Discovery of a long half-life AURKA inhibitor to treat MYC-amplified solid tumors as a monotherapy and in combination with everolimus.

Aurora kinase inhibitors such as alisertib can destabilize MYC-family oncoproteins and have demonstrated compelling anti-tumor efficacy. In this study, we report 6K465, a novel pyrimidine-based Aurora A (AURKA) inhibitor that reduces levels of c-MYC and N-MYC oncoproteins more potently than alisertib. In an analysis of the antiproliferative effect of 6K465, the sensitivities of small cell lung cancer (SCLC) and breast cancer (BC) cell lines to 6K465 were strongly associated with the protein levels of c-MYC and/or N-MYC. We also report DBPR728, an acyl-based prodrug of 6K465 bearing fewer hydrogen-bond donors that exhibited 10-fold improved oral bioavailability. DBPR728 induced durable tumor regression of c-MYC- and/or N-MYC- overexpressing xenografts including SCLC, triple-negative breast cancer (TNBC), hepatocellular carcinoma and medulloblastoma using a 5-on-2-off or once-a-week dosing regimen on a 21-day cycle. A single oral dose of DBPR728 at 300 mg/kg induced c-MYC reduction and cell apoptosis in the tumor xenografts for more than 7 days. The inhibitory effect of DBPR728 at a reduced dosing frequency was attributed to its uniquely high tumor/plasma ratio (3.6-fold within 7 days) and the long tumor half-life of active moiety 6K465. Furthermore, DBPR728 was found to synergize with the mTOR inhibitor everolimus to suppress c-MYC- or N-MYC- driven SCLC. Collectively, these results suggest DBPR728 has the potential to treat cancers overexpressing c-MYC- and/or N-MYC.

Natural fucoidans inhibit coronaviruses by targeting viral spike protein and host cell furin.

Fucoidans are a class of long chain sulfated polysaccharides and have multiple biological functions. Herein, four natural fucoidans extracted from Fucus vesiculosus, F. serratus, Laminaria japonica and Undaria pinnatifida, were tested for their HCoV-OC43 inhibition and found to demonstrate EC50 values ranging from 0.15 to 0.61 µg/mL. That from U. pinnatifida exhibited the most potent anti-HCoV-OC43 activity with an EC50 value of 0.15 ± 0.02 µg/mL, a potency largely independent of its sulfate content. Comparison of the gene expression profiles of fucoidan-treated and untreated cells infected with HCoV-OC43 revealed that fucoidan treatment effectively diminished HCoV-OC43 gene expressions associated with induced chemokines, cytokines and viral activities. Further studies using a highly fucoidan-resistant HCoV-OC43 determined that fucoidan inhibited HCoV-OC43 infection via interfering with viral entry and led to the identification of the specific site on the N-terminal region of spike protein, that located adjacent to the host cell receptor binding domain, targeted by the virus. Furthermore, in a SARS-CoV-2 pseudovirus neutralization assay, fucoidan also blocked SARS-CoV-2 entry. In vitro and in vivo, fucoidan decreased SARS-CoV-2 viral loads and inhibited viral infection in Calu-3 or Vero E6 cells and SARS-CoV-2 infected hamsters, respectively. Fucoidan was also found to inhibit furin activity, and reported furin inhibitors were found to inhibit viral infection by wild type HCoV-OC43 or SARS-CoV-2. Accordingly, we conclude that fucoidans inhibit coronaviral infection by targeting viral spike protein and host cell furin to interfere with viral entry.

Correction: Lee et al. The Synergistic Inhibition of Coronavirus Replication and Induced Cytokine Production by Ciclesonide and the Tylophorine-Based Compound Dbq33b. Pharmaceutics 2022, 14, 1511.

In the original publication [...].

Development of Furanopyrimidine-Based Orally Active Third-Generation EGFR Inhibitors for the Treatment of Non-Small Cell Lung Cancer.

