Epigenetic modification of ferroptosis by non-coding RNAs in cancer drug resistance.

The development of drug resistance remains a major challenge in cancer treatment. Ferroptosis, a unique type of regulated cell death, plays a pivotal role in inhibiting tumour growth, presenting new opportunities in treating chemotherapeutic resistance. Accumulating studies indicate that epigenetic modifications by non-coding RNAs (ncRNA) can determine cancer cell vulnerability to ferroptosis. In this review, we first summarize the role of chemotherapeutic resistance in cancer growth/development. Then, we summarize the core molecular mechanisms of ferroptosis, its upstream epigenetic regulation, and its downstream effects on chemotherapeutic resistance. Finally, we review recent advances in understanding how ncRNAs regulate ferroptosis and from such modulate chemotherapeutic resistance. This review aims to enhance general understanding of the ncRNA-mediated epigenetic regulatory mechanisms which modulate ferroptosis, highlighting the ncRNA-ferroptosis axis as a key druggable target in overcoming chemotherapeutic resistance.

Chemotherapy re-use versus anti-angiogenic monotherapy as the third-line treatment of patients with metastatic colorectal cancer: a real-world cohort study.

BACKGROUND: There are various recommendations for third-line treatment in mCRC, however, there is no consensus on who is more suitable for particular strategy. Chemotherapy re-use in third-line setting is a common option in clinical practice. This study aimed to investigate the efficacy of third-line chemotherapy re-use by the comparison with that of anti-angiogenic monotherapy, and further find the population more suitable for third-line chemotherapy. METHODS: Using electronic medical records of patients with mCRC, a retrospective cohort study was conducted. A total of 143 patients receiving chemotherapy and 40 patients receiving anti-angiogenic monotherapy in third-line setting as control group were retrospectively collected. Baseline characteristics were analyzed using the χ² test or the Fisher's exact test. ROC curve and surv_cutpoint function of 'survminer' package in R software were used to calculate the cut-off value. Survival curves were plotted with the Kaplan-Meier method and were compared using the log-rank test. The Cox proportional hazard regression model was used to analyze the potential risk factors. RESULTS: A total of 143 patients receiving chemotherapy and 40 patients receiving anti-angiogenic monotherapy in third-line setting were retrospectively collected. Chemotherapy rechallenge was recorded in 93 patients (93/143, 65.0%), and the remaining patients chose new chemotherapeutic drugs that had not been previously used, including irinotecan-based (22/50), oxaliplatin-based (9/50), raltitrexed (9/50), gemcitabine (5/50) and other agents (5/50). The ORR and DCR of third-line chemotherapy reached 8.8%, 61.3%, respectively (anti-angiogenic monotherapy group: ORR 2.6%, DCR 47.4%). The mPFS and mOS of patients receiving chemotherapy were 4.9 and 12.0 m, respectively (anti-angiogenic monotherapy group: mPFS 2.7 m, mOS 5.2 m). Subgroup analyses found that patients with RAS/RAF mutation, longer PFS (greater than 10.6 m) in front-line treatment or larger tumor burden had better prognosis with third-line chemotherapy rather than anti-angiogenic monotherapy. CONCLUSIONS: Third-line chemotherapy re-use was effective in mCRC. Those with more aggressive characteristics (RAS/RAF mutant, larger tumor burden) or better efficacy of previous chemotherapy (longer PFS) were more appropriate for third-line chemotherapy, rather than anti-angiogenic monotherapy.

Sintilimab Plus Apatinib and Chemotherapy as Second­/Third-Line treatment for Advanced Gastric or Gastroesophageal Junction Adenocarcinoma: a prospective, Single-Arm, phase II trial.

