Agglutination Of Erythrocytes Research Articles

Evaluation of mRNA lipoplexes prepared using modified ethanol injection method as a tumor vaccine

We have previously demonstrated that messenger RNA (mRNA) lipoplexes composed of N-hexadecyl-N,N-dimethylhexadecan-1-aminium bromide (DC-1-16), 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE), and polyethylene glycol-cholesteryl ether (PEG-Chol) exhibited high protein expression in the lungs and spleen of mice after intravenous injection and induced high levels of antigen-specific IgG1 upon immunization. In this study, we optimized PEG modification in mRNA lipoplexes to reduce mRNA accumulation in the lungs and evaluated the suppression of tumor growth in mice bearing mouse lymphoma E.G7-ovalbumin (OVA) tumors by immunizing them with an intravenous injection of OVA mRNA lipoplexes. PEGylation of mRNA lipoplexes with 3 mol% PEG-Chol (LP-DC-1-16-3PCL) prevented agglutination of erythrocytes and reduced accumulation in the lungs. Intravenous injection of LP-DC-1-16-3PCL lipoplexes containing OVA mRNA into mice induced high levels of anti-OVA IgG1 (83,000 mU/mL) in serum, and exhibited a high cytotoxic activity (97%) against E.G7-OVA cells by the splenocytes of mice. Furthermore, immunization with LP-DC-1-16-3PCL lipoplexes containing OVA mRNA suppressed E.G7-OVA tumor growth compared to control mRNA. Based on these results, LP-DC-1-16-3PCL lipoplexes may be an effective mRNA vaccine for inducing antibody- and cytotoxic cell-mediated immune responses to tumors through intravenous injection.

In silico discovery and anti-tumor bioactivities validation of an algal lectin from Kappaphycus alvarezii genome

Lectins are proteins that bind specifically and reversibly to carbohydrates, and some of them have significant anti-tumor activities. Compared to those of lectins from land plants, there are far fewer studies on algal lectins, despite of the high biodiversity of algae. However, canonical strategies based on chromatographic feature-oriented screening cannot satisfy the requirement for algal lectin discovery. In this study, prospecting for novel OAAH family lectins throughout 358 genomes of red algae and cyanobacteria was conducted. Then 35 candidate lectins and 1843 of their simulated mutated forms were virtually screened based on predicted binding specificities to characteristic carbohydrates on cancer cells inferred by a deep learning model. A new lectin, named Siye, was discovered in Kappaphycus alvarezii genome and further verified on different cancer cells. Without causing agglutination of erythrocytes, Siye showed significant cytotoxicity to four human cancer cell lines (IC50 values ranging from 0.11 to 3.95 μg/mL), including breast adenocarcinoma HCC1937, lung carcinoma A549, liver cancer HepG2 and romyelocytic leukemia HL60. And the cytotoxicity was induced through promoting apoptosis by regulating the caspase and the p53 pathway within 24 h. This study testifies the feasibility and efficiency of the genome mining guided by evolutionary theory and artificial intelligence in the discovery of algal lectins.

Laboratory perspective of Cold Agglutinin syndrome: A case study

Cold Agglutinin Disease (CAD) represents a rare hemolytic disorder, constituting 15-30% of cases within the spectrum of Autoimmune Hemolytic Anemias (AIHAs) [1,2]. The pathogenesis of CAD revolves around autoantibodies known as Cold Agglutinins (CAs), named for their capacity to induce erythrocyte agglutination at temperatures below 37°C. Those autoantibodies typically consist of monoclonal Immunoglobulin (Ig) M type, less commonly IgA or IgG, and they target the carbohydrate antigens on the erythrocyte surface, predominantly antigen I [3-5].