The development of orally bioavailable, furanopyrimidine-based double-mutant (L858R/T790M) EGFR inhibitors is described. First, selectivity for mutant EGFR was accomplished by replacing the (S)-2-phenylglycinol moiety of 12 with either an ethanol or an alkyl substituent. Then, the cellular potency and physicochemical properties were optimized through insights from molecular modeling studies by implanting various solubilizing groups in phenyl rings A and B. Optimized lead 52 shows 8-fold selective inhibition of H1975 (EGFRL858R/T790M overexpressing) cancer cells over A431 (EGFRWT overexpressing) cancer cells; western blot analysis further confirmed EGFR mutant-selective target modulation inside the cancer cells by 52. Notably, 52 displayed in vivo antitumor effects in two different mouse xenograft models (BaF3 transfected with mutant EGFR and H1975 tumors) with TGI = 74.9 and 97.5% after oral administration (F = 27%), respectively. With an extraordinary kinome selectivity (S(10) score of 0.017), 52 undergoes detailed preclinical development.

Omicron-specific mRNA vaccine induced cross-protective immunity against ancestral SARS-CoV-2 infection with low neutralizing antibodies.

The major challenge in COVID-19 vaccine effectiveness is immune escape by SARS-CoV-2 variants. To overcome this, an Omicron-specific messenger RNA (mRNA) vaccine was designed. The extracellular domain of the spike of the Omicron variant was fused with a modified GCN4 trimerization domain with low immunogenicity (TSomi). After immunization with TSomi mRNA in hamsters, animals were challenged with SARS-CoV-2 virus. The raised nonneutralizing antibodies or cytokine secretion responses can recognize both Wuhan S and Omicron S. However, the raised antibodies neutralized SARS-CoV-2 Omicron virus infection but failed to generate Wuhan virus neutralizing antibodies. Surprisingly, TSomi mRNA immunization protected animals from Wuhan virus challenge. These data indicated that non-neutralizing antibodies or cellular immunity may play a more important role in vaccine-induced protection than previously believed. Next-generation COVID-19 vaccines using the Omicron S antigen may provide sufficient protection against ancestral or current SARS-CoV-2 variants.

BPR0C261, An Analogous of Microtubule Disrupting Agent D-24851 Enhances the Radiosensitivity of Human Non-Small Cell Lung Cancer Cells via p53-Dependent and p53-Independent Pathways.

(1) Destabilization of microtubule dynamics is a primary strategy to inhibit fast growing tumor cells. The low cytotoxic derivative of microtubule inhibitor D-24851, named BPR0C261 exhibits antitumor activity via oral administration. In this study, we investigated if BPR0C261 could modulate the radiation response of human non-small cell lung cancer (NSCLC) cells with or without p53 expression. (2) Different doses of BPR0C261 was used to treat human NSCLC A549 (p53+/+) cells and H1299 (p53-/-) cells. The cytotoxicity, radiosensitivity, cell cycle distribution, DNA damage, and protein expression were evaluated using an MTT assay, a colony formation assay, flow cytometry, a comet assay, and an immunoblotting analysis, respectively. (3) BPR0C261 showed a dose-dependent cytotoxicity on A549 cells and H1299 cells with IC50 at 0.38 µM and 0.86 µM, respectively. BPR0C261 also induced maximum G2/M phase arrest and apoptosis in both cell lines after 24 h of treatment with a dose-dependent manner. The colony formation analysis demonstrated that a combination of low concentration of BPR0C261 and X-rays caused a synergistic radiosensitizing effect on NSCLC cells. Additionally, we found that a low concentration of BPR0C261 was sufficient to induce DNA damage in these cells, and it increased the level of DNA damage induced by a fractionation radiation dose (2 Gy) of conventional radiotherapy. Furthermore, the p53 protein level of A549 cell line was upregulated by BPR0C261. On the other hand, the expression of PTEN tumor suppressor was found to be upregulated in H1299 cells but not in A549 cells under the same treatment. Although radiation could not induce PTEN in H1299 cells, a combination of low concentration of BPR0C261 and radiation could reverse this situation. (4) BPR0C261 exhibits specific anticancer effects on NSCLC cells by the enhancement of DNA damage and radiosensitivity with p53-dependent and p53-independent/PTEN-dependent manners. The combination of radiation and BPR0C261 may provide an important strategy for the improvement of radiotherapeutic treatment.

