抗P-糖蛋白单克隆抗体JSB-1和三氟拉嗪协同逆转人肾癌MDR的实验研究

目的 为证明钙调蛋白的抑制剂三氟拉嗪和抗P—糖蛋白单克隆抗体(JSB-l)能协同逆转人肾癌的天然多药耐药(MDR)性。方法 我们对经过病理证实36例肾透明细胞癌采用RT-PCR方法检测mdr—l表达水平，高表达的为耐药组，无或低表达的为敏感组，给予阿霉素与三氟拉嗪和JSB-l处理，并采用MTT法评价细胞毒作用，流式细胞仪检测细胞内ADR浓度聚集。结果 三氟拉嗪(TFP)和JSB-l对肿瘤细胞无明显的细胞毒作用；但具有逆转肾癌对ADR的耐药，并且在耐药(P—gp过表达MDR)的肾癌细胞中，效果更明显(P＜0．01)。结论 三氟拉嗪和JSB-l可作为肾癌化疗的辅助用药，联合应用效果更佳，它的低毒副作用使其具有应用价值和可行性。     

新辅助化疗时加用曲妥珠单抗对HER2阳性乳腺癌的临床疗效观察

目的探讨新辅助化疗加用曲妥珠单抗对HER2阳性乳腺癌的疗效。方法回顾性研究了我院2013年5月~2015年3月,对224例HER2阳性可手术乳腺癌病例,主要有两组:术前仅蒽环类化疗组(对照组),共120例;术前蒽环类化疗加用曲妥珠单抗组。两组化疗结束后再予手术。结果两组的手术难度,手术时间,术中出血量,术后引流量及平均住院天数均无显著性差异(P0.05)。但是术前加用曲妥珠单抗组生存率明显高于术前仅蒽环类化疗组。结论对HER2阳性乳腺癌患者行术前新辅助化疗时加用曲妥珠单抗可以提高术前化疗的疗效,尤其能延长患者远期生存率。     

抗人H血型物质单克隆抗体的研制

经H血型物质免疫的BALB/C小鼠脾细胞与SP2/0细胞用PEG融合,获得一株分泌抗人H血型物质单克隆抗体的杂交瘤细胞株—6H_3,为IgG_1类。其效价,细胞培养上清液为512,腹水达16384。凝血特异性试验、吸收放散试验、血凝抑制试验证明其仅对人H血型物质特异。杂交瘤细胞在液氮中贮存1年,分泌抗体能力不变。抗体经56℃保温30min及-20℃和37℃反复冻融试验,凝血活性保持稳定。可望能取代传统的抗H试剂而用于临床血型分型。     

抗EGFR单克隆抗体治疗肿瘤进展

表皮生长因子受体(epidermal growth factor receptor,EGFR)突变、失调或过表达于许多上皮恶性肿瘤,在肿瘤的生长和分化过程中起重要作用。抗EGFR的单克隆抗体是针对于胞外域EFGR的靶向性抗体,临床应用显示了良好的抗肿瘤活性,而且并不产生严重副反应。本文对3种抗EGFR单克隆抗体(cetuximab,panitumumab和nimotuzomab)的药代动力学及其应用研究进行了综述。     

抗人胰腺癌单克隆抗体的制备及其鉴定

目的：制备抗人胰腺癌单克隆抗体,并对其特异性进行鉴定。方法：以人分化胰腺癌细胞株8988为抗原,免疫BALB/c小鼠,常规融合,运用ELISA方法筛选;流式细胞仪及免疫组化鉴定其组织特异性,West blot检测其特异性结合抗原的相对分子量。结果：得到一株高效价的稳定分泌IgG1抗人胰腺癌的单克隆抗体的细胞株,命名为SZ121,腹水效价为2×10^-5。运用ELISA方法及流式细胞仪方法检测证实SZ121特异地识别胰腺癌细胞。West blot检测显示SZ121特异性结合的抗原相对分子量为66 000左右。结论：实验证实SZ121是特异结合胰腺癌细胞的单克隆抗体。     

抗人胰腺癌细胞单克隆抗体的研制及初步鉴定

目的制备抗人胰腺癌细胞单克隆抗体(McAb),并对其特性进行鉴定。方法以胰腺癌细胞系Patu8988免疫Balb／ c小鼠,取其脾细胞与小鼠骨髓瘤细胞SP2／0进行融合,经选择培养,筛选及克隆化后,建立杂交瘤细胞系。以ELISA flow cytometry等方法研究抗体特性。以MTT法研究细胞毒作用。结果得到一株能够稳定分泌特异性抗体的杂交瘤细胞系 A7。用琼脂糖双扩散鉴定其为IgG1亚类,ELISA测定其腹水效价为:1:16000。流式细胞仪检测表明:该抗体与Patu8988 细胞呈阳性反应(>90％),与肝癌、结肠癌、胃癌、肺癌、乳腺癌呈阴性反应(<5％)。MTT法示该抗体对胰腺癌细胞的杀伤率明显高于对照组。结论 McAbA7对胰腺癌的诊断和治疗可能具有一定的临床意义。     