BACKGROUND: The prognosis of patients with previously treated advanced gastric or gastroesophageal junction (GEJ) cancer remains poor. Given the robust development of immunotherapy and targeted therapy during the last decades, we aimed to investigate if the combination of traditional second-line chemotherapy with sintilimab and apatinib could bring survival benefits for these patients. METHODS: In this single-center, single-arm, phase II trial, patients with previously treated advanced gastric or GEJ adenocarcinoma received specific dose level of intravenous paclitaxel or irinotecan (investigator's choice), 200 mg intravenous sintilimab on day 1, and 250 mg oral apatinib once daily continuously in each cycle until disease progression, intolerable toxicity, or withdrawal of consent. The primary endpoints were objective response rate and progression-free survival. The secondary endpoints were mainly overall survival and safety. RESULTS: From May 2019 to May 2021, 30 patients were enrolled. At the data cutoff date (March 19, 2022), the median follow-up duration was 12.3 months and 53.6% (95% CI, 33.9-72.5%) patients achieved objective response. The median progression-free survival and overall survival were 8.5 months (95% CI, 5.4-11.5) and 12.5 months (95% CI, 3.7-21.3), respectively. Grade 3-4 adverse events included hematological toxicities, elevated alanine aminotransferase, elevated aspartate aminotransferase, elevated alkaline phosphatase, elevated gamma-glutamyl transpeptidase, hyperbilirubinemia and proteinuria. The most frequent grade 3-4 adverse event was neutropenia (13.3%). No serious treatment-related adverse events or treatment-related deaths occurred. CONCLUSION: Sintilimab plus apatinib and chemotherapy demonstrates promising anti-tumor activity with manageable safety profile in patients with previously treated advanced gastric or GEJ cancer. TRIAL REGISTRATION: ClinicalTrials.gov: NCT05025033, 27/08/2021.

Synthesis of biotinylated-LPG as a chemical biology tool enabling discovery of ALCAT1 modulators.

As a mitochondrial signature phospholipid, cardiolipin (CL) is required for membrane structure, respiration, dynamics, fragmentation, and mitophagy. Alteration of CL by reactive oxygen species (ROS) can cause mitochondrial dysfunction, which is implicated in the pathogenesis of many diseases. The enzyme ALCAT1 (acyl-CoA: lysocardiolipin acyltransferase-1) facilitates the conversion of CL by incorporating polyunsaturated fatty acids into lysocardiolipin. Accumulating evidence suggests that overexpression of ALCAT1 is involved in pathological cardiolipin remodeling and mitochondrial bioenergetics. Few ALCAT1 modulators are reported in the literature, and the enzymatic activity was tested via a low-throughput TLC (thin layer chromatography) assay. To identify small molecule ALCAT1 inhibitors, a robust assay was needed to enable a full deck high throughput screen. Scintillation proximity assay (SPA) was the method of choice because it permits the rapid and sensitive measurement of a broad range of biological processes in a homogeneous system. A biotinylated ALCAT1 substrate was required as a chemical biology tool in developing SPA. Among a panel of phospholipids, lysophosphatidyl glycerol (LPG) was identified as the best substrate for ALCAT1. Herein we report the synthesis of biotinylated-LPG analogs with varied linker lengths and their activity towards ALCAT1.

Discovery of an Allosteric, Inactive Conformation-Selective Inhibitor of Full-Length HPK1 Utilizing a Kinase Cascade Assay.

Achieving selectivity across the human kinome is a major hurdle in kinase inhibitor drug discovery. Assays using active, phosphorylated protein kinases bias hits toward poorly selective inhibitors that bind within the highly conserved adenosine triphosphate (ATP) pocket. Targeting inactive (vs active) kinase conformations offers advantages in achieving selectivity because of their more diversified structures. Kinase cascade assays are typically initiated with target kinases in their unphosphorylated inactive forms, which are activated during the assays. Therefore, these assays are capable of identifying inhibitors that preferentially bind to the unphosphorylated form of the enzyme in addition to those that bind to the active form. We applied this cascade assay to the emerging cancer immunotherapy target hematopoietic progenitor kinase 1 (HPK1), a serine/threonine kinase that negatively regulates T cell receptor signaling. Using this approach, we discovered an allosteric, inactive conformation-selective triazolopyrimidinone HPK1 inhibitor, compound 1. Compound 1 binds to unphosphorylated HPK1 >24-fold more potently than active HPK1, is not competitive with ATP, and is highly selective against kinases critical for T cell signaling. Furthermore, compound 1 does not bind to the isolated HPK1 kinase domain alone but requires other domains. Together, these data indicate that 1 is an allosteric HPK1 inhibitor that attenuates kinase autophosphorylation by binding to a pocket consisting of residues within and outside of the kinase domain. Our study demonstrates that cascade assays can lead to the discovery of highly selective kinase inhibitors. The triazolopyrimidinone described in this study may represent a privileged chemical scaffold for further development of potent and selective HPK1 inhibitors.

