Blockade of Rho-associated kinase prevents inhibition of axon regeneration of peripheral nerves induced by anti-ganglioside antibodies

Anti-ganglioside antibodies are associated with delayed/poor clinical recovery in Guillain-Barrèsyndrome,mostly related to halted axon regeneration.Cross-linking of cell surface gangliosides by anti-ganglioside antibodies triggers inhibition of nerve repair in in vitro and in vivo paradigms of axon regeneration.These effects involve the activation of the small GTPase Rho A/ROCK signaling pathways,which negatively modulate growth cone cytoskeleton,similarly to well stablished inhibitors of axon regeneration described so far.The aim of this work was to perform a proof of concept study to demonstrate the effectiveness of Y-27632,a selective pharmacological inhibitor of ROCK,in a mouse model of axon regeneration of peripheral nerves,where the passive immunization with a monoclonal antibody targeting gangliosides GD1a and GT1b was previously reported to exert a potent inhibitory effect on regeneration of both myelinated and unmyelinated fibers.Our results demonstrate a differential sensitivity of myelinated and unmyelinated axons to the pro-regenerative effect of Y-27632.Treatment with a total dosage of 9 mg/kg of Y-27632 resulted in a complete prevention of anti-GD1a/GT1b monoclonal antibody-mediated inhibition of axon regeneration of unmyelinated fibers to skin and the functional recovery of mechanical cutaneous sensitivity.In contrast,the same dose showed toxic effects on the regeneration of myelinated fibers.Interestingly,scale down of the dosage of Y-27632 to 5 mg/kg resulted in a significant although not complete recovery of regenerated myelinated axons exposed to anti-GD1a/GT1b monoclonal antibody in the absence of toxicity in animals exposed to only Y-27632.Overall,these findings confirm the in vivo participation of Rho A/ROCK signaling pathways in the molecular mechanisms associated with the inhibition of axon regeneration induced by anti-GD1a/GT1b monoclonal antibody.Our findings open the possibility of therapeutic pharmacological intervention targeting Rho A/Rock pathway in immune neuropathies associated with the presence of anti-ganglioside antibodies and delayed or incomplete clinical recovery after injury in the peripheral nervous system.

Atypical Guillain-Barrésyndrome with positive anti-sulfatide,anti-GT1b,and anti-GT1a antibodies:A case report

BACKGROUND The role of diverse antibodies in mediating peripheral nerve injury in Guillain-Barrésyndrome(GBS)is becoming clearer,but positivity for multiple antibodies in one case is uncommon.To our knowledge,this is the first case involving GBS with positive anti-sulfatide,anti-GT1a,and anti-GT1b antibodies.CASE SUMMARY A 20-year-old female patient was admitted to the hospital due to weakness of limbs for 5 d,and deterioration of the weakness and muscle aches for 1 d.The patient's limbs were weak,but the tendon reflexes in the part of the limbs were normal.There was no comorbid peripheral nociception or deep sensory dysfunction.She was diagnosed with GBS and was discharged after receiving intravenous human immunoglobulin pulse therapy.CONCLUSION In this article,the clinical manifestations,neurophysiological examination,and auxiliary examination findings of a GBS patient positive for multiple antibodies were analyzed to improve the identification of the disease by clinical physicians at an early stage.

Anti-PD1 antibody and not anti-LAG-3 antibody improves the antitumor effect of photodynamic therapy for treating metastatic breast cancer