Protective Effect of CXCR4 Antagonist DBPR807 against Ischemia-Reperfusion Injury in a Rat and Porcine Model of Myocardial Infarction: Potential Adjunctive Therapy for Percutaneous Coronary Intervention.

CXCR4 antagonists have been claimed to reduce mortality after myocardial infarction in myocardial infarction (MI) animals, presumably due to suppressing inflammatory responses caused by myocardial ischemia-reperfusion injury, thus, subsequently facilitating tissue repair and cardiac function recovery. This study aims to determine whether a newly designed CXCR4 antagonist DBPR807 could exert better vascular-protective effects than other clinical counterparts (e.g., AMD3100) to alleviate cardiac damage further exacerbated by reperfusion. Consequently, we find that instead of traditional continuous treatment or multiple-dose treatment at different intervals of time, a single-dose treatment of DBPR807 before reperfusion in MI animals could attenuate inflammation via protecting oxidative stress damage and preserve vascular/capillary density and integrity via mobilizing endothelial progenitor cells, leading to a desirable fibrosis reduction and recovery of cardiac function, as evaluated with the LVEF (left ventricular ejection fraction) in infarcted hearts in rats and mini-pigs, respectively. Thus, it is highly suggested that CXCR4 antagonists should be given at a single high dose prior to reperfusion to provide the maximal cardiac functional improvement. Based on its favorable efficacy and safety profiles indicated in tested animals, DBPR807 has a great potential to serve as an adjunctive medicine for percutaneous coronary intervention (PCI) therapies in acute MI patients.

Benzo[b]thiophene-2-carboxamides as novel opioid receptor agonists with potent analgesic effect and reduced constipation.

Currently, there is a significant unmet need for novel analgesics with fewer side effects. In this study, we carried out structural modification of a hit compound previously identified in an artificial-intelligence (AI) virtual screening and discovered the potent analgesic, benzo[b]thiophene-2-carboxamide analog (compound 25) with new structural scaffold. We investigated the signaling pathways of opioid receptors mediated by compound 25, and found this racemic compound activated mu-opioid receptor through the cyclic adenosine monophosphate (cAMP) and ß-arrestin-2-mediated pathways with strong potency and efficacy, and accompanying nociceptin-orphanin FQ opioid peptide and delta-opioid receptors through the cAMP pathway with weak potencies. Compound 25 elicited potent antinociception in thermal-stimulated pain (ED50 value of 127.1 ± 34.65 µg/kg) and inflammatory-induced allodynia models with less gastrointestinal transit inhibition and antinociceptive tolerance than morphine. Overall, this study revealed a novel analgesic with reduced risks of side effects.

Targeting the Phosphatidylserine-Immune Checkpoint with a Small-Molecule Maytansinoid Conjugate.

Ligand-targeting drug delivery systems have made significant strides for disease treatments with numerous clinical approvals in this era of precision medicine. Herein, we report a class of small molecule-based immune checkpoint-targeting maytansinoid conjugates. From the ligand targeting ability, pharmacokinetics profiling, in vivo anti-pancreatic cancer, triple-negative breast cancer, and sorafenib-resistant liver cancer efficacies with quantitative mRNA analysis of treated-tumor tissues, we demonstrated that conjugate 40a not only induced lasting regression of tumor growth, but it also rejuvenated the once immunosuppressive tumor microenvironment to an "inflamed hot tumor" with significant elevation of gene expressions that were not accessible in the vehicle-treated tumor. In turn, the immune checkpoint-targeting small molecule drug conjugate from this work represents a new pharmacodelivery strategy that can be expanded with combination therapy with existing immune-oncology treatment options.

