Requirements for human antibody cocktails for oncology

Cancer patients receiving antibodies as monotherapy have benefited from these treatments. However, significant improvements can be made that should make the therapy more effective. Applying lessons learned from the natural oligoclonal antibody response that cancer patients mount to their own tumours suggests that cocktails of monoclonal antibodies could be formulated, which may be more effective in treating cancers. The next phase of antibody immunotherapy will include cocktails of monoclonal antibodies. Various requirements for human antibody cocktails are discussed, as well as potential limitations of this approach.     

人源抗CEA噬菌体抗体库的构建及筛选

目的构建人癌胚抗原(CEA)特异性噬菌体抗体库,制备人源抗CEA单克隆抗体(mAb)。方法用常规RT-PCR法,直接从10例结肠癌患者肠系膜淋巴结中的淋巴细胞克隆出免疫球蛋白Fd段和κ链基因,建成噬菌体抗体库。对抗体库进行5轮吸附-洗脱-扩增的亲和选择后,以ELISA法鉴定抗CEA噬菌体。结果RT-PCR可有效地扩增出Fd和κ基因并以此构建成容量为5.2×106的噬菌体抗体库。经5轮亲和选择可使特异性噬菌体抗体得到高度富集,并成功获得抗CEA噬菌体抗体阳性克隆。结论抗CEA噬菌体抗体库的构建和人源抗CEA mAb的制备,为CEA阳性表达肿瘤的靶向治疗奠定基础。     

Clinical application of SARS-CoV-2 antibody detection and monoclonal antibody therapies against COVID-19

The purpose of this study was to investigate the clinical application of severe acute respiratory distress syndrome coronavirus-2(SARS-CoV-2) specific antibody detection and anti-SARS-CoV-2 specific monoclonal antibodies(m Abs) in the treatment of coronavirus infectious disease 2019(COVID-19). The dynamic changes of SARS-CoV-2 specific antibodies during COVID-19 were studied. Immunoglobulin M(Ig M) appeared earlier and lasted for a short time, while immunoglobulin G(Ig G) appeared later and last...     

Lessons learned from immunoadsorption for hyperviscosity in IgM multiple myeloma—A case report

We report the case of a 63-year-old Caucasian woman with multiple relapsed IgM multiple myeloma (MM) and elevated free kappa light chains (fκLC). Due to hyperviscosity syndrome with visual impairment, regular plasma exchanges were performed. As part of her 11th line of therapy, an experimental protocol consisting of pembrolizumab, pomalidomide, and dexamethasone was initiated. To reduce fκLC and immunoglobulin (Ig) M, we performed immunoadsorption (IA) using columns containing recombinant single domain camelid antibody fragments as ligands. We measured pembrolizumab (humanized IgG4 kappa anti-PD1 antibody) levels before and after each IA session and found a 98.1% reduction from baseline with five sessions of IA. Comparable elimination kinetics were observed for serum IgG, whereas fκLC and IgM were eliminated to a substantially lesser extent. These findings highlight that in hyperviscosity syndrome due to IgM MM, broad spectrum IA columns might be only moderately effective compared to total plasma exchange or double filtration plasmapheresis. Monoclonal antibodies are efficiently reduced by extracorporeal therapies and re-dosing is necessary to provide sufficient efficacy. In the case of serious adverse events such as immune-related adverse events, IA might be used to eliminate the monoclonal antibody. Measuring IgG levels might be a reasonable strategy for monitoring drug levels of monoclonal antibodies during IA.     

抗人CD20单克隆抗体治疗B细胞淋巴瘤研究进展

近年来,对B细胞淋巴瘤的抗原靶向性治疗已取得了巨大进展。出现了一种新的非常有发展前景的治疗方法。即以B细胞分化抗原CD20为靶抗原的单克隆抗体（McAb)疗法。CD20是非糖基化的跨膜磷酸蛋白,分子质量为35kD。表达在大多数成熟B细胞表达,分化为浆细胞后,CD20表达消失。CD20可能参与B细胞成熟与分化的调节,作为钙离子通道发挥某些生物学作用。更为重要的是,95%以上的B细胞淋巴瘤表达CD20,且无显内化及脱落,这些特点使其成为单克隆抗体疗法的理想靶抗原,临床试验结果表明,抗CD20单克隆抗体既可单独应用,亦可作为放射性同位素或细胞毒制剂的载体,其疗效显,且使用安全,副反应小,许多研究报道已阐明了抗CD20单克隆抗体治疗B细胞淋巴瘤的几种可能的效应机制。目前,人们正致力于探索一些新的治疗策略以期提高这种疗法的特异性,减少其非特异性的毒副作用。     