Photodynamic therapy(PDT)has limited effects in treating metastatic breast cancer.Immune checkpoints can deplete the function of immune cells;however,the expression of immune checkpoints after PDT is unclear.This study investigates whether the limited e±cacy of PDT is due to upregulated immune checkpoints and tries to combine the PDT and immune checkpoint inhibitor to observe the e±cacy.A metastatic breast cancer model was treated by PDT mediated by hematoporphyrin derivatives(HpD-PDT).The anti-tumor effect of HpD-PDT was observed,as well as CD4þT,CD8þT and calreticulin(CRT)by immunohistochemistry and immunofluorescence.Immune checkpoints on T cells were analyzed byflow cytometry after HpD-PDT.When combining PDT with immune checkpoint inhibitors,the antitumor effect and immune effect were assessed.For HpD-PDT at 100 mW/cm2 and 40,60 and 80 J/cm2,primary tumors were suppressed and CD4þT,CD8þT and CRT were elevated;however,distant tumors couldn't be inhibited and survival could not be prolonged.Immune checkpoints on T cells,especially PD1 and LAG-3 after HpD-PDT,were upregulated,which may explain the reason for the limited HpD-PDT effect.After PDT combined with anti-PD1 antibody,but not with anti-LAG-3 antibody,both the primary and distant tumors were signi-cantly inhibited and the survival time was prolonged,additionally,CD4þT,CD8þT,IFN-þCD4þT and TNF-þCD4þT cells were signi-cantly increased compared with HpD-PDT.HpD-PDT could not combat metastatic breast cancer.PD1 and LAG-3 were upregulated after HpD-PDT.Anti-PD1 antibody,but not anti-LAG-3 antibody,could augment the antitumor effect of HpD-PDT for treating metastatic breast cancer.

Miller fisher syndrome with positive anti-GQ1b/GT1a antibodies associated with COVID-19 infection:A case report

BACKGROUND Miller fisher syndrome(MFS)is a variant of Guillain-Barrésyndrome,an acute immune-mediated peripheral neuropathy that is often secondary to viral infections.Anti-ganglioside antibodies play crucial roles in the development of MFS.The positive rate of ganglioside antibodies is exceptionally high in MFS patients,particularly for anti-GQ1b antibodies.However,the presence of other ganglioside antibodies does not exclude MFS.CASE SUMMARY We present a 56-year-old female patient who suddenly developed right blepharoptosis and progressively worsening vision in both eyes.There were flu symptoms prior to onset,and a coronavirus disease 2019 test was positive.On physical examination,the patient exhibited bilateral extraocular muscle paralysis,weakened reflexes in both limbs,and impaired coordination.The cerebrospinal fluid examination results showed no obvious abnormalities.Bilateral peroneal nerve F-waves were not extracted.Serum anti-GD1b IgG and anti-GT1a IgG antibodies were positive.The patient received intravenous methylprednisolone(1000 mg/day),with the dosage gradually decreased.Additionally,intravenous high-dose immunoglobulin treatment was administered for 5 days(0.4 g/kg/day)from day 2 to day 6 of hospitalization.The patient’s symptoms improved after treatment with immunoglobulins and hormones.CONCLUSION Positive ganglioside antibodies may be used as supporting evidence for the diagnosis;however,the diagnosis of MFS is more reliant on clinical symptoms.

Preparation of anti-canine interleukin-31 receptor alpha polyclonal antibody and evaluation of its therapeutic efect in canine atopic dermatitis

Canine atopic dermatitis(CAD)is a prevalent genetically susceptible infammatory and pruritic allergic skin condition afecting not only the health of dogs but also the quality of life of their owners.Interleukin-31(IL-31)and interleukin-31 receptor alpha(IL-31RA)are essential for the development of pruritus in primates and mice.Hence,it is expected that inhibiting IL-31RA will be an efective approach to alleviate pruritus.The purpose of the study was to produce anti-canine IL-31RA polyclonal antibodies(anti-IL-31RA pAbs)and evaluate their efcacy in inhibiting house dust mite(HDM)-evoked pruritic responses.Dogs were immunized with antigens formed by IL-31RA recombinant short peptides coupled to BSA to produce anti-IL-31RA pAbs.The CAD model was developed by using HDM allergen stimulation,and the efects of IL-31RA pAbs on the reduction of pruritus in CAD model dogs were examined.The Canine Atopic Dermatitis Extent and Severity Index(CADESI)-4 and pruritus Visual Analog Scale(pVAS)were utilized to evaluate pruritic responses,and skin tissue samples were collected from the inguinal area for pathological assessment of skin infammatory cell infltration.The results showed that anti-IL-31RA pAbs with high titers(1:128,000)and specifcity were efectively produced.In the CAD model group,the severity of skin damage,pruritus score,infammatory cell infltration and level of infammatory factors were considerably elevated.Anti-IL-31RA pAbs relieved pruritic behavior and dermatitis in dogs compared to placebo-treated dogs.In conclusion,anti-IL-31RA pAbs efectively suppressed CAD in vivo and are anticipated to be an efective novel treatment for pruritic skin disorders such as CAD.