Peritoneal effluent MicroRNA profile for detection of encapsulating peritoneal sclerosis.

BACKGROUND: Encapsulating peritoneal sclerosis (EPS) is a catastrophic complication of peritoneal dialysis (PD) with high mortality. Our aim is to develop a novel noninvasive microRNA (miRNA) test for EPS. METHODS: We collected 142 PD effluents (EPS: 62 and non-EPS:80). MiRNA profiles of PD effluents were examined by a high-throughput real-time polymerase chain reaction (PCR) array to first screen. Candidate miRNAs were verified by single real-time PCR. The model for EPS prediction was evaluated by multiple logistic regression and machine learning. RESULTS: Seven candidate miRNAs were identified from the screening of PCR-array of 377 miRNAs. The top five area under the curve (AUC) values with 5 miRNA-ratios were selected using 127 samples (EPS: 56 vs non-EPS: 71) to produce a receiver operating characteristic curve. After considering clinical characteristics and 5 miRNA-ratios, the accuracies of the machine learning model of Random Forest and multiple logistic regression were boosted to AUC 0.97 and 0.99, respectively. Furthermore, the pathway analysis of miRNA associated targeting genes and miRNA-compound interaction network revealed that these five miRNAs played the roles in TGF-ß signaling pathway. CONCLUSION: The model-based miRNA expressions in PD effluents may help determine the probability of EPS and provide further therapeutic opinion for EPS.

Synthesis and Evaluation of Small Molecule Drug Conjugates Harnessing Thioester-Linked Maytansinoids.

Ligand-targeting drug conjugates are a class of clinically validated biopharmaceutical drugs constructed by conjugating cytotoxic drugs with specific disease antigen targeting ligands through appropriate linkers. The integrated linker-drug motif embedded within such a system can prevent the premature release during systemic circulation, thereby allowing the targeting ligand to engage with the disease antigen and selective accumulation. We have designed and synthesized new thioester-linked maytansinoid conjugates. By performing in vitro cytotoxicity, targeting ligand binding assay, and in vivo pharmacokinetic studies, we investigated the utility of this new linker-drug moiety in the small molecule drug conjugate (SMDC) system. In particular, we conjugated the thioester-linked maytansinoids to the phosphatidylserine-targeting small molecule zinc dipicolylamine and showed that Zn8_DM1 induced tumor regression in the HCC1806 triple-negative breast cancer xenograft model. Moreover, in a spontaneous sorafenib-resistant liver cancer model, Zn8_DM1 exhibited potent antitumor growth efficacy. From quantitative mRNA analysis of Zn8_DM1 treated-tumor tissues, we observed the elevation of gene expressions associated with a "hot inflamed tumor" state. With the identification and validation of a plethora of cancer-associated antigens in the "omics" era, this work provided the insight that antibody- or small molecule-based targeting ligands can be conjugated similarly to generate new ligand-targeting drug conjugates.

The Synergistic Inhibition of Coronavirus Replication and Induced Cytokine Production by Ciclesonide and the Tylophorine-Based Compound Dbq33b.

Ciclesonide is an inhaled corticosteroid used to treat asthma and has been repurposed as a treatment for mildly ill COVID-19 patients, but its precise mechanism of action is unclear. Herein, we report that ciclesonide blocks the coronavirus-induced production of the cytokines IL-6, IL-8, and MCP-1 by increasing IκBα protein levels and significantly decreasing p65 nuclear translocation. Furthermore, we found that the combination of ciclesonide and dbq33b, a potent tylophorine-based coronavirus inhibitor that affects coronavirus-induced NF-κB activation a little, additively and synergistically decreased coronavirus-induced IL-6, IL-8, and MCP-1 cytokine levels, and synergistically inhibited the replication of both HCoV-OC43 and SARS-CoV-2. Collectively, the combination of ciclesonide and dbq33b merits consideration as a treatment for COVID-19 patients who may otherwise be overwhelmed by high viral loads and an NF-κB-mediated cytokine storm.

