Near-infrared active ferrocenyl porous organic polymer with photothermal enhanced enzymatic activity for combination antibacterial application.

As a severe ongoing global problem, bacterial contamination exists in every aspect of human life and the search for new antibacterial agents is urgently needed. Herein, a ferrocenyl porous organic polymer (FMC-POP) broad-spectrum antibacterial agent based on synergistic photothermal and peroxidase-like activity was prepared in a facile manner via the copolymerization of ferrocene diformaldehyde and cinnamaldehyde with mannitol through the acid-responsive acetal bond. The photoactive FMC-POP, with high photothermal conversion efficiency (41.45%), could convert not only the near-infrared laser irradiation into local heat to eradicate bacteria, but also low-concentration H2O2 into radical oxygen species (˙OH) that are effective against bacteria. Compared with single-mode photothermal (PTT) and enzymatic therapies, this combination therapy could significantly improve the bactericidal effect, exhibiting a germicidal efficiency of up to 99% (vs. 80.42% for PTT and 70% for enzyme). Thus, our work paves the way for a synergistic non-invasive antimicrobial therapy, which could expand the applications of POP-based artificial enzymes in biomedicine.

Relatlimab: a novel drug targeting immune checkpoint LAG-3 in melanoma therapy.

Relatlimab is a type of human immunoglobulin G4 monoclonal blocking antibody. It is the world's first Lymphocyte-Activation Gene-3 (LAG-3) inhibitor and the third immune checkpoint inhibitor with clinical application, following PD-1 and CTLA-4. Relatlimab can bind to the LAG-3 receptor which blocks the interaction between LAG-3 and its ligand to reduce LAG-3 pathway-mediated immunosuppression and promote T-cell proliferation, inducing tumor cell death. On 18 March 2022, the U.S. FDA approved the fixed-dose combination of relatlimab developed by Bristol Myers Squibb with nivolumab, under the brand name Opdualag for the treatment of unresectable or metastatic melanoma in adult and pediatric patients aged 12 and older. This study comprehensively describes the mechanism of action and clinical trials of relatlimab and a brief overview of immune checkpoint drugs currently used for the treatment of melanoma.

Geared Toward Applications: A Perspective on Functional Sequence-Controlled Polymers.

Sequence-controlled polymers are an emerging class of synthetic polymers with a regulated sequence of monomers. In the past decade, tremendous progress has been made in the synthesis of polymers with the sophisticated sequence control approaching the level manifested in biopolymers. In contrast, the exploration of novel functions that can be achieved by controlling synthetic polymer sequences represents an emerging focus in polymer science. This Viewpoint will survey recent advances in the functional applications of sequence-controlled polymers and provide a perspective on the challenges and outlook for pursuing future applications of this fascinating class of macromolecules.

Metathesis Cascade-Triggered Depolymerization of Enyne Self-Immolative Polymers*.

A novel class of enyne self-immolative polymers (SIPs) capable of metathesis cascade-triggered depolymerization is reported. Studies on model compounds established 1,6-enyne structures for efficient metathesis cascade reactions. SIPs incorporating the optimized 1,6-enyne motif were prepared via both polycondensation and iterative exponential growth approaches. These SIPs demonstrated excellent stability in strong acid, base, nucleophiles, or at elevated temperatures, and can undergo efficient and complete depolymerization once triggered by a metathesis catalyst. Further studies revealed that introducing a terminal alkene to the chain end of the enyne SIPs improved the depolymerization efficiency, and established their potential as stimuli-responsive materials.

Antibiotic use in pig farming and its associated factors in L County in Yunnan, China.