急性巨核细胞白血病的免疫表型分析

目的 探讨急性巨核细胞白血病（ＡＭＫＬ）的免疫表型特点及其意义。方法 利用多种单克隆抗体对５例急性巨核白血病患的白血病细胞进行标记，应用间接免疫荧光技术或流式细胞术检测各单抗的表达率。２例通过电镜观察血小板过氧化物酶的表达。结果 ４／５例表达巨核系单抗，所有病例均不表达淋巴系相关单抗，但３例有髓系单抗的表达。ＨＬＡ－Ｄｒ、ＣＤ３４、ＣＤ３８也表达于不同的病例。结论 ＡＭＫＬ可能起源于较早期的干细     

细胞角蛋白20单克隆抗体纯化方法的探讨

[目的]探讨细胞角蛋白20（CK20）单克隆抗体纯化的方法。[方法]采用SPA-Sepharose CL-4B为固定相的亲和层析法，将含有CK20单抗的小鼠腹水过柱后，用pH6．0、4．0和3．03个范围0．1mol／L枸橼酸分步洗脱，并以双向琼脂扩散法和Western-blotting对获得的单抗加以鉴定。[结果]以枸橼酸3个pH范围洗柱时分别收集5ml、4ml和20ml洗脱液，双向琼脂扩散法鉴定结果是：pH6．0时洗脱液中单抗亚型是IgG1、pH4．0时为IgG2a、pH3．0时为IgG2b，Western-blotting结果表明所获抗体具有较高免疫活性。[结论]该方法简便、特异，并可直接获得不同亚型单抗，适合腹水中CK20单克隆抗体纯化。     

Circumsporozoite protein as a potential target for antimalarials

Since the discovery of circumsporozoite protein (CSP), a major sporozoite surface antigen, by Ruth Nussenzweig and Victor Nussenzweig in the early 1980s, the role of CSP in protection against malaria has been extensively investigated. Several monoclonal antibodies against CSP have been generated to date, with some of them mediating antimalarial protection upon passive transfer into animals. Genetically engineered transgenic mosquitoes producing the anti-CSP antibody have recently been generated to reduce malarial transmission. A monoclonal anti-CSP antibody was produced in mice by adeno-associated virus vector, which protected them from malaria. Phase III trials with RTS,S vaccine that targets CSP of Plasmodium falciparum have shown modest efficacy. Polyclonal anti-CSP antibodies derived from children who received the RTS,S vaccine failed to block malarial transmission through mosquitoes, but passive transfer of monoclonal antibodies raised from RTS,S-vaccinated recipient conferred protection against malaria in mice. Taken together, these findings may imply CSP as an antimalarial target.     

抗人甲状腺球蛋白人源化单克隆抗体的制备

目的获得抗人甲状腺球蛋白（hTg）人源化单克隆抗体。方法从分泌抗人甲状腺球蛋白的杂交瘤细胞株hTg-60中提取总RNA,逆转录-酶链聚合反应（RT-PCR）扩增出鼠源性单克隆抗体hTg-60的VH及VL基因,经基因测序分析,设计出人源化抗体的基因引物,获得抗Tg单克隆抗体人源化VH及VL基因并将其克隆入真核表达载体,转染中国仓鼠卵巢细胞（CHO）,表达人源化抗hTg抗体。结果成功获得了6株能稳定分泌抗hTg人源化抗体的细胞株。结论为进一步获得人源化的hTg抗体及临床应用奠定了基础。     

Monoclonal antibody therapies for solid tumors

Background: About 1.4 million new cancer cases are expected to be diagnosed in the US in 2008 and the overall costs of cancer treatment is estimated at > $200 billion per year. Over half a million people will die from cancer each year. Monoclonal antibodies have attracted attention because of their potential clinical benefits. Currently, there are four monoclonal antibodies that have been approved by the FDA for the treatment of solid tumor malignancies. Over 100 other antibodies are in early- and late-phase clinical testing. Objectives/methods: Clinical efficacy of FDA-approved and investigational monoclonal antibody therapies are summarized. Results/conclusions: Anti-tumor responses have been modest. However, the advent of novel antibody engineering and combination therapy strategies has generated much optimism for current and future antibody-based therapeutics in solid tumors.     

