1 Wu, Y. M., F. Su, S. Kalyana-Sundaram, N. Khazanov, B. Ateeq, X. Cao, R. J. Lonigro, P. Vats, R. Wang, S. F. Lin, A. J. Cheng, L. P. Kunju, J. Siddiqui, S. A. Tomlins, P. Wyngaard, S. Sadis, S. Roychowdhury, M. H. Hussain, F. Y. Feng, M. M. Zalupski, M. Talpaz, K. J. Pienta, D. R. Rhodes, D. R. Robinson, and A. M. Chinnaiyan. Identification of targetable FGFR gene fusions in diverse cancers. (2013). Cancer discovery. 3: 636-47.
2 Arai, Y., Y. Totoki, F. Hosoda, T. Shirota, N. Hama, H. Nakamura, H. Ojima, K. Furuta, K. Shimada, T. Okusaka, A. Kosuge, and T. Shibata. FGFR2 tyrosine kinase fusions define a unique molecular subtype of cholangiocarcinoma. (2014). Hepatology. 59, 1427-34.
3 Ross, J. S., K. Wang, L. Gay, R. Al-Rohil, J. V. Rand, D. M. Jones, H. J. Lee, C. E. Sheehan, G. A. Otto, G. Palmer, R. Yelensky, D. Lipson, D. Morosini, M. Hawryluk, D. V. Catenacci, V. A. Miller, C. Churi, S. Ali, and P. J. Stephens. New routes to targeted therapy of intrahepatic cholangiocarcinomas revealed by next-generation sequencing. (2014). Oncologist. 19: 235-42.
4 Borad, M. J., M. D. Champion, J. B. Egan, W. S. Liang, R. Fonseca, A. H. Bryce, A. E. McCullough, M. T. Barrett, K. Hunt, M. D. Patel, S. W. Young, J. M. Collins, A. C. Silva, R. M. Condjella, M. Block, R. R. McWilliams, K. N. Lazaridis, E. W. Klee, K. C. Bible, P. Harris, G. R. Oliver, J. D. Bhavsar, A. A. Nair, S. Middha, Y. Asmann, J. P. Kocher, K. Schahl, B. R. Kipp, E. G. Barr Fritcher, A. Baker, J. Aldrich, A. Kurdoglu, T. Izatt, A. Christoforides, I. Cherni, S. Nasser, R. Reiman, L. Phillips, J. McDonald, J. Adkins, S. D. Mastrian, P. Placek, A. T. Watanabe, J. Lobello, H. Han, D. Von Hoff, D. W. Craig, A. K. Stewart, and J. D. Carpten. (2014) (Mayo Clinic Cancer Center, Arizon and Mayo Clinic, Minnesota) Integrated genomic characterization reveals novel, therapeutically relevant drug targets in FGFR and EGFR pathways in sporadic intrahepatic cholangiocarcinoma. . PLoS Genet. 10: e1004135.
5 Graham, R. P., E. G. Barr Fritcher, E. Pestova, J. Schulz, L. A. Sitailo, G. Vasmatzis, S. J. Murphy, R. R. McWilliams, S. N. Hart, K. C. Halling, L. R. Roberts, G. J. Gores, F. J. Couch, L. Zhang, M. J. Borad, and B. R. Kipp. Fibroblast growth factor receptor 2 translocations in intrahepatic cholangiocarcinoma. (2014). Human pathology. 45: 1630-8.
6 Sia D, Losic B, Moeini A, Cabellos L, Hao K, et al. (2015) Massive parallel sequencing uncovers actionable FGFR2-PPHLN1 fusion and ARAF mutations in intrahepatic cholangiocarcinoma. Nat Commun 6: 6087.
7 Shaw AT, Hsu, PP, Awad, MM & Engelman, JA. Tyrosine kinase gene rearrangements in epithelial malignancies. Nat Rev Cancer 2013; 13, 772-787.
8 Zheng Z, Liebers M, Zhelyazkova B, Cao Y, Panditi D, et al. (2014) Anchored multiplex PCR for targeted next-generation sequencing. Nat Med 20: 1479-1484.
9. Yoshihara K, Wang Q, Torres-Garcia W, Zheng S, Vegesna R, et al. (2014) The landscape and therapeutic relevance of cancer-associated transcript fusions. Oncogene 10.1038/onc.2014.406.
10 Klijn C, Durinck S, Stawiski EW, Haverty PM, Jiang Z, et al. (2015) A comprehensive transcriptional portrait of human cancer cell lines. Nat Biotechnol 33: 306-312.
#1由 sufang 在 日, 05/31/2015 – 10:54 發表。
2015/05
Nat Rev Cancer. 2015 May 22;15(6):371-81. doi: 10.1038/nrc3947.
The emerging complexity of gene fusions in cancer. (PubMed Link)
Mertens F1, Johansson B1, Fioretos T1, Mitelman F1.
