Protein Degradation Research Articles

Preclinical Assessment of the PI3Kα Selective Inhibitor Inavolisib and Prediction of Its Pharmacokinetics and Efficacious Dose in Human.

1. Small molecule inhibitors of the PI3K pathway have been extensively investigated as potential anticancer agents. Among the effectors in this pathway, PI3Kα is the kinase most frequently associated with the development of tumors, through mutations and amplifications of the PIK3CA gene encoding the p110α catalytic subunit.2. Inavolisib (GDC-0077) is a potent and PI3Kα-selective inhibitor that also specifically triggers the degradation of the mutant p110α protein.3. We characterized inavolisib ADME properties in preclinical in vitro and in vivo studies, assessed its efficacy in the PIK3CA mutant KPL-4 breast cancer xenograft model, and predicted its pharmacokinetics and efficacious dose in humans.4. Inavolisib had a moderate permeability (1.9•10-6 cm/s) in MDCK cells and was a P-gp and Bcrp1 substrate. It appeared metabolically stable in hepatocytes incubations from human and preclinical species. The systemic clearance was low in mouse, monkey and dog and high in rat. Oral bioavailability ranged from 57.5% to 100%. Inavolisib was efficacious in the KPL-4 sub-cutaneous xenograft model.5. The PK/PD model parameters estimated from the efficacy study, combined with PBPK model-predicted human PK profiles, projected that a dose of 3 mg could lead to clinical response. Inavolisib is currently being tested in phase 3 trials.

BCKDH kinase promotes hepatic gluconeogenesis independent of BCKDHA

AbstractElevated circulating branched-chain amino acids (BCAAs) are tightly linked to an increased risk in the development of type 2 diabetes mellitus. The rate limiting enzyme of BCAA catabolism branched-chain α-ketoacid dehydrogenase (BCKDH) is phosphorylated at E1α subunit (BCKDHA) by its kinase (BCKDK) and inactivated. Here, the liver-specific BCKDK or BCKDHA knockout mice displayed normal glucose tolerance and insulin sensitivity. However, knockout of BCKDK in the liver inhibited hepatic glucose production as well as the expression of key gluconeogenic enzymes. No abnormal gluconeogenesis was found in mice lacking hepatic BCKDHA. Consistent with the vivo results, BT2-mediated inhibition or genetic knockdown of BCKDK decreased hepatic glucose production and gluconeogenic gene expressions in primary mouse hepatocytes while BCKDK overexpression exhibited an opposite effect. Whereas, gluconeogenic gene expressions were not altered in BCKDHA-silenced hepatocytes. Mechanistically, BT2 treatment attenuated the interaction of cAMP response element binding protein (CREB) with CREB-binding protein and promoted FOXO1 protein degradation by increasing its ubiquitination. Our findings suggest that BCKDK regulates hepatic gluconeogenesis through CREB and FOXO1 signalings, independent of BCKDHA-mediated BCAA catabolism.

MicroRNA-Triggered Programmable DNA-Encoded Pre-PROTACs for Cell-Selective and Controlled Protein Degradation.

Proteolysis-targeting chimeras (PROTACs) have accelerated drug development; however, some challenges still exist owing to their lack of tumor selectivity and on-demand protein degradation. Here, we developed a miRNA-initiated assembled pre-PROTAC (miRiaTAC) platform that enables the on-demand activation and termination of target degradation in a cell type-specific manner. Using miRNA-21 as a model, we engineered DNA hairpins labeled with JQ-1 and pomalidomide and facilitated the modular assembly of DNA-encoded pre-PROTACs through a hybridization chain reaction. This configuration promoted the selective polyubiquitination and degradation of BRD4 upon miR-21 initiation, highlighting significant tumor selectivity and minimal systemic toxicity. Furthermore, the platform incorporates photolabile groups, enabling the precise optical control of pre-PROTACs during DNA assembly/disassembly, mitigating the risk of excessive protein degradation. Additionally, by introducing a secondary ligand targeting CDK6, these pre-PROTACs were used as a modular scaffold for the programmable assembly of active miRiaTACs containing two different warheads in exact stoichiometry, enabling orthogonal multitarget degradation. The integration of near-infrared light-mediated photodynamic therapy through an upconversion nanosystem further enhanced the efficacy of the platform with potent in vivo anticancer activity. We anticipate that miRiaTAC represents a significant intersection between dynamic DNA nanotechnology and PROTAC, potentially expanding the versatility of PROTAC toolkit for cancer therapy.