Identification and characterization of an L-fucose specific lectin GVSL involved in gamete recognition of Gracilaria vermiculophylla

The lectin-carbohydrate recognition system is especially important for red algae which female and male gametes are both non-motile. A sex-related lectin gene GvSL from Gracilaria vermiculophylla was cloned and expressed heterologously in Escherichia coli. ORF of GvSL was 684 bp in length and encoded a polypeptide of 228 amino acids with a deduced molecular weight of 25.8 kDa. Real-time PCR analysis showed that it up-regulated most in mature male gametophytes which was 15.4-fold and 21.94-fold higher than mature female gametophytes and immature male gametophytes respectively. The immunohistochemical staining also demonstrated that GvSL was mainly expressed in the spermatangium crypts of mature male gametophytes. Purified rGVSL showed significant agglutination of rabbit blood erythrocytes and only L-fucose showed the glycoinhibitory effect among the eight monosaccharides tested. The inhibitory tests further proved that GVSL and its complementary carbohydrate L-fucose composed a lectin-carbohydrate recognition system affecting the fertilization efficiency of G. vermiculophylla.

Hemostatic activity <i>in vitro</i> of lectin-containing substances of bryathypes on the example of some Antarctic and Belarus representatives

BACKGROUND: The modern market of medical devices in Belarus and Russia is represented by a wide range of hemostatic agents, of which the most popular are local hemostatics of plant origin possessing the significant technological potential for renewal and improvement. A promising reserve for this may be biologically active compounds of mosses, which are characterized by anti-inflammatory, antibacterial and antifungal effects. However, their hemostatic effect is almost not studied, which determines the relevance of this work.
 AIM: The aim of this work is to study the effect of lectin-containing substances from mosses of three species collected in East Antarctica and Belarus on the parameters of human blood hemostasis in vitro.
 MATERIALS AND METHODS: We studied mosses of the genera Bryum, Ceratodon, and Coscinodon, collected in the area of the Belarusian Antarctic station Gora Vechernyaya in East Antarctica and in Belarus. Lectin-containing substances of mosses were obtained by extracting shoots in 0.05 M tris-HCl buffer (pH 8.0), centrifugation, filtration. The assessment of the biological activity of lectin-containing substances in mosses was carried out by the agglutination reaction of rabbit erythrocytes, as well as the effect on human platelet aggregation and in the test for activated partial thromboplastin time.
 RESULTS: It was established that lectin-containing substances of the studied moss species had agglutinating activity against erythrocytes in the range from 11708.28 (Belarusian samples) to 1333979.59 U/mg of protein (Antarctic samples) depending on the species and localization; initiated the aggregation of human platelets (25–80% of the effect of thrombin) regardless of blood group, Rh and gender of donors; influenced the plasma link of hemostasis, reducing activated partial thromboplastin time (by 15–18%).
 CONCLUSIONS: It was found that some species of mosses of the genera Bryum, Ceratodon and Coscinodon of Antarctica and Belarus had an agglutinating and hemostatic effect on erythrocytes and platelets, with the greatest activity noted for Antarctic species. A hypothesis has been put forward that the observed phenomenon is due to the structural features of proteins, including lectins. It is assumed that lectins are possible inducers of erythrocyte agglutination and platelet aggregation in mosses. It is shown that the moss species Bryum pseudotriquetrum and Ceratodon purpureus have a certain resource potential in Belarus for their annual harvest. The results obtained expand the list of moss species with hemostatic activity, and can be used to develop new hemostatics of plant origin for local use from Belarusian plant materials.

Dissociation Phenomenon of Erythrocyte Agglutination and Its Application to Assay of Functional Activity of the Complement System in Clinical Laboratory.