Author information
1Department of Clinical Genetics, Lund University and Skåne University Hospital, SE-221 85 Lund, Sweden.
Unbiased gene fusion detection
In contrast to the guided approaches for gene fusion detection described above, the introduction of deep-sequencing technologies (also known as massively parallel sequencing and next-generation sequencing) some 10 years ago provided a radically new means to identify fusions either at the DNA or RNA levels in a single experiment (Box 1). By simultaneously providing detailed (nucleotide-level) and comprehensive (genome-wide) information on the genome or transcriptome, structural variants and fusion transcripts could now be identified without any prior information on the cytogenetic features of the neoplastic cells. The first studies using deep sequencing to detect neoplasia-associated gene fusions were carried out on established cell lines45, and these were quickly followed by numerous investigations of primary samples from common cancer types, such as carcinomas of the breast, colon, lung, prostate and uterus46, 47, 48, 49, 50, 52, as well as leukaemias and lymphomas52, 53, 54. Recently, a further leap was taken when Yoshihara and co-workers queried transcriptome data from 4,366 neoplasms, from 13 different tumour types, that had been studied within the Cancer Genome Atlas (TCGA) network; using stringent bioinformatic criteria, more than 8,600 different fusion transcripts were detected55. These results have dramatically changed the gene fusion landscape; a plethora of gene fusions (more than 9,000), the great majority involving previously unsuspected genes, has now been identified through deep sequencing5 (Fig. 4;Table 1).
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#2由 sufang 在 六, 01/10/2015 – 20:07 發表。
Fusion transcript discovery in formalin-fixed paraffin-embedded
PLoS One. 2014 Apr 11;9(4):e94202. doi: 10.1371/journal.pone.0094202. eCollection 2014.
Fusion transcript discovery in formalin-fixed paraffin-embedded human breast cancer tissues reveals a link to tumor progression.
PMID: 24727804
pdf: 4208
Ma Y1, Ambannavar R1, Stephans J1, Jeong J1, Dei Rossi A1, Liu ML1, Friedman AJ1, Londry JJ1, Abramson R1, Beasley EM1, Baker J1, Levy S1, Qu K1.
Author information
Abstract
The identification of gene fusions promises to play an important role in personalized cancer treatment decisions. Many rare gene fusion events have been identified in fresh frozen solid tumors from common cancers employing next-generation sequencing technology. However the ability to detect transcripts from gene fusions in RNA isolated from formalin-fixed paraffin-embedded (FFPE) tumor tissues, which exist in very large sample repositories for which disease outcome is known, is still limited due to the low complexity of FFPE libraries and the lack of appropriate bioinformatics methods. We sought to develop a bioinformatics method, named gFuse, to detect fusion transcripts in FFPE tumor tissues. An integrated, cohort based strategy has been used in gFuse to examine single-end 50 base pair (bp) reads generated from FFPE RNA-Sequencing (RNA-Seq) datasets employing two breast cancer cohorts of 136 and 76 patients. In total, 118 fusion events were detected transcriptome-wide at base-pair resolution across the 212 samples. We selected 77 candidate fusions based on their biological relevance to cancer and supported 61% of these using TaqMan assays. Direct sequencing of 19 of the fusion sequences identified by TaqMan confirmed them. Three unique fused gene pairs were recurrent across the 212 patients with 6, 3, 2 individuals harboring these fusions respectively. We show here that a high frequency of fusion transcripts detected at the whole transcriptome level correlates with poor outcome (P<0.0005) in human breast cancer patients. This study demonstrates the ability to detect fusion transcripts as biomarkers from archival FFPE tissues, and the potential prognostic value of the fusion transcripts detected.
Briefly, total RNA was isolated from three 10-um FFPE tissue sections per patient using Epicentre’s MasterPure Purification Kit (Epicenter Biotechnologies, Madison, WI). Paraf- fin was first removed by xylene extraction followed by ethanol wash.
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#3由 sufang 在 五, 12/05/2014 – 10:28 發表。
所謂的Biliary Tract Cancer (BTC)
IHCC
EHCC
Gallbladder
Ampulla of vater
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#4由 sufang 在 四, 12/04/2014 – 12:46 發表。
Therapeutics: Targeting an oncometabolite
McCarthy, N. Therapeutics: Targeting an oncometabolite. (2013). Nat Rev Cancer. IDH1 and IDH2 related. AGI-5198 Therapeutics: Targeting an oncometabolite
Nicola McCarthy
TherapeuticsTargeting an oncometabolite
NPG
Specific mutations in isocitrate dehydrogenase 1 (IDH1) and IDH2 result in the generation of a particular metabolite — the (R)-enantiomer of 2-hydroxyglutarate ((R)-2HG). The production of high concentrations of (R)-2HG is associated with biological outcomes that promote tumour progression. Thus, there has been considerable interest in developing drugs that target mutant IDH1 and IDH2.