抗体依赖细胞介导的细胞毒性抗肿瘤研究进展

近年来,随着单克隆抗体临床应用的逐渐增加,抗体依赖细胞介导的细胞毒性(antibody-dependent cell-mediated cytotoxicity,ADCC)的抗肿瘤效果日益受到重视。自然杀伤细胞(NK细胞)是介导ADCC特异性抗肿瘤最主要的效应细胞,而吞噬细胞、T细胞及粒细胞对ADCC抗肿瘤效应也有一定作用。ADCC被证明是单克隆抗体临床治疗肿瘤的重要机制和手段,IgG型抗体首先通过抗原结合部位与靶细胞(肿瘤细胞)结合,再由效应细胞上的FcγR识别其Fc段,介导ADCC。已投入临床应用的单克隆抗体有:利妥昔单克隆抗体及新型的抗CD20单克隆抗体、曲妥珠单克隆抗体、西妥昔单克隆抗体、依决洛单克隆抗体、尼妥珠单克隆抗体及吉妥单克隆抗体,这些单克隆抗体均具有ADCC抗肿瘤作用。单克隆抗体介导的ADCC抗肿瘤疗效受多方面因素影响,包括:IgGFc受体的基因多态、肿瘤细胞抗原、血清抗体水平、细胞因子及药物等。FcγRIIIa-158V/V基因型和FcγRIIa-131H/H基因型比其他基因型的外周血单个核细胞的ADCC作用更强。提高肿瘤抗原水平,降低血清抗体水平或增加某些细胞因子(如IL-2,IL-21,IL-15等)均可提高单克隆抗体的ADCC作用,而避免使用某些药物(如地塞米松、肿瘤坏死因子拮抗剂)或适当使用昂丹司琼和氯马斯汀也可增强单克隆抗体的ADCC抗肿瘤作用。总之,ADCC抗肿瘤的临床应用十分重要,但效果会受多方面因素的影响。本文就ADCC杀伤机理、临床常用的和已投入临床使用的具有ADCC抗肿瘤效应的单克隆抗体,影响ADCC抗肿瘤效率的因素及其他细胞ADCC的抗肿瘤作用进行了综述。     

Clinical uses of monoclonal antibodies

Immunization of animals for the purpose of generating antisera for clinical and experimental use results in the production of antibodies with multiple specificities directed at the parent antigen. Furthermore, if the immunogen contains contaminants of a highly immunogenic nature, large amounts of irrelevant antibody will be generated and which must subsequently be absorbed. The unpredictable nature of heteroantisera can be overcome by adopting the monoclonal antibody technique which allows one to produce standardized antibody with well defined and selected specificity toward a given epitope on a macromolecule. Cross-reactions are not eliminated by monoclonal antibodies.     

Development of a recombinant hepatitis B immunoglobulin derived from B cells collected from healthy individuals administered with hepatitis B virus vaccines: A feasibility study

Background

In Japan, plasma with a high concentration of Hepatitis B Virus (HBV) antibodies for hepatitis B immunoglobulin (HBIG) is almost entirely imported. We aimed to produce recombinant HBIG by isolating immunoglobulin cDNAs against the HBV surface antigen (HBsAg).

Study Design and Methods

B cells expressing HBsAg antibodies were obtained from blood center personnel who had been administered HB vaccine booster and then isolated by either an Epstein–Barr virus hybridoma or an antigen-specific memory B cell sorting method. Each cDNA of the heavy and light chains of the target antibody was cloned into an IgG₁ expression vector and transfected into Expi293F cells to produce a recombinant monoclonal antibody (mAb), which was screened by ELISA and in vitro HBV neutralizing assays. The cross-reactivity of the mAbs to normal human molecules was evaluated by ELISA and immunohistochemistry.