Prevalence and Factors Associated with Positivity of Antinuclear Antibodies (ANA) Patterns, Native Anti-DNA and Extractable Nuclear Antigens (ENA) Antibodies: Experience from a Laboratory in Dakar

Background: Diagnosis of autoimmune diseases (AID) is challenging, due to overlapping features with other non-immune disorders. Anti-nuclear antibodies (ANA) are sensitive screening tests but anti-deoxyribonucleic acid-antibody (anti-DNA), and anti-extractable nuclear antigens (anti-ENA) are specific for AIDs. We aimed to look at ANA patterns in our patients and correlated them with anti-ENA for proper interpretation and better patient management cost-effectively. Methods: A retrospective study was conducted over 1 year from January to December 2022 who were tested for ANA at biology medical laboratory of Pasteur Institute of Dakar. Anti-ENA and anti-DNA results were also analyzed for ANA-positive patients. Statistical analysis was performed using STATA 14.0, p Results: 216 patients were analyzed. Women predominated at 79.2% and mean age was 48 years [CI 95%, 46 - 50], with extremes of 10 and 89. Most represented age group was [41 - 60] with 38%. ANA was positive in 27 (12.5%) of patients, 59.2% of whom were strongly positive (titer of 1/1000, 1/3200 or 1/6400). The most common pattern was nuclear speckled, which was found in 77.8% of samples. Anti-ENA and anti-DNA positivity in ANA-positive patients was found respectively in 63% (17/27) and 1.4% (3/27) of the samples analyzed. Most commonly identified anti-ENA was anti-Sm 29.6%, anti-SSA 29.6%, anti-Ro-52 25.9%, anti-RNP 18.5% and anti-SSB 14.8% which was associated with speckled pattern. Association results indicated a significant relationship between both tests and between ANA titer in the anti-ENA- and ANA-positive patients (p 0.001). Conclusions: ANA, Anti-ENA and anti-DNA antibodies are essential for AIDS diagnosis. However, the testing repertoire should follow an algorithm comprising of clinical features, followed by ANA results with nuclear, mitotic, and cytoplasmic patterns, anti-ENA, and anti-DNA for a more meaningful, and cost-effective diagnostic approach.

Anti-OX40 Antibody Combined with HBc VLPs Delays Tumor Growth in a Mouse Colon Cancer Model

Objective Combination immunotherapy strategies targeting OX40,a co-stimulatory molecule that can enhance antitumor immunity by modulating the proliferation,differentiation,and effector function of tumor-infiltrating T cells,have attracted much attention for their excellent therapeutic effects.In this study,we aimed to evaluate the antitumor efficacy of combined anti-OX40 and hepatitis B core viruslike particles(HBc VLPs)therapy using a mouse colon cancer model.Methods Humanized B-h OX40 mice were injected subcutaneously with MC38 colon tumor cells and treated with HBc VLPs+anti-h OX40 antibody.Tumor growth was monitored.Flow cytometric analysis was performed to evaluate the populations of T cell subsets in the tumors.Results The combination of anti-OX40 with HBc VLPs resulted in a significant delay in tumor growth,suggesting that a potent antitumor immunity was induced by the combination therapy.Further studies revealed that HBc VLPs+anti-OX40 treatment induced a significant increase in effector T cells(Teffs)and a significant decrease in regulatory T cells(Tregs)in the tumor microenvironment(TME),which accounted for the synergistic antitumor effect of anti-OX40 in combination with HBc VLPs.Conclusion Combination therapy of anti-h OX40 and HBc VLPs provides synergistic antitumor activity in colon cancer-bearing mice,which may represent a potential design strategy for cancer immunotherapy.

