BacPROTACs mediate targeted protein degradation in bacteria.

Hijacking the cellular protein degradation system offers unique opportunities for drug discovery, as exemplified by proteolysis-targeting chimeras. Despite their great promise for medical chemistry, so far, it has not been possible to reprogram the bacterial degradation machinery to interfere with microbial infections. Here, we develop small-molecule degraders, so-called BacPROTACs, that bind to the substrate receptor of the ClpC:ClpP protease, priming neo-substrates for degradation. In addition to their targeting function, BacPROTACs activate ClpC, transforming the resting unfoldase into its functional state. The induced higher-order oligomer was visualized by cryo-EM analysis, providing a structural snapshot of activated ClpC unfolding a protein substrate. Finally, drug susceptibility and degradation assays performed in mycobacteria demonstrate in vivo activity of BacPROTACs, allowing selective targeting of endogenous proteins via fusion to an established degron. In addition to guiding antibiotic discovery, the BacPROTAC technology presents a versatile research tool enabling the inducible degradation of bacterial proteins.

Stepwise activities of mSWI/SNF family chromatin remodeling complexes direct T cell activation and exhaustion.

Highly coordinated changes in gene expression underlie T cell activation and exhaustion. However, the mechanisms by which such programs are regulated and how these may be targeted for therapeutic benefit remain poorly understood. Here, we comprehensively profile the genomic occupancy of mSWI/SNF chromatin remodeling complexes throughout acute and chronic T cell stimulation, finding that stepwise changes in localization over transcription factor binding sites direct site-specific chromatin accessibility and gene activation leading to distinct phenotypes. Notably, perturbation of mSWI/SNF complexes using genetic and clinically relevant chemical strategies enhances the persistence of T cells with attenuated exhaustion hallmarks and increased memory features in vitro and in vivo. Finally, pharmacologic mSWI/SNF inhibition improves CAR-T expansion and results in improved anti-tumor control in vivo. These findings reveal the central role of mSWI/SNF complexes in the coordination of T cell activation and exhaustion and nominate small-molecule-based strategies for the improvement of current immunotherapy protocols.

Mini PROTACs: N-end rule-mediated degradation on the horizon.

Heterobifunctional proteolysis-targeting chimeras (PROTACs) offer a promising cancer treatment avenue by efficiently degrading unwanted cellular proteins. A recent study from Zhang et al. demonstrated the successful utilization of the N-end rule in PROTAC design, allowing for a modular degradation rate tailored to the oncogenic driver BCR-ABL.

Plasticity of the Cullin-RING Ligase Repertoire Shapes Sensitivity to Ligand-Induced Protein Degradation.

Inducing protein degradation via small molecules is a transformative therapeutic paradigm. Although structural requirements of target degradation are emerging, mechanisms determining the cellular response to small-molecule degraders remain poorly understood. To systematically delineate effectors required for targeted protein degradation, we applied genome-scale CRISPR/Cas9 screens for five drugs that hijack different substrate receptors (SRs) of cullin RING ligases (CRLs) to induce target proteolysis. We found that sensitivity to small-molecule degraders is dictated by shared and drug-specific modulator networks, including the COP9 signalosome and the SR exchange factor CAND1. Genetic or pharmacologic perturbation of these effectors impairs CRL plasticity and arrests a wide array of ligases in a constitutively active state. Resulting defects in CRL decommissioning prompt widespread CRL auto-degradation that confers resistance to multiple degraders. Collectively, our study informs on regulation and architecture of CRLs amenable for targeted protein degradation and outlines biomarkers and putative resistance mechanisms for upcoming clinical investigation.

AUTACs: Cargo-Specific Degraders Using Selective Autophagy.

Protein silencing represents an essential tool in biomedical research. Targeted protein degradation (TPD) strategies exemplified by PROTACs are rapidly emerging as modalities in drug discovery. However, the scope of current TPD techniques is limited because many intracellular materials are not substrates of proteasomal clearance. Here, we described a novel targeted-clearance strategy (autophagy-targeting chimera [AUTAC]) that contains a degradation tag (guanine derivatives) and a warhead to provide target specificity. As expected from the substrate scope of autophagy, AUTAC degraded fragmented mitochondria as well as proteins. Mitochondria-targeted AUTAC accelerated both the removal of dysfunctional fragmented mitochondria and the biogenesis of functionally normal mitochondria in patient-derived fibroblast cells. Cytoprotective effects against acute mitochondrial injuries were also seen. Canonical autophagy is viewed as a nonselective bulk decomposition system, and none of the available autophagy-inducing agents exhibit useful cargo selectivity. With its target specificity, AUTAC provides a new modality for research on autophagy-based drugs.