China has a long history of pig rearing, and it currently raises and consumes approximately half of the pigs in the world. Major improvements have been made in pig farming in China in the last four decades with the growing application of new livestock farming technologies. Among the new improvements, the use of antibiotics in pig farming is a common but not well-documented practise. In order to understand the behaviour of the farmers regarding antibiotic use in pig farming, we conducted a household survey in four townships of L County in Yunnan Province, China, during August 2014 and April 2015. In this survey, 404 farmer households were interviewed using a questionnaire. Among the farmers interviewed, 89% reported easy access to antibiotics, 83.7% reported experience of self-purchasing antibiotics, and 40.3% reported that they often used antibiotics in pig farming mainly for the prevention and treatment of pig diseases. These farmers identified 20 antibiotics that they had used in pig farming 6 months before the survey. Of these, 11 and 8 antibiotics have been categorised under 'critically important' and 'highly important' antimicrobial groups, respectively, by the World Health Organization (WHO), and 12 and 8 have been categorised under the 'Watch' and 'Access' groups, respectively, as per the 2019 WHO AWaRe classification of antibiotics. Factors associated with the behaviour of self-purchasing antibiotics included types of farms, sources of antibiotics, and previous experiences of pig diseases: those who were smallholders, buying antibiotics from veterinary drugstores and village vets, and whose pigs had suffered diseases previously were more likely to self-purchase antibiotics for their pigs. Farmers who cleaned their pigsties less frequently and those whose pigs had suffered from diseases used antibiotics more frequently as compared to their peer farmers.

4-Hydroxyproline-Derived Sustainable Polythioesters: Controlled Ring-Opening Polymerization, Complete Recyclability, and Facile Functionalization.

The sustainable production of chemically recyclable polymers presents a significant opportunity to polymer scientists to tackle the growing environmental and energy problems of current petroleum-based plastics. Despite recent advances, however, there are still pressing needs for an expanded horizon of chemically recyclable polymers. Herein, we introduce a new paradigm of biosourced polythioesters (PTEs) with high polymerizability and complete recyclability under mild and economical conditions. The thiolactone monomers with a high ring strain can be easily prepared in a two-step process from 4-hydroxyproline. Controlled ring-opening polymerizations (ROP) using inexpensive and weak bases afford PTEs with high molar masses ( Mn) up to 259 kg mol-1 and narrow dispersities generally below 1.15. The properties of PTEs can be readily adjusted by copolymerization and/or pre/post-functionalization on the side chains. Selective and complete depolymerizations of the PTEs in dilute solution at ambient to modest temperatures recycle clean monomers. Density functional theory (DFT) calculation of model reactions provides mechanistic insights and highlights the importance of judicious molecular design. Taken together, the unique ROP/depolymerization chemistry of such PTEs may offer a sustainable solution for creating and manufacturing high-value materials such as optical/photochemical plastics, self-immolative polymers, and degradable biomaterials under situations where recycle and reuse are indispensable.

Therapeutic Protein PEPylation: The Helix of Nonfouling Synthetic Polypeptides Minimizes Antidrug Antibody Generation.

Polymer conjugation is a clinically proven approach to generate long acting protein drugs with decreased immune responses. Although poly(ethylene glycol) (PEG) is one of the most commonly used conjugation partners due to its unstructured conformation, its therapeutic application is limited by its poor biodegradability, propensity to induce an anti-PEG immune response, and the resultant accelerated blood clearance (ABC) effect. Moreover, the prevailing preference of unstructured polymers for protein conjugation still lacks strong animal data support with appropriate control reagents. By using two biodegradable synthetic polypeptides with similar structural compositions (l-P(EG3Glu) and dl-P(EG3Glu)) for site-specific protein modification, in the current study, we systematically investigate the effect of the polymer conformation on the in vivo pharmacological performances of the resulting conjugates. Our results reveal that the conjugate l20K-IFN, interferon (IFN) modified with the helical polypeptide l-P(EG3Glu) shows improved binding affinity, in vitro antiproliferative activity, and in vivo efficacy compared to those modified with the unstructured polypeptide analogue dl-P(EG3Glu) or PEG. Moreover, l20K-IFN triggered significantly less antidrug and antipolymer antibodies than the other two. Importantly, the unusual findings observed in the IFN series are reproduced in a human growth hormone (GH) conjugate series. Subcutaneously infused l20K-GH, GH modified with l-P(EG3Glu), evokes considerably less anti-GH and antipolymer antibodies compared to those modified with dl-P(EG3Glu) or PEG (dl20K-GH or PEG20K-GH). As a result, repeated injections of dl20K-GH or PEG20K-GH, but not l20K-GH, result in a clear ABC effect and significantly diminished drug availability in the blood. Meanwhile, immature mouse bone marrow cells incubated with the helical l20K-GH exhibit decreased drug uptake and secretion of proinflammatory cytokines compared to those treated with one of the other two GH conjugates bearing unstructured polymers. Taken together, the current study highlights an urgent necessity to systematically reassess the pros and cons of choosing unstructured polymers for protein conjugation. Furthermore, our results also lay the foundation for the development of next-generation biohybrid drugs based on helical synthetic polypeptides.

