Molecular alterations in prostate carcinomas that associate with in vivo exposure to chemotherapy. Homo sapiens

Patients with high-risk localized prostate cancer (tumor-node-metastasis ≥ T2b or prostate-specific antigen ≥ 15 ng/mL or Gleason glade ≥ 4+3) were enrolled into a phase II clinical trial of neoadjuvant chemotherapy with docetaxel and mitoxantrone followed by prostatectomy. Pretreatment prostate tissue was acquired by needle biopsy and posttreatment tissue was acquired by prostatectomy. Prostate epithelium was captured by microdissection, and transcript levels were quantitated by cDNA microarray hybridization. Overall design: To evaluate gene expression alterations after chemotherapy, neoplastic epithelium from pretreated biopsy and posttreated prostatectomy specimens were laser captured separately (∼3,000 cells per sample). cDNA synthesized from amplified total RNA of pre-treated biopsy samples and post-treatment surgical samples were hybridized head-to-head, alternating Cy3 and Cy5 to custom-made microarrays composed of 6,200 clones derived from the Prostate Expression Database.

纳入本研究的受试者为高危局限性前列腺癌患者，满足以下任一入组标准：肿瘤-淋巴结-转移（tumor-node-metastasis, TNM）分期≥T2b、前列腺特异性抗原（prostate-specific antigen, PSA）≥15 ng/mL，或格里森评分（Gleason grade）≥4+3。所有患者均接受多西他赛联合米托蒽醌的新辅助化疗II期临床试验，随后行前列腺切除术。
治疗前前列腺组织通过穿刺活检获取，治疗后前列腺组织则通过前列腺切除术获取。通过显微切割捕获前列腺上皮细胞，并采用cDNA微阵列杂交技术对转录本水平进行定量检测。
实验设计概述：为评估化疗后的基因表达变化，分别激光捕获预处理活检标本与术后前列腺切除标本中的肿瘤上皮细胞（每份样本约3000个细胞）。将预处理活检样本与术后手术样本的扩增总RNA反转录得到的cDNA进行头对头杂交，通过交替使用Cy3与Cy5荧光标记，与由前列腺表达数据库来源的6200个克隆构建的定制微阵列进行杂交。