Interplay of precision therapeutics and MD study: Calocybe indica's potentials against cervical cancer and its interaction with VEGF via octadecanoic acid.

The evolving landscape of personalized medicine necessitates a shift from traditional therapeutic interventions towards precision-driven approaches. Embracing this paradigm, our research probes the therapeutic efficacy of the aqueous crude extract (ACE) of Calocybe indica in cervical cancer treatment, merging botanical insights with advanced molecular research. We observed that ACE exerts significant influences on nuclear morphology and cell cycle modulation, further inducing early apoptosis and showcasing prebiotic attributes. Characterization of ACE have identified several phytochemicals including significant presence of octadeconoic acid. Simultaneously, utilizing advanced Molecular Dynamics (MD) simulations, we deciphered the intricate molecular interactions between Vascular Endothelial Growth Factor (VEGF) and Octadecanoic acid to establish C.indica's role as an anticancer agent. Our study delineates Octadecanoic acid's potential as a robust binding partner for VEGF, with comprehensive analyses from RMSD and RMSF profiles highlighting the stability and adaptability of the protein-ligand interactions. Further in-depth thermodynamic explorations via MM-GBSA calculations reveal the binding landscape of the VEGF-Octadecanoic acid complex. Emerging therapeutic innovations, encompassing proteolysis-targeting chimeras (PROTACs) and avant-garde nanocarriers, are discussed in the context of their synergy with compounds like Calocybe indica P&C. This convergence underscores the profound therapeutic potential awaiting clinical exploration. This study offers a holistic perspective on the promising therapeutic avenues facilitated by C. indica against cervical cancer, intricately woven with advanced molecular interactions and the prospective integration of precision therapeutics in modern oncology.

Restraining the power of Proteolysis Targeting Chimeras in the cage: A necessary and important refinement for therapeutic safety.

Proteolysis Targeting Chimeras (PROTACs) represent a significant advancement in therapeutic drug development by leveraging the ubiquitin-proteasome system to enable targeted protein degradation, particularly impacting oncology. This review delves into the various types of PROTACs, such as peptide-based, nucleic acid-based, and small molecule PROTACs, each addressing distinct challenges in protein degradation. It also discusses innovative strategies like bridged PROTACs and conditional switch-activated PROTACs, offering precise targeting of previously "undruggable" proteins. The potential of PROTACs extends beyond oncology, with ongoing research and technological advancements needed to maximize their therapeutic potential. Future progress in this field relies on interdisciplinary collaboration and the integration of advanced computational tools to open new treatment avenues across various diseases.

New-generation advanced PROTACs as potential therapeutic agents in cancer therapy.

Proteolysis-targeting chimeras (PROTACs) technology has garnered significant attention over the last 10 years, representing a burgeoning therapeutic approach with the potential to address pathogenic proteins that have historically posed challenges for traditional small-molecule inhibitors. PROTACs exploit the endogenous E3 ubiquitin ligases to facilitate degradation of the proteins of interest (POIs) through the ubiquitin-proteasome system (UPS) in a cyclic catalytic manner. Despite recent endeavors to advance the utilization of PROTACs in clinical settings, the majority of PROTACs fail to progress beyond the preclinical phase of drug development. There are multiple factors impeding the market entry of PROTACs, with the insufficiently precise degradation of favorable POIs standing out as one of the most formidable obstacles. Recently, there has been exploration of new-generation advanced PROTACs, including small-molecule PROTAC prodrugs, biomacromolecule-PROTAC conjugates, and nano-PROTACs, to improve the in vivo efficacy of PROTACs. These improved PROTACs possess the capability to mitigate undesirable physicochemical characteristics inherent in traditional PROTACs, thereby enhancing their targetability and reducing off-target side effects. The new-generation of advanced PROTACs will mark a pivotal turning point in the realm of targeted protein degradation. In this comprehensive review, we have meticulously summarized the state-of-the-art advancements achieved by these cutting-edge PROTACs, elucidated their underlying design principles, deliberated upon the prevailing challenges encountered, and provided an insightful outlook on future prospects within this burgeoning field.

Dual-Programmable Semiconducting Polymer NanoPROTACs for Deep-Tissue Sonodynamic-Ferroptosis Activatable Immunotherapy.

