mRNA PROTACs: engineering PROTACs for high-efficiency targeted protein degradation.

Proteolysis-targeting chimeras (PROTACs) are essential bifunctional molecules that target proteins of interest (POIs) for degradation by cellular ubiquitination machinery. Despite significant progress made in understanding PROTACs' functions, their therapeutic potential remains largely untapped. As a result of the success of highly flexible, scalable, and low-cost mRNA therapies, as well as the advantages of the first generation of peptide PROTACs (p-PROTACs), we present for the first time an engineering mRNA PROTACs (m-PROTACs) strategy. This design combines von Hippel-Lindau (VHL) recruiting peptide encoding mRNA and POI-binding peptide encoding mRNA to form m-PROTAC and promote cellular POI degradation. We then performed proof-of-concept experiments using two m-PROTACs targeting two cancer-related proteins, estrogen receptor alpha and B-cell lymphoma-extra large protein. Our results demonstrated that m-PROTACs could successfully degrade the POIs in different cell lines and more effectively inhibit cell proliferation than the traditional p-PROTACs. Moreover, the in vivo experiment demonstrated that m-PROTAC led to significant tumor regression in the 4T1 mouse xenograft model. This finding highlights the enormous potential of m-PROTAC as a promising approach for targeted protein degradation therapy.

Universal smartphone-assisted label-free CRISPR/Cas12a-DNAzyme chemiluminescence biosensing platform for on-site detection of nucleic acid and non-nucleic acid targets.

The clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein (Cas) (CRISPR/Cas) system enables sensitive and specific detection of biomolecules, thanks to its programmability, high fidelity, and powerful signal amplification capabilities. Herein, a universal smartphone-assisted label-free G-quadruplex (G4) DNAzyme-based chemiluminescence CRISPR/Cas12a biosensing platform (G4CLCas) is firstly described that achieves on-site, ultrasensitive visual detection of nucleic acid and non-nucleic acid targets. The G4CLCas-based sensing platform relies on Cas12a trans-cleavage activation that triggers the cleavage of the G4 DNAzyme, resulting in chemiluminescence signals off/on compared to that of the control. Chemiluminescence signals are captured as images that are quantitatively analyzed and visualized using a smartphone-assisted imaging cartridge. Under optimal conditions, G4CLCas achieves a low limit of detection (LOD) of 8.6 aM (∼5.2 copies/µL) for monkeypox virus (MPXV) DNA within the linear concentration range of 10-300 aM and can accurately quantify viral DNA in spiked samples. G4CLCas can also detect non-nucleic acid targets, whereby it achieves a low LOD value of 84.3 nM for adenosine triphosphate (ATP) within the linear concentration range of 2-2000 µM. Here, a label-free, portable, on-site CRISPR/Cas12a chemiluminescence biosensing platform based on the G4 DNAzyme substrates is proposed with potential applications in clinical detection and bioanalytical chemistry research.

Conjugated Polyelectrolyte/Single Strand DNA Hybrid Polyplexes for Efficient Nucleic Acid Delivery and Targeted Protein Degradation.

Nucleic acid-based therapeutics have gained increasing attention due to their ability to regulate various genetic disorders. However, the safe and effective delivery of nucleic acids to their intended cellular sites remains a challenge, primarily due to poor cell membrane permeation and low in vivo stability. Limitations associated with the commonly used nucleic acid delivering agent viral vectors such as carcinogenesis and immunogenicity have driven scientists to develop various nonviral vectors. In this study, we present a highly efficient nucleic acid delivery system based on cationic conjugated polyelectrolytes and single-strand DNA polyplexes with further application in efficient ubiquitin-regulated targeting protein degradation. These polyplexes, formed by 9TC, an aptamer sequence for estrogen receptor (ERα), and cationic PPET3N2 through electrostatic and hydrophobic interactions, demonstrate improved cellular uptake efficiency as well as enhanced stability against nuclease degradation. Furthermore, by incorporation of 9TC into a proteolysis targeting chimera (PROTAC) molecule (P9TC), PPET3N2/P9TC polyplexes significantly enhance the target protein ERα degradation efficiency. Collectively, our findings suggest that PPET3N2 provides a versatile, low cytotoxicity platform for safe, efficient, and simplified delivery of nucleic acids.