A carboxylic acid isostere screen of the DHODH inhibitor Brequinar.

Dihydroorotate dehydrogenase (DHODH) enzymatic activity impacts many aspects critical to cell proliferation and survival. Recently, DHODH has been identified as a target for acute myeloid differentiation therapy. In preclinical models of AML, the DHODH inhibitor Brequinar (BRQ) demonstrated potent anti-leukemic activity. Herein we describe a carboxylic acid isostere study of Brequinar which revealed a more potent non-carboxylic acid derivative with improved cellular potency and good pharmacokinetic properties.

Hit-to-lead optimization and discovery of a potent, and orally bioavailable G protein coupled receptor kinase 2 (GRK2) inhibitor.

G-protein coupled receptor kinase 2 (GRK2), which is upregulated in the failing heart, appears to play a critical role in heart failure (HF) progression in part because enhanced GRK2 activity promotes dysfunction of ß-adrenergic signaling and myocyte death. An orally bioavailable GRK2 inhibitor could offer unique therapeutic outcomes that cannot be attained by current heart failure treatments that directly target GPCRs or angiotensin-converting enzyme. Herein, we describe the discovery of a potent, selective, and orally bioavailable GRK2 inhibitor, 8h, through high-throughput screening, hit-to-lead optimization, structure-based design, molecular modelling, synthesis, and biological evaluation. In the cellular target engagement assays, 8h enhances isoproterenol-mediated cyclic adenosine 3',5'-monophosphate (cAMP) production in HEK293 cells overexpressing GRK2. Compound 8h was further evaluated in a human stem cell-derived cardiomyocyte (HSC-CM) contractility assay and potentiated isoproterenol-induced beating rate in HSC-CMs.

FGF23 regulates atrial fibrosis in atrial fibrillation by mediating the STAT3 and SMAD3 pathways.

High fibroblast growth factor 23 (FGF23) concentrations are a strong predictor of atrial fibrillation (AF), but researchers have not clearly determined the mechanism by which FGF23 causes atrial fibrosis in patients with AF. This study aims to elucidate the mechanism by which FGF23 induces atrial fibrosis in patients with AF. Immunohistochemistry was used to study the expression of FGF23, FGFR4, and fibrotic factors in patients with a normal sinus rhythm (SR) and patients with AF. Cardiac fibroblasts (CFs) were cocultured with different concentrations of the recombinant FGF23 protein. Compared with the SR group, the levels of FGF23, FGFR4, α-smooth muscle actin (α-SMA), and collagen-1 were significantly increased in the AF group. Exposure to high concentrations of the recombinant FGF23 protein increased the accumulation of reactive oxygen species (ROS) and activated α-SMA, collagen-1, signal transducer and activator of transcription 3 (STAT3) and SMAD3 signaling in cultured CFs. The levels of fibrotic proteins in CFs stimulated with high concentrations of the recombinant FGF23 protein were reversed by N-acetylcysteine (NAC, a ROS inhibitor), ship information system 3 (a SMAD3 inhibitor), and Stattic (a STAT3 inhibitor). Furthermore, compared to untreated CFs, CFs treated with the recombinant FGF23 protein were characterized by an increased interaction between STAT3 and SMAD3. Based on these results, FGF23 induces atrial fibrosis in patients with AF by increasing ROS production and subsequently activating STAT3 and SMAD3 signaling.

Pd Nanoparticles Confined in the Porous Graphene-like Carbon Nanosheets for Olefin Hydrogenation.