Proteolysis-targeting chimeras (PROTACs) can provide promising opportunities for cancer treatment, while precise regulation of their activities remains challenging to achieve effective and safe therapeutic outcomes. A semiconducting polymer nanoPROTAC (SPNFeP ) is reported that can achieve ultrasound (US) and tumor microenvironment dual-programmable PROTAC activity for deep-tissue sonodynamic-ferroptosis activatable immunotherapy. SPNFeP is formed through a nano-precipitation of a sonodynamic semiconducting polymer, a ferroptosis inducer, and a newly synthesized PROTAC molecule. The semiconducting polymers work as sonosensitizers to produce singlet oxygen (1 O2 ) via sonodynamic effect under US irradiation, and ferroptosis inducers react with intratumoral hydrogen peroxide (H2 O2 ) to generate hydroxyl radical (·OH). Such a dual-programmable reactive oxygen species (ROS) generation not only triggers ferroptosis and immunogenic cell death (ICD), but also induces on-demand activatable delivery of PROTAC molecules into tumor sites. The effectively activated nanoPROTACs degrade nicotinamide phosphoribosyl transferase (NAMPT) to suppress tumor infiltration of myeloid-derived suppressive cells (MDSCs), thus promoting antitumor immunity. In such a way, SPNFeP mediates sonodynamic-ferroptosis activatable immunotherapy for entirely inhibiting tumor growths in both subcutaneous and 2-cm tissue-covered deep tumor mouse models. This study presents a dual-programmable activatable strategy based on PROTACs for effective and precise cancer combinational therapy.

IKZF2 Degradation: It's Time to Take into Account it When Designing Cereblon-Based PROTACs.

Proteolysis-targeting chimera (PROTAC) has become a very important means of protein degradation and a new way of disease treatment. In particular, PROTACs constructed with ligands for E3 ligase cereblon account for more than 90 % of the PROTACs currently in clinical research. Notably, CRBN ligands themselves are a class of molecular glue compounds capable of degrading neo-substrate proteins. Compared to the target proteins degradation, the degradation of neo-substrates, especially IKZF2, has not received enough attention. Therefore, this review summarizes the currently published IKZF2 degraders derived from articles and patents, which are conducive to the design of PROTACs with desired IKZF2 degradation from the perspective of medicinal chemistry.

Design and Evaluation of PROTACs Targeting Acyl Protein Thioesterase 1.

PROTAC linker design remains mostly an empirical task. We employed the PRosettaC computational software in the design of sulfonyl-fluoride-based PROTACs targeting acyl protein thioesterase 1 (APT1). The software efficiently generated ternary complex models from empirically-designed PROTACs and suggested alkyl linkers to be the preferred type of linker to target APT1. Western blotting analysis revealed efficient degradation of APT1 and activity-based protein profiling showed remarkable selectivity of an alkyl linker-based PROTAC amongst serine hydrolases. Collectively, our data suggests that combining PRosettaC and chemoproteomics can effectively assist in triaging PROTACs for synthesis and providing early data on their potency and selectivity.

TAC-tics for Leveraging Proximity Biology in Drug Discovery.

Chemically induced proximity (CIP) refers to co-opting naturally occurring biological pathways using synthetic molecules to recruit neosubstrates that are not normally encountered or to enhance the affinity of naturally occurring interactions. Leveraging proximity biology through CIPs has become a rapidly evolving field and has garnered considerable interest in basic research and drug discovery. PROteolysis TArgeting Chimera (PROTAC) is a well-established CIP modality that induces the proximity between a target protein and an E3 ubiquitin ligase, causing target protein degradation via the ubiquitin-proteasome system. Inspired by PROTACs, several other induced proximity modalities have emerged to modulate both proteins and RNA over recent years. In this review, we summarize the critical advances and opportunities in the field, focusing on protein degraders, RNA degraders and non-degrader modalities such as post-translational modification (PTM) and protein-protein interaction (PPI) modulators. We envision that these emerging proximity-based drug modalities will be valuable resources for both biological research and therapeutic discovery in the future.

Deciphering the mechanism of E3 ubiquitin ligases in plant responses to abiotic and biotic stresses and perspectives on PROTACs for crop resistance.