Recent Advances in Optically Controlled PROTAC.

Proteolysis-targeting chimera (PROTAC) technology is a groundbreaking therapeutic approach with significant clinical potential for degrading disease-inducing proteins within targeted cells. However, challenges related to insufficient target selectivity raise concerns about PROTAC toxicity toward normal cells. To address this issue, researchers are modifying PROTACs using various approaches to enhance their target specificity. This review highlights innovative optically controlled PROTACs as anti-cancer therapies currently used in clinical practice and explores the challenges associated with their efficacy and safety. The development of optically controlled PROTACs holds the potential to significantly expand the clinical applicability of PROTAC-based technology within the realm of drug discovery.

A universal platform for one-pot detection of circulating non-coding RNA combining CRISPR-Cas12a and branched rolling circle amplification.

Multiple circulating non-coding RNAs (ncRNAs) in serum may serve as vital biomarkers for use in diagnosing early-stage colorectal cancer (CRC). Herein, a universal platform for one-pot detection of CRC-related ncRNAs was developed based on branched rolling circle amplification and CRISPR-Cas12a (BRCACas). For the implementation of the method, primers incorporating ncRNA sequences of circulating CRC-associated RNAs (piRNA or miRNA) were designed that could specifically hybridize with circular probes to initiate the BRCA process. Thereafter, the generation of dendritic DNA products triggered Cas12a trans-cleavage activity to elicit a fluorescent signal. The proposed method, combining high BRCA reaction efficiency with powerful Cas12a trans-cleavage activity, provided greatly enhanced detection sensitivity, as reflected by limits of detection (LODs) for model piRNA (piR-54265) and model miRNA (miR21) of 0.76 fM and 0.87 fM, respectively. Notably, the proposed BRCACas platform, assaying two different types of CRC-associated ncRNAs in patient samples, produced consistent results with the conventional reverse transcription-quantitative PCR (RT-qPCR) method. Therefore, the one-pot, isothermal, and specific BRCACas platform provided excellent performance, thus demonstrating its promise as a rapid, adaptable, and practical diagnostic/prognostic cancer screening method.

PROTAC Degrader of Estrogen Receptor α Targeting DNA-Binding Domain in Breast Cancer.

PROteolysis-TArgeting Chimeras (PROTACs) are a powerful class of drugs that selectively degrade the proteins of interest (POIs) through cellular ubiquitination mechanisms. Estrogen receptor α (ERα) plays a vital role in the pathogenesis and treatment of breast cancer. In this work, the DNA-binding domain (DBD) of ERα was selected as the target to avoid drug resistance caused by the ligand-binding domain (LBD) of ERα. The estrogen response element (ERE), a natural DNA sequence binding with DBD of ERα, was chosen as a recognized unit of PROTAC. Therefore, we designed a nucleic acid-conjugated PROTAC, ERE-PROTAC, via a click reaction, in which the ERE sequence recruits ERα and the typical small molecule VH032 recruits the von Hippel-Lindau (VHL) E3 ligase. The proposed ERE-PROTAC showed to efficiently and reversibly degrade ERα in different breast cancer cells by targeting the DBD, indicating its potential to overcome the current resistance caused by LBD mutations.

Molecular Design and Photothermal Application of Thienoisoindigo Dyes with Aggregation-Induced Emission.

Organic fluorescent dyes with aggregation-induced emission (AIE) property have an extensive application range, especially in the fields of imaging, labeling, and adjusting microprocesses in aggregated environments. In particular, the thienoisoindigo skeleton, which exhibits an outstanding electron-withdrawing capacity in optoelectronic materials, has been defined as a promising AIE candidate. For instance, by installing AIE blocks or other rotatable groups at two terminal sites, such as various arylamine groups, thienoisoindigo derivatives can be efficiently turned to be functional AIE structures. In this work, a thienoisoindigo derivative with AIE characteristics, namely, TII-TPE, was developed. This AIE system was expanded by linking typical AIE fragments, namely, tetraphenylethene, with the proposed thienoisoindigo derivative, which exhibited typical AIE fluorescence in the 600-850 nm range and maintained high photostability. Then, employing the reported derivative TII-TPA coating thienoisoindigo and triphenylamine as a contrast, aggregated TII-TPE and TII-TPA nanoparticles were prepared and demonstrated photothermal conversion efficiencies of 36.2 and 35.6%, respectively. Moreover, both nanoparticles were evaluated as photothermal therapeutic (PTT) agents in a tumor mouse model, which showed to significantly inhibit tumor growth after four treatment cycles in vivo. This work not only presents an enriched thienoisoindigo system but also provides a pattern for subsequent construction of functional AIE molecules.