抗尿苷二磷酸葡萄糖焦磷酸化酶2单克隆抗体的制备与鉴定

本研究制备抗人尿苷二磷酸葡萄糖焦磷酸化酶2（UDP-glucose pyrophosphorylase2）的单克隆抗体（mAb）并进行初步鉴定。将正常成人肝组织匀浆离心并分离肝脏胞浆总蛋白,用肝脏胞浆总蛋白免疫BALB/c小鼠,采用杂交瘤技术制备单克隆抗体,通过间接ELISA法、Western blot及免疫组织化学的方法对单克隆抗体进行特性鉴定,通过Uni-ZAP XR肝脏cDNA表达文库筛选鉴定抗体特异性。结果表明：通过间接ELISA筛选获得1株可稳定分泌抗人UGP2mAb的杂交瘤细胞系BAD062,其分泌的单克隆抗体的Ig亚类（型）为IgG2b（κ）,Western blot结果显示该抗体可以特异地识别分子量为56kD的蛋白;应用单克隆抗体BAD062对人肝脏cDNA表达文库（Uni-ZAPXR）进行筛选,阳性噬菌斑测序结果显示2个阳性克隆插入序列均为UGP2。结论：单克隆抗体BAD062特异地识别抗原UGP2,该抗体可用于ELISA检测、Western blot、免疫组织化学实验,为UGP2的研究提供了有力的工具。     

Metnase Mediates Resistance to Topoisomerase II Inhibitors in Breast Cancer Cells

DNA replication produces tangled, or catenated, chromatids, that must be decatenated prior to mitosis or catastrophic genomic damage will occur. Topoisomerase IIα (Topo IIα) is the primary decatenating enzyme. Cells monitor catenation status and activate decatenation checkpoints when decatenation is incomplete, which occurs when Topo IIα is inhibited by chemotherapy agents such as the anthracyclines and epididophyllotoxins. We recently demonstrated that the DNA repair component Metnase (also called SETMAR) enhances Topo IIα-mediated decatenation, and hypothesized that Metnase could mediate resistance to Topo IIα inhibitors. Here we show that Metnase interacts with Topo IIα in breast cancer cells, and that reducing Metnase expression significantly increases metaphase decatenation checkpoint arrest. Repression of Metnase sensitizes breast cancer cells to Topo IIα inhibitors, and directly blocks the inhibitory effect of the anthracycline adriamycin on Topo IIα-mediated decatenation in vitro. Thus, Metnase may mediate resistance to Topo IIα inhibitors, and could be a biomarker for clinical sensitivity to anthracyclines. Metnase could also become an important target for combination chemotherapy with current Topo IIα inhibitors, specifically in anthracycline-resistant breast cancer.     

乳腺癌细胞多药耐药基因（MDR1）表达的检测及其临床意义的探讨

目的：探讨多耐药基因MDR1在乳腺癌中的表达及其临床意义,方法：用Pgp单抗C219,C49,JSB1免疫组化方法检测42例乳腺癌组织中的MDR1表达水平,并行化疗疗效的判定,将各指标对比分析,结果,MDR1表达的阳性率为38．1％,MDR1 达与化疗疗效呈负相关,MDR1表达水平与肿瘤大小,病理组织学分化程度及ER情况均有相关性,结论：MDR1在乳腺癌中的普遍存在,MDR1不仅是耐药性指标,也可能是肿瘤细胞的一个生物学指标。     

生长激素释放肽对乳腺癌耐药蛋白表达的影响及意义

[目的]探讨生长激素释放肽(ghrelin)对乳腺癌耐药蛋白(BCRP)表达的影响.[方法]培养人乳腺癌细胞MDA-MB-231细胞,分别加入ghrelin、多柔比星、ghrelin联合多柔比星,采用tunnel方法检测细胞凋亡,western-blot方法检测细胞丝裂原活化蛋白激酶P38(P38)、磷酸化P38 (p-P38)、BCRP的表达并比较.[结果]tunnel 法检测显示人乳腺癌细胞MDA-MB-231在ghrelin的干预下增殖,而凋亡率在多柔比星的干预下显著降低,ghrelin联合多柔比星能够抑制多柔比星对人乳腺癌细胞MDA-MB-231的杀伤作用(P＜0.001).ghrelin干预组P38、p-P38蛋白表达增加、且BCRP蛋白表达明显上升;多柔比星干预组P38、p-P38蛋白表达下降;ghrelin联合多柔比星组较多柔比星组P38、p-P38蛋白表达增加,且BCRP蛋白表达明显上升(P＜0.05).[结论]ghrelin激活P38MAPK信号通路促进乳腺癌细胞BCRP表达增加从而抑制多柔比星诱导乳腺癌细胞凋亡.     