As a novel type of defective graphene, porous graphene has been considered an excellent support material for metal clusters, as the interaction between defective carbon atoms surrounded with the metal nanoparticles (NPs) is very different from that on the ordinary supported catalyst. In this work, we reported a facile three-step method to confine the Pd NPs and grow the graphene-like carbon nanosheets (GLCs) in the same interlayer space of the layered silicate, generating embedded Pd NPs in the pores of porous GLCs in situ. The Pd@GLC nanocomposite exhibited not only high activity and stability than the common commercial Pd/C catalyst for the hydrogenation of olefins but also superior ability of resisting high temperature, which benefitted from the two-dimensional structure of layered GLCs, the confinement of Pd, and the increased edge and defect of the unsaturated carbon atoms in GLCs.

Discovery of Inactive Conformation-Selective Kinase Inhibitors by Utilizing Cascade Assays.

Achieving selectivity across the human kinome is a major hurdle in kinase inhibitor drug discovery. Targeting inactive (vs active) kinase conformations offers advantages in achieving selectivity because of their more diversified structures. Discovery of inactive conformation-selective inhibitors, however, has been hampered partly by the lack of general assay methods. Herein, we show that such inhibitors can be discovered by utilizing kinase cascade assays. This type of assay is initiated with the target kinase in its unphosphorylated, inactive conformation, which is activated during the assay. Inactive conformation-selective inhibitors stabilize the inactive kinase, block activation, and yield reduced kinase activity. We investigate the properties of the assay by mathematical modeling, as well as by proof-of-concept experiments using the BRAF-MEK1 cascade. This study demonstrates effective identification of inactive conformation-selective inhibitors by cascade assays, reveals key factors that impact results, and provides guidelines for successful cascade assay development.

Spectroscopic and Computational Investigations of Ligand Binding to IspH: Discovery of Non-diphosphate Inhibitors.

Isoprenoid biosynthesis is an important area for anti-infective drug development. One isoprenoid target is (E)-1-hydroxy-2-methyl-but-2-enyl 4-diphosphate (HMBPP) reductase (IspH), which forms isopentenyl diphosphate and dimethylallyl diphosphate from HMBPP in a 2H+ /2e- reduction. IspH contains a 4 Fe-4 S cluster, and in this work, we first investigated how small molecules bound to the cluster by using HYSCORE and NRVS spectroscopies. The results of these, as well as other structural and spectroscopic investigations, led to the conclusion that, in most cases, ligands bound to IspH 4 Fe-4 S clusters by η1 coordination, forming tetrahedral geometries at the unique fourth Fe, ligand side chains preventing further ligand (e.g., H2 O, O2 ) binding. Based on these ideas, we used in silico methods to find drug-like inhibitors that might occupy the HMBPP substrate binding pocket and bind to Fe, leading to the discovery of a barbituric acid analogue with a Ki value of ≈500 nm against Pseudomonas aeruginosa IspH.

The Streptococcus suis transcriptional landscape reveals adaptation mechanisms in pig blood and cerebrospinal fluid.

Streptococcus suis (SS) is an important pathogen of pigs, and it is also recognized as a zoonotic agent for humans. SS infection may result in septicemia or meningitis in the host. However, little is known about genes that contribute to the virulence process and survival within host blood or cerebrospinal fluid (CSF). Small RNAs (sRNA) have emerged as key regulators of virulence in several bacteria, but they have not been investigated in SS. Here, using a differential RNA-sequencing approach and RNAs from SS strain P1/7 grown in rich medium, pig blood, or CSF, we present the SS genome-wide map of 793 transcriptional start sites and 370 operons. In addition to identifying 29 sRNAs, we show that five sRNA deletion mutants attenuate SS virulence in a zebrafish infection model. Homology searches revealed that 10 sRNAs were predicted to be present in other pathogenic Streptococcus species. Compared with wild-type strain P1/7, sRNAs rss03, rss05, and rss06 deletion mutants were significantly more sensitive to killing by pig blood. It is possible that rss06 contributes to SS virulence by indirectly activating expression of SSU0308, a virulence gene encoding a zinc-binding lipoprotein. In blood, genes involved in the synthesis of capsular polysaccharide (CPS) and subversion of host defenses were up-regulated. In contrast, in CSF, genes for CPS synthesis were down-regulated. Our study is the first analysis of SS sRNAs involved in virulence and has both improved our understanding of SS pathogenesis and increased the number of sRNAs known to play definitive roles in bacterial virulence.