Results

Antibody cDNAs were cloned from 11 hybridoma cell lines and 204 HBsAg-bound memory B cells. Three of the resulting recombinant mAbs showed stronger neutralizing activity in vitro than the currently used HBIG. All three bind to the conformational epitope(s) of HBsAg but not to human DNA or cells.

Discussion

We successfully isolated HBV-neutralizing monoclonal antibodies from B cells collected from healthy plasma donors boosted against the HBV. To obtain an alternative source for HBIG, HBV-neutralizing monoclonal antibodies from B cells collected from healthy plasma donors boosted against the HBV may be useful.     

Idiotypic markers of polyclonal B cell activation. Public idiotypes shared by monoclonal antibodies derived from patients with systemic lupus erythematosus or leprosy.

We investigated idiotypic markers of monoclonal antibodies derived from patients with polyclonal B-cell activation. Four monoclonal antibodies with different ligand binding specificities derived from a patient with lepromatous leprosy and three monoclonal anti-DNA antibodies from two patients with SLE were studied. Three new public idiotopes, which were common to monoclonal antibodies from all three patients, were defined by five polyclonal rabbit antiidiotypes, two monoclonal mouse antiidiotopes, and a monoclonal mouse antibody against a synthetic peptide that contains residues of the heavy chain CDR-1 of a monoclonal lupus anti-DNA antibody. The antibody against the synthetic idiotype was found to react with native immunoglobulins in solution. One idiotope was found to be consistently immunogenic in all animals tested. Since the three patients are of different ethnic origins, these shared idiotypes are probably encoded by germline V genes. These genes may be recurrently expressed in states of polyclonal B-cell activation, regardless of etiology. The results suggest that some autoantibodies arise by expansion of a pool of precursors in the normal antibody repertoire.     

Monoclonal and polyclonal antibodies compared for radioimmunoassay of somatomedin-C in patients with acromegaly or hypopituitarism

We used a synthetic recombinant analog of somatomedin-C (Sm-C) to directly compare the performance of polyclonal and monoclonal antibodies for measuring Sm-C in serum of normal persons and in acromegalic or hypopituitary patients. Mean concentrations of Sm-C in healthy adults were 181 (SD 42) micrograms/L as measured with the monoclonal antibody, 194 (SD 61) micrograms/L with the polyclonal antibody. Both antisera gave excellent discrimination between acromegalics and normals. However, the assay with the polyclonal antibody was more sensitive than that with the monoclonal antibody (lower detection limits: 5 vs 100 micrograms/L) and thus better suited for quantifying Sm-C in samples from hypopituitary patients.     

Monoclonal antibody for the detection of Clostridium botulinum type A toxin

Hybridomas producing monoclonal antibodies against type A Hall strain Clostridium botulinum toxin were generated by fusing mouse myeloma cell line P3-X63-Ag8.653 with spleen cells of Balb/c mice immunized with C. botulinum type A toxoid. The monoclonal antibody from one hybridoma, identified as No. 424, was selected from 61 others for its high antibody titre. This monoclonal antibody was used in a double-sandwich enzyme-linked immunosorbent assay (ELISA) system to detect type A toxin in culture fluids and in foods. The monoclonal antibody did not react with either C. botulinum toxin types B, C, D, E and F or with other clostridial species tested. This particular monoclonal antibody (No. 424) did not neutralize type A toxin in the mouse bioassay procedure but detected approximately 10 mouse lethal doses of type A toxin/ml culture fluid. Monoclonal antibody and rabbit antitoxin to type A C. botulinum toxin were useful in a double-sandwich ELISA for the rapid and specific detection of type A toxic fluids in culture and in food samples.     