Contribution of Anti-p63 Antibodies in the Interpretation of Benign Label Prostatic Biopsies

Introduction: Prostate cancer is the second most common cancer in men. The diagnosis is most often based on the prostate biopsies’ analysis and on histological criteria recognizable on standard coloring. In some cases, the use of immunohistochemistry is important. Objectives: This paper aims to specify the p63 phenotypic profile of lesions diagnosed benign, with minimal suspect foci, difficult to interpret, HGPIN (high grade intraepithelial neoplasia) and LGPIN (low-grade prostatic intraepithelial neoplasia) and evaluate the manual technique of p63 immunohistochemistry. Patients and Method: This was a retrospective, descriptive study of prostate biopsies recorded in the PAC service of the HALD from January 1st, 2018 to December 31st, 2018. It was completed by a manual immunohistochemical study of the blocks enrolled from November 19th to December 4th, 2020 in the PAC department of the HPD. The studied parameters were: registry number, age, clinical stage, prostate volume, PSA level, microscopic appearance and p63 immunohistochemical profile. Results: Our study included 60 prostate biopsies. The ages of our patients varied from 45 to 77 years, with an average of 64.2 years and a standard deviation of 6.2. The majority of patients were at clinical stage cT2b (33%) with a prostate volume varying between 33.15 and 169.4 cc. The minimum value of PSA in our series is 5 ng/ml, the maximum being 100 ng/ml with an average level of 24.1 ng/ml and a standard deviation of 21.2. Our series included 50 adenomyomatous hyperplasias, 7 adenomyomatous hyperplasias associated with chronic prostatitis, 2 HGPIN and 1 LGPIN. After re-reading we found 5 discordant cases, which corresponded to minimal suspect foci (kappa = 0.5098). The p63 marking was informative in 53 cases, i.e. 88%, and non-informative in 7 cases, i.e. 12%. Among the uninformative markings, 2 were due to lack of tissue adhesion to the slides. Among the informative markings, 11 were negative. p63 immunohistochemistry was useful in all suspected foci and detected 6 other minimal foci of adenocarcinoma. Conclusion: The immunostaining with the anti-p63 antibody in the prostate cancer diagnosis is of considerable benefit. It made it possible to correct 11.3% of benign diagnosis in minimal malignant focus in our context. Despite the difficulties associated with the manual technique, it is possible to have an informative rate, similar to the automatic technique.

Two Cases of Autoimmune Encephalitis with Multiple Anti-neuronal Antibodies

Autoimmune encephalitis(AE)can arise from various etiologies and present with complex clinical manifestations,especially in cases involving multiple anti-neuronal antibodies.This report presents two cases of AE with multiple anti-neuronal antibodies admitted to Ningbo Medical Center Li Huili Hospital on October 9,2020,and March 12,2024.Case 1 is a 15-year-old boy with positive anti-N-methyl-D-aspartate receptor(NMDAR)and anti-metabotropic glutamate receptor 5(mGluR5)antibodies in his serum and cerebrospinal fluid(CSF).Case 2 is a 14-year-old boy with positive NMDAR and myelin oligodendrocyte glycoprotein(MOG)antibodies in his CSF.Patients with AE who have multiple anti-neuronal antibodies present significant diagnostic and therapeutic challenges,warranting close clinical attention.