Fluorescence Analysis of Circulating Exosomes for Breast Cancer Diagnosis Using a Sensor Array and Deep Learning.

Emerging liquid biopsy methods for investigating biomarkers in bodily fluids such as blood, saliva, or urine can be used to perform noninvasive cancer detection. However, the complexity and heterogeneity of exosomes require improved methods to achieve the desired sensitivity and accuracy. Herein, we report our study on developing a breast cancer liquid biopsy system, including a fluorescence sensor array and deep learning (DL) tool AggMapNet. In particular, we used a 12-unit sensor array composed of conjugated polyelectrolytes, fluorophore-labeled peptides, and monosaccharides or glycans to collect fluorescence signals from cells and exosomes. Linear discriminant analysis (LDA) processed the fluorescence spectral data of cells and cell-derived exosomes, demonstrating successful discrimination between normal and different cancerous cells and 100% accurate classification of different BC cells. For heterogeneous plasma-derived exosome analysis, CNN-based DL tool AggMapNet was applied to transform the unordered fluorescence spectra into feature maps (Fmaps), which gave a straightforward visual demonstration of the difference between healthy donors and BC patients with 100% prediction accuracy. Our work indicates that our fluorescent sensor array and DL model can be used as a promising noninvasive method for BC diagnosis.

Design, synthesis and evaluation of novel ErbB/HDAC multitargeted inhibitors with selectivity in EGFRT790M mutant cell lines.

Acquired resistance leads to the failure of EGFR TKIs in NSCLC treatment. A novel series of hydroxamic acid-containing 4-aminoquinazoline derivatives as irreversible ErbB/HDAC multitargeted inhibitors for NSCLC therapy had been designed and synthesized, which displayed weak anti-proliferative activity in several EGFR wild-type cancer cell lines (NCI-H838, SK-BR-3, A549, A431) yet retained moderate activity to EGFRT790M resistance mutation harboring NCI-H1975 cells. The mechanistic studies revealed that the representative compound 11e was able to inhibit the phosphorylation of EGFR, up-regulate hyperacetylation of histone H3 and even reduce the expression of EGFR and Akt in NCI-H1975 cells. In further assays, compound 11e also showed moderate anti-proliferative activity in other EGFRT790M harboring tumor cell lines (NCI-H820, Ba/F3_EGFR_Del19-T790M-C797S) and low toxicities in normal cell lines (HL-7702, FHC). This selectivity of designed multitargeted compounds could serve as a potential strategy to circumvent multiple mechanisms of acquired resistance to EGFR-targeted therapy without severe toxicities and side effects resulting from broad inhibition.

Conjugated Polymer Nanoparticles Based on Copper Coordination for Real-Time Monitoring of pH-Responsive Drug Delivery.

Metal coordination-driven composite systems have excellent stability and pH-responsive ability, making them suitable for specific drug delivery in physiological conditions. In this study, an anionic conjugated polymer PPEIDA with a poly(p-phenylene ethynylene) backbone and iminodiacetic acid (IDA) side chains is used as a drug carrier to construct a class of pH-responsive nanoparticles, PPEIDA-Cu-DOX conjugated polymer nanoparticles (CPNs), by taking advantage of the metal coordination interaction of Cu2+ with PPEIDA and the drug doxorubicin (DOX). PPEIDA-Cu-DOX CPNs have high drug loading and encapsulation efficiency (EE), calculated to be 54.30 ± 1.10 and 95.80 ± 0.84%, respectively. Due to the good spectral overlap, Förster resonance energy transfer (FRET) takes place between PPEIDA and the drug DOX, which enables the observation of the loading and the release of DOX from CPNs in an acidic environment by monitoring fluorescence emission changes. Therefore, PPEIDA-Cu-DOX CPNs can also be used in real-time cell imaging to monitor drug release in addition to delivering DOX targeting tumor cells. Compared with free DOX, PPEIDA-Cu-DOX CPNs show a similar inhibition to tumor cells and lower toxicity to normal cells. Our results demonstrate the feasibility and potential of constructing pH-responsive CPNs via metal-ligand coordination interactions for cancer treatment.