MicroRNA‐Triggered Programmable DNA‐Encoded Pre‐PROTACs for Cell‐Selective and Controlled Protein Degradation

Proteolysis‐targeting chimeras (PROTACs) have accelerated drug development; however, some challenges still exist owing to their lack of tumor selectivity and on‐demand protein degradation. Here, we developed a miRNA‐initiated assembled pre‐PROTAC (miRiaTAC) platform that enables the on‐demand activation and termination of target degradation in a cell type‐specific manner. Using miRNA‐21 as a model, we engineered DNA hairpins labeled with JQ‐1 and pomalidomide and facilitated the modular assembly of DNA‐encoded pre‐PROTACs through a hybridization chain reaction. This configuration promoted the selective polyubiquitination and degradation of BRD4 upon miR‐21 initiation, highlighting significant tumor selectivity and minimal systemic toxicity. Furthermore, the platform incorporates photolabile groups, enabling the precise optical control of pre‐PROTACs during DNA assembly/disassembly, mitigating the risk of excessive protein degradation. Additionally, by introducing a secondary ligand targeting CDK6, these pre‐PROTACs were used as a modular scaffold for the programmable assembly of active miRiaTACs containing two different warheads in exact stoichiometry, enabling orthogonal multitarget degradation. The integration of near‐infrared light‐mediated photodynamic therapy through an upconversion nanosystem further enhanced the efficacy of the platform with potent in vivo anticancer activity. We anticipate that miRiaTAC represents a significant intersection between dynamic DNA nanotechnology and PROTAC, potentially expanding the versatility of PROTAC toolkit for cancer therapy.

Hepatocyte-derived Igκ promotes HCC progression by stabilizing electron transfer flavoprotein subunit α to facilitate fatty acid β-oxidation

BackgroundLipid metabolism dysregulation is a key characteristic of hepatocellular carcinoma (HCC) onset and progression. Elevated expression of immunoglobulin (Ig), especially the Igκ free light chain with a unique Vκ4-1/Jκ3 rearrangement in cancer cells, is linked to increased malignancy and has been implicated in colon cancer tumorigenesis. However, the role of Igκ in HCC carcinogenesis remains unclear. The aim of this study was to elucidate the pivotal roles of hepatocyte-derived Igκ in HCC development.MethodsThe rearrangement sequence and expression level of hepatocyte-derived Igκ in HCC cells were determined via RT-PCR, Sanger sequencing, immunohistochemistry, and western blot analysis. The function of Igκ in HCC tumorigenesis was assessed by silencing Igκ using siRNA or gRNA in various HCC cell lines. To assess the role of Igκ in HCC pathogenesis in vivo, a mouse model with hepatocyte-specific Igκ knockout and diethylnitrosamine (DEN) and carbon tetrachloride (CCL4)-induced HCC was utilized. The molecular mechanism by which Igκ affects HCC tumorigenesis was investigated through multiomics analyses, quantitative real-time PCR, immunoprecipitation, mass spectrometry, immunofluorescence, and metabolite detection.ResultsWe confirmed that Igκ, especially Vκ4-1/Jκ3-Igκ, is highly expressed in human HCC cells. Igκ depletion inhibited HCC cell proliferation and migration in vitro, and hepatocyte-specific Igκ deficiency ameliorated HCC progression in mice with DEN and CCL4-induced HCC in vivo. Mechanistically, Vκ4-1/Jκ3-Igκ interacts with electron transfer flavoprotein subunit α (ETFA), delaying its protein degradation. Loss of Igκ led to a decrease in the expression of mitochondrial respiratory chain complexes III and IV, resulting in aberrant fatty acid β-oxidation (FAO) and lipid accumulation, which in turn inhibited HCC cell proliferation and migration.ConclusionOur findings indicate that the Igκ/ETFA axis deregulates fatty acid β-oxidation, contributing to HCC progression, which suggests that targeting fatty acid metabolism may be an effective HCC treatment strategy. The results of this study suggest that hepatocyte-derived Vκ4-1/Jκ3-Igκ may serve as a promising therapeutic target for HCC.