Production and Characterization of Anti-estrone Monoclonal Antibody

Objective Determination of estrone (E1) levels has a significant meaning in evaluating physiological effect and diagnosing some diseases. In order to detect free E1 in biological fluids, a monoclonal antibody specific for E1 was prepared after the complete antigen of E1 was synthesized. The purified monoclonal antibody was fully characterized for later immunoassay. Methods 3-O-carboxymethyl ether derivative of E1 was synthesized and in turn coupled to bovine serum albumin (BSA) to form complete antigen E1-BSA. A monoclonal antibody (McAb) specific for E1 was produced both in vitro and in vivo by a hybridoma anti-E1. Anti-E1 was prepared by fusion of SP2/0 murine myeloma cells with spleen cells isolated from immunized BALB/c mouse. The McAb was characterized by enzyme-linked immunosorbent assay (ELISA), SDS-PAGE and Western-blotting. The specificity of the immunoassay was investigated by determining the cross-reactions of E1 analogs when free E1 was detected by competitive indirect enzyme-linked immunosorbent assay (CI-ELISA). Results Analysis revealed that anti-E1 McAb (E1-McAb) was of the IgG1 type, the molecular weight of E1-McAb was 164 000 daltons. The affinity constant of E1-McAb with coated complete antigen was 8.2108L/mol. The linear range for free E1 determined by CI-ELISA was 10pg/mL^10ng/mL. The detection limit was 21.4 pg/mL (defined as twice the standard deviation of the blank). Conclusion The CI-ELISA developed with E1-McAb was both sensitive and specific. The prepared E1-McAb can be used in some immunoassays.

Mechanisms of resistance to anti-EGFR monoclonal antibody treatment in metastatic colorectal cancer

Metastatic colorectal cancer (mCRC) continues to be counted as a major health problem. The introduction of newer cytotoxics, irinotecan and oxaliplatin, has achieved a significant improvement in survival rates. Novel targeted therapies (bevacizumab, and cetux-imab) in combination with most efficient chemotherapy regimens have pushed the median survival beyond the 2-year mark and increased the proportion of patients which could benefit from resection of metastatic lesions. In addition, several studies have proved that the CRC mutation profiles should influence patient selection or stratif ication in prospective trials. KRAS mutational status represents a paradigm for biomarker development in the era of molecular targeted therapies. The present article is an overview of the most important studies in the development of biomarkers for the optimization of anti-epidermal growth factor receptor (anti-EGFR) treatment in mCRC, beyond KRAS mutations, which is a work in progress. The aim will be to identify molecular markers that might be used to select patients with a higher probability of response to anti-EGFR monoclonal antibodies. Overall the accumulating evidence of the molecular biology of CRC has substantially changed the approach to mCRC treatment and has given clinicians more rational options for treating this illness.

Identification and epitope mapping of anti-p72 single-chain antibody against African swine fever virus based on phage display antibody library

African swine fever virus(ASFV)is a lethal pathogen that causes severe threats to the global swine industry and it has already had catastrophic socio-economic effects.To date,no licensed prophylactic vaccine exists.Limited knowledge exists about the major immunogens of ASFV and the epitope mapping of the key antigens.As such,there is a considerable requirement to understand the functional monoclonal antibodies(mAbs)and the epitope mapping may be of utmost importance in our understanding of immune responses and designing improved vaccines,therapeutics,and diagnostics.In this study,we generated an ASFV antibody phage-display library from ASFV convalescent swine PBMCs,further screened a specific ASFV major capsid protein(p72)single-chain antibody and fused with an IgG Fc fragment(scFv-83-Fc),which is a specific recognition antibody against ASFV Pig/HLJ/2018 strain.Using the scFv-83-Fc mAb,we selected a conserved epitope peptide(221MTGYKH226)of p72 retrieved from a phage-displayed random peptide library.Moreover,flow cytometry and cell uptake experiments demonstrated that the epitope peptide can significantly promote BMDCs maturation in vitro and could be effectively uptaken by DCs,which indicated its potential application in vaccine and diagnostic reagent development.Overall,this study provided a valuable platform for identifying targets for ASFV vaccine development,as well as to facilitate the optimization design of subunit vaccine and diagnostic reagents.

