Trastuzumab Resistance Breast Cancer Xenograft

Breast cancer cell line BT474 cells were administered subcutaneously into NCr nude mice. After randomization, mice received vehicle or trastuzumab (15mg/kg twice weekly). Tumors were categorized as control, responders, intrinsic resistance, and acquired resistance. RNA isolation was performed using RNeasy Plus kit (Qiagen) using manufacturer’s instructions. Sequencing libraries were created using Truseq stranded mRNA library preparation method from Illumina and sequenced on an Illumina NextSeq2000 sequencer. Reads were aligned to reference genome GRCh38 using HISAT2, then SAMTools was used sort and merge the alignments from multiple flow cells.

将BT474乳腺癌细胞系细胞皮下接种于NCr裸小鼠体内。经随机分组后，小鼠分别接受溶剂对照或曲妥珠单抗（trastuzumab）给药，剂量为15mg/kg，每周两次。根据肿瘤应答情况将其分为对照组、应答组、原发性耐药组与获得性耐药组。采用Qiagen公司的RNeasy Plus试剂盒（RNeasy Plus kit），严格按照制造商说明书完成RNA分离提取。使用Illumina公司的Truseq链特异性mRNA建库方法（Truseq stranded mRNA library preparation method）构建测序文库，并在Illumina NextSeq2000测序仪（Illumina NextSeq2000 sequencer）上完成测序。利用HISAT2工具将测序读段比对至GRCh38参考基因组，随后通过SAMTools对多个流动池（flow cells）产生的比对结果进行排序与合并。