Deciphering Hyperammonia-Producing Bacteria (HAB) in the Rumen of Water Buffaloes (Bubalus bubalis) and Their Inhibition through Plant Extracts and Essential Oils

Hyperammonia-producing bacteria (HAB) are a class of microbes present in the stomach of ruminants, responsible for the rapid rate of ammonia production from protein degradation beyond the capacity of these animals for their utilization. Thus, ruminant nutritionists are interested in decreasing ruminal protein degradation and ammonia genesis by focusing on controlling the activity of HAB. The investigations of the present study were carried out to determine predominant hyperammonia-producing bacteria in the rumen of buffaloes, their isolation and characterization, as well as the inhibition of these isolates with various sources of plant secondary compounds (tannins, saponins, and essential oils). Studies employing high-throughput sequencing of amplicons of the 16S rRNA gene from genomic DNA recovered from enrichment culture of HAB of buffalo rumina indicated that, at the phylum level, Proteobacteria (61.1 to 68.2%) was the most predominant HAB. Acidaminococcus was most predominant among the identified genera. In vitro experiments were conducted with enrichment culture of buffalo rumen contents incubated with different types of feed additives such as essential oils (eucalyptus oil, lemon grass oil, and clove oil) and extracts of plants (Sapindus mukorossi fruits and Ficus bengalensis leaves), each at graded dose levels. The reduction in ammonia production by clove and lemon grass oils was evident due to the presence of major bioactive compounds, especially eugenol and limonene, which have strong antimicrobial activity. However, clove oil and Indian soapberry/reetha (Sapindus mukorossi) fruit were found to be promising and effective in reducing the growth, protease production, and ammonia production of HAB culture.

The NEDD4/FLRT2 axis regulates NSCLC cell stemness

IntroductionLung cancer is the leading cause of cancer-related death worldwide. The treatment for lung cancer, particularly for non-small cell lung cancer (NSCLC), remains a clinical challenge. Cancer stem cells are vital for lung cancer development. This study aimed to determine the influence of the neuronally expressed developmentally downregulated 4-fibronectin leucine-rich transmembrane 2 (NEDD4-FLRT2) axis on cancer cell stemness in NSCLC.MethodsFLRT2 expression in NSCLC tissues and stem cells was investigated using western blot and RT-qPCR. The sphere formation assay and the abundance of stemness markers were employed to confirm the stemness of NSCLC stem cells. The CCK-8, colony formation, and Trans-well assays, as well as flow cytometry, were used to determine NSCLC stem cell growth, metastasis, and apoptosis, respectively. The Co-IP assay was used to confirm the binding between NEDD4 and FLRT2. Xenograft tumor mouse models were used to investigate tumorigenesis in vivo.ResultsHere, we reported that FLRT2 expression was reduced in NSCLC tissues, cells, and NSCLC stem cells. FLRT2 upregulation inhibited NSCLC stem cell proliferation, sphere formation, and drug resistance and promoted drug-resistant cell apoptosis. Furthermore, FLRT2 overexpression demonstrated antitumor effects in a xenograft tumor mouse model. Mechanically, FLRT2 was ubiquitinated and degraded by E3 ligase NEDD4. NEDD4 overexpression significantly abolished the inhibitory effects of FLRT2 on NSCLC stemness, as evidenced by in vitro and in vivo experiments. DiscussionThis study revealed that FLRT2 acted as a tumor suppressor by inhibiting cancer cell stemness in NSCLC. NEDD4 promoted ubiquitination degradation of FLRT2 protein. NEDD4 counteracted the inhibitory effects of FLRT2 on NSCLC stem cell tumorigenesis.

A structure-based virtual screening identifies a novel MDM2 antagonist in the activation of the p53 signaling and inhibition of tumor growth.

p53, a tumor suppressor protein, has a vital role in the regulation of the cell cycle, apoptosis, and DNA damage repair. The degradation of p53 is predominantly controlled by the murine double minute 2 (MDM2) protein, a ubiquitin E3 ligase. The overexpression or amplification of MDM2 is commonly observed in various human cancers bearing wild-type p53 alleles, leading to the rapid degradation of the p53 protein and the attenuation of p53 tumor suppression functions. Thus, a major effort in p53-based cancer therapy has been to research MDM2 antagonists that specifically stabilize and activate p53, leading to the suppression of tumor growth. However, despite numerous efforts to develop MDM2 antagonists, to date they have failed to reach clinical use, largely because of the cytotoxicity associated with these small molecules. This study used our newly designed structure-based virtual screening approach on a commercial compound library to identify a novel compound, CGMA-Q18, which directly binds to MDM2, leading to the activation of p53, the induction of apoptosis, and cell cycle arrest in cancer cells. Notably, CGMA-Q18 significantly inhibited tumor xenograft growth in nude mice without observable toxicity. These findings highlight our useful virtual screening protocol and CGMA-Q18 as a putative MDM2 antagonist.