血清anti-β2GPI、FSTL1、PD-1对系统性红斑狼疮免疫功能、疾病活动度的影响

目的探究血清抗β2糖蛋白I抗体(anti-β2GPI)、卵泡抑素样蛋白(FSTL1)、程序性死亡受体1(PD-1)对系统性红斑狼疮(SLE)患者免疫功能、疾病活动度的影响。方法选取我院2019年1月~2022年6月SLE患者113例作为观察组,另选取同期健康体检者105例作为对照组。比较两组、不同疾病活动度患者血清anti-β2GPI、FSTL1、PD-1水平,评价各血清指标与疾病活动度的关系,并根据观察组血清表达水平分为高表达者、低表达者,对比两者免疫功能(CD4+、CD8+、CD4+/CD8+),分析各血清指标与免疫功能相关性。结果观察组血清anti-β2GPI、FSTL1、PD-1高于对照组(P<0.05);血清anti-β2GPI、FSTL1、PD-1高表达患者CD4+、CD4+/CD8+低于低表达患者,CD8+高于低表达患者(P<0.05);血清anti-β2GPI、FSTL1、PD-1与CD4+、CD4+/CD8+呈负相关,与CD8+呈正相关(P<0.05);血清anti-β2GPI、FSTL1、PD-1水平随疾病活动度增加呈升高趋势,且重度活动患者>中度活动患者>轻度活动患者>病情稳定患者(P<0.05);血清anti-β2GPI、FSTL1、PD-1与疾病活动度显著相关(P<0.05)。结论血清anti-β2GPI、FSTL1、PD-1高表达可能参与SLE患者细胞免疫功能紊乱、疾病活动度加重的发生过程。

Prevalence of Anti-endothelial Cell Antibodies in Patients with Pulmonary Arterial Hypertension Associated with Connective Tissue Diseases

Objective To investigate the prevalence of anti-endothelial cell antibodies (AECAs) in the sera of connective tissue diseases (CTD) patients with pulmonary arterial hypertension (PAH) and its correlation with clinical manifestations. Methods AECAs in sera of 39 CTD patients with PAH,22 CTD patients without PAH,and 10 healthy donors as controls were detected with Western blotting. The prevalence of different AECAs in different groups was compared and its correlation with clinical manifestations was also investigated. Results The prevalence of AECAs was 82.1% in CTD patients with PAH,72.7% in CTD patients without PAH,and 20.0% in healthy donors. Anti-22 kD AECA was only detected in CTD patients with PAH (15.4%). Anti-75 kD AECA was more frequently detected in CTD patients with PAH than in those without PAH (51.3% vs. 22.7%,P<0.05). In CTD patients with PAH,anti-75 kD AECA was more frequently detected in those with Raynaud’s phenomenon or with positive anti-RNP antibody. Conclusion AECAs could be frequently detected in CTD patients with or without PAH,while anti-22 kD and anti-75 kD AECA might be specific in CTD patients with PAH.

Organizing Pneumonia in A Patient Double-Positive for ANCA and Anti-GBM Antibodies:A Case Report

Both anti-glomerular basement membrane(GBM)disease and the anti-neutrophil cytoplasmic antibody(ANCA)-associated vasculitis(AAV)are common causes of pulmonary-renal syndrome.Organizing pneumonia(OP),a special pattern of interstitial lung disease,is extremely rare either in AAV or anti-GBM disease.We report an old woman presented with OP on a background of co-presentation with both ANCA and anti-GBM antibodies.

Animal models for anti-neutrophil cytoplasmic antibody-associated vasculitis:Are current models good enough?

Antineutrophil cytoplasmic autoantibody(ANCA)-associated vasculitis(AAV)is a rare and severe systemic autoimmune disease characterized by pauci-immune necrotizing inflammation of small blood vessels.AAV involves multiple organ systems throughout the body.Our knowledge of the pathogenesis of AAV has increased considerably in recent years,involving cellular,molecular and genetic factors.Because of the controlled environment with no other confounding factors,animal models are beneficial for studying the mechanistic details of disease development and for providing novel therapeutic targets with fewer toxic side effects.However,the complexity and heterogeneity of AAV make it very difficult to establish a single animal model that can fully represent the entire clinical spectrum found in patients.The aim of this review is to overview the current status of animal models for AAV,outline the pros and cons of methods,and propose potential directions for future research.