Selective and antagonist-dependent µ-opioid receptor activation by the combination of 2-{[2-(6-chloro-3,4-dihydro-1(2H)-quinolinyl)-2-oxoethyl]sulfanyl}-5-phenyl-4,6-(1H,5H)-pyrimidinedione and naloxone/naltrexone.

We identified, via high-throughput screening using a FLIPR® calcium assay, compound 1, which incorporated a dihydroquinolinyl-2-oxoethylsulfanyl-(1H,5H)-pyrimidinedione core and activated the µ-opioid receptor (MOR) in the presence of naloxone or naltrexone. A structure-activity relationship study of the analogs of 1 led to the design of compound 21, which activated MOR in the presence of naloxone with an EC50 of 3.3 ± 0.2 µM. MOR activation by the compound 21-antagonist pair was antagonist-dependent. Compound 21 did not affect the potency of the orthosteric agonist, morphine, toward MOR, indicating that it affected the function of MOR antagonists rather than that of the agonists. Computer modeling of the compound 21-MOR-naloxone complex revealed major interactions between compound 21 and MOR, including hydrogen bonding with Ser196, π-π stacking with Tyr149, and sulfur-aromatic interaction with Trp192. This study may pave the way for developing agents capable of safe and effective MOR modulation.

Furin and TMPRSS2 Resistant Spike Induces Robust Humoral and Cellular Immunity Against SARS-CoV-2 Lethal Infection.

An effective COVID-19 vaccine against broad SARS-CoV-2 variants is still an unmet need. In the study, the vesicular stomatitis virus (VSV)-based vector was used to express the SARS-CoV-2 Spike protein to identify better vaccine designs. The replication-competent of the recombinant VSV-spike virus with C-terminal 19 amino acid truncation (SΔ19 Rep) was generated. A single dose of SΔ19 Rep intranasal vaccination is sufficient to induce protective immunity against SARS-CoV-2 infection in hamsters. All the clones isolated from the SΔ19 Rep virus contained R682G mutation located at the Furin cleavage site. An additional S813Y mutation close to the TMPRSS2 cleavage site was identified in some clones. The enzymatic processing of S protein was blocked by these mutations. The vaccination of the R682G-S813Y virus produced a high antibody response against S protein and a robust S protein-specific CD8+ T cell response. The vaccinated animals were protected from the lethal SARS-CoV-2 (delta variant) challenge. The S antigen with resistance to enzymatic processes by Furin and TMPRSS2 will provide better immunogenicity for vaccine design.

Discovery of Potential Neuroprotective Agents against Paclitaxel-Induced Peripheral Neuropathy.

Chemotherapy-induced neurotoxicity is a common adverse effect of cancer treatment. No medication has been shown to be effective in the prevention or treatment of chemotherapy-induced neurotoxicity. Using minoxidil as an initial template for structural modifications in conjunction with an in vitro neurite outgrowth assay, an image-based high-content screening platform, and mouse behavior models, an effective neuroprotective agent CN016 was discovered. Our results showed that CN016 could inhibit paclitaxel-induced inflammatory responses and infiltration of immune cells into sensory neurons significantly. Thus, the suppression of proinflammatory factors elucidates, in part, the mechanism of action of CN016 on alleviating paclitaxel-induced peripheral neuropathy. Based on excellent efficacy in improving behavioral functions, high safety profiles (MTD > 500 mg/kg), and a large therapeutic window (MTD/MED > 50) in mice, CN016 might have great potential to become a peripherally neuroprotective agent to prevent neurotoxicity caused by chemotherapeutics as typified by paclitaxel.