Changes of gluten protein composition during sourdough fermentation in rye flour

AbstractBackground and ObjectivesCeliac disease (CD) is an immune‐mediated condition triggered by gluten consumption. Although more studies stated that gluten proteins are partially degrading during sourdough fermentation, it is still unclear whether the CD toxic epitopes are affected. Our aim was to examine the effect of controlled sourdough fermentation on the gluten content and composition with different sourdough starters in rye flour.FindingsDespite significant changes in protein size distribution and a decrease in secalin types confirmed by high‐performance liquid chromatography methods, the enzyme‐linked immunosorbent assay results showed increased gluten content after fermentation. The incomplete degradation of gluten and the potential resistance of celiac toxic epitopes may explain our results. As part of the R5 antibody binding, epitopes are supposed to be unavailable in the native form of secalins; they might be released and become accessible in the smaller protein fragments.ConclusionsIn comparison of the effect of different starter cultures, the results show that the different microbiological composition of the sourdough samples might be the reason for the differences in the protein degradation.Significance and NoveltyResults highlight the importance of a deeper investigation of the effects of gluten protein degradation on CD toxicity and the standardization of the fermentation process.

CYpHER: catalytic extracellular targeted protein degradation with high potency and durable effect

Many disease-causing proteins have multiple pathogenic mechanisms, and conventional inhibitors struggle to reliably disrupt more than one. Targeted protein degradation (TPD) can eliminate the protein, and thus all its functions, by directing a cell’s protein turnover machinery towards it. Two established strategies either engage catalytic E3 ligases or drive uptake towards the endolysosomal pathway. Here we describe CYpHER (CatalYtic pH-dependent Endolysosomal delivery with Recycling) technology with potency and durability from a catalytic mechanism that shares the specificity and straightforward modular design of endolysosomal uptake. By bestowing pH-dependent release on the target engager and using the rapid-cycling transferrin receptor as the uptake receptor, CYpHER induces endolysosomal delivery of surface and extracellular targets while re-using drug, potentially yielding increased potency and reduced off-target tissue exposure risks. The TfR-based approach allows targeting to tumors that overexpress this receptor and offers the potential for transport to the CNS. CYpHER function was demonstrated in vitro with EGFR and PD-L1, and in vivo with EGFR in a model of EGFR-driven non-small cell lung cancer.

New Approaches to Herbicide and Bioherbicide Discovery

Abstract During the past 30 yr an impasse has developed in the discovery and commercialization of synthetic herbicides with new molecular targets and novel chemistries. Similarly, there has been little success with bioherbicides, both microbial and chemical. These bioherbicides are needed to combat fast-growing herbicide resistance and to fulfill the need for more environmentally and toxicologically safe herbicides. In response to this substantial and growing opportunity, numerous start-up companies are utilizing novel approaches to provide new tools for weed management. These diverse new tools broaden the scope of discovery, encompassing advanced computational, bioinformatic, and imaging platforms; plant genome–editing and targeted protein degradation technologies; and machine learning and artificial intelligence (AI)-based strategies. This review contains summaries of the presentations of 10 such companies that took part in a symposium held at the WSSA annual meeting in 2024. Four of the companies are developing microbial bioherbicides or natural product–based herbicides, and the other six are using advanced technologies, such as AI, to accelerate the discovery of herbicides with novel molecular target sites or to develop non-GMO, herbicide-resistant crops.