Janeta Popovici-Muller and colleagues have developed a selective inhibitor (AGI-5198) of R132H-IDH1, which is the most common mutant of IDH1 in glioma. Along with colleagues from the laboratories of Katharine Yen, Thomas Graeber and Ingo Mellinghoff, they tested the activity of this drug in a patient-derived glioma cell line that has an endogenous R132H-IDH1 mutation. AGI-5198 inhibited, in a dose-dependent manner, the production of (R)-2HG and also limited colony formation in soft agar by up to 60%. By contrast, it had no effect on two patient-derived glioma cell lines with wild-type IDH1. In mice, orally administered AGI-5198 limited the growth of human R132H-IDH1 glioma xenografts by 50–60%. Proliferation rates were reduced in these xenografts, but apoptosis rates were unaffected. Gene expression studies on mRNA isolated from treated xenografts indicated that the transcription of genes involved in the differentiation of oligodendrocytes and astrocytes was increased. This and other analyses indicated that R132H-IDH1 restricts glial differentiation and that this is restored when R132H-IDH1 is inhibited.
orally administered AGI-5198 limited the growth of human R132H-IDH1glioma xenografts by 50–60%
Mutant IDH1 has been associated with specific epigenetic changes, such as DNA and histone methylation, owing to the inhibitory effects of (R)-2HG on α-ketoglutarate (αKG)-dependent dioxygenases, including TET methyl cytosine hydroxylases and Jumonji-C domain histone demethylases. The authors used two different concentrations of AGI-5198 that either partially reduced or almost fully reduced the concentration of (R)-2HG to physiological levels. Inhibition of xenograft growth occurred as a result of partial (R)-2HG blockade, and further experiments showed that this was uncoupled from effects on histone and DNA methylation. Thus, the authors conclude that the roles of additional αKG-dependent dioxygenases in the maintenance of IDH1-mutant glioma need to be better understood.
Jeremy Travins and colleagues have also developed a drug, AGI-6780, that targets R140Q-IDH2, a mutation that is present in approximately 9% of patients with acute myeloid leukaemia (AML). Expression of R140Q-IDH2 in the human erythroleukaemia cell line, TF-1, which is dependent on granulocyte–macrophage colony-stimulating factor (GM-CSF) for survival and proliferation, resulted in the production of high concentrations of (R)-2HG, GM-CSF-independent growth and the attainment of a stem cell and/or progenitor cell phenotype. Moreover, erythropoietin failed to induce the differentiation of TF-1 cells that expressed R140Q-IDH2, but this was restored in the presence of AGI-6780 at concentrations that reduced the expression of (R)-2HG to near physiological concentrations. Additional ex vivo experiments that used patient-derived AML samples showed that AGI-6780 induces the differentiation of leukaemic blasts specifically in patient samples that have the R140Q-IDH2 mutation. Whether this translates to clinical efficacy has not yet been determined.
Both papers indicate that (R)-2HG is associated with a block in differentiation in glioma and AML cells, as several studies have previously indicated, and that this can be reversed by specific inhibitors that reduce the production of (R)-2HG by mutant IDH.
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References and links
ORIGINAL RESEARCH PAPERS
Rohle, D. et al. An inhibitor of mutant IDH1 delays growth and promotes differentiation of glioma cells. Science 4 Apr 2013 (doi: 10.1126/science.1236062)
Article
Wang, F. et al. Targeted inhibition of mutant IDH2 in leukemia cells induces cellular differentiation. Science 4 Apr 2013 (doi: 10.1126/science.1234769)
Article.
4 comments on PubPeer (by: Unregistered Submission, Peer 2, Potentilla Turfosa, Enoplognatha Caricis)
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______________
#5由 sufang 在 三, 11/12/2014 – 13:38 發表。
2014/07/03 其它文獻
¶ Gu TL, Deng X, Huang F, Tucker M, Crosby K, Rimkunas V et al. Survey of tyrosine kinase signaling reveals ROS kinase fusions in human cholangiocarcinoma. PLoS One 2011; 6: e15640. (PubMed Link)
¶ Ong CK, Subimerb C, Pairojkul C, Wongkham S, Cutcutache I, Yu W et al. Exome sequencing of liver fluke-associated cholangiocarcinoma. Nat Genet 2012; 44: 690-693. (PubMed Link)
¶ Davare MA, Saborowski A, Eide CA, Tognon C, Smith RL, Elferich J et al. Foretinib is a potent inhibitor of oncogenic ROS1 fusion proteins. Proc Natl Acad Sci USA 2013; 110: 19519-19524. (PubMed Link)
¶ Rizvi S, Gores GJ. Pathogenesis, diagnosis, and management of cholangiocarcinoma. Gastroenterology 