Discovery of BPR1R024, an Orally Active and Selective CSF1R Inhibitor that Exhibits Antitumor and Immunomodulatory Activity in a Murine Colon Tumor Model.

Colony-stimulating factor-1 receptor (CSF1R) is implicated in tumor-associated macrophage (TAM) repolarization and has emerged as a promising target for cancer immunotherapy. Herein, we describe the discovery of orally active and selective CSF1R inhibitors by property-driven optimization of BPR1K871 (9), our clinical multitargeting kinase inhibitor. Molecular docking revealed an additional nonclassical hydrogen-bonding (NCHB) interaction between the unique 7-aminoquinazoline scaffold and the CSF1R hinge region, contributing to CSF1R potency enhancement. Structural studies of CSF1R and Aurora kinase B (AURB) demonstrated the differences in their back pockets, which inspired the use of a chain extension strategy to diminish the AURA/B activities. A lead compound BPR1R024 (12) exhibited potent CSF1R activity (IC50 = 0.53 nM) and specifically inhibited protumor M2-like macrophage survival with a minimal effect on antitumor M1-like macrophage growth. In vivo, oral administration of 12 mesylate delayed the MC38 murine colon tumor growth and reversed the immunosuppressive tumor microenvironment with the increased M1/M2 ratio.

Engineering Nano- and Microparticles as Oral Delivery Vehicles to Promote Intestinal Lymphatic Drug Transport.

Targeted oral delivery of a drug via the intestinal lymphatic system (ILS) has the advantages of protecting against hepatic first-pass metabolism of the drug and improving its pharmacokinetic performance. It is also a promising route for the oral delivery of vaccines and therapeutic agents to induce mucosal immune responses and treat lymphatic diseases, respectively. This article describes the anatomical structures and physiological characteristics of the ILS, with an emphasis on enterocytes and microfold (M) cells, which are the main gateways for the transport of particulate delivery vehicles across the intestinal epithelium into the lymphatics. A comprehensive overview of recent advances in the rational engineering of particulate vehicles, along with the challenges and opportunities that they present for improving ILS drug delivery, is provided, and the mechanisms by which such vehicles target and transport through enterocytes or M cells are discussed. The use of naturally sourced materials, such as yeast microcapsules and their derived polymeric ß-glucans, as novel ILS-targeting delivery vehicles is also reviewed. Such use is the focus of an emerging field of research. Their potential use in the oral delivery of nucleic acids, such as mRNA vaccines, is proposed.

Remdesivir and Cyclosporine Synergistically Inhibit the Human Coronaviruses OC43 and SARS-CoV-2.

Remdesivir, a prodrug targeting RNA-dependent-RNA-polymerase, and cyclosporine, a calcineurin inhibitor, individually exerted inhibitory activity against human coronavirus OC43 (HCoV-OC43) in HCT-8 and MRC-5 cells at EC50 values of 96 ± 34 ∼ 85 ± 23 nM and 2,920 ± 364 ∼ 4,419 ± 490 nM, respectively. When combined, these two drugs synergistically inhibited HCoV-OC43 in both HCT-8 and MRC-5 cells assayed by immunofluorescence assay (IFA). Remdesivir and cyclosporine also separately reduced IL-6 production induced by HCoV-OC43 in human lung fibroblasts MRC-5 cells with EC50 values of 224 ± 53 nM and 1,292 ± 352 nM, respectively; and synergistically reduced it when combined. Similar trends were observed for SARS-CoV-2, which were 1) separately inhibited by remdesivir and cyclosporine with respective EC50 values of 3,962 ± 303 nM and 7,213 ± 143 nM by IFA, and 291 ± 91 nM and 6,767 ± 1,827 nM by a plaque-formation assay; and 2) synergistically inhibited by their combination, again by IFA and plaque-formation assay. Collectively, these results suggest that the combination of remdesivir and cyclosporine merits further study as a possible treatment for COVID-19 complexed with a cytokine storm.