Acceleration of Ethanol Metabolism by a Patented Bos taurus Isolated Alcohol Degradation Protein (ADP) on Acute Alcohol Consumption

Alcoholic beverages are among the most widely enjoyed leisure drinks around the world. However, irresponsible drinking habits can have detrimental effects on human health. Therefore, exploring strategies to alleviate discomfort following alcohol consumption would be beneficial for individuals who inevitably need to consume alcohol. In this study, three different models were used to determine the efficacy of a patented alcohol degradation protein (ADP) extracted from Bos taurus on ethanol metabolism. In an ethanol-challenged HepG2 cell model, ADP significantly protected the cell from ethanol-induced toxicity. Subsequently, results demonstrated that ADP significantly alleviated the effect of ethanol, as reflected by the increased distance and activity time of zebrafish during the testing period. Additionally, in a rat model, ADP promoted ethanol degradation at 1 and 2 h after ethanol consumption. Mechanistic studies found that ADP treatment increased ADH and ALDH activity in the gastrointestinal tract. ADP also exhibited potent antioxidation effects by lowering HO-1 expression in the liver. In conclusion, we believe that ADP is a promising product for relieving hangover symptoms after ethanol consumption, with demonstrated safety and effectiveness at the recommended dosage.

Development of folate receptor targeting chimeras for cancer selective degradation of extracellular proteins.

Targeted protein degradation has emerged as a novel therapeutic modality to treat human diseases by utilizing the cell's own disposal systems to remove protein target. Significant clinical benefits have been observed for degrading many intracellular proteins. Recently, the degradation of extracellular proteins in the lysosome has been developed. However, there have been limited successes in selectively degrading protein targets in disease-relevant cells or tissues, which would greatly enhance the development of precision medicine. Additionally, most degraders are not readily available due to their complexity. We report a class of easily accessible Folate Receptor TArgeting Chimeras (FRTACs) to recruit the folate receptor, primarily expressed on malignant cells, to degrade extracellular soluble and membrane cancer-related proteins in vitro and in vivo. Our results indicate that FRTAC is a general platform for developing more precise and effective chemical probes and therapeutics for the study and treatment of cancers.

Chemical inhibition of the integrated stress response impairs the ubiquitin-proteasome system.

Inhibitors of the integrated stress response (ISR) have been used to explore the potential beneficial effects of reducing the activation of this pathway in diseases. As the ISR is in essence a protective response, there is, however, a risk that inhibition may compromise the cell's ability to restore protein homeostasis. Here, we show that the experimental compound ISRIB impairs degradation of proteins by the ubiquitin-proteasome system (UPS) during proteotoxic stress in the cytosolic, but not nuclear, compartment. Accumulation of a UPS reporter substrate that is intercepted by ribosome quality control was comparable to the level observed after blocking the UPS with a proteasome inhibitor. Consistent with impairment of the cytosolic UPS, ISRIB treatment caused an accumulation of polyubiquitylated and detergent insoluble defective ribosome products (DRiPs) in the presence of puromycin. Our data suggest that the persistent protein translation during proteotoxic stress in the absence of a functional ISR increases the pool of DRiPs, thereby hindering the efficient clearance of cytosolic substrates by the UPS.

Advancing Proteolysis Targeting Chimera (PROTAC) Nanotechnology in Protein Homeostasis Reprograming for Disease Treatment.

Proteolysis targeting chimeras (PROTACs) represent a transformative class of therapeutic agents that leverage the intrinsic protein degradation machinery to modulate the hemostasis of key disease-associated proteins selectively. Although several PROTACs have been approved for clinical application, suboptimal therapeutic efficacy and potential adverse side effects remain challenging. Benefiting from the enhanced targeted delivery, reduced systemic toxicity, and improved bioavailability, nanomedicines can be tailored with precision to integrate with PROTACs which hold significant potential to facilitate PROTAC nanomedicines (nano-PROTACs) for clinical translation with enhanced efficacy and reduced side effects. In this review, we provide an overview of the recent progress in the convergence of nanotechnology with PROTAC design, leveraging the inherent properties of nanomaterials, such as lipids, polymers, inorganic nanoparticles, nanohydrogels, proteins, and nucleic acids, for precise PROTAC delivery. Additionally, we discuss the various categories of PROTAC targets and provide insights into their clinical translational potential, alongside the challenges that need to be addressed.

Glucose Transporter-Targeting Chimeras Enabling Tumor-Selective Degradation of Secreted and Membrane Proteins.