With global climate change, it is essential to find strategies to make crops more resistant to different stresses and guarantee food security worldwide. E3 ubiquitin ligases are critical regulatory elements that are gaining importance due to their role in selecting proteins for degradation in the ubiquitin-proteasome proteolysis pathway. The role of E3 Ub ligases has been demonstrated in numerous cellular processes in plants responding to biotic and abiotic stresses. E3 Ub ligases are considered a class of proteins that are difficult to control by conventional inhibitors, as they lack a standard active site with pocket, and their biological activity is mainly due to protein-protein interactions with transient conformational changes. Proteolysis-targeted chimeras (PROTACs) are a new class of heterobifunctional molecules that have emerged in recent years as relevant alternatives for incurable human diseases like cancer because they can target recalcitrant proteins for destruction. PROTACs interact with the ubiquitin-proteasome system, principally the E3 Ub ligase in the cell, and facilitate proteasome turnover of the proteins of interest. PROTAC strategies harness the essential functions of E3 Ub ligases for proteasomal degradation of proteins involved in dysfunction. This review examines critical advances in E3 Ub ligase research in plant responses to biotic and abiotic stresses. It highlights how PROTACs can be applied to target proteins involved in plant stress response to mitigate pathogenic agents and environmental adversities.

Design, synthesis, and evaluation of BCL-2 targeting PROTACs.

BCL-2, a member of the BCL-2 protein family, is an antiapoptotic factor that regulates the intrinsic pathway of apoptosis. Due to its aberrant activity, it is frequently implicated in haematopoietic cancers and represents an attractive target for the development of therapeutics that antagonize its activity. A selective BCL-2 inhibitor, venetoclax, was approved for treating chronic lymphocytic leukaemia, acute myeloid leukemia, and other hematologic malignancies, validating BCL-2 as an anticancer target. Since then, alternative therapeutic approaches to modulate the activity of BCL-2 have been explored, such as antibody-drug conjugates and proteolysis-targeting chimeras. Despite numerous research groups focusing on developing degraders of BCL-2 family member proteins, selective BCL-2 PROTACs remain elusive, as disclosed compounds only show dual BCL-xL/BCL-2 degradation. Herein, we report our efforts to develop BCL-2 degraders by incorporating two BCL-2 binding moieties into chimeric compounds that aim to hijack one of three E3 ligases: CRBN, VHL, and IAPs. Even though our project did not result in obtaining a potent and selective BCL-2 PROTAC, our research will aid in understanding the narrow chemical space of BCL-2 degraders.

Discovery of a DCAF11-dependent cyanoacrylamide-containing covalent degrader of BET-proteins.

Targeted protein degradation is mediated by small molecules that induce or stabilize protein-protein interactions between targets and the ubiquitin-proteasome machinery. Currently, there remains a need to expand the repertoire of viable E3 ligases available for hijacking. Notably, covalent chemistry has been employed to engage a handful of E3 ligases, including DCAF11. Here, we disclose a covalent PROTAC that enables DCAF11-dependent degradation, featuring a cyanoacrylamide warhead. Our findings underscore DCAF11 as an interesting candidate with a capacity to accommodate diverse electrophilic chemistries compatible with targeted protein degradation.

Design, synthesis and biological evaluation of new RNF126-based p300/CBP degraders.

Histone acetyltransferase CREB-binding protein (CBP) and its homologous protein p300 are key transcriptional activators that can activate oncogene transcription, which present promising targets for cancer therapy. Here, we designed and synthesized a series of p300/CBP targeted low molecular weight PROTACs by assembling the covalent ligand of RNF126 E3 ubiquitin ligase and the bromodomain ligand of the p300/CBP. The optimal molecule A8 could effectively degrade p300 and CBP through the ubiquitin-proteasome system in time- and concentration-dependent manners, with half-maximal degradation (DC50) concentrations of 208.35/454.35 nM and 82.24/79.45 nM for p300/CBP in MV4-11 and Molm13 cell lines after 72 h of treatment. And the degradation of p300/CBP by A8 is dependent on the ubiquitin-proteasome pathway and its simultaneous interactions with the target proteins and RNF126. A8 exhibits good antiproliferative activity in a series of p300/CBP-dependent cancer cells. It could transcriptionally inhibit the expression of c-Myc, induce cell cycle arrest in the G0/G1 phase and apoptosis in MV4-11 cells. This study thus provided us a new chemotype for the development of drug-like PROTACs targeting p300/CBP, which is expected to be applied in cancer therapy.

The design, synthesis and bioactivity evaluation of novel androgen receptor degraders based on hydrophobic tagging.