Biological Sensing and Imaging Using Conjugated Polymers and Peptide Substrates.

Peptides have been widely applied as targeting elements or enzyme-substrates in biological sensing and imaging. Conjugated Polymers (CPs) have emerged as a novel biosensing material and received considerable attention due to their excellent light absorption, strong fluorescence emission, as well as amplified quenching properties. In this review, we summarize the recent advances of using CPs and peptide substrates in biosensing and bioimaging. After a brief introduction of the advantages of CPs and peptide substrates, different sensing designs and mechanisms are discussed based on peptides' structures and functions, including targeting recognition elements, enzyme-substrates, and cell-penetrating elements. Applications of CPs and peptides in fluorescent imaging and Raman imaging in living cells are subsequently reviewed.

Highly Selective Oxidation of Organic Sulfides by a Conjugated Polymer as the Photosensitizer for Singlet Oxygen Generation.

A cationic conjugated polyelectrolyte PPET3-N2 was used as a photosensitizer for photocatalytic oxidation of organic sulfides, including thioanisole, ethyl phenyl sulfide, 4-methylphenyl methyl sulfide, etc., to form sulfoxides with good yields and high selectivity. Oxidation reactions were performed in both batch and microfluidic reactors, where the microfluidic reactor can significantly promote the conversion of photocatalytic oxidation reaction to over 98% in about 8 min. Further studies of the photocatalytic oxidation of the antitumor drug ricobendazole in the microfluidic reactor demonstrate the potential application of the polymer material in organic reactions given its high selectivity, good efficiency, and operation convenience.

The efficacy and safety of panitumumab supplementation for colorectal cancer: A meta-analysis of randomized controlled studies.

BACKGROUND: The efficacy of panitumumab supplementation for colorectal cancer remains controversial. We conduct a systematic review and meta-analysis to explore the influence of panitumumab supplementation on treatment efficacy of colorectal cancer. METHODS: We search PubMed, EMbase, Web of science, EBSCO, and Cochrane library databases through June 2019 for randomized controlled trials (RCTs) assessing the efficacy of panitumumab supplementation for colorectal cancer. This meta-analysis is performed using the random-effect model. RESULTS: Five RCTs are included in the meta-analysis. Overall, compared with control group for colorectal cancer, panitumumab supplementation is associated with the increase in objective response for wild-type (WT) KRAS (RR = 1.70; 95% CI = 1.07-2.69; P = .03), but has no remarkable influence on objective response for mutant KRAS (RR = 0.92; 95% CI = 0.79-1.08; P = .32), objective response (RR = 1.35; 95% CI = 1.00-1.83; P = 0.05), progressive disease for WT KRAS (RR = 0.94; 95% CI = 0.85-1.02; P = .15), mortality (RR = 0.86; 95% CI = 0.69-1.08; P = .20), or mortality for WT KRAS (RR = 0.94; 95% CI = 0.84-1.05; P = .28). In addition, grade 3 and 4 adverse events are found to be higher in panitumumab group than those in control group (RR = 1.17; 95% CI = 1.08-1.27; P = .0001; ). CONCLUSIONS: Panitumumab supplementation can provide some improvement in objective response for colorectal cancer patients with WT KRAS, but results in the increase in grade 3 and 4 adverse events.

Magnetic bead-gold nanoparticle hybrids probe based on optically countable gold nanoparticles with dark-field microscope for T4 polynucleotide kinase activity assay.

