protein phosphorylation with dose and temporal control. We demonstrated the utility of this platform for controlling phosphorylations that underlie two dynamic cellular processes, namely oncogenic signaling and phase separation. An AMPK PHICS directed against Brutons Tyrosine Kinase (BTK), which is a driver of several B cell malignancies, was effective at inducing the death of drug-resistant cancer cells. Here, PHICS induced inhibitory phosphorylations on BTK and attenuated its oncogenic pathway. Phosphorylation of Liprin-3 induces phase separation in neurons and is critical for neurotransmitter release. Using AMPK PHICS that phosphorylated Liprin-3, we induced Liprin-3 phase separation with dose and temporal control. We envision this PHICS platform to find utility in inducing and controlling protein phosphorylations for basic research and biomedicine. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=102 SRC="FIGDIR/small/652895v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@16bbb20org.highwire.dtl.DTLVardef@1ca5267org.highwire.dtl.DTLVardef@1e805d5org.highwire.dtl.DTLVardef@1d6ec22_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2023

Photoswitchable Molecular Glues Enable Optical Control of Transcription Factor Degradation

Immunomodulatory drugs (IMiDs), which include thalidomide and its derivatives, have emerged as the standard of care against multiple myeloma. They function as molecular glues that bind to the E3 ligase cereblon (CRBN) and induce protein interactions with neosubstrates, including the transcription factors Ikaros (IKZF1) and Aiolos (IKZF3). The subsequent ubiquitylation and degradation of these transcription factors underlies the antiproliferative activity of IMiDs. Here, we introduce photoswitchable immunomodulatory drugs (PHOIMiDs) that can be used to degrade Ikaros and Aiolos in a light-dependent fashion. Our lead compound shows minimal activity in the dark and becomes an active degrader upon irradiation with violet light. It shows high selectivity over other transcription factors, regardless of its state, and could therefore be used to control the levels of Ikaros and Aiolos with high spatiotemporal precision.

biochemistry↗

bioRxiv2022

Diversity-oriented synthesis encoded by deoxyoligonucleotides

Diversity-oriented synthesis (DOS)is a powerful strategy to prepare molecules with underrepresented features in commercial screening collections, resulting in the elucidation of novel biological mechanisms. In parallel to the development of DOS, DNA-encoded libraries (DELs) have emerged as an effective, efficient screening strategy to identify protein binders. Despite recent advancements in this field, most DEL syntheses are limited by the presence of sensitive DNA-based constructs. Here, we describe the design, synthesis, and validation experiments performed for a 3.7 million-member DEL, generated using diverse skeleton architectures with varying exit vectors, derived from DOS, to achieve structural diversity beyond what is possible by varying appendages alone. We will make this DEL available to the academic scientific community to increase access to novel structural features and accelerate early-phase drug discovery.

biochemistry↗

bioRxiv2021

Proteolysis Targeting Chimeras With Reduced Off-targets

Proteolysis Targeting Chimeras (PROTACs), a class of heterobifunctional molecules that recruit target proteins to E3 ligases, have gained traction for targeted protein degradation. However, pomalidomide, a widely used E3 ligase recruiter in PROTACs, can independently degrade other targets, such as zinc-finger (ZF) proteins, that hold key functions in normal development and disease progression. This off-target degradation of pomalidomide-based PROTACs raises concerns about their therapeutic applicability and long-term side effects. Therefore, there is a crucial need to develop rules for PROTAC design that minimize off-target degradation. In this study, we developed a high-throughput platform that interrogates the off-target degradation of ZF domains and discovered, using this platform, that PROTACs with the current design paradigm induce degradation of several ZF proteins. To identify new rules for PROTAC design, we generated a library of pomalidomide analogs that allowed systematic exploration of the impact of positional isomerism (e.g., C4 and C5 positions of the phthalimide ring), hydrogen bonding, steric and hydrophobic effects on propensities for ZF protein degradation. We found that modifications of appropriate size on the C5 position reduced off-target ZF degradation. We validated these results using immunoblotting, target engagement, and global mass spectrometric studies. We applied our newfound design principles on a previously developed ALK oncoprotein-targeting PROTAC and generated PROTACs with enhanced potency and minimal off-target degradation. We envision the reported off-target profiling platform and pomalidomide analogs will find utility in design of specific PROTACs.

pharmacology and toxicology↗