Prostate Cancer (PCa) easily progress to metastatic Castration-Resistant Prostate Cancer (mCRPC) that remains a significant cause of cancer-related death. Androgen receptor (AR)-dependent transcription is a major driver of prostate tumor cell proliferation. Proteolysis-targeting chimaera (PROTAC) technology based on Hydrophobic Tagging (HyT) represents an intriguing strategy to regulate the function of therapeutically androgen receptor proteins. In the present study, we have designed, synthesized, and evaluated a series of PROTAC-HyT AR degraders using AR antagonists, RU59063, which were connected with adamantane-based hydrophobic moieties by different alkyl chains. Compound D-4-6 exhibited significant AR protein degradation activity, with a degradation rate of 57 % at 5 µM and nearly 90 % at 20 µM in 24 h, and inhibited the proliferation of LNCaP cells significantly with an IC50 value of 4.77 ± 0.26 µM in a time-concentration-dependent manner. In conclusion, the present study lays the foundation for the development of a completely new class of therapeutic agents for the treatment of mCRPC, and further design and synthesis of AR-targeting degraders are currently in progress for better degradation rate.

Unveiling a new strategy for PDIA1 inhibition: Integration of activity-based probes profiling and targeted degradation.

The overexpression of PDIA1 in cancer has spurred the quest for effective inhibitors. However, existing inhibitors often bind to only one active site, limiting their efficacy. In our study, we developed a PROTAC-mimetic probe dPA by combining PACMA31 (PA) analogs with cereblon-directed pomalidomide. Through protein profiling and analysis, we confirmed dPA's specific interaction with PDIA1's active site cysteines. We further synthesized PROTAC variants with a thiophene ring and various linkers to enhance degradation efficiency. Notably, H4, featuring a PEG linker, induced significant PDIA1 degradation and inhibited cancer cell proliferation similarly to PA. The biosafety profile of H4 is comparable to that of PA, highlighting its potential for further development in cancer therapy. Our findings highlight a novel strategy for PDIA1 inhibition via targeted degradation, offering promising prospects in cancer therapeutics. This approach may overcome limitations of conventional inhibitors, presenting new avenues for advancing anti-cancer interventions.

Discovery of a potent and selective PARP1 degrader promoting cell cycle arrest via intercepting CDC25C-CDK1 axis for treating triple-negative breast cancer.

PARP1 is a multifaceted component of DNA repair and chromatin remodeling, making it an effective therapeutic target for cancer therapy. The recently reported proteolytic targeting chimera (PROTAC) could effectively degrade PARP1 through the ubiquitin-proteasome pathway, expanding the therapeutic application of PARP1 blocking. In this study, a series of nitrogen heterocyclic PROTACs were designed and synthesized through ternary complex simulation analysis based on our previous work. Our efforts have resulted in a potent PARP1 degrader D6 (DC50 = 25.23 nM) with high selectivity due to nitrogen heterocyclic linker generating multiple interactions with the PARP1-CRBN PPI surface, specifically. Moreover, D6 exhibited strong cytotoxicity to triple negative breast cancer cell line MDA-MB-231 (IC50 = 1.04 µM). And the proteomic results showed that the antitumor mechanism of D6 was found that intensifies DNA damage by intercepting the CDC25C-CDK1 axis to halt cell cycle transition in triple-negative breast cancer cells. Furthermore, in vivo study, D6 showed a promising PK property with moderate oral absorption activity. And D6 could effectively inhibit tumor growth (TGI rate = 71.4 % at 40 mg/kg) without other signs of toxicity in MDA-MB-321 tumor-bearing mice. In summary, we have identified an original scaffold and potent PARP1 PROTAC that provided a novel intervention strategy for the treatment of triple-negative breast cancer.

RAS degraders: The new frontier for RAS-driven cancers.

The function and significance of RAS proteins in cancer have been widely studied for decades. In 2013, the National Cancer Institute established the RAS Initiative to explore innovative approaches for attacking the proteins encoded by mutant forms of RAS genes and to create effective therapies for RAS-driven cancers. This initiative spurred researchers to develop novel approaches and to discover small molecules targeting this protein that was at one time termed "undruggable." More recently, advanced efforts in RAS degraders including PROTACs, linker-based degraders, and direct proteolysis degraders have been explored as novel strategies to target RAS for cancer treatment. These RAS degraders present new opportunities for RAS therapies and may prove fruitful in understanding basic cell biology. Novel delivery strategies will further enhance the efficacy of these therapeutics. In this review, we summarize recent efforts to develop RAS degraders, including PROTACs and E3 adaptor and ligase fusions as cancer therapies. This review also details the direct RAS protease degrader, RAS/RAP1-specific endopeptidase that directly and specifically cleaves RAS.