Coupling Oxygen Consumption with Hydrocarbon Oxidation in Bacterial Multicomponent Monooxygenases.

A fundamental goal in catalysis is the coupling of multiple reactions to yield a desired product. Enzymes have evolved elegant approaches to address this grand challenge. A salient example is the biological conversion of methane to methanol catalyzed by soluble methane monooxygenase (sMMO), a member of the bacterial multicomponent monooxygenase (BMM) superfamily. sMMO is a dynamic protein complex of three components: a hydroxylase, a reductase, and a regulatory protein. The active site, a carboxylate-rich non-heme diiron center, is buried inside the 251 kDa hydroxylase component. The enzyme processes four substrates: O2, protons, electrons, and methane. To couple O2 activation to methane oxidation, timely control of substrate access to the active site is critical. Recent studies of sMMO, as well as its homologues in the BMM superfamily, have begun to unravel the mechanism. The emerging and unifying picture reveals that each substrate gains access to the active site along a specific pathway through the hydroxylase. Electrons and protons are delivered via a three-amino-acid pore located adjacent to the diiron center; O2 migrates via a series of hydrophobic cavities; and hydrocarbon substrates reach the active site through a channel or linked set of cavities. The gating of these pathways mediates entry of each substrate to the diiron active site in a timed sequence and is coordinated by dynamic interactions with the other component proteins. The result is coupling of dioxygen consumption with hydrocarbon oxidation, avoiding unproductive oxidation of the reductant rather than the desired hydrocarbon. To initiate catalysis, the reductase delivers two electrons to the diiron(III) center by binding over the pore of the hydroxylase. The regulatory component then displaces the reductase, docking onto the same surface of the hydroxylase. Formation of the hydroxylase-regulatory component complex (i) induces conformational changes of pore residues that may bring protons to the active site; (ii) connects hydrophobic cavities in the hydroxylase leading from the exterior to the diiron active site, providing a pathway for O2 and methane, in the case of sMMO, to the reduced diiron center for O2 activation and substrate hydroxylation; (iii) closes the pore, as well as a channel in the case of four-component BMM enzymes, restricting proton access to the diiron center during formation of "Fe2O2" intermediates required for hydrocarbon oxidation; and (iv) inhibits undesired electron transfer to the Fe2O2 intermediates by blocking reductase binding during O2 activation. This mechanism is quite different from that adopted by cytochromes P450, a large class of heme-containing monooxygenases that catalyze reactions very similar to those catalyzed by the BMM enzymes. Understanding the timed enzyme control of substrate access has implications for designing artificial catalysts. To achieve multiple turnovers and tight coupling, synthetic models must also control substrate access, a major challenge considering that nature requires large, multimeric, dynamic protein complexes to accomplish this feat.

Impact of incomplete revascularization of coronary artery disease on long-term cardiac outcomes. Retrospective comparison of angiographic and myocardial perfusion imaging criteria for completeness.

BACKGROUND: Coronary revascularization in patients with coronary artery disease may be guided by coronary angiography (CA) or alternatively by ischemia on stress myocardial perfusion imaging (MPI). Which strategy leads to optimal cardiac outcomes is uncertain. METHODS: We performed a retrospective analysis of 170 patients with MPI ischemia and percutaneous coronary intervention. The primary endpoint was all-cause mortality at a mean follow-up of 47 ± 21 months; the secondary end point was the composite of deaths, nonfatal myocardial infarction, and repeat coronary revascularization (MACE). The coronary revascularization was defined as complete (CCR) or incomplete (ICR) as judged by CA criteria and by MPI ischemia matched with CA criteria. RESULTS: Nighty-two patients (54%) had ICR by CA criteria (ICR-CA) and 84 (49%) had ICR by MPI criteria (ICR-MPI). Mortality and MACE were lower in patients with CCR-MPI than with ICR-MPI (P = .048, and P = .025). Survival of patients with CCR-CA and ICR-CA was not different (P = .081). Patients with both ICR-MPI and ICR-CA had the worst survival, whereas patients with CCR-MPI and CCR-CA had the best survival (P = .047). By multivariate analysis, ICR-MPI + ICR-CA was an independent predictor of death (P = .025). CONCLUSION: Patients with ICR by MPI were at higher risk than those with CCR. Patients with both ICR by MPI and CA were at the highest risk, while patients with CCR by both MPI and CA had the best long-term event-free survival.

