儿童急性B淋巴细胞白血病免疫球蛋白重链可变区基因的分子特征

目的 探讨儿童急性B淋巴细胞白血病(B—ALL)的起源及其免疫球蛋白重链可变区(IgHV)上可被细胞毒性T淋巴细胞(cTL)识别的表位(epitope)。方法 PCR扩增108例儿童ALL的7个IgHV基因家族；PCR产物直接测序并翻译成氨基酸序列；利用生物信息资源分析B—ALL IgHV基因重排类型、胚系基因片段利用及体细胞突变情况，并预测CTL细胞识别的表位。结果 66％的ALL患儿检测到IgHV基因重排，其中单等位基因重排37例(52．1％)，双等位基因重排26例(36.6％)，寡克隆基因重排8例(11．3％)。40份B-ALL IgHV基因序列中，不含终止密码子的读码框内(in frame)重排8份(20．0％)。重排利用最多的VH家族为VH3、VH4和VH1，占75%；VH4—59和VH4—34是VH4家族中利用频率最高的片段。优先利用的D家族为D3(35．9％)和D2(28.2％),D7—27的利用率(15．4％)明显高于正常外周血淋巴细胞(P=0．02)。JH6是B-ALL利用最多的JH基因片段(47．5％)，JH4(27．5％)次之。20％的B—ALL在DJH连接区缺乏N核苷酸的插入，高于正常外周血淋巴细胞(P=0．02)。17．5％B—ALL IgHV基因发生替代突变，突变碱基散布于IgHV全长，互补决定区替代突变与静寂突变的比例≤1。26例B—ALL IgHV的HIA—I类分子结合肽80％以上位于框架区，同一IgHV家族的B-ALL有相同的1～2条九肽。结论 B—ALL起源于前B祖细胞或前B细胞，恶性转化往往发生于胚胎期，它们的IgHV基因具有胚系特征；生物信息分析B-ALL IgHV的T细胞表位主要位于框架1区和框架3区。

Characteristics of immunoglobulin heavy chain variable region genes in childhood B-cell acute lymphoblastic leukemia

OBJECTIVE: To investigate the origin of childhood B-cell acute lymphoblastic leukemia (B-ALL) and its epitopes recognized by cytotoxic T lymphocytes (CTL) in immunoglobulin heavy chain variable region (IgHV). METHODS: Seven IgHV gene families were respectively amplified by PCR and directly sequenced in 108 childhood ALL. The amino acid sequences were deducted from sequenced nucleotides. Bioinformatics was applied to analyses of recombination patterns, somatic mutations and germline gene segments usage, and to prediction of epitopes recognized by CTL. RESULTS: IgHV gene rearrangements were identified in 66% of the cases, including 37 (52.1%) monoallelic rearrangements, 26 (36.6%) biallelic rearrangements and 8 (11.3%) oligoclonal rearrangements. Among the obtained 40 B-ALL IgHV gene sequences, 8 (20.0%) were in frame rearrangements without stop codons. V(H3) (11/40), V(H4) (11/40) and V(H1) (8/40) amounted to 75% rearranged V(H) families. V(H)(4-59) and V(H)(4-34) were the most frequently rearranged V(H)(4) family gene segments. Usage of D2 and D3 families was most prominent (35.9% and 28.2%, respectively). Increased frequency of D7-27 (15.4%) was found as compared to that of normal peripheral B lymphocytes (P = 0.02). J(H)(6) was found in 47.5% rearrangements followed by J(H)(4) (27.5%). 8/40 (20.0%) DJ(H) junctions lacked N nucleotides, which was higher than that reported for normal peripheral B lymphocytes (P = 0.02). 17.5% B-ALL IgHV contained scattered replacement mutations with replacement (R) to silent (S) substitution ratio (R/S ratio)