Beck W T, Grogan T M, Willman C L, Cordon-Cardo C, Parham D M, Kuttesch J F, Andreeff M, Bates S E, Berard C W, Boyett J M, Brophy N A, Broxterman H J, Chan H S, Dalton W S, Dietel M, Fojo A T, Gascoyne R D, Head D, Houghton P J, Srivastava D K, Lehnert M, Leith C P, Paietta E, Pavelic Z P, Weinstein R
St Jude Children's Research Hospital, Memphis, Tennessee 38105, USA.
Cancer Res. 1996 Jul 1;56(13):3010-20.
Multidrug resistance (MDR), especially that associated with overexpression of MDR1 and its product, P-glycoprotein (Pgp), is thought to play a role in the outcome of therapy for some human tumors; however, a consensus conclusion has been difficult to reach, owing to the variable results published by different laboratories. Many factors appear to influence the detection of Pgp in clinical specimens, including its low and heterogeneous expression; conflicting definitions of detection end points; differences in methods of sample preparation, fixation, and analysis; use of immunological reagents with variable Pgp specificity and avidity and with different recognition epitopes; use of secondary reagents and chromogens; and differences in clinical end points. Also, mechanisms other than Pgp overexpression may contribute to clinical MDR. The combined effect of these factors is clearly important, especially among tumors with low expression of Pgp. Thus, a workshop was organized in Memphis, Tennessee, to promote the standardization of approaches to MDR1 and Pgp detection in clinical specimens. The 15 North American and European institutions that agreed to participate conducted three preworkshop trials with well-characterized MDR myeloma and carcinoma cell lines that expressed increasing amounts of Pgp. The intent was to establish standard materials and methods for a fourth trial, assays of Pgp and MDR1 in clinical specimens. The general conclusions emerging from these efforts led to a number of recommendations for future studies: (a) although detection of Pgp and MDR1 is at present likely to be more reliable in leukemias and lymphomas than in solid tumors, accurate measurement of low levels of Pgp expression under most conditions remains an elusive goal; (b) tissue-specific controls, antibody controls, and standardized MDR cell lines are essential for calibrating any detection method and for subsequent analyses of clinical samples; (c) use of two or more vendor-standardized anti-Pgp antibody reagents that recognize different epitopes improves the reliability of immunological detection of Pgp; (d) sample fixation and antigen preservation must be carefully controlled; (e) multiparameter analysis is useful in clinical assays of MDR1/Pgp expression; (f) immunostaining data are best reported as staining intensity and the percentage of positive cells; and (g) arbitrary minimal cutoff points for analysis compromise the reliability of conclusions. The recommendations made by workshop participants should enhance the quality of research on the role of Pgp in clinical MDR development and provide a paradigm for investigations of other drug resistance-associated proteins.
多药耐药（MDR），尤其是与MDR1及其产物P-糖蛋白（Pgp）过表达相关的多药耐药，被认为在某些人类肿瘤的治疗结果中起作用；然而，由于不同实验室发表的结果存在差异，很难达成共识性结论。许多因素似乎会影响临床标本中Pgp的检测，包括其低表达和异质性表达；检测终点的定义相互矛盾；样品制备、固定和分析方法的差异；使用具有不同Pgp特异性和亲和力以及不同识别表位的免疫试剂；使用二抗试剂和显色剂；以及临床终点的差异。此外，除了Pgp过表达之外的其他机制可能也会导致临床多药耐药。这些因素的综合作用显然很重要，尤其是在Pgp低表达的肿瘤中。因此，在田纳西州孟菲斯组织了一次研讨会，以促进临床标本中MDR1和Pgp检测方法的标准化。同意参与的15家北美和欧洲机构对表达越来越多Pgp的特征明确的MDR骨髓瘤和癌细胞系进行了三项研讨会前试验。目的是为第四项试验（临床标本中Pgp和MDR1的检测）建立标准材料和方法。这些努力得出的总体结论产生了一些对未来研究的建议：（a）尽管目前在白血病和淋巴瘤中检测Pgp和MDR1可能比在实体瘤中更可靠，但在大多数情况下准确测量低水平的Pgp表达仍然是一个难以实现的目标；（b）组织特异性对照、抗体对照和标准化的MDR细胞系对于校准任何检测方法以及随后对临床样本的分析至关重要；（c）使用两种或更多种识别不同表位的供应商标准化抗Pgp抗体试剂可提高Pgp免疫检测的可靠性；（d）必须仔细控制样品固定和抗原保存；（e）多参数分析在MDR1/Pgp表达的临床检测中很有用；（f）免疫染色数据最好报告为染色强度和阳性细胞百分比；（g）任意设定的分析最小临界值会损害结论的可靠性。研讨会参与者提出的建议应提高关于Pgp在临床多药耐药发展中作用的研究质量，并为其他与耐药相关蛋白的研究提供范例。
