Srivastava Alok, Fuchs Bruno, Zhang Kunbo, Ruan Ming, Halder Chandralekha, Mahlum Eric, Weber Kristin, Bolander Mark E, Sarkar Gobinda
Department of Orthopedics, Mayo Clinic and Foundation, Rochester, Minnesota 55905, USA.
Clin Cancer Res. 2006 Jul 15;12(14 Pt 1):4237-43. doi: 10.1158/1078-0432.CCR-05-2307.
Although metastasis is the primary determinant of poor survival of patients with osteogenic sarcoma, some patients live much longer than others, indicating metastatic heterogeneity underlying survival outcome. The purpose of the investigation was to identify genes underlying survival outcome of patients with osteogenic sarcoma metastasis.
We have used microarray to first compare mRNA expression between normal bone and osteogenic sarcoma specimens, identified genes overexpressed in osteogenic sarcoma, and compared expression of the selected gene between a poorly metastatic (SAOS) and two highly metastatic cell lines (LM8 and 143B). Finally, expression of the selected gene was assessed by immunostaining of osteogenic sarcoma samples with known survival outcome.
Microarray analysis revealed 5.3-fold more expression of WT1 mRNA in osteogenic sarcoma compared with normal bone and >2-fold overexpression in 143B and LM8 cells compared with SAOS. Furthermore, WT1 mRNA was absent in normal bone (10 of 10) by reverse transcription-PCR but present in osteogenic sarcoma-derived cell lines (5 of 8). One hundred percent (42 of 42) of low-grade osteogenic sarcoma specimens expressed no WT1 as determined by immunostaining; however, 24% (12 of 49) of the high-grade specimens showed intense staining. Mean survival of patients with high-grade metastatic osteogenic sarcoma but low WT1 staining (27 of 37) was 96.5 +/- 129.3 months, whereas mean survival of patients with high-grade metastatic osteogenic sarcoma having intense staining (10 of 37) was 18.3 +/- 12.3 months (P > 0.0143). All splice variants of WT1 mRNA, including a hitherto unknown variant (lacking exons 4 and 5), were found to be expressed in osteogenic sarcoma.
WT1 seems to be associated with very poor survival of patients with osteogenic sarcoma metastasis.
尽管转移是骨肉瘤患者生存预后差的主要决定因素，但有些患者的生存期比其他患者长得多，这表明生存结果存在转移异质性。本研究的目的是确定骨肉瘤转移患者生存结果的潜在基因。
我们首先使用微阵列比较正常骨组织和骨肉瘤标本之间的mRNA表达，鉴定出在骨肉瘤中过表达的基因，并比较了低转移细胞系（SAOS）与两个高转移细胞系（LM8和143B）中所选基因的表达。最后，通过对已知生存结果的骨肉瘤样本进行免疫染色来评估所选基因的表达。
微阵列分析显示，与正常骨组织相比，骨肉瘤中WT1 mRNA的表达高5.3倍，与SAOS细胞系相比，143B和LM8细胞系中WT1 mRNA的表达高2倍以上。此外，通过逆转录PCR检测，正常骨组织（10例中有10例）中未检测到WT1 mRNA，但在骨肉瘤来源的细胞系中（8例中有5例）可检测到。免疫染色结果显示，100%（42例中有42例）的低级别骨肉瘤标本未表达WT1；然而，24%（49例中有12例）的高级别标本显示强染色。高级别转移性骨肉瘤且WT1染色低的患者（37例中有27例）的平均生存期为96.5±129.3个月，而高级别转移性骨肉瘤且染色强的患者（37例中有10例）的平均生存期为18.3±12.3个月（P>0.0143）。WT1 mRNA的所有剪接变体，包括一种迄今未知的变体（缺少外显子4和5），均在骨肉瘤中表达。
WT1似乎与骨肉瘤转移患者的极差生存预后相关。