Mechanistic Studies of the Anticancer Activity of An Octahedral Hexanuclear Pt(II) Cage.

The cellular response evoked by a hexanuclear platinum complex, Pt6L4 (1), is reported. Compound 1, a 3-nm octahedral cage formed by self-assembly of six Pt(II) centers and four 2,4,6-tris(4-pyridyl)-1,3,5-triazine ligands (L), exhibits promising in vitro potency against a panel of human cancer cell lines. Unlike classical platinum-based anticancer agents, 1 interacts with DNA in a non-covalent, intercalative manner and promotes DNA condensation. In cancer cells, 1 induces DNA damage, upregulates p53, its phosphorylated form phospho-p53 and its downstream effector, p21, as well as both apoptosis and senescence.

Structure, function and inhibition of the two- and three-domain 4Fe-4S IspG proteins.

IspG is a 4Fe4S protein involved in isoprenoid biosynthesis. Most bacterial IspGs contain two domains: a TIM barrel (A) and a 4Fe4S domain (B), but in plants and malaria parasites, there is a large insert domain (A*) whose structure and function are unknown. We show that bacterial IspGs function in solution as (AB)(2) dimers and that mutations in either both A or both B domains block activity. Chimeras harboring an A-mutation in one chain and a B-mutation in the other have 50% of the activity seen in wild-type protein, because there is still one catalytically active AB domain. However, a plant IspG functions as an AA*B monomer. We propose, using computational modeling and electron microscopy, that the A* insert domain has a TIM barrel structure that interacts with the A domain. This structural arrangement enables the A and B domains to interact in a "cup and ball" manner during catalysis, just as in the bacterial systems. EPR/HYSCORE spectra of reaction intermediate, product, and inhibitor ligands bound to both two and three domain proteins are identical, indicating the same local electronic structure, and computational docking indicates these ligands bridge both A and B domains. Overall, the results are of broad general interest because they indicate the insert domain in three-domain IspGs is a second TIM barrel that plays a structural role and that the pattern of inhibition of both two and three domain proteins are the same, results that can be expected to be of use in drug design.

Diiron oxidation state control of substrate access to the active site of soluble methane monooxygenase mediated by the regulatory component.

The regulatory component (MMOB) of soluble methane monooxygenase (sMMO) has a unique N-terminal tail not found in regulatory proteins of other bacterial multicomponent monooxygenases. This N-terminal tail is indispensable for proper function, yet its solution structure and role in catalysis remain elusive. Here, by using double electron-electron resonance (DEER) spectroscopy, we show that the oxidation state of the hydroxylase component, MMOH, modulates the conformation of the N-terminal tail in the MMOH-2MMOB complex, which in turn facilitates catalysis. The results reveal that the N-terminal tail switches from a relaxed, flexible conformational state to an ordered state upon MMOH reduction from the diiron(III) to the diiron(II) state. This observation suggests that some of the crystallographically observed allosteric effects that result in the connection of substrate ingress cavities in the MMOH-2MMOB complex may not occur in solution in the diiron(III) state. Thus, O2 may not have easy access to the active site until after reduction of the diiron center. The observed conformational change is also consistent with a higher binding affinity of MMOB to MMOH in the diiron(II) state, which may allow MMOB to displace more readily the reductase component (MMOR) from MMOH following reduction.

Electron transfer control in soluble methane monooxygenase.