Mitosis-specific MRN complex promotes a mitotic signaling cascade to regulate spindle dynamics and chromosome segregation.

The MRE11-RAD50-NBS1 (MRN) complex is well known for participating in DNA damage response pathways in all phases of cell cycle. Here, we show that MRN constitutes a mitosis-specific complex, named mMRN, with a protein, MMAP. MMAP directly interacts with MRE11 and is required for optimal stability of the MRN complex during mitosis. MMAP colocalizes with MRN in mitotic spindles, and MMAP-deficient cells display abnormal spindle dynamics and chromosome segregation similar to MRN-deficient cells. Mechanistically, both MMAP and MRE11 are hyperphosphorylated by the mitotic kinase, PLK1; and the phosphorylation is required for assembly of the mMRN complex. The assembled mMRN complex enables PLK1 to interact with and activate the microtubule depolymerase, KIF2A, leading to spindle turnover and chromosome segregation. Our study identifies a mitosis-specific version of the MRN complex that acts in the PLK1-KIF2A signaling cascade to regulate spindle dynamics and chromosome distribution.

RNF169 limits 53BP1 deposition at DSBs to stimulate single-strand annealing repair.

Unrestrained 53BP1 activity at DNA double-strand breaks (DSBs) hampers DNA end resection and upsets DSB repair pathway choice. RNF169 acts as a molecular rheostat to limit 53BP1 deposition at DSBs, but how this fine balance translates to DSB repair control remains undefined. In striking contrast to 53BP1, ChIP analyses of AsiSI-induced DSBs unveiled that RNF169 exhibits robust accumulation at DNA end-proximal regions and preferentially targets resected, RPA-bound DSBs. Accordingly, we found that RNF169 promotes CtIP-dependent DSB resection and favors homology-mediated DSB repair, and further showed that RNF169 dose-dependently stimulates single-strand annealing repair, in part, by alleviating the 53BP1-imposed barrier to DSB end resection. Our results highlight the interplay of RNF169 with 53BP1 in fine-tuning choice of DSB repair pathways.

Salt- and pH-Triggered Helix-Coil Transition of Ionic Polypeptides under Physiology Conditions.

Controlling the helix-coil transition of polypeptides under physiological conditions is an attractive way toward smart functional materials. Here, we report the synthesis of a series of tertiary amine-functionalized ethylene glycol (EG x)-linked polypeptide electrolytes with their secondary structures tunable under physiological conditions. The resultant polymers, denoted as P(EG xDMA-Glu) ( x = 1, 2, and 3), show excellent aqueous solubility (>20 mg/mL) regardless of their charge states. Unlike poly-l-lysine that can form a helix only at pH above 10, P(EG xDMA-Glu) undergo a pH-dependent helix-coil switch with their transition points within the physiological range (pH ∼5.3-6.5). Meanwhile, P(EG xDMA-Glu) exhibit an unusual salt-induced helical conformation presumably owing to the unique properties of EG x linkers. Together, the current work highlights the importance of fine-tuning the linker chemistry in achieving conformation-switchable polypeptides and represents a facile approach toward stimuli-responsive biopolymers for advanced biological applications.

From neutral to zwitterionic poly(α-amino acid) nonfouling surfaces: Effects of helical conformation and anchoring orientation.