PNAS-2蛋白单克隆抗体的制备和初步鉴定

本研究制备凋亡相关蛋白PNAS-2的单克隆抗体,并对其进行初步鉴定,为PNAS-2蛋白功能研究提供必需工具。以PNAS-2的合成肽为免疫原,通过动物免疫、细胞融合、克隆化制备抗PNAS-2蛋白的单克隆抗体,并用Western blot、免疫荧光、ELISA实验对单克隆抗体的特异性、效价、亚型进行了检测。结果表明：获得了稳定的分泌型杂交瘤细胞株S-31-7,属IgG1λ亚型,腹水效价为1：8000;Western blot测得分子量为28kD的单条特异性条带;免疫荧光实验显示胞浆中有阳性反应。结论：制备的单克隆抗体特异性好、效价高,能够作为深入研究PNAS-2蛋白功能的有利工具。     

Dacomitinib (PF-00299804), an Irreversible Pan-HER Inhibitor, Inhibits Proliferation of HER2-Amplified Breast Cancer Cell Lines Resistant to Trastuzumab and Lapatinib

The human EGF (HER) family of receptors has been pursued as therapeutic targets in breast cancer and other malignancies. Trastuzumab and lapatinib are standard treatments for HER2-amplified breast cancer, but a significant number of patients do not respond or develop resistance to these drugs. Here we evaluate the in vitro activity of dacomitinib (PF-00299804), an irreversible small molecule pan-HER inhibitor, in a large panel of human breast cancer cell lines with variable expression of the HER family receptors and ligands, and with variable sensitivity to trastuzumab and lapatinib. Forty-seven human breast cancer and immortalized breast epithelial lines representing the known molecular subgroups of breast cancer were treated with dacomitinib to determine IC(50) values. HER2-amplified lines were far more likely to respond to dacomitinib than nonamplified lines (RR, 3.39; P < 0.0001). Furthermore, HER2 mRNA and protein expression were quantitatively associated with response. Dacomitinib reduced the phosphorylation of HER2, EGFR, HER4, AKT, and ERK in the majority of sensitive lines. Dacomitinib exerted its antiproliferative effect through a combined G(0)-G(1) arrest and an induction of apoptosis. Dacomitinib inhibited growth in several HER2-amplified lines with de novo and acquired resistance to trastuzumab. Dacomitinib maintained a high activity in lines with acquired resistance to lapatinib. This study identifies HER2-amplified breast cancer lines as most sensitive to the antiproliferative effect of dacomitinib and provides a strong rationale for its clinical testing in HER2-amplified breast cancers resistant to trastuzumab and lapatinib. Mol Cancer Ther; 11(9); 1978-87. ?2012 AACR.     

乳腺癌三苯氧胺耐药机制的研究进展

三苯氧胺作为一线药物应用于乳腺癌内分泌治疗已有多年,但长期应用后的耐药现象仍然是一个问题.本文拟通过回顾近年来有关乳腺癌三苯氧胺耐药机制的研究进展,为阐明三苯氧胺耐药机理及克服耐药性提供有价值的信息和思路.     

Phase 2a Study of the CCR5 Monoclonal Antibody PRO 140 Administered Intravenously to HIV-Infected Adults