The objective of the study was to validate the dissociation phenomenon of erythrocyte agglutination which is based on erythrocyte fragments and to apply it in the functional activity assay of the complement system. The dissociation-agglutination effect of erythrocyte fragments was validated by detecting the number of free erythrocytes after the action of erythrocyte fragments on agglutinated erythrocytes. The number of free erythrocytes produced after hemolysis of agglutinated erythrocytes caused by complements and complement activators(CAs) was detected by auto hematology analyzer and the results were indicated by mean hemoglobin concentration of erythrocytes (MCHC). We optimized the test conditions and validated the inter-batch stability, explored the resolution of the assay method, and assayed for the total complement activity (AC) and the CAs activated complement activity (ACA) in serum from patients and healthy individual groups. Erythrocyte fragments have a dissociative effect on agglutinated erythrocytes. The auto hematology analyzer was able to detect AC and ACA, where AC showed an inverse correlation with MCHC, and ACA demonstrated a positive correlation with MCHC. The inter-batch CV of AC, ACA, and ACA/AC was found to be 5%, 9%, and 11.7%, respectively, with good stability. The study found that serum samples from acute phase reaction patients showed significant differences in ACA compared with healthy individuals, with a p value of 0.018; serum samples from patients with nephrotic syndrome showed significant differences in AC, ACA, and ACA/AC compared with healthy individuals, with p values of 0.014, 0.002, and 0.041, respectively. Erythrocyte fragments have dissociation-agglutination effect. The complement system immunological functional detection method, based on this effect, has potential clinical application value due to its sensitivity and accuracy.

Generation and Characterization of Monoclonal Antibodies against Swine Acute Diarrhea Syndrome Coronavirus Spike Protein

Swine acute diarrhea syndrome coronavirus (SADS-CoV), a member of the family Coronaviridae and the genus Alphacoronavirus, primarily affects piglets under 7 days old, causing symptoms such as diarrhea, vomiting, and dehydration. It has the potential to infect human primary and passaged cells in vitro, indicating a potential risk of zoonotic transmission. In this study, we successfully generated and purified six monoclonal antibodies (mAbs) specifically targeting the spike protein of SADS-CoV, whose epitope were demonstrated specificity to the S1A or S1B region by immunofluorescence assay and enzyme-linked immunosorbent assay. Three of these mAbs were capable of neutralizing SADS-CoV infection on HeLa-R19 and A549. Furthermore, we observed that SADS-CoV induced the agglutination of erythrocytes from both humans and rats, and the hemagglutination inhibition capacity and antigen–antibody binding capacity of the antibodies were assessed. Our study reveals that mAbs specifically targeting the S1A domain demonstrated notable efficacy in suppressing the hemagglutination phenomenon induced by SADS-CoV. This finding represents the first instance of narrowing down the protein region responsible for SADS-CoV-mediated hemagglutination to the S1A domain, and reveals that the cell attachment domains S1A and S1B are the main targets of neutralizing antibodies.

Screening of antigenic epitopes related to the adhesion of the avian Escherichia coli Type 1 Fimbrial Agglutinin Domain

BackgroundAvian Escherichia coli (E.coli) type 1 fimbriae adhere to avian tracheal epithelial cells through the FimH protein. However, the adhesion-related antigen is still unknown. The purpose of this study was to analyze the antigenicity of the type 1 fimbrial FimH protein of wild-type avian E. coli, screen antigen epitopes, and prepare monoclonal antibodies (mAbs) that can block the adhesion of avian E. coli.ResultsIn this study, the nucleic acid homologies of MG2 (O11), TS12 (O18), and YR5 (O78) with K12 were 97.7%, 99.6%, and 97.7%, respectively, and the amino acid sequence similarity reached 98.7%, 99.3%, and 98.0%, respectively. The epitopes and hydrophilicities of the FimH proteins of these three strains were similar. The more obvious lectin domain epitopes were located at FimH protein positions 111–124 and 154–162. The mAbs 7C2 and 7D8 against these two epitopes were prepared. An adhesion inhibition test showed that 7C2 and 7D8 blocked bacterial adhesion to avian tracheal epithelial cells. The mAb 7C2 against the 111–124 epitope inhibited O78 strain adhesion by 93%, and the mAb 7D8 against the 154–162 epitope inhibited O78 strain adhesion by 49%, indicating that these two epitopes are closely related to the adhesion of type 1 fimbriae. However, only the 111–124 epitope-recognizing mAb 7C2 inhibited bacterial agglutination of erythrocytes, indicating that host cell receptor binding and erythrocyte agglutination are not mediated by the same spatial locations within the FimH protein.ConclusionsThe results demonstrate that the mAbs 7C2 and 7D8 against FimH protein positions 111–124 and 154–162 could inhibit the adhesion of E.coli to the chicken trachea.