Platelet-specific antibodies in HLA-immunized patients receiving chronic platelet support

BACKGROUND: In HLA-alloimmunized patients, the unexpected failure of HLA-matched platelet transfusions usually raises the suspicion about concomitant platelet-specific antibodies. As the reported frequency of platelet-specific antibodies in multitransfused patients varies widely, the aim of this study was to determine the prevalence of such antibodies in a population of chronic thrombocytopenic patients with HLA antibodies. STUDY DESIGN AND METHODS: From 1985 to 1997, 11,777 determinations of HLA antibodies were performed in 1330 hematologic patients receiving chronic platelet support. Fifty-two patients with HLA alloimmunization that lasted more than 1 month were selected. The search for platelet-specific antibodies was performed by using a monoclonal antibody immobilization of platelet antigens assay, thus allowing the identification of platelet-specific antibodies directed against the platelet glycoproteins (GP) Ib/IX, GPIIb/IIIa, and GPIa/IIa. Specificity of the platelet-specific antibodies was further investigated by using a solid-phase assay with chloroquine-treated platelets. RESULTS: Only 2 (3.8%) of the 52 patients had platelet-specific antibodies. One antibody reacted with an epitope of the GPIIb/IIIa that was present in all the panel platelets, and that probably was an autoantibody. The other was an anti-HPA-5b. CONCLUSIONS: The prevalence of platelet-specific antibodies in patients with HLA alloimmunization is very small. The search for concomitant platelet-specific antibodies would be indicated only when other causes of refractoriness to HLA-matched platelets are ruled out.     

Ligelizumab for the treatment of chronic spontaneous urticaria

ABSTRACT

Introduction

Due to daily hives with itch, sleeplessness, and unforeseen development of angioedema, chronic spontaneous urticaria significantly impairs quality of life, often for years. Its management is challenging. In most cases, H1-antihistamines are not effective. Although the disease is not characterized by specific IgE antibodies against allergens, the last decade demonstrated that neutralizing IgE by using the monoclonal anti-IgE antibody Omalizumab is safe and effective. Nevertheless, symptoms are not controlled by Omalizumab in approximately one-fourth of patients.     

抗人血浆蛋白C单克隆抗体研制的初步报告

用物理化学方法纯化的人血浆蛋白C(HPC)免疫小鼠,制备了8株分泌抗HPC单克隆抗体的杂交瘤细胞株,分别命名为PC01～PC08。各株杂交瘤细胞都具有长期稳定的抗体分泌能力,其中PC03为构象特异性单克隆抗体,二价金属离子对其与HPC的结合有明显影响;PC02和PC03对活性蛋白C的酶活性有促进作用;而PC04具有抑制作用。     

IAA、ICA和GADA的年龄分布及与临床指标的关系研究

目的分析抗胰岛素抗体(IAA)、抗胰岛细胞抗体(ICA)和抗谷氨酸脱羧酶抗体(GADA)在患者中的分布情况及实验室指标的变化,探讨三种糖尿病相关抗体对疾病发生、发展变化的预示价值,为疾病治疗提供依据。方法分析2008-2018年在四川大学华西医院联合检测了IAA、ICA和GADA三种抗体患者的相关信息。将抗体阳性患者分为3个不同的年龄组(<40岁、40~59岁、>59岁),比较抗体检测结果组合在各年龄组间分布差异,分析抗体阳性患者疾病分布情况。以抗体阳性分组,分析三种抗体单项阳性组(单阳组)患者实验室指标检测结果的差异。结果三种抗体在不同年龄组间的分布差异无统计学意义(P>0.05);糖尿病患者人群IAA抗体单项阳性占65.08%;三种抗体主要存在于糖尿病患者中(占92.03%),其他疾病(如癌症、低血糖、系统性红斑狼疮)患者也可出现。碱性磷酸酶在IAA单阳组中的水平显著低于ICA单阳组(P<0.05);尿酸和肌酐在IAA单阳组中的水平显著高于GADA单阳组(P<0.05);球蛋白和肾小球滤过率在IAA单阳组水平显著低于GADA单阳组(P<0.05);空腹胰岛素在IAA单阳组水平显著高于ICA和GADA单阳组(P<0.05)。结论IAA、ICA和GADA三种抗体在各年龄组分布相近。IAA可作为糖尿病抗体检测的首选指标,IAA阳性糖尿病患者更应该注意其并发肾功能损害的风险。