Waning of Anti-Spike IgG Antibody Titer after COVID-19 Vaccination: Myth or Reality?

This observational prospective study was conducted on 25 patients who had received a full 3-dose COVID-19 vaccination scheme with a follow-up ranging from 12 to 19 months after the last injection. The aim of the study was focused on a single biological indicator the anti-spike IgG antibody titer. The age of the patients ranged from 51 to 85 years old. 15 out 25 patients (60%) presented a comorbidity. Our data showed a persistent positive anti-spike IgG antibodies titer ranging from 105 to 5680 BAU/mL (mean: 2661 BAU/mL) in all patients. In view of these results, systematic administration of a SARS-CoV-2 vaccine booster is questionable and should be individually tailored according to the patient medical condition and the anti-spike IgG antibody level.

Molecular Markers for the Prediction of Anti-EGFR Monoclonal Antibody Treatment Efficacy in Metastatic Colorectal Cancer

The implementation of individualized targeted therapy for metastatic colorectal cancer (mCRC), in addition to standard chemotherapeutic regimens, currently is a topic under debate. Approximately 35% - 45% of mCRC patients exhibit mutated KRAS, which is considered to be an independent predictor of poor response to treatment with epidermal growth factor receptor (EGFR) monoclonal antibody. However, only about 50% of patients with wild-type KRAS respond to anti-EGFR therapy. Two major EGFR-dependent signaling pathways, RAS-RAF-MAPK and PI3K-PTEN-AKT, may be involved in the poor response to anti-EGFR. Increased EGFR gene copy number as detected by fluorescence in situ hybridization, but not increased EGFR protein expression, correlates with efficacy of anti-EGFR treatment. The identification of mutations in BRAF and PIK3CA (exon 20) and deletions in PTEN also may help clinicians screen for anti-EGFR resistance in mCRC patients with wild-type KRAS. To guide health professionals through the realm of individualized targeted therapies for mCRC, we review recent progress on identifying negative predictors and prognostic markers of anti-EGFR treatment efficacy.

Buffalo anti-PDC-109 antibodies improve the semen quality profiles and in-vitro zona binding index and minimize the cryoinjury of sperm in cryopreserved buffalo semen

Objective:To optimize the concentration of PDC-109 protein in semen for higher cryopreservability and fertility by sequestration or neutralization of PDC-109 by its antibodies(anti-PDC-109 antibodies)in bubaline species.Methods:PDC-109 protein was purified by applying two-step chromatography procedures.Purified protein was injected in rabbits to raise antibodies.These raised anti-PDC-109 antibodies were used in neutralization or sequestration of PDC-109 in in-vitro model.Ejaculates were collected from buffaloes and splited for four groups.Group 1 received egg yolk Tris glycerol extender,without anti-PDC-109 antibodies,while group 2 to 4 received anti-PDC-109 antibodies 266μg/mL,80μg/mL,and 26μg/mL in Tris-fructose-citrate buffer,respectively.Semen quality parameters viz.,forward progressive motility,viability,total morphological abnormality,acrosomal integrity,plasma membrane integrity,cryoinjury and in-vitro zona binding index were evaluated.Results:Semen quality parameters of neat semen were within the normal range of bubaline species.Sperm motility,livability,acrosomal integrity,plasma membrane integrity,and cholesterol content of sperm were decreased and total sperm abnormality was increased significantly in post-thaw semen compared to those in pre-freeze and fresh semen(P<0.05).Semen in group 2 had higher sperm motility,livability,acrosomal integrity,plasma membrane integrity,and cholesterol content of sperm and lower total sperm abnormality significantly compared to those in group 1,3 and 4 at pre-freeze and post-thaw stages(P<0.05).Conclusions:Sequestration or neutralization of PDC-109 by its antibodies significantly improves pre-freeze,and post-thaw semen quality parameters and in-vitro zona binding index with simultaneously reducing cryoinjury or cryodamage in the sperm of bubaline species.