T4 polynucleotide kinase (T4 PNK) plays an essential role in DNA phosphorylation during the DNA repair process. Therefore, the sensitive, selective and convenient detection of T4 PNK activity is of great significance. In this work, we proposed a sensitive non-amplification strategy for the sensing of T4 PNK activity via dark field microscope (DFM) based on magnetic bead (MB)-gold nanoparticle (AuNP) hybrids probe, MB-dsDNA-AuNP (MDA). In the presence of T4 PNK, the 5'-OH termini of DNA are phosphorylated and cleaved into oligonucleotides by lambda exonuclease (λexo), resulting in the destruction of the MDA probe and the separation of AuNP from the MB. Through automatic counting of AuNPs from DFM images, T4 PNK activity can be quantitatively measured. This strategy revealed a limit of detection (LOD) as low as 0.0058 U/mL and exhibited a dynamic range from 0.01 to 1 U/mL. The strategy presents an excellent ability to discriminate T4 PNK from the other proteins and enzymes. Notably, this strategy was applied to screen the T4 PNK inhibitors and test the activity in cell lysates, showing great potential for the discovery of new anticancer drugs and other related research field.

Gadolinium-based contrast agents: Stimulators of myeloid-induced renal fibrosis and major metabolic disruptors.

Evidence for gadolinium-based contrast agent- (GBCA-) induced disease continues to mount. Risk factors for gadolinium-induced systemic fibrosis are entirely unexplored. Obesity-related renal injury is characterized by activation of glomerular mesangial cells and podocyte damage with alteration of lipid metabolism/lipid accumulation in both cell types resulting in matrix accumulation and eventual progression to glomerulosclerosis. We examined the consequences of GBCA treatment in the kidneys from mice with normal kidney function and the potential interplay between obesity and gadolinium exposure. We found that administration of GBCA (4 weeks) causes significant renal fibrosis and podocyte injury that are associated with metabolic disorders as evidenced by dyslipidemia. Metabolomic analysis demonstrated that renal lipid metabolism and metabolic markers of collagen turnover are significantly altered by gadolinium. GBCA stimulates myeloid-derived fibrocytes to the kidney. Obesity was induced by feeding a group of mice a high fat diet (HFD) for 22 weeks. Groups were sub-randomized to GBCA treatment versus none for 4 weeks before sacrifice. HFD-induced fibrosis and podocyte injury were worsened by GBCA. Similarly, HFD-mediated hyperlipidemia and lipid metabolites were exacerbated by gadolinium. This is the first evidence that GBCA causes significant metabolic disorders and kidney injury in mice without renal insufficiency and that the injurious actions of GBCA are amplified by obesity. The understanding of the functional interplay between gadolinium and obesity will allow the development of therapeutic interventions or the establishment of effective preventive measures to reduce gadolinium- and obesity-mediated renal pathologies.

Nephrogenic Systemic Fibrosis Is Mediated by Myeloid C-C Chemokine Receptor 2.

Gadolinium-based contrast agents are implicated in several pathologic abnormalities (long-term retention in vital organs such as the skin and the brain) and are the cause of a sometimes fatal condition in patients, nephrogenic systemic fibrosis. Bone marrow-derived fibrocytes and the monocyte chemoattractant protein-1 inflammatory pathway have been implicated as mediators of the adverse effects induced by gadolinium-based contrast agents. Mechanistic studies are scant; therefore, a mouse model of nephrogenic systemic fibrosis was established. Dermal cellularity was increased in contrast-treated green fluorescent protein (GFP) chimeric mice. GFP in the skin and fibrosis were increased in the contrast-treated chimeric animals. Monocyte chemoattractant protein-1 and C-C chemokine receptor 2 were increased in the tissues from contrast-treated mice. C-C chemokine receptor 2-deficient recipients of GFP-expressing marrow had an abrogation of gadolinium-induced pathology and displayed less GFP-positive cells in the skin. Wild-type animals that received C-C chemokine receptor 2-deficient bone marrow had a complete abrogation of dermal pathology. That GFP levels and expression increase in the skin, in tandem with a fibrocyte marker, supports the blood-borne circulating fibrocyte hypothesis of the disease. As of now, fibrocyte trafficking has yet to be demonstrated. Importantly, our data demonstrate that the monocyte chemoattractant protein-1/C-C chemokine receptor 2 axis plays a critical role in the pathogenesis of nephrogenic systemic fibrosis.