The development of high-performance nonfouling polymer surfaces for implantable medical devices and therapeutic nanomaterials is of great importance. Elaborating the relationship of polymer structural characteristics and the resulted surface properties can offer useful guidance toward ideal biointerfaces. In this work, we investigate the effects of the helical conformation and anchoring orientation of poly(α-amino acid)s (PαAAs) to produce advanced nonfouling surfaces. By using the neutral poly(γ-(2-(2-(2-methoxyethoxy)ethoxy)ethoxy)esteryl glutamates) (P(EG3Glu)s) as a model system, the adsorption kinetics are monitored by ex-situ variable angle spectroscopic ellipsometry and in-situ quartz crystal microbalance with dissipation. It is found that the polymers adopting a rigid rod-like α-helical conformation can self-assemble more rapidly to produce denser adlayers, and generate significantly improved nonfouling surfaces compared to those flexible polymer analogues including the widely used antifouling polymer PEG. Moreover, the surface properties can be further enhanced by using the antiparallel orientated helical P(EG3Glu)s. Most importantly, the insights gained from the P(EG3Glu) model system are successfully applied to the generation of ultra-low-fouling surfaces using zwitterionic PαAAs brushes, underscoring the generality of the approach. Particularly, the surface based on the antiparallel aligned zwitterionic helical PαAAs exhibits ∼98-99% reduction of human serum adsorption relative to the bare gold, and gives almost no adhesion of mouse platelet. Taken together, this work depicts an extremely simple yet highly effective approach to manipulate surface properties for numerous applications in biomaterial interfaces, diagnostics, and biosensors.

Macrocyclization of Interferon-Poly(α-amino acid) Conjugates Significantly Improves the Tumor Retention, Penetration, and Antitumor Efficacy.

Cyclization and polymer conjugation are two commonly used approaches for enhancing the pharmacological properties of protein drugs. However, cyclization of parental proteins often only affords a modest improvement in biochemical or cell-based in vitro assays. Moreover, very few studies have included a systematic pharmacological evaluation of cyclized protein-based therapeutics in live animals. On the other hand, polymer-conjugated proteins have longer circulation half-lives but usually show poor tumor penetration and suboptimal pharmacodynamics due to increased steric hindrance. We herein report the generation of a head-to-tail interferon-poly(α-amino acid) macrocycle conjugate circ-P(EG3Glu)20-IFN by combining the aforementioned two approaches. We then compared the antitumor pharmacological activity of this macrocycle conjugate against its linear counterparts, N-P(EG3Glu)20-IFN, C-IFN-P(EG3Glu)20, and C-IFN-PEG. Our results found circ-P(EG3Glu)20-IFN to show considerably greater stability, binding affinity, and in vitro antiproliferative activity toward OVCAR3 cells than the three linear conjugates. More importantly, circ-P(EG3Glu)20-IFN exhibited longer circulation half-life, remarkably higher tumor retention, and deeper tumor penetration in vivo. As a result, administration of the macrocyclic conjugate could effectively inhibit tumor progression and extend survival in mice bearing established xenograft human OVCAR3 or SKOV3 tumors without causing severe paraneoplastic syndromes. Taken together, our study provided until now the most relevant experimental evidence in strong support of the in vivo benefit of macrocyclization of protein-polymer conjugates and for its application in next-generation therapeutics.

A S-Sn Lewis Pair-Mediated Ring-Opening Polymerization of α-Amino Acid N-Carboxyanhydrides: Fast Kinetics, High Molecular Weight, and Facile Bioconjugation.

The rapid and controlled generation of polypeptides with ultrahigh molecular weight (MW) and well-defined chain end functionality has been a great challenge. To tackle this problem, we report here an initiation system based on a S-Sn Lewis pair, trimethylstannyl phenyl sulfide (PhS-SnMe3), for the ring-opening polymerization (ROP) of α-amino acid N-carboxyanhydrides (NCAs). This initiator displays a strong solvent effect, and can yield polypeptides with high MW (>1.0 × 105 g·mol-1) and low polydispersity index within a few hours. The MWs of the obtained polypeptides are strongly dependent on the THF/DMF ratio. The polymerization follows a typical first-order kinetic character with respect to the monomer concentration in mixed THF and DMF. Moreover, a highly reactive phenyl thioester is in situ generated at the C-terminus of the polypeptides, which is readily accessible for native chemical ligation affording high MW and site-specific protein-polypeptide conjugates. Together, this initiator sheds light on regulating the ROP of NCAs via appropriate Lewis pair and solvent selection, and is particularly useful in preparing ultrahigh MW polypeptides within a short period of time.