The anti-CCR5 antibody PRO 140 has shown potent and prolonged antiretroviral activity in subjects infected with CCR5-tropic (R5) HIV-1. Prior studies have examined single intravenous doses ranging up to 5 mg/kg of body weight or up to three subcutaneous doses ranging up to 324 mg. Here we report the results of a randomized, double-blind, placebo-controlled trial that examined the antiviral activity, tolerability, and pharmacokinetics of single 5-mg/kg and 10-mg/kg intravenous infusions of PRO 140 in 31 treated subjects. Eligibility criteria included HIV-1 RNA levels of >5,000 copies/ml, CD4⁺ cell counts of >300/μl, no antiretroviral therapy for ≥12 weeks, and detection of only R5 HIV-1 in the original Trofile assay. Following poststudy testing with an enhanced-sensitivity Trofile assay, one subject treated with 10 mg/kg was reclassified as having dual/mixed-tropic virus at screening, and the data for that subject were censored from efficacy analyses. The mean maximum reduction of the HIV-1 RNA level from the baseline level was 1.8 log₁₀ units for both the 5-mg/kg and 10-mg/kg doses (P < 0.0001 relative to placebo). Viral loads reached their nadir at day 12 posttreatment and remained significantly (P < 0.01) reduced through day 29 for both PRO 140 dose groups. Treatment was generally well tolerated, with no dose-limiting toxicity being observed. Peak serum concentrations and overall exposures increased proportionally with dose. In summary, single 5-mg/kg and 10-mg/kg doses of PRO 140 exhibited potent, long-lived antiviral activity and were generally well tolerated. The findings further delineate the safety and antiviral properties of this novel, long-acting antiretroviral agent.The chemokine receptor CCR5 plays a physiological role in the activation and migration of T cells and other leukocytes. CCR5 also binds to the HIV-1 envelope glycoprotein gp120 and serves as a coreceptor for HIV-1 entry into CD4⁺ cells (11). Certain strains of HIV-1 can use the chemokine receptor CXCR4 either exclusively (X4 viruses) or in addition to CCR5 (R5X4 or dual-tropic viruses). Viruses that use CCR5 exclusively (R5 viruses) are the only strains detected in most individuals during the initial to middle stages of disease. CXCR4-using virus can be detected in an increasing percentage of individuals as disease progresses (1, 2, 14, 24). A small-molecule CCR5 antagonist (maraviroc; Pfizer/ViiV Healthcare) has been approved by the U.S. Food and Drug Administration (FDA) for use in patients with only R5 virus detectable (5) and serves to validate CCR5 as a target for new anti-HIV-1 therapies.PRO 140 is a humanized monoclonal antibody that binds to CCR5 and potently inhibits R5 but not CXCR4-using viruses in laboratory studies (15, 22). PRO 140 or its murine counterpart shows synergy and limited cross-resistance with small-molecule CCR5 antagonists in vitro (9, 13, 15). Both intravenous (i.v.) and subcutaneous (s.c.) forms of PRO 140 have previously been evaluated in short-term monotherapy studies with HIV-1-infected subjects with only R5 virus detectable. Both dosage forms of PRO 140 were generally well tolerated relative to placebo and demonstrated potent, prolonged, and dose-dependent antiretroviral activity (6, 7).In a prior study, intravenous PRO 140 was evaluated as single doses of 0.5 mg/kg of body weight, 2 mg/kg, or 5 mg/kg. Antiviral effects increased in a dose-dependent manner, with a 1.83-log₁₀-unit mean reduction in the HIV-1 RNA level being observed with the 5-mg/kg dose. On the basis of these findings, the present study was conducted to evaluate single 5-mg/kg and 10-mg/kg intravenous doses for their antiviral effects, tolerability, and pharmacokinetics (PK) in individuals infected with R5 HIV.     

自噬对B细胞淋巴瘤细胞耐药性的影响

目的:探讨细胞自噬对不同种人源淋巴瘤细胞耐药性的影响.方法:以人Burkitt's淋巴瘤细胞Daudi、人B淋巴瘤细胞SUDHL-4和人套细胞淋巴瘤细胞JeKo-1为研究对象,通过自噬抑制剂处理细胞和慢病毒稳定感染干扰Atg5基因的表达,改变B细胞淋巴瘤细胞的自噬活性,并观察淋巴瘤细胞对ADR和VCR的耐药性的变化.结...     

自制与进口肺炎衣原体单克隆抗体的特性比较

目的:自制肺炎衣原体单克隆抗体(Cpn-McAb)与进口Cpn-McAb的特性比较及临床应用。方法:双向琼脂扩散法鉴定自制Cpn-McAb的效价和IgG亚型,用显色法检测单抗与Cpn-MOMP的免疫反应,然后用异硫氰酸荧光素(FITC)标记纯化的IgG。为了评估Cpn-McAb,用自制和进口Cpn-McAb同时检测外周血标本454例。另外,分别从Cpn特异性抗原(Cpn-Ag)阳性的外周血单核细胞(PBMC)标本(A组)中和Cpn-Ag阴性的对照组(B组)中随机各挑选40例,用PCR检测PBMC中的Cpn-DNA。结果:免疫琼脂扩散结果表明,Cpn单抗为IgG2b亚型,扩散效价为1:128。在1:30工作浓度下,微量免疫荧光检测的结果表明,自制Cpn-McAb与鹦鹉热衣原体有较弱的交叉反应;与沙眼衣原体无交叉反应,特性与进口Cpn-McAb一致。自制Cpn-McAb检测PBMC的Cpn-Ag阳性率为53.3%(242/454),进口Cpn-McAb检测PBMC的阳性率为52.6%(239/454),两种单抗检测同时阳性的标本207例,Kappa值是0.704,根据其判断标准提示两者有较高的一致性。A、B两组Cpn-DNA阳性标本分别为38例和2例。结论:自制Cpn-McAb几乎和进口Cpn-McAb一样有较高的特异性和敏感性,有可能替代进口单抗用于临床Cpn感染的检测,有助于相关疾病的诊治。