Gustatory Receptor 206 Participates in the Foraging Behavior of Larvae of Polyphagous Pest Spodoptera litura.

Insect gustatory receptors (GRs) aid in the precise identification of deterrent or stimulant compounds associated with food, mating, and egg-laying. Thus, they are promising targets for developing efficient insecticides. Here, 61 GRs in the chemosensory organs of Spodoptera litura larvae and adults were identified. Among them, SlitGR206 exhibited larval labium (LL)-specific expression characteristics. To explore the role of SlitGR206, a bacterial expression system was established to produce high-quality double-stranded RNA (dsRNA) and suppress SlitGR206 expression in LL. Subsequent behavioral assessments revealed that SlitGR206 silencing influenced larval feeding preferences and absorption. Moreover, it was found to reduce the ability of larvae to forage the five crucial host odorants. These findings demonstrate that SlitGR206 likely plays an indirect regulatory role in host recognition, consequently affecting foraging behavior. This provides a crucial foundation for the analysis of functional diversity among insect GRs and the precise development of nucleic acid pesticides in the future.

Development of a Novel DNA Mono-alkylator Platform for Antibody-Drug Conjugates.

Although microtubule inhibitors (MTI) remain a therapeutically valuable payload option for antibody-drug conjugates (ADC), some cancers do not respond to MTI-based ADCs. Efforts to fill this therapeutic gap have led to a recent expansion of the ADC payload "toolbox" to include payloads with novel mechanisms of action such as topoisomerase inhibition and DNA cross-linking. We present here the development of a novel DNA mono-alkylator ADC platform that exhibits sustained tumor growth suppression at single doses in MTI-resistant tumors and is well tolerated in the rat upon repeat dosing. A phosphoramidate prodrug of the payload enables low ADC aggregation even at drug-to-antibody ratios of 5:1 while still delivering a bystander-capable payload that is effective in multidrug resistant (MDR)-overexpressing cell lines. The platform was comparable in xenograft studies to the clinical benchmark DNA mono-alkylator ADC platform DGN459 but with a significantly better tolerability profile in rats. Thus, the activity and tolerability profile of this new platform make it a viable option for the development of ADCs.

Discovery and Preclinical Characterization of XMT-1660, an Optimized B7-H4-Targeted Antibody-Drug Conjugate for the Treatment of Cancer.

Antibody-drug conjugates (ADC) achieve targeted drug delivery to a tumor and have demonstrated clinical success in many tumor types. The activity and safety profile of an ADC depends on its construction: antibody, payload, linker, and conjugation method, as well as the number of payload drugs per antibody [drug-to-antibody ratio (DAR)]. To allow for ADC optimization for a given target antigen, we developed Dolasynthen (DS), a novel ADC platform based on the payload auristatin hydroxypropylamide, that enables precise DAR-ranging and site-specific conjugation. We used the new platform to optimize an ADC that targets B7-H4 (VTCN1), an immune-suppressive protein that is overexpressed in breast, ovarian, and endometrial cancers. XMT-1660 is a site-specific DS DAR 6 ADC that induced complete tumor regressions in xenograft models of breast and ovarian cancer as well as in a syngeneic breast cancer model that is refractory to PD-1 immune checkpoint inhibition. In a panel of 28 breast cancer PDXs, XMT-1660 demonstrated activity that correlated with B7-H4 expression. XMT-1660 has recently entered clinical development in a phase I study (NCT05377996) in patients with cancer.

Functional differentiation of two general odorant-binding proteins to sex pheromones in Spodoptera frugiperda.

A precise chemosensory system can help insects complete various important behavioral responses by accurately identifying different external odorants. Therefore, deeply understanding the mechanism of insect recognition of important odorants will help us develop efficient and environmentally-friendly behavioral inhibitors. Spodoptera frugiperda is a polyphagous pest that feeds on >350 different host plants worldwide and also harms maize production in China. However, the molecular mechanism of the first step for males to use odorant-binding proteins (OBPs) to recognize sex pheromones remains unclear. Here, we obtained 50 OBPs from the S. frugiperda genome, and the expression level of SfruGOBP1 in females was significantly higher than that in males, whereas SfruGOBP2 displayed male-biased expression. Fluorescence competitive binding assays showed that only SfruGOBP2 showed binding affinities for the four sex pheromones of female S. frugiperda. Subsequently, we identified some key amino acid residues that can participate in the interaction between SfruGOBP2 and sex pheromones using molecular docking and site-directed mutagenesis methods. These findings will help us explore the interaction mechanism between GOBPs and sex pheromones in moths, and provide important target genes for developing new mating inhibitors of S. frugiperda in the future.

Discovery of novel polyamide-pyrrolobenzodiazepine hybrids for antibody-drug conjugates.

Pyrrolobenzodiazepine (PBD) dimers are well-known highly potent antibody drug conjugate (ADC) payloads. The corresponding PBD monomers, in contrast, have received much less attention from the ADC community. We prepared several novel polyamide-linked PBD monomers and evaluated their utility as ADC payloads. The unconjugated polyamide-PBD hybrids exhibited potent antiproliferative activity (IC50 range: 10-11-10-8 M) against a variety of HER2-expressing cancer cell lines. Several peptide-linked variants of the lead compound were prepared and conjugated to trastuzumab to afford ADCs with drug-to-antibody (DAR) ratios ranging from 3 to 5. The ADCs exhibited antigen-dependent cytotoxicity in vitro and potently suppressed tumor xenograft growth in vivo in a target-dependent manner. Moreover, the ADCs were well-tolerated in both mouse and rat. This work demonstrates for the first time that PBD polyamide hybrids can serve as effective ADC payloads.

Salvianolic acid B regulates collagen synthesis: Indirect influence on human dermal fibroblasts through the microvascular endothelial cell pathway.

BACKGROUND: Salvianolic acid B (SAB) is one of the main active ingredients of Salvia Miltiorrhiza. It has significant skin anti-aging, whitening, and sun protection properties. AIMS: The study aimed at studying the mechanism underlying the effect of salvianolic acid Bon collagen synthesis, which has good anti-aging efficacy and modulates microcirculation. METHODS: This study employed available public databases, bioinformatics methodologies, and the inverse docking approach to explore the effectiveness of SAB in the regulating collagen synthesis, and then used an human dermal fibroblast (HDF)- Human dermal microvascular endothelial cell (HDMEC) in vitro model to validate the predicted mechanism of SAB in influencing collagen synthesis. RESULTS: The results showed that NO production in SAB-treated HDMEC-conditioned medium was increased compared to that in control media, and the same tendency was also observed for growth factor production. SAB also upregulated HDMEC cellular eNOS and VEGF. When SAB-treated HDMEC conditioned medium was transferred to HDFs, the expression of collagen I, collagen III, and elastin in HDFs was upregulated and MMP-1 was downregulated. CONCLUSIONS: The results show that SAB regulates collagen through the HDMEC-HDF pathway. Furthermore, the mechanisms might be closely related to the microcirculation factors NO and VEGF.

Influence of different nitrogen sources on carbon and nitrogen metabolism and gene expression in tea plants (Camellia sinensis L.).

Nitrogen plays an important role in plant growth and development, with different nitrogen forms also having an impact on carbon/nitrogen metabolism. Unlike most plants, tea plants prefer ammonium over nitrate. In this paper, we focused on how different nitrogen sources regulate the carbon/nitrogen metabolism in tea plants. Tea seedlings of 'Longjing 43' were cultivated hydroponically in four different solutions (zero-nitrogen, only NH4+, only NO3- and mixed nitrogen (NH4+: NO3- = 1:1). We analyzed characteristic components of tea plants and related genes in carbon and nitrogen metabolism. Tea polyphenols and catechins representing carbon pool, increased when NO3- was supplied as the nitrogen source, and similar findings were recorded in the zero-nitrogen treatment. The expression of most catechins biosynthesis-related genes was up regulated under NO3- and zero-N treatment, that was associated with tea polyphenols and catechins changes. Compared with NO3- as the nitrogen source, NH4+ and mixed nitrogen treatments had a positive effect on the accumulation of amino acids, especially theanine, glutamate and arginine, and these components contribute to the freshness flavor of tea. The expression of ammonium-assimilation genes was also up-regulated with NH4+ supply. Under mixed nitrogen treatment, the ratio of total polyphenols to free amino acids (PP/AA) was between sole NH4+ and NO3- supply. Therefore, compared with single nitrogen source, carbon and nitrogen metabolism of tea plant was more balanced under mixed nitrogen treatment. The results suggested that NO3- as the nitrogen source promoted the biosynthesis of catechins enriching the carbon pool, whereas NH4+ supply was more conducive to nitrogen metabolism, indicating that different nitrogen sources could affect the carbon and nitrogen balance.

Dolaflexin: A Novel Antibody-Drug Conjugate Platform Featuring High Drug Loading and a Controlled Bystander Effect.

After significant effort over the last 30 years, antibody-drug conjugates (ADC) have recently gained momentum as a therapeutic modality, and nine ADCs have been approved by the FDA to date, with additional ADCs in late stages of development. Here, we introduce dolaflexin, a novel ADC technology that overcomes key limitations of the most common ADC platforms with two key features: a higher drug-to-antibody ratio and a novel auristatin with a controlled bystander effect. The novel, cell permeable payload, auristatin F-hydroxypropylamide, undergoes metabolic conversion to the highly potent, but less cell permeable auristatin F to balance the bystander effect through drug trapping within target cells. We conducted studies in mice, rats, and cynomolgus monkeys to complement in vitro characterization and contrasted the performance of dolaflexin with regard to antitumor activity, pharmacokinetic properties, and safety in comparison with the ADC platform utilized in the approved ADC ado-trastuzumab emtansine (T-DM1). A HER2-targeted dolaflexin ADC was shown to have a much lower threshold of antigen expression for potent cell killing in vitro, was effective in vivo in tumors with low HER2 expression, and induced tumor regressions in a xenograft model that is resistant to T-DM1.

The Dolaflexin-based Antibody-Drug Conjugate XMT-1536 Targets the Solid Tumor Lineage Antigen SLC34A2/NaPi2b.

Target selection for antibody-drug conjugates (ADC) frequently focuses on identifying antigens with differential expression in tumor and normal tissue, to mitigate the risk of on-target toxicity. However, this strategy restricts the possible target space. SLC34A2/NaPi2b is a sodium phosphate transporter expressed in a variety of human tumors including lung and ovarian carcinoma, as well as the normal tissues from which these tumors arise. Previous clinical trials with a NaPi2b targeting MMAE-ADCs have shown objective durable responses. However, the protein-based biomarker assay developed for use in that study was unable to discern a statistically significant relationship between NaPi2b protein expression and the probability of response. XMT-1536 is a NaPi2b targeting ADC comprised of a unique humanized antibody conjugated with 10-15 auristatin F- hydroxypropylamide (AF-HPA) payload molecules via the Dolaflexin platform. AF-HPA is a cell-permeable, antimitotic compound that is slowly metabolized intratumorally to an active, very low-permeable metabolite, auristatin F (AF), resulting in controlled bystander killing. We describe the preclinical in vitro and in vivo antitumor effects of XMT-1536 in models of ovarian and lung adenocarcinoma. Pharmacokinetic analysis showed approximately proportional increases in exposure in rat and monkey. Systemic free AF-HPA and AF concentrations were observed to be low in all animal species. Finally, we describe a unique IHC reagent, generated from a chimeric construct of the therapeutic antibody, that was used to derive a target expression and efficacy relationship in a series of ovarian primary xenograft cancer models.

Isoquercitrin suppresses the expression of histamine and pro-inflammatory cytokines by inhibiting the activation of MAP Kinases and NF-κB in human KU812 cells.

Mast cells and basophils are multifunctional effector cells that contain abundant secretory granules in their cytoplasm. Both cell types are involved in a variety of inflammatory and immune events, producing an array of inflammatory mediators, such as cytokines. The aim of the study was to examine whether isoquercitrin modulates allergic and inflammatory reactions in the human basophilic KU812 cells and to elucidate its influence on the phosphorylation of mitogen-activated protein kinase (MAPK) and nuclear factor (NF)-κB activation. The KU812 cells were stimulated with phorbol-12-myristate 13-acetate plus the calcium ionophore A23187 (PMACI). The inhibitory effects of isoquercitrin on the productions of histamine and pro-inflammatory cytokines in the stimulated KU812 cells were measured using cytokine-specific enzyme-linked immunosorbent (ELISA) assays. Western blotting analysis was used to assess the effects of isoquercitrin on the MAPKs and NF-κB protein levels. Our results indicated that the isoquercitrin treatment of PMACI-stimulated KU812 cells significantly reduced the production of histamine and the pro-inflammatory cytokines, such as interleukin (IL)-6, IL-8, IL-1ß, and tumor necrosis factor (TNF)-α. The treated cells exhibited decreased phosphorylation of extracellular signal-regulated kinase (ERK), revealing the role of ERK MAPK in isoquercitrin-mediated allergy inhibition. Furthermore, isoquercitrin suppressed the PMACI-mediated activation of NF-κB in the human basophil cells. In conclusion, the results from the present study provide insights into the potential therapeutic use of isoquercitrin for the treatment of inflammatory and allergic reactions.

Methylamine irisolidone, a novel compound, increases total ATPase activity and inhibits apoptosis in vivo and in vitro.

We propose to further research the protective effect of MMI on myocardium ischemic rat model and H9c2 cells that underwent cell apoptosis induced by hypoxia. We established the myocardium ischemic rat model via the cardiac surgical procedures in vivo and treated the model rats with different concentration of MMI. In vitro, with the pretreatment of MMI for 12 h in the model of Na2S2O4-induced hypoxia injury, the H9c2 cells viability was determined by MTT assay. We found that MMI had significantly improved cardiac function of the myocardial ischemia, and significantly decreased the reactive oxygen species level. The expression of P53, Bcl-2, Bax, and caspase-9 was also induced by MMI. In vitro study revealed a concentration-dependent increase in cell viability associated with MMI pretreatment. Annexin V-FITC and PI staining results showed that MMI had a preventive effect on hypoxia-induced apoptosis in H9c2 cells. MMI also inhibited the mitochondrial membrane potential decrease and increased total ATPase activity during hypoxia in H9c2 cells. In conclusion, MMI can enhance the cardiac function in myocardial ischemic rat and increase cell viability and attenuate the apoptosis in H9c2 cells induced by hypoxia, which was associated with inhibiting MMP decreasion and increasing total ATPase activity.

A Polymer-Based Antibody-Vinca Drug Conjugate Platform: Characterization and Preclinical Efficacy.

Antibody-drug conjugates (ADC) are an emerging drug class that uses antibodies to improve cytotoxic drug targeting for cancer treatment. ADCs in current clinical trials achieve a compromise between potency and physicochemical/pharmacokinetic properties by conjugating potent cytotoxins directly to an antibody at a 4:1 or less stoichiometric ratio. Herein, we report a novel, polyacetal polymer-based platform for creating ADC that use poly-1-hydroxymethylethylene hydroxymethyl-formal (PHF), also known as Fleximer. The high hydrophilicity and polyvalency properties of the Fleximer polymer can be used to produce ADC with high drug loading without compromising physicochemical and pharmacokinetic properties. Using trastuzumab and a vinca drug derivative to demonstrate the utility of this platform, a novel Fleximer-based ADC was prepared and characterized in vivo. The ADC prepared had a vinca-antibody ratio of 20:1. It exhibited a high antigen-binding affinity, an excellent pharmacokinetic profile and antigen-dependent efficacy, and tumor accumulation in multiple tumor xenograft models. Our findings illustrate the robust utility of the Fleximer platform as a highly differentiated alternative to the conjugation platforms used to create ADC currently in clinical development.

Activity of factor VII in patients with isolated blunt traumatic brain injury: association with coagulopathy and progressive hemorrhagic injury.

BACKGROUND: Given the importance of factor VII (FVII) in extrinsic pathway of coagulation cascade, we sought to elucidate the relationship between FVII and traumatic brain injury-induced coagulopathy and progressive hemorrhagic injury (PHI). METHODS: Eighty-one patients with isolated traumatic brain injury, 16 years or older, were recruited between 2010 and 2012. Blood was collected on arrival in the emergency department and analyzed with activated partial thromboplastic time, international normalized ratio, platelet count, and activity of FVII. Coagulopathy was defined as thrombocytopenia (platelet count < 120,000/µL) or elevated international normalized ratio of greater than 1.2 or prolonged activated partial thromboplastin time greater than 40 seconds at admission. PHI was present when the follow-up computed tomographic scan reported any increase in size or number of the hemorrhagic lesions. Logistic regression examined the risks for coagulopathy and PHI. RESULTS: Mean (SD) FVII activity in patients with coagulopathy was 85.69% (34.88%), which was significantly lower than patients without coagulopathy (99.57% [29.37%], p = 0.04). Isolated traumatic brain injury patients with FVII activity less than 77.5% have an odds ratio for coagulopathy of 5.52 (95% confidence interval, 1.82-16.68; p = 0.03) relative to patients with FVII activity of 77.5% or greater. Mean (SD) FVII activity in patients with PHI was 70.76% (18.21%), which was significantly lower than patients without PHI (105.76% [32.27%], p < 0.001). A stepwise logistic regression analysis identified FVII less than 77.5% (odds ratio, 4.53; 95% confidence interval, 1.62-12.67; p = 0.004) as a predisposing risk factors independently associated with the presence of PHI. The overall mortality rate in the study population was 7.4% (n = 6). The plasma FVII in death patients (91.44% [47.19%]) was slightly lower than that in survival patients (92.01% [32.04%]). However, there was no statistical difference between the two groups (p = 0.95). CONCLUSION: A decrease of FVII activity significantly contributes to the coagulopathy and PHI in patients with isolated traumatic brain injury. LEVEL OF EVIDENCE: Prognostic study, level III.