Transitional basal cells at the squamous-columnar junction generate Barrett's oesophagus.

In several organ systems, the transitional zone between different types of epithelium is a hotspot for pre-neoplastic metaplasia and malignancy, but the cells of origin for these metaplastic epithelia and subsequent malignancies remain unknown. In the case of Barrett's oesophagus, intestinal metaplasia occurs at the gastro-oesophageal junction, where stratified squamous epithelium transitions into simple columnar cells. On the basis of a number of experimental models, several alternative cell types have been proposed as the source of this metaplasia but in all cases the evidence is inconclusive: no model completely mimics Barrett's oesophagus in terms of the presence of intestinal goblet cells. Here we describe a transitional columnar epithelium with distinct basal progenitor cells (p63+KRT5+KRT7+) at the squamous-columnar junction of the upper gastrointestinal tract in a mouse model. We use multiple models and lineage tracing strategies to show that this squamous-columnar junction basal cell population serves as a source of progenitors for the transitional epithelium. On ectopic expression of CDX2, these transitional basal progenitors differentiate into intestinal-like epithelium (including goblet cells) and thereby reproduce Barrett's metaplasia. A similar transitional columnar epithelium is present at the transitional zones of other mouse tissues (including the anorectal junction) as well as in the gastro-oesophageal junction in the human gut. Acid reflux-induced oesophagitis and the multilayered epithelium (believed to be a precursor of Barrett's oesophagus) are both characterized by the expansion of the transitional basal progenitor cells. Our findings reveal a previously unidentified transitional zone in the epithelium of the upper gastrointestinal tract and provide evidence that the p63+KRT5+KRT7+ basal cells in this zone are the cells of origin for multi-layered epithelium and Barrett's oesophagus.

Clustered, Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas9-coupled Affinity Purification/Mass Spectrometry Analysis Revealed a Novel Role of Neurofibromin in mTOR Signaling.

Neurofibromin (NF1) is a well known tumor suppressor that is commonly mutated in cancer patients. It physically interacts with RAS and negatively regulates RAS GTPase activity. Despite the importance of NF1 in cancer, a high quality endogenous NF1 interactome has yet to be established. In this study, we combined clustered, regularly interspaced short palindromic repeats (CRISPR)/Cas9-mediated gene knock-out technology with affinity purification using antibodies against endogenous proteins, followed by mass spectrometry analysis, to sensitively and accurately detect NF1 protein-protein interactions in unaltered in vivo settings. Using this system, we analyzed endogenous NF1-associated protein complexes and identified 49 high-confidence candidate interaction proteins, including RAS and other functionally relevant proteins. Through functional validation, we found that NF1 negatively regulates mechanistic target of rapamycin signaling (mTOR) in a LAMTOR1-dependent manner. In addition, the cell growth and survival of NF1-deficient cells have become dependent on hyperactivation of the mTOR pathway, and the tumorigenic properties of these cells have become dependent on LAMTOR1. Taken together, our findings may provide novel insights into therapeutic approaches targeting NF1-deficient tumors.

RPA-Binding Protein ETAA1 Is an ATR Activator Involved in DNA Replication Stress Response.

ETAA1 (Ewing tumor-associated antigen 1), also known as ETAA16, was identified as a tumor-specific antigen in the Ewing family of tumors. However, the biological function of this protein remains unknown. Here, we report the identification of ETAA1 as a DNA replication stress response protein. ETAA1 specifically interacts with RPA (Replication protein A) via two conserved RPA-binding domains and is therefore recruited to stalled replication forks. Interestingly, further analysis of ETAA1 function revealed that ETAA1 participates in the activation of ATR signaling pathway via a conserved ATR-activating domain (AAD) located near its N terminus. Importantly, we demonstrate that both RPA binding and ATR activation are required for ETAA1 function at stalled replication forks to maintain genome stability. Therefore, our data suggest that ETAA1 is a new ATR activator involved in replication checkpoint control.

A Concise Approach to Site-Specific Topological Protein-Poly(amino acid) Conjugates Enabled by in Situ-Generated Functionalities.

Controlling the topology of polymer-modified proteins has attracted growing interest. However, one of the main challenges in this field is the lack of efficient and site-specific methods for installing multiple bioorthogonal functionalities on substrate polymers. We report here an orchestrating strategy that provides easy access to various topological protein-poly(amino acid) (PAA) conjugates in high yields. This method features the in situ installation of two "chemical handles", including a thioester for native chemical ligation and a polyglycine nucleophile for sortase A-mediated ligation, at both ends of substrate PAAs. As a result, neither pre-functionalization of initiator or monomer units, nor post-polymerization modification of the resultant polymers, is necessary. Site-specific topological conjugates, particularly circular conjugates, can be conveniently synthesized under mild conditions from the functionalized PAAs. The biomedical utility of our method is demonstrated by the rapid and efficient generation of several therapeutic interferon-α conjugates, which exhibit significantly enhanced protease resistance and thermostability. Given the versatility of both PAAs and proteins, the method offers a convenient approach to producing libraries of conjugates for biological applications.

Phenyl Trimethylsilyl Sulfide-Mediated Controlled Ring-Opening Polymerization of α-Amino Acid N-Carboxyanhydrides.

We describe here the first example of trimethylsilyl sulfide (S-TMS) mediated controlled ring-opening polymerization (ROP) of α-amino acid N-carboxyanhydrides (NCAs). We show that phenyl trimethylsilyl sulfide (PhS-TMS), an inexpensive and commercially available compound, mediates rapid ROP of a broad scope of NCA monomers, produces functional poly(amino acids) (PAAs) with controllable molecular weights (MWs), narrow polydispersity index (PDI), and an in situ generated phenyl thioester group at the C-terminus (PAA-SPhs). PhS-TMS offers more rapid chain initiation than previously reported hexamethyldisilazane (HMDS) initiator, ensuring a living polymerization with better control. Mechanistic studies suggest that a reactive trimethylsilyl carbamate (TMSC) was generated during the chain initiation and continued to regulate the chain propagation through a TMS transfer process. Considering the versatility of NCAs, and the potential of leveraging the C-terminal phenyl thioester for native chemical ligation (NCL), we believe this method may offer a powerful platform enabling the rapid generation of functional PAAs and their C-terminal conjugates for numerous biological applications.

Identification of hallmarks of lung adenocarcinoma prognosis using whole genome sequencing.

In conjunction with clinical characteristics, prognostic biomarkers are essential for choosing optimal therapies to lower the mortality of lung adenocarcinoma. Whole genome sequencing (WGS) of 7 cancerous-noncancerous tissue pairs was performed to explore the comparative copy number variations (CNVs) associated with lung adenocarcinoma. The frequencies of top ranked CNVs were verified in an independent set of 114 patients and then the roles of target CNVs in disease prognosis were assessed in 313 patients. The WGS yielded 2604 CNVs. After frequency validation and biological function screening of top 10 CNVs, 9 mutant driver genes from 7 CNVs were further analyzed for an association with survival. Compared with the PBXIP1 amplified copy number, unamplified carriers had a 0.62-fold (95%CI = 0.43-0.91) decreased risk of death. Compared with an amplified TERT, those with an unamplified TERT had a 35% reduction (95% CI = 3%-56%) in risk of lung adenocarcinoma progression. Cases with both unamplified PBXIP1 and TERT had a median 34.32-month extension of overall survival and 34.55-month delay in disease progression when compared with both amplified CNVs. This study demonstrates that CNVs of TERT and PBXIP1 have the potential to translate into the clinic and be used to improve outcomes for patients with this fatal disease.

N-Fused BDOPV: a tetralactam derivative as a building block for polymer field-effect transistors.

An N-fused BDOPV derivative, NBDOPV, was designed and synthesized. The photophysical and electrochemical properties of NBDOPV were systematically investigated. The NBDOPV-based conjugated polymer PITET shows a large hole mobility of 1.92 cm(2) V(-1) s(-1).