Tumor-selective degradation of target proteins has the potential to offer superior therapeutic benefits with maximized therapeutic windows and minimized off-target effects. However, the development of effective lysosome-targeted degradation platforms for achieving selective protein degradation in tumors remains a substantial challenge. Cancer cells depend on certain solute carrier (SLC) transporters to acquire extracellular nutrients to sustain their metabolism and growth. This current study exploits facilitative glucose transporters (GLUTs), a group of SLC transporters widely overexpressed in numerous types of cancer, to drive the endocytosis and lysosomal degradation of target proteins in tumor cells. GLUT-targeting chimeras (GTACs) were generated by conjugating multiple glucose ligands to an antibody specific for the target protein. We demonstrate that the constructed GTACs can induce the internalization and lysosomal degradation of the extracellular and membrane proteins streptavidin, tumor necrosis factor-alpha (TNF-α), and human epidermal growth factor receptor 2 (HER2). Compared with the parent antibody, the GTAC exhibited higher potency in inhibiting the growth of tumor cells in vitro and enhanced tumor-targeting capacity in a tumor-bearing mouse model. Thus, the GTAC platform represents a novel degradation strategy that harnesses an SLC transporter for tumor-selective depletion of secreted and membrane proteins of interest.

Defining Alzheimer's Disease through Proteomic CSF Profiling.

Alzheimer disease (AD) is the main cause of dementia, and its complexity is not yet completely understood. Proteomic profiles can provide useful information to explore the pathways involved and the heterogeneity among AD patients. A proteomic analysis was performed in cerebrospinal fluid (CSF) samples from mild cognitive impairment due to AD (MCI-AD) and control individuals; both groups were classified by amyloid β42/amyloid β40 levels in CSF (data available in BioStudies database (S-BSST1456)). The analysis based on PLS regression and volcano plot identified 7 proteins (FOLR2, PPP3CA, SMOC2, STMN1, TAGLN3, TMEM132B, and UCHL1) mainly related to protein phosphorylation, structure maintenance, inflammation, and protein degradation. Enrichment analysis revealed the involvement of different biological processes related to neuronal mechanisms and synapses, lipid and carbohydrate metabolism, immune system and inflammation, vascular, hormones, and response to stimuli, and cell signaling and adhesion. In addition, the proteomic profile showed some association with the levels of AD biomarkers in CSF. Regarding the subtypes, two MCI-AD subgroups were identified: one could be related to synapsis and neuronal functions and the other to innate immunity. The study of the proteomic profile in the CSF of AD patients reflects the heterogeneity of biochemical pathways involved in AD.

Discovery of the First Clinical Protein Degrader for the Treatment of Autoimmune Indications: Orally Bioavailable and Selective IRAK4 Degrader KT-474.

IRAK4 inhibitors have been sought for the treatment of a host of diseases, however, recent evidence suggests a protein degradation approach might have advantages over an inhibitor. This viewpoint summarizes the discovery of KT-474─a selective and orally bioavailable interleuken receptor-associated kinase 4 proteolysis-targeting chimera in Phase 2 clinical trials for autoimmune indications.

DNA Nanotechnology for Application in Targeted Protein Degradation.

DNA is a kind of flexible and versatile biomaterial for constructing nanostructures and nanodevices. Due to high biocompatibility and programmability and easy modification and fabrication, DNA nanotechnology has emerged as a powerful tool for application in intracellular targeted protein degradation. In this review, we summarize the recent advances in the design and mechanism of targeted protein degradation technologies such as protein hydrolysis targeted chimeras, lysosomal targeted chimeras, and autophagy based protein degradation. Subsequently, we introduce the DNA nanotechnologies of DNA cascade circuits, DNA nanostructures, and dynamic machines. Moreover, we present the latest developments in DNA nanotechnologies in targeted protein degradation. Finally, the vision and challenges are discussed.

Precise Modulation of Protein Degradation by Smart PROTACs.

Proteolysis-targeting chimera (PROTAC)hasemergedas an attractive therapeutic modalityin drug discovery.PROTACs are bifunctional moleculesthat effectivelybridge proteins of interest(POIs)withE3 ubiquitin ligases, such that,the target proteinsaretagged with ubiquitin and subsequently degraded viathe proteasome. Despite significant progressin the field of targeted protein degradation (TPD), the application of conventional PROTAC degradersstill faces significantchallenges,includingsystemic toxicity induced by non-tissue-specific targeting.To address this issue, a variety ofsmart PROTACs that can be activated byspecific stimuli, have been developed for achievingconditional andspatiotemporal modulationof protein levels.Here, on the basis of our contributions, we overview recent advances of smart PROTACs, including tumor microenvironment-, photo-, and X-ray radiation-responsivePROTACs, that enable controllableTPD. The design strategy, case studies, potential applications and challengeswill be focused on.