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Jiang W , Dulaimi E , Devarajan K , Parsons T , Wang Q , Liao L , Cho EA , O'Neill R , Solomides C , Peiper SC , Testa JR , Uzzo R , Yang H
Immunohistochemistry Successfully Uncovers Intratumoral Heterogeneity and Widespread Co-Losses of Chromatin Regulators in Clear Cell Renal Cell Carcinoma
PLoS One. 2016 Oct;11(10) :e0164554
PMID: 27764136    PMCID: PMC5072613    URL: https://www.ncbi.nlm.nih.gov/pubmed/27764136
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Recent studies have shown that intratumoral heterogeneity (ITH) is prevalent in clear cell renal cell carcinoma (ccRCC), based on DNA sequencing and chromosome aberration analysis of multiple regions from the same tumor. VHL mutations were found to be universal throughout individual tumors when it occurred (ubiquitous), while the mutations in other tumor suppressor genes tended to be detected only in parts of the tumors (subclonal). ITH has been studied mostly by DNA sequencing in limited numbers of samples, either by whole genome sequencing or by targeted sequencing. It is not known whether immunohistochemistry (IHC) can be used as a tool to study ITH. To address this question, we examined the protein expression of PBRM1, and PBRM1-related proteins such as ARID1A, SETD2, BRG1, and BRM. Altogether, 160 ccRCC (40 per stage) were used to generate a tissue microarray (TMA), with four foci from each tumor included. Loss of expression was defined as 0-5% of tumor cells with positive nuclear staining in an individual focus. We found that 49/160 (31%), 81/160 (51%), 23/160 (14%), 24/160 (15%), and 61/160 (38%) of ccRCC showed loss of expression of PBRM1, ARID1A, SETD2, BRG1, and BRM, respectively, and that IHC could successfully detect a high prevalence of ITH. Phylogenetic trees were constructed that reflected the ITH. Striking co-losses among proteins were also observed. For instance, ARID1A loss almost always accompanied PBRM1 loss, whereas BRM loss accompanied loss of BRG1, PBRM1 or ARID1A. SETD2 loss frequently occurred with loss of one or more of the other four proteins. Finally, in order to learn the impact of combined losses, we compared the tumor growth after cells acquired losses of ARID1A, PBRM1, or both in a xenograft model. The results suggest that ARID1A loss has a greater tumor-promoting effect than PBRM1 loss, indicating that xenograft analysis is a useful tool to investigate how these losses impact on tumor behavior, either alone or in combination.
1932-6203 Jiang, Wei Dulaimi, Essel Devarajan, Karthik Parsons, Theodore Wang, Qiong Liao, Lili Cho, Eun-Ah O'Neill, Raymond Solomides, Charalambos Peiper, Stephen C Testa, Joseph R Uzzo, Robert Yang, Haifeng Journal article United States PLoS One. 2016 Oct 20;11(10):e0164554. doi: 10.1371/journal.pone.0164554. eCollection 2016.