The hydroxylation or epoxidation of hydrocarbons by bacterial multicomponent monooxygenases (BMMs) requires the interplay of three or four protein components. How component protein interactions control catalysis, however, is not well understood. In particular, the binding sites of the reductase components on the surface of their cognate hydroxylases and the role(s) that the regulatory proteins play during intermolecular electron transfer leading to the hydroxylase reduction have been enigmatic. Here we determine the reductase binding site on the hydroxylase of a BMM enzyme, soluble methane monooxygenase (sMMO) from Methylococcus capsulatus (Bath). We present evidence that the ferredoxin domain of the reductase binds to the canyon region of the hydroxylase, previously determined to be the regulatory protein binding site as well. The latter thus inhibits reductase binding to the hydroxylase and, consequently, intermolecular electron transfer from the reductase to the hydroxylase diiron active site. The binding competition between the regulatory protein and the reductase may serve as a control mechanism for regulating electron transfer, and other BMM enzymes are likely to adopt the same mechanism.

Bioorganometallic chemistry with IspG and IspH: structure, function, and inhibition of the [Fe(4)S(4)] proteins involved in isoprenoid biosynthesis.

Enzymes of the methylerythritol phosphate pathway of isoprenoid biosynthesis are attractive anti-infective drug targets. The last two enzymes of this pathway, IspG and IspH, are [Fe4 S4 ] proteins that are not produced by humans and catalyze 2 H(+) / 2 e(-) reductions with novel mechanisms. In this Review, we summarize recent advances in structural, mechanistic, and inhibitory studies of these two enzymes. In particular, mechanistic proposals involving bioorganometallic intermediates are presented, and compared with other mechanistic possibilities. In addition, inhibitors based on substrate analogues as well as developed by rational design and compound-library screening, are discussed. The results presented support bioorganometallic catalytic mechanisms for IspG and IspH, and open up new routes to anti-infective drug design targeting [Fe4 S4 ] clusters in proteins.

Feasibility and Safety of PD-1 Blockades Among Elderly Patients with Metastatic Esophageal Squamous Cell Carcinoma: A Real-World Study.

Objective: This study aimed to identify the effectiveness and safety of PD-1 blockades among elderly patients with metastatic esophageal squamous cell carcinoma (ESCC) clinically. Methods: A total of 78 elderly patients with previously treated metastatic ESCC aged ≥65 years who received PD-1 blockades monotherapy were included retrospectively. Demographic characteristics, therapeutic effectiveness and adverse reactions of the elderly patients who underwent PD-1 blockade therapy were recorded. Regular follow-up was conducted for all patients. The analysis aimed to identify potential risk factors for OS by examining the correlation between prognosis and subgroups based on baseline characteristics. Results: The median age of the 78 elderly patients was 73 years, ranging from 65 to 87 years. Among the 78 patients, 18 cases showed partial response, 26 cases had stable disease, 29 cases experienced progressive disease and 5 cases were not assessable for response, yielding an ORR of 23.1%, a DCR of 56.4%. The prognostic outcomes indicated that among the 78 patients with metastatic ESCC who received PD-1 blockades, the median PFS was 3.1 months [95% confidence interval (CI): 1.64-4.56], and the median OS was 10.9 months (95% CI: 6.02-15.78), 24-month OS rate was 22.7% (95% CI: 12.8-34.2%). In terms of the safety profile, among the 78 patients with metastatic ESCC during PD-1 blockades single-agent treatment, a total of 61 patients (78.2%) experienced any grade adverse reactions and the incidence of grade ≥3 adverse reactions were 20.5%. Briefly, the common adverse reactions manifested as fatigue (32.1%), gastrointestinal reaction (24.4%), diarrhea (19.2%), anemia (17.9%) and rash (16.7%). Overall tolerability of PD-1 blockade monotherapy in elderly patients with metastatic ESCC was acceptable and manageable. Conclusion: PD-1 blockades single agent demonstrated encouraging effectiveness and acceptable safety profile for elderly patients with previously treated metastatic ESCC in clinical practice. Prospective study should be performed to elucidate the conclusion in this study subsequently.