The role of acidification of the medium on the erythrocyte agglutinating and adsorbing properties of blood group-specific monoclonal antibodies.

Studies on the specificity of ABH antigen-antibody interactions at different pH values are rare. Therefore, the aim of this study was to estimate the effect of acidification of the reacting medium on the agglutinating ability of anti-A monoclonal antibodies (mABs) and their inhibition by glycoconjugates of red blood cell membranes. Anti-A mABs were obtained from the Fourth International Workshop on Monoclonal Antibodies Against Human Red Blood Cell and Related Antigens (on July 19-20, 2002, in Paris, France). The glycoconjugates were isolated from erythrocytes' membranes. The inhibition of the lipid and protein isotypes of the blood group A antigen was assessed. The acidic medium decreased the agglutinating ability of acid immunoglobulin M (IgM) anti-A mABs, in contrast to alkaline immunoglobulin G (IgG) mABs. Meanwhile, at pH 6.5, IgM antibodies exhibited a high adsorption capacity, while IgG antibodies demonstrated a strong adsorption capacity at an alkaline pH. mABs 2-19, 2-28, 2-22, and 2-8 were inhibited by the acidic lipid and protein glycotopes of erythrocyte membranes. However, mAB 2-8 was inhibited by both acidic and alkaline glycotope variants. The agglutinating and adsorbing abilities of mABs, which are revealed at opposing pH values, should be taken into account during studies of antigen-antibody interactions.

Novel mono- and multivalent N-acetylneuraminic acid glycoclusters as potential broad-spectrum entry inhibitors for influenza and coronavirus infection

N-acetylneuraminic acid (Neu5Ac) is a glycan receptor of viruses spread in many eukaryotic cells. The present work aimed to design, synthesis and biological evaluation of a panel of Neu5Ac derivatives based on a cyclodextrin (CD) scaffold for targeting influenza and coronavirus membrane proteins. The multivalent Neu5Ac glycoclusters efficiently inhibited chicken erythrocyte agglutination induced by intact influenza virus in a Neu5Ac density–dependent fashion. Compared with inhibition by Neu5Ac, the multivalent inhibitor with 21 Neu5Ac residues on the primary face of the β-CD scaffold afforded 1788-fold higher binding affinity inhibition for influenza virus hemagglutinin with a dissociation constant (KD) of 3.87 × 10−7 M. It showed moderate binding affinity to influenza virus neuraminidase, but with only about one-thirtieth the potency of that with the HA protein. It also exhibited strong binding affinity to the spike protein of three human coronaviruses (severe acute respiratory syndrome coronavirus, Middle East respiratory syndrome coronavirus, and severe acute respiratory syndrome coronavirus 2), with KD values in the low micromolar range, which is about 10-time weaker than that of HA. Therefore, these multivalent sialylated CD derivatives have possible therapeutic application as broad-spectrum antiviral entry inhibitors for many viruses by targeting the Neu5Ac of host cells.

Glutathione disrupts galectin-10 Charcot-Leyden crystal formation to possibly ameliorate eosinophil-based diseases such as asthma.

Charcot-Leyden crystals (CLCs) are the hallmark of many eosinophilic-based diseases, such as asthma. Here, we report that reduced glutathione (GSH) disrupts CLCs and inhibits crystallization of human galectin-10 (Gal-10). GSH has no effect on CLCs from monkeys ( Macaca fascicularis or M. mulatta), even though monkey Gal-10s contain Cys29 and Cys32. Interestingly, human Gal-10 contains another cysteine residue (Cys57). Because GSH cannot disrupt CLCs formed by the human Gal-10 variant C57A or inhibit its crystallization, the effects of GSH on human Gal-10 or CLCs most likely occur by chemical modification of Cys57. We further report the crystal structures of Gal-10 from M. fascicularis and M. mulatta, along with their ability to bind to lactose and inhibit erythrocyte agglutination. Structural comparison with human Gal-10 shows that Cys57 and Gln75 within the ligand binding site are responsible for the loss of lactose binding. Pull-down experiments and mass spectrometry show that human Gal-10 interacts with tubulin α-1B, with GSH, GTP and Mg 2+ stabilizing this interaction and colchicine inhibiting it. Overall, this study enhances our understanding of Gal-10 function and CLC formation and suggests that GSH may be used as a pharmaceutical agent to ameliorate CLC-induced diseases.

Recombinant Phaseolus vulgaris phytohemagglutinin L-form expressed in the Bacillus brevis exerts in vitro and in vivo anti-tumor activity through potentiation of apoptosis and immunomodulation

Leukocyte phytohemagglutinin (PHA-L) is a tetrameric isomer of phytohemagglutinin (PHA) purified from the red kidney bean (Phaseolus vulgaris) and is a well-known human lymphocyte mitogen. Due to its antitumor and immunomodulatory effects, PHA-L may serve as a potential antineoplastic agent in future cancer therapeutics. However, various negative consequences of PHA have been reported in the literature as a result of the restricted acquisition methods, including oral toxicity, hemagglutinating activity, and immunogenicity. There is a critical need to explore a new method to obtain PHA-L with high purity, high activity and low toxicity. In this report active recombinant PHA-L protein was successfully prepared by Bacillus brevius expression system, and the antitumor and immunomodulatory activities of recombinant PHA-L were characterized by in vitro and in vivo experiments. The results showed that recombinant PHA-L protein had stronger antitumor effect, and its anti-tumor mechanism was realized through direct cytotoxicity and immune regulation. Importantly, compared with natural PHA-L, the recombinant PHA-L protein showed the lower erythrocyte agglutination toxicity in vitro and immunogenicity in mice. Altogether, our study provides a new strategy and important experimental basis for the development of drugs with dual effects of immune regulation and direct antitumor activity.

SCREENING OF AGGLUTINATION ACTIVITIES OF COMMON VEGETABLES IN BANGLADESH

Lectins are capable of cell agglutination by interacting with the cell surface exposed glycans. Lectins that agglutinate erythrocytes are called hemagglutinin. These compounds occur in various organisms including plants. In this study, twenty edible vegetables were subjected to screen for haemagglutination and lectin activity. Lectin activity was assessed by measuring the inhibition ability of haemagglutination by different sugars. PBS (Phosphate Buffer Saline) treated homogenate was tested for haemagglutination against erythrocytes suspension, and inhibition of agglutination was determined by serially diluted sugar solution. The results of this study demonstrated that one homogenate among the homogenates of 20 vegetables was found to be capable of human erythrocyte agglutination. The Ipomoea aquatica homogenate shown positive haemagglutination reaction against red blood cells (RBCs) with HA titer value of 4 HA-U/25uL, and four out of seven sugars were able to inhibit RBCs agglutination. Among the four sugars, two are potent inhibitors of agglutination. The neutralization of haemagglutination by four monosaccharides provides a suggestion that the lectin present in I. aquatica may be multivalent carbohydrate binding hemagglutinin. This is the first report to identify that Ipomoea aquatica contains lectin protein which is a polyvalent sugar binding lectin and specific for glucose, galactose, mannose and N-Acetyl-D-Glucosamine.

Region Met225 to Ile412 of Bacillus cereus Hemolysin II Is Capable to Agglutinate Red Blood Cells.

Hemolysin II (HlyII) is one of the virulence factors of the opportunistic bacterium Bacillus cereus belonging to the group of β-pore-forming toxins. This work created a genetic construct encoding a large C-terminal fragment of the toxin (HlyIILCTD, M225-I412 according to the numbering of amino acid residues in HlyII). A soluble form of HlyIILCTD was obtained using the SlyD chaperone protein. HlyIILCTD was first shown to be capable of agglutinating rabbit erythrocytes. Monoclonal antibodies against HlyIILCTD were obtained by hybridoma technology. We also proposed a mode of rabbit erythrocyte agglutination by HlyIILCTD and selected three anti-HlyIILCTD monoclonal antibodies that inhibited the agglutination.

Collection and processing of hematopoietic progenitor cell products at risk of presenting with cold agglutination

Background aimsCold agglutinins are commonly identified in transfusion laboratories and are defined by their ability to agglutinate erythrocytes at 3–4°C, with most demonstrating a titer >64. Similarly, cryoglobulins can precipitate from plasma when temperatures drop below central body temperature, resulting in erythrocyte agglutination. Thankfully, disease associated from these autoantibodies is rare, but unfortunately, such temperature ranges are routinely encountered outside of the body's circulation, as in an extracorporeal circuit during hematopoietic progenitor cell (HPC) collection or human cell therapy laboratory processing. When agglutination occurs ex vivo, complications with the collection and product may be encountered, resulting in adverse events or product loss. Here, we endeavor to share our experience in preventing and responding to known cases at risk of or spontaneous HPC agglutination in our human cell therapy laboratory. Case reportsFour cases of HPC products at risk for, or spontaneously, agglutinating were seen at our institution from 2018 to 2020. Planned modifications occurred, including ambient room temperature increases, tandem draw and return blood warmers, warm product transport and extended post-thaw warming occurred. In addition, unplanned modifications were undertaken, including warm HPC product processing and plasma replacement of the product when spontaneous agglutination of the product was identified. All recipients successfully engrafted after infusion. ConclusionsWhile uncommon, cold agglutination of HPC products can disrupt standard processes of collection and processing. Protocol modifications can circumvent adverse events for the donor and minimize product loss. Such process modifications should be considered in individuals with known risks for agglutination going to HPC donation/collection.

Photosynthesis of hybrid silver-based nanoparticles mediated lectins and evaluation of their hemagglutinating properties.

Lectins are carbohydrate-binding proteins belonging to the Leguminosae family. In this family stand out proteins extracted from species belonging to Diocleinae subtribe, which includes, for example, the seed lectin from Dioclea violacea (DVL) and the jack bean lectin Concanavalin A (ConA). Here, we report the photosynthesis of silver/silver chloride nanoparticles (NPs) assisted by ConA and DVL. The syntheses were simple processes using a green-chemistry approach. Under electron microscopy, NPs heterogeneous in size, nearly spherical and covered by a thin lectin corona, were observed. Both NPs assisted by lectins were capable to cause strong rabbit erythrocytes agglutination with the same titers of hemagglutinating activities. These results indicate that both lectins maintained their biological activities even after association with the NPs and therefore are able to interact with biological membrane carbohydrates. However, for rabbit erythrocytes treated with proteolytic enzymes were observed different titers of hemagglutinating activities, suggesting differences in the spatial arrangement of the lectins on the surface of the NPs. This study provides evidences that these hybrid lectin-coated silver/silver chloride NPs can be used for selective recognition and interaction with membrane carbohydrates and others biotechnological applications.

Novel Sirpα-Fc Fusion Protein IMM01 Exhibits Dual Anti-Tumor Activities By Targeting CD47/Sirpα Signal Pathway

Background: This study aims to evaluate the anti-tumor mechanism of IMM01, a novel recombinant SIRPα-Fc fusion protein targeting CD47-SIRPα pathway. Methods: IMM01 was a recombinant fusion protein of SIRPα (V2) extracellular segment domain 1 and human IgG1 Fc, with the N glycosylation site N80 mutated to A (Alanine) in D1 region. IMM01 fusion proteins were produced using in house developed CHO-K1 cell expression system. IMM01 blocking activities on Jurkat-CSR (Jurkat-CD47KO-CARSIRPα) cells, antibody-dependent cellular phagocytosis (ADCP), antibody-dependent cell-mediated cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC) assays were done by flow cytometry. Detection of cytokines of IL-1β, IL-2, IL-4, IL-5, IL-6, GM-CSF, IFN-γ, IL-10 and TNF production by PBMC was done CBA assay. In vivo mechanism of action study was done in different mouse models including HL-60 CB17-SCID, Daudi CB17-SCID, SNU-1 CB17-SCID, Raji CB17-SCID and CT26-hPDL1(Tg) hCD47(Tg) mPDL1(KO) mCD47(KO). In addition, human red blood cells binding and erythrocyte agglutination of IMM01 were also evaluated. Results:In vitro binding assay revealed that IMM01 has a strong binding affinity to CD47 with lowest EC50 at 0.4967 nM. The removal of N-glycosylation modification improves the consistence and purity of the IMM01 protein expression, and the potential mitigation of immunogenicity. Using a chimeric SIRPα receptor protein expressing cell line, IMM01 not only can block the CD47 at the protein level, but also block the signal transduction at the cellular levels. Compared to negative (hIgG1-Fc) controls, IMM01 stimulated secretion of minimal amount of IL-10 and TNF but inhibited IL-8 production by PBMC. IMM01 can induce strong ADCP and moderate ADCC, but not the CDC activities. The strong phagocytosis against tumor cells is dependent on interaction of Fc with activating FcγRs on the presence of macrophages. Mechanism of action studies demonstrated that IMM01 induced strong phagocytosis against tumor cells was attributed to dual activities of blocking "don't eat me" signal and activating "eat me" signal. IMM01 has strong and robust in vivo anti-tumor activities in different mouse tumor models either as monotherapy on hematological malignancies, or in combination therapy with PD-L1 mAb, PD-1 mAb, CD33 mAb, and HER-2 mAb, respectively. The in vivo therapeutic efficacy of IMM01 was done in HL-60 xenograft model showed that 100% of the mice achieved CR after administration of IMM01 at 5mg/kg for 2 weeks. Also, 100% of mice achieved CR after IMM01 combined with the above mentioned mAbs in different mouse models, respectively. This illustrated that IMM01 has a great therapeutic potential in combination with these commonly used immunotherapy or targeted therapy agents to be used in the treatment of solid tumors and hematological malignancies. Finally, IMM01 demonstrated a favorable safety profile with no human RBC binding, nor hemagglutination induction. Conclusion: Our study results demonstrated that, as a novel bi-functional SIRPα-Fc recombinant protein, IMM01 has very strong anti-tumor activities with a good curative effect on mouse xenograft tumor models with dual anti-tumor activities by blocking CD47 "don't eat me" signal and activating the phagocytosis "eat me" signal. Furthermore, IMM01 demonstrated a well synergistic effect when combined with different immunotherapeutic agents. Importantly, our results revealed that IMM01 has a favorable safety profile with no human RBC binding activity and no hemagglutination induction. Phase I clinical study is ongoing, and the clinical trial results will be reported soon. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Binding of Glycerol to Human Galectin-7 Expands Stability and Modulates Its Functions.

Glycerol is seen in biological systems as an intermediate in lipid metabolism. In recent years, glycerol has been reported to act as a chemical chaperone to correct the conformation of proteins. Here, we investigate the role of glycerol in galectin-7 (Gal-7). The thermal shift and CD assays showed that the thermal stability of Gal-7 increased with glycerol concentration but with little secondary structure changes induced by glycerol. In addition, glycerol can inhibit Gal-7-mediated erythrocyte agglutination. We also solved the crystal structures of human Gal-7 in complex with glycerol in two different conditions. Glycerol binds at the carbohydrate-recognition binding sites of Gal-7, which indicates glycerol as a small ligand for Gal-7. Surprisingly, glycerol can bind a new pocket near the N-terminus of Gal-7, which can greatly reduce the flexibility and improve the stability of this region. Moreover, overexpression of Gal-7 decreased the intracellular triglyceride levels and increased mRNA expression of aquaporin-3 (AQP-3) when HeLa cells were incubated with glycerol. These findings indicate that Gal-7 might regulate glycerol metabolism. Overall, our results on human Gal-7 raise the perspective to systematically explore this so far unrecognized phenomenon for Gal-7 in glycerol metabolism.

Autoimmune haemolytic anemias

Autoimmune haemolytic anemia (AIHA) is defined as the immune-mediated destruction of red blood cells. In most cases, antibodies that target surface antigens on erythrocytes lead to their premature degradation in the spleen or, less commonly, in the liver. The term includes a heterogenous group of diseases, which differ largely in pathophysiology and treatment. The two most common entities are warm AIHA and cold AIHA. Diagnostic testing involves the analysis of haemolytic markers like lactate dehydrogenase, haptoglobin and unconjugated bilirubin as well as a hemoglobin and reticulocytes. In case of a haemolytic anemia, further testing like a blood smear and a direct antiglobulin test should follow. As diagnostic testing and treatment of AIHA are complex, affected patients should always be referred to a hematologist.In warm AIHA, mainly IgG autoantibodies bind to their antigen on the erythrocyte surface at body temperature, leading to their premature destruction in the spleen. First line treatment options include the administration of steroids which mitigate the destruction of red blood cells by macrophages in the spleen. In contrast, IgM autoantibodies in cold AIHA lead to intravasal agglutination of erythrocytes and complement activation. The IgM antibodies have their highest affinity below body temperature which is why patients experience symptoms mainly in cold-exposed body areas. Although the IgM antibodies dissolve at body temperature, the complement-loaded erythrocytes are destroyed in the liver. Therapeutic options include protection from cold and immunosuppressive agents or complement inhibition.

Effect of New 2-Thioxoimidazolidin-4-one Compounds against Staphylococcus aureus Clinical Strains and Immunological Markers' Combinations.

Although the structure-activity relationship indicates that the 4-thioxoimidazolidin ring is essential for antibacterial activities and pharmaceutical applications, there were no enough studies on the derivatives of this compound. Evaluating the new hydantoin compounds C5 (3-((2-bromobenzylidene) amino)-2- thioxoimidazolidin-4-one) and C6 (3-((4- methoxybenzylidene) amino)-2-thioxoimidazolidin-4-one) that were prepared against clinical Staphylococcus aureus isolates for antibacterial, antibiofilm, and antihemagglutination activities is the aim of this study. Therefore, the potential clinical resistance of the strains was evaluated by their ability to form biofilms, antibiotic resistance, and agglutinate erythrocytes macroscopically and microscopically; besides, the bacterial biofilm was screened for any association with the patient's serum immunoglobulin levels and complements. Despite the effective concentration for C5 and C6 compounds, which is ≤ 31.25 μg/ml, the reduction rate is not concentration-dependent; it depends on the molecular docking of the hydantoin compounds. Hence, the effect of the minimal inhibitory concentrations (MICs) is variable. In this study, the results for the compounds (with the concentration of 31.25–62.5 μg/mL for C5 and 62.5–125 μg/mL for C6) significantly manifest the antibacteria, antibiofilm, and antihemagglutination effects against the virulent strains of S. aureus due to the high percentage of biofilm inhibition that was caused by the new hydantoin compounds. Besides, time-kill kinetics studies showed that these compounds pose bactericidal action. Overall, this study revealed that the new hydantoin derivatives have an interesting potential as new antibacterial drugs through the inhibition of bacterial adhesion. The infections of these isolates activate the complement system through the lectin pathway. Nevertheless, these compounds can be improved in order to be used at even lower concentrations.