On Demand Bioorthogonal Switching of an Antibody-Conjugated SPECT Probe to a Cytotoxic Payload: from Imaging to Therapy.

Antibody-drug conjugates (ADCs) for the treatment of cancer aim to achieve selective delivery of a cytotoxic payload to tumor cells while sparing normal tissue. In vivo, multiple tumor-dependent and -independent processes act on ADCs and their released payloads to impact tumor-versus-normal delivery, often resulting in a poor therapeutic window. An ADC with a labeled payload would make synchronous correlations between distribution and tissue-specific pharmacological effects possible, empowering preclinical and clinical efforts to improve tumor-selective delivery; however, few methods to label small molecules without destroying their pharmacological activity exist. Herein, we present a bioorthogonal switch approach that allows a radiolabel attached to an ADC payload to be removed tracelessly at will. We exemplify this approach with a potent DNA-damaging agent, the pyrrolobenzodiazepine (PBD) dimer, delivered as an antibody conjugate targeted to lung tumor cells. The radiometal chelating group, DOTA, was attached via a novel trans-cyclooctene (TCO)-caged self-immolative para-aminobenzyl (PAB) linker to the PBD, stably attenuating payload activity and allowing tracking of biodistribution in tumor-bearing mice via SPECT-CT imaging (live) or gamma counting (post-mortem). Following TCO-PAB-DOTA reaction with tetrazines optimized for extra- and intracellular reactivity, the label was removed to reveal the unmodified PBD dimer capable of inducing potent tumor cell killing in vitro and in mouse xenografts. The switchable antibody radio-drug conjugate (ArDC) we describe integrates, but decouples, the two functions of a theranostic given that it can serve as a diagnostic for payload delivery in the labeled state, but can be switched on demand to a therapeutic agent (an ADC).

The relativity analysis of hypoxia inducible factor-1α in pulmonary arterial hypertension (ascites syndrome) in broilers: a review.

Ascites syndrome (AS) in broiler chickens, also known as pulmonary arterial hypertension (PAH), is a significant disease in the poultry industry. It is a nutritional metabolic disease that is closely associated with hypoxia-inducible factors and rapid growth. The rise in pulmonary artery pressure is a crucial characteristic of AS and is instrumental in its development. Hypoxia-inducible factor 1α (HIF-1α) is an active subunit of a key transcription factor in the oxygen-sensing pathway. HIF-1α plays a vital role in oxygen homeostasis and the development of pulmonary hypertension. Studying the effects of HIF-1α on pulmonary hypertension in humans or mammals, as well as ascites in broilers, can help us understand the pathogenesis of AS. Therefore, this review aims to (1) summarize the mechanism of HIF-1α in the development of pulmonary hypertension, (2) provide theoretical significance in explaining the mechanism of HIF-1α in the development of pulmonary arterial hypertension (ascites syndrome) in broilers, and (3) establish the correlation between HIF-1α and pulmonary arterial hypertension (ascites syndrome) in broilers. HIGHLIGHTSExplains the hypoxic mechanism of HIF-1α.Linking HIF-1α to pulmonary hypertension in broilers.Explains the role of microRNAs in pulmonary arterial hypertension in broilers.

Transradial and Transfemoral Approaches for Vertebral Artery Stenting: A Single-Center, Prospective, Non-Inferiority Comparison.

BACKGROUND Vertebral artery origin stenosis (VAOS) has recently gained increased attention, with endovascular treatments like stent implantation showing high success and low complication rates, although less is known about VAOS compared to carotid artery stenosis. This study evaluated the safety and effectiveness of transradial (TRA) and transfemoral (TFA) approaches for VAOS stent placement. MATERIAL AND METHODS We recruited a total of 102 patients undergoing vertebral artery stenting in our hospital between January 2020 and November 2022. Patients were randomly assigned to undergo either radial or femoral approach for stent implantation in the vertebral artery, and the radial approach group secondary divided into 2 groups by patients' consent: ipsilateral or contralateral radial approach. The success rates of VAOS stent implantation, operation time, and postoperative hospitalization time were compared between the 3 groups. In addition, we compared the outcomes of stroke within 30 days, transient ischemic attack (TIA) within 30 days, and other indicators. RESULTS Of the 102 patients, the final success rate of stent implantation was not significantly different between the 3 groups. The time from sheath insertion to stent insertion in the ipsilateral TRA group (median time: 19 min [interquartile range (IQR): 12-24.5 min]) was significantly shorter than in the transfemoral approach (TFA) group (median time: 29 min [IQR: 21-35.5 min]) (P<0.01; 95% confidence interval (95% CI): 10 min [6-14 min]). There were no statistically significant differences between the 3 groups in terms of cerebrovascular events within 1 month, and patient satisfaction and preference favored the radial approach. CONCLUSIONS The postoperative hospitalization time and operation time associated with the ipsilateral TRA were shorter, and patient acceptance and satisfaction were higher.

Design, Synthesis, and Antitumor Activity Evaluation of Artemisinin Bivalent Ligands.

Five artemisinin bivalent ligands molecules 4a-4e were designed, synthesized, and confirmed by 1H NMR, 13C NMR, and low-resolution mass spectrometry, and the bioactivities of the target compounds were investigated against four human tumor cell lines in vitro, including BGC-823, HepG-2, MCF-7, and HCT-116. The results showed 4a, 4d, and 4e exhibited significantly tumor cell inhibitory activity compared with the artemisinin and dihydroartemisinin; compound 4e has good biological activity inhibiting BGC-823 with an IC50 value of 8.30 µmol/L. Then, the good correlations with biological results were validated by molecular docking through the established bivalent ligands multi-target model, which showed that 4e could bind well with the antitumor protein MMP-9.

[Different concentrations of adapalene induce differentiation and apoptosis of SH-SY5Y cells]. / 不同浓度阿达帕林诱导SH-SY5Y细胞的分化及凋亡.

OBJECTIVES: To investigate the effects of different concentrations of adapalene on the morphology and functions of neuroblastoma cell line SH-SY5Y, as well as its role in inducing cell differentiation and apoptosis. METHODS: SH-SY5Y cells were divided into control group, low concentration (0.1 µM and 1 µM) adapalene groups, and high concentration (10 µM) adapalene group. Time-lapse microscopy was used to observe the morphological changes of SH-SY5Y cells. Immunofluorescence staining was performed to detect the expression of neuronal specific marker ßIII-tubulin and mature neuronal marker neurofilament heavy polypeptide (NFH). Multi-electrode array was used to record the electrophysiological features of SH-SY5Y cells. Cell apoptosis was evaluated using a cell apoptosis detection kit. RESULTS: Low concentrations of adapalene promoted the formation of neurite outgrowth in SH-SY5Y cells, with the neurites interconnected to form a network. Spontaneous discharge activity was observed in SH-SY5Y cells treated with low concentrations of adapalene. Compared to the control group, the expression of ßIII-tubulin and NFH increased in the 1 µM adapalene group, while the level of cell apoptosis increased in the high concentration adapalene group (P<0.05). CONCLUSIONS: Low concentrations of adapalene can induce differentiation of SH-SY5Y cells into mature functional neurons, while high concentrations of adapalene can induce apoptosis in SH-SY5Y cells.

Translatability of In Vitro Stress for Predicting Deamidation and Oxidation Biotransformation on Biotherapeutics.

Biotransformation leading to single residue modifications (e.g., deamidation, oxidation) can contribute to decreased efficacy/potency, poor pharmacokinetics, and/or toxicity/immunogenicity for protein therapeutics. Identifying and characterizing such liabilities in vivo are emerging needs for biologics drug discovery. In vitro stress assays involving PBS for deamidation or AAPH for oxidation are commonly used for predicting liabilities in manufacturing and storage and are sometimes considered a predictive tool for in vivo liabilities. However, reports discussing their in vivo translatability are limited. Herein, we introduce a mass spectrometry workflow that characterizes in vivo oxidation and deamidation in pharmacokinetically relevant compartments for diverse protein therapeutic modalities. The workflow has low bias of <10% in quantitating degradation in the relevant pharmacokinetic concentration range for monkey and rabbit serum/plasma (1-100 µg/mL) and allows for high sequence coverage (∼85%) for discovery/monitoring of amino acid modifications. For oxidation and deamidation, the assay was precise, with percent coefficient of variation of <8% at 1-100 µg/mL and ≤6% method-induced artifacts. A high degree of in vitro and in vivo correlation was observed for deamidation on the six diverse protein therapeutics (seven liability sites) tested. In vivo translatability for oxidation liabilities were not observed for the 11 molecules tested using in vitro AAPH stress. One of the molecules dosed in eyes resulted in a false positive and a false negative prediction for in vivo oxidation following AAPH stress. Finally, peroxide stress was also tested but resulted in limited success (1 out of 4 molecules) in predicting oxidation liabilities.

The 16S rDNA high-throughput sequencing correlation analysis of milk and gut microbial communities in mastitis Holstein cows.

This study aimed to understand the changes in the milk and gut microbiota of dairy cows with mastitis, and to further explore the relationship between mastitis and the microbiota. In this study, we extracted microbial DNA from healthy and mastitis cows and performed high-throughput sequencing using the Illumina NovaSeq sequencing platform. OTU clustering was performed to analyze complexity, multi-sample comparisons, differences in community structure between groups, and differential analysis of species composition and abundance. The results showed that there were differences in microbial diversity and community composition in the milk and feces of normal and mastitis cows, where the diversity of microbiota decreased and species abundance increased in the mastitis group. There was a significant difference in the flora composition of the two groups of samples (P < 0.05), especially at the genus level, the difference in the milk samples was Sphingomonas (P < 0.05) and Stenotrophomonas (P < 0.05), the differences in stool samples were Alistipes (P < 0.05), Flavonifractor (P < 0.05), Agathobacter (P < 0.05) and Pygmaiobacter (P < 0.05). In conclusion, the microbiota of the udder and intestinal tissues of dairy cows suffering from mastitis will change significantly. This suggests that the development of mastitis is related to the endogenous pathway of microbial intestinal mammary glands, but the mechanisms involved need further study.

Fabricating Robust Pt Clusters on Sn-Doped CeO2 for CO Oxidation: A Deep Insight into Support Engineering and Surface Structural Evolution.

The size effect on nanoparticles, which affects the catalysis performance in a significant way, is crucial. The tuning of oxygen vacancies on metal-oxide support can help reduce the size of the particles in active clusters of Pt, thus improving catalysis performance of the supported catalyst. Herein, Ce-Sn solid solutions (CSO) with abundant oxygen vacancies have been synthesized. Activated by simple CO reduction after loading Pt species, the catalytic CO oxidation performance of Pt/CSO was significantly better than that of Pt/CeO2 . The reasons for the elevated activity were further explored regarding ionic Pt single sites being transformed into active Pt clusters after CO reduction. Due to more exposed oxygen vacancies, much smaller Pt clusters were created on CSO (ca. 1.2 nm) than on CeO2 (ca. 1.8 nm). Consequently, more exposed active Pt clusters significantly improved the ability to activate oxygen and directly translated to the higher catalytic oxidation performance of activated Pt/CSO catalysts in vehicle emission control applications.

Induced pluripotent stem cell-derived extracellular vesicles overexpressing SFPQ protect retinal Müller cells against hypoxia-induced injury.

Splicing factor proline/glutamine-rich (SFPQ) is expressed in induced pluripotent stem cells (iPSCs), which are reported to orchestrate hypoxic injury responses and release extracellular vesicles (EVs). Therefore, this study sought to explore the role of iPSC-derived EVs carrying SFPQ in hypoxia-induced injury to retinal Müller cells. We induced oxygen-glucose deprivation/reoxygenation (OGD/R) in Müller cells. SFPQ was overexpressed or knocked down in iPSCs, from which EVs were extracted. Müller cells were co-cultured with EVs, and the results indicated that SFPQ protein was transferred into retinal Müller cells by iPSC-derived EVs. We identified an interaction of SFPQ with HDAC1 in retinal Müller cells. Specifically, SFPQ recruited HDAC1 to downregulate HIF-2α by regulating its acetylation. The in vitro studies suggested that iPSC-derived EVs, SFPQ or HDAC1 overexpression, or HIF-2α silencing diminished cell injury and apoptosis but elevated proliferation in retinal Müller cells. The in vivo studies indicated that iPSC-derived EVs containing SFPQ curtailed apoptosis of retinal Müller cells, thus alleviating retinal ischemia/reperfusion (I/R) injury of rat model. Taken together, iPSC-derived EVs containing SFPQ upregulated HDAC1 to attenuate OGD/R-induced Müller cell injury via downregulation of HIF-2α.

Rapid mitogenic regulation of the mTORC1 inhibitor, DEPTOR, by phosphatidic acid.

The mammalian target of rapamycin complex 1 (mTORC1) is regulated, in part, by the endogenous inhibitor DEPTOR. However, the mechanism of DEPTOR regulation with regard to rapid mTORC1 activation remains unknown. We report that DEPTOR is rapidly and temporarily dissociated from mTORC1 upon mitogenic stimulation, suggesting a mechanism underlying acute mTORC1 activation. This mitogen-stimulated DEPTOR dissociation is blocked by inhibition or depletion of the mTORC1 regulator, phospholipase D (PLD), and recapitulated with the addition of the PLD product phosphatidic acid (PA). Our mass spectrometry analysis has independently identified DEPTOR as an mTOR binding partner dissociated by PA. Interestingly, only PA species with unsaturated fatty acid chains, such as those produced by PLD, are capable of displacing DEPTOR and activating mTORC1, with high affinity for the FRB domain of mTOR. Our findings reveal a mechanism of mTOR regulation and provide a molecular explanation for the exquisite specificity of PA function.

Overexpression of LAG3, TIM3, and A2aR in adenoid cystic carcinoma and mucoepidermoid carcinoma.

OBJECTIVE: Adenoid cystic carcinoma (AdCC) and mucoepidermoid carcinoma (MEC) are the two most frequent malignancies of salivary glands. This study aims to explore the expression and migration of LAG3, TIM3, and A2aR in AdCC and MEC, and the potential relationship with oncogenic signaling molecules and immunosuppressive cytokines. MATERIALS AND METHODS: Custom made human salivary gland tissue microarrays included 81 AdCCs, 52 MECs, 76 normal salivary glands (NSG), and 14 pleomorphic adenoma (PMA) samples. Immunohistochemical analysis of lymphocyte activation gene 3 (LAG3), T-cell immunoglobulin and mucin domain-containing protein 3 (TIM3), adenosine 2a receptor (A2aR), oncogenic phosphorylated S6 kinase (p-S6) and ERK1/2 (p-ERK1/2 ), and TGF-ß1 was performed with salivary gland tissue microarrays of human samples. The correlation of the immunostaining was analyzed based on a digital pathological system, and data were evaluated by hierarchical cluster. Further in vitro studies of knockdown immune checkpoints LAG3, TIM3, and A2aR were carried out by siRNA transfection. RESULTS: The expression levels of LAG3, TIM3, and A2aR were remarkably increased in AdCC and MEC, compared with NSG and PMA samples, but were independent of pathology grade. They were closely correlated with TGF-ß1, slightly related to p-ERK1/2 and p-S6. After the knockdown of immune checkpoints LAG3, TIM3, and A2aR, the migration of SACC-LM cell line was significantly reduced. CONCLUSIONS: These results suggested that LAG3, TIM3, and A2aR are overexpressed in AdCC and MEC, may promote migration of SACC-LM cell and correlated with TGF-ß1 and oncogenic signaling pathways.

Fluoroscopy-guided direct anterior approach total hip arthroplasty provides more accurate component positions in the supine position than in the lateral position.

PURPOSE: The position of the acetabular and femoral components is critical for stability and wear resistance. The aim of this study is to investigate whether the fluoroscopy-guided direct anterior approach in the supine position (S-DAA) is more helpful in improving the position of acetabular and femoral components than the fluoroscopy-guided direct anterior approach in the lateral decubitus position (L-DAA). METHODS: A retrospective analysis of 76 cases of fluoroscopy-guided direct anterior approach total hip arthroplasty (38 cases in the S-DAA and 38 cases in the L-DAA group) was performed in one hospital from 2019 to 2021. The differences in inclination, anteversion, femoral offset (FO), global offset (GO), and leg length discrepancy (LLD) measurements during and after surgery were analyzed. The postoperative femoral offset (FO), global offset (GO), leg length discrepancy (LLD), and preoperative and postoperative Harris hip score were compared between the two groups. RESULTS: In the S-DAA group, there were no significant differences in the mean intraoperative inclination angle anteversion angle, FO, GO, and LLD compared to the postoperative values, whereas in the L-DAA group, there were significant differences between the intraoperative and postoperative measurements (P < 0.001, P = 0.009, P＜0.001, P＜0.001 and P = 0.008, respectively). Additionally, there were significant differences in the accuracy of LLD, FO, and GO between the two groups (P < 0.001). Compared with the L-DAA group, the average differences of inclination, anteversion, LLD, FO, and GO during and after operation in the S-DAA group were smaller, and the consistency was higher. There was a significant difference in Harris hip score between the two groups at 1 week after surgery (P = 0.033). There was no significant difference in Harris hip score between 1 month and 3 months after surgery (P = 0.482 and P = 0.797, respectively). CONCLUSIONS: In the supine group, the direct anterior approach (DAA) provides more accurate positioning of the acetabular and femoral components. However, there was no significant difference in hip joint function and activity between the two groups at follow-up.

High-throughput antibody screening from complex matrices using intact protein electrospray mass spectrometry.

Toward the goal of increasing the throughput of high-resolution mass characterization of intact antibodies, we developed a RapidFire-mass spectrometry (MS) assay using electrospray ionization. We achieved unprecedented screening throughput as fast as 15 s/sample, which is an order of magnitude improvement over conventional liquid chromatography (LC)-MS approaches. The screening enabled intact mass determination as accurate as 7 ppm with baseline resolution at the glycoform level for intact antibodies. We utilized this assay to characterize and perform relative quantitation of antibody species from 248 samples of 62 different cell line clones at four time points in 2 h using RapidFire-time-of-flight MS screening. The screening enabled selection of clones with the highest purity of bispecific antibody production and the results significantly correlated with conventional LC-MS results. In addition, analyzing antibodies from a complex plasma sample using affinity-RapidFire-MS was also demonstrated and qualified. In summary, the platform affords high-throughput analyses of antibodies, including bispecific antibodies and potential mispaired side products, in cell culture media, or other complex matrices.

ent-Pimarane Diterpenes from the Aerial Parts of Siegesbeckia pubescens.

Five new ent-pimarane diterpenes (1-5) and five known analogs (6-10) were isolated from the aerial parts of Siegesbeckia pubescens. Their structures, including absolute configurations, were determined by comprehensive spectroscopic methods especially 1D and 2D NMR and quantum chemical electronic circular dichroism calculations. All the isolated compounds were evaluated for their cytotoxicity against human BT549, A549 and H157 cancer cell lines. Among them, compounds 1 and 2 showed mild cytotoxicity against lung cancer cell lines H157 with IC50 values of 16.35±2.59 and 18.86±4.83 µM, respectively.

A ratiometric fluorescence sensor for tetracycline detection based on two fluorophores derived from Partridge tea.

Blue fluorescent carbon dots (PCDs) were prepared by hydrothermal method with Partridge tea. The ethanol extract of Partridge tea (PEE) was found to emit red fluorescence. Thus, a novel ratiometric sensor was constructed by simply mixing the two fluorophores derived from Partridge tea. The presence of tetracycline (TET) at lower concentrations enhanced the emission peak at 508 nm of PCDs and had a negligible effect on the emission peak at 680 nm of PEE. TET at higher concentrations led to  quenching  both the fluorescence of PCDs and PEE via inner filter effect and fluorescence resonance energy transfer, separately. Good linearities for the detection of TET were obtained in the ranges 0.67 to 15.00 µM and 33.33 to 266.67 µM, with limit of detection of 0.095 µM. The sensor was successfully applied to detect TET in lake water and milk samples with good recoveries ranging from 93.27 ± 4.04% to 107.30 ± 6.16%. This study provided a simple, selective, sensitive, rapid, and environmentally friendly method of monitoring TET residues in the environment and food.

High-Performance Flexible Polymer Memristor Based on Stable Filamentary Switching.

Polymer-based atomic switch memristors via the formation/dissolution of atomic-scale conductive filaments are considered as the leading candidate for next-generation nonvolatile memory. However, the instability of conductive filaments of incomplete bridge makes their switching performances unsatisfied. In this work, we report a flexible polymeric memristor using polyethylenimine incorporated with silver salt. The memristor device exhibited superior performances at room temperature with a favorable endurance, high ON/OFF ratio, good retention, and low operating voltage. These satisfactory performances are attributed to the pre-existing Ag ions in the polymer, guiding the formation of a robust Ag filament. In addition, the device shows stable bipolar switching behavior in bending conditions or after hundreds of bending cycles. In our work, we provide a simple and efficient method to construct robust filament-based memristors for flexible electronics.

Role of interfacial electron transfer reactions on sulfamethoxazole degradation by reduced nontronite activating H2O2.

It has been documented that organic contaminants can be degraded by hydroxyl radicals (•OH) produced by the activation of H2O2 by Fe(II)-bearing clay. However, the interfacial electron transfer reactions between structural Fe(II) and H2O2 for •OH generation and its effects on contaminant remediation are unclear. In this study, we first investigated the relation between •OH generation sites and sulfamethoxazole (SMX) degradation by activating H2O2 using nontronite with different reduction extents. SMX (5.2-16.9 µmol/L) degradation first increased and then decreased with an increase in the reduction extent of nontronite from 22% to 62%, while the •OH production increased continually. Passivization treatment of edge sites and structural variation results revealed that interfacial electron transfer reactions between Fe(II) and H2O2 occur at both the edge and basal plane. The enhancement on basal plane interfacial electron transfer reactions in a high reduction extent rNAu-2 leads to the enhancement on utilization efficiencies of structural Fe(II) and H2O2 for •OH generation. However, the •OH produced at the basal planes is less efficient in oxidizing SMX than that of at edge sites. Oxidation of SMX could be sustainable in the H2O2/rNAu-2 system through chemically reduction. The results of this study show the importance role of •OH generation sites on antibiotic degradation and provide guidance and potential strategies for antibiotic degradation by Fe(II)-bearing clay minerals in H2O2-based treatments.

PEGylation of anti-MerTK Antibody Modulates Ocular Biodistribution.

Here, we explore whether PEGylation of antibodies can modulate their biodistribution to the eye, an organ once thought to be immune privileged but has recently been shown to be accessible to IV-administered large molecules, such as antibodies. We chose to PEGylate an anti-MerTK antibody, a target with known potential for ocular toxicity, to minimize biodistribution to retinal pigment epithelial cells (RPEs) in the eye by increasing the hydrodynamic volume of the antibody. We used site-specific conjugation to an engineered cysteine on anti-MerTK antibody to chemically attach 40-kDa branched or linear PEG polymers. Despite reduced binding to MerTK on cells, site-specifically PEGylated anti-MerTK retained similar potency in inhibiting MerTK-mediated macrophage efferocytosis of apoptotic cells. Importantly, we found that PEGylation of anti-MerTK significantly reduced MerTK receptor occupancy in RPE cells in both naïve mice and MC-38 tumor-bearing mice, with the branched PEG exhibiting a greater effect than linear PEG. Furthermore, similar to unconjugated anti-MerTK, PEGylated anti-MerTK antibody triggered type I IFN response and exhibited antitumor effect in syngeneic mouse tumor studies. Our results demonstrate the potential of PEGylation to control ocular biodistribution of antibodies.

Using a peptide-based mass spectrometry approach to quantitate proteolysis of an intact heterogeneous procollagen substrate by BMP1 for antagonistic antibody screening.

Proteases are critical proteins involved in cleaving substrates that may impact biological pathways, cellular processes, or disease progression. In the biopharmaceutical industry, modulating the levels of protease activity is an important strategy for mitigating many types of diseases. While a variety of analytical tools exist for characterizing substrate cleavages, in vitro functional screening for antibody inhibitors of protease activity using physiologically relevant intact protein substrates remains challenging. In addition, detecting such large protein substrates with high heterogeneity using high-throughput mass spectrometry screening has rarely been reported in the literature with concerns for assay robustness and sensitivity. In this study, we established a peptide-based in vitro functional screening assay for antibody inhibitors of mouse bone morphogenic protein 1 (mBMP1) metalloprotease using a heterogeneous recombinant 66-kDa mouse Procollagen I alpha 1 chain (mProcollagen) substrate. We compared several analytical tools including capillary gel electrophoresis Western blot (CE-Western blot), as well as both intact protein and peptide-based mass spectrometry (MS) to quantitate the mBMP1 proteolytic activity and its inhibition by antibodies using this heterogeneous mProcollagen substrate. We concluded that the peptide-based mass spectrometry screening assay was the most suitable approach in terms of throughput, sensitivity, and assay robustness. We then optimized our mBMP1 proteolysis reaction after characterizing the enzyme kinetics using the peptide-based MS assay. This assay resulted in Z' values ranging from 0.6 to 0.8 from the screening campaign. Among over 1200 antibodies screened, IC50 characterization was performed on the top candidate hits, which showed partial or complete inhibitory activities against mBMP1.

Expression of HHLA2, TMIGD2, and GITR in salivary gland adenoid cystic carcinoma and mucoepidermoid carcinoma.

BACKGROUND: Mucoepidermoid carcinoma and adenoid cystic carcinoma are the two most common malignancies of salivary gland. Our study aims to explore the role of human endogenous Retrovirus-H long terminal repeat-associating protein 2, transmembrane and immunoglobulin domain-containing 2, and glucocorticoid-induced tumor necrosis factor receptor in adenoid cystic carcinoma and mucoepidermoid carcinoma, and the relationship between human endogenous Retrovirus-H long terminal repeat-associating protein 2, transmembrane and immunoglobulin domain-containing 2, glucocorticoid-induced TNF receptor, oncogenic signaling molecules, and cluster of differentiation 8. METHODS: Custom-made human salivary gland tissue microarrays included 81 Adenoid cystic carcinoma, 52 mucoepidermoid carcinoma, 76 normal salivary gland, and 14 pleomorphic adenoma samples. Immunohistochemical analysis of human endogenous Retrovirus-H long terminal repeat-associating protein 2, transmembrane and immunoglobulin domain-containing 2, and glucocorticoid-induced TNF receptor, oncogenic phosphorylated Erk1/2 , the epithelial-mesenchymal transition (EMT) molecule transforming growth factor ß1, and cluster of differentiation 8 was performed with salivary gland tissue microarray of human samples. RESULTS: According to a digital pathological system, we analyzed the correlation of immunostaining. The expression levels of human endogenous Retrovirus-H long terminal repeat-associating protein 2, transmembrane and immunoglobulin domain-containing 2, and glucocorticoid-induced TNF receptor were significantly enhanced in the adenoid cystic carcinoma and mucoepidermoid carcinoma, compared with those of pleomorphic adenoma and NSG samples. However, the expression levels of human endogenous Retrovirus-H long terminal repeat-associating protein 2, transmembrane and immunoglobulin domain-containing 2, and glucocorticoid-induced TNF receptor were independent of the pathological grade of malignancy of mucoepidermoid carcinoma and histological pattern of adenoid cystic carcinoma. They were closely related to phosphorylated Erk1/2 and transforming growth factor ß1, but negligibly related to cluster of differentiation 8. CONCLUSIONS: These results described that certain immune checkpoint molecules, namely, human endogenous Retrovirus-H long terminal repeat-associating protein 2, transmembrane and immunoglobulin domain-containing 2, and glucocorticoid-induced TNF receptor were overexpressed in Adenoid cystic carcinoma and mucoepidermoid carcinoma, but were independent of pathological grade, and may relate to transforming growth factor ß1, phosphorylated Erk1/2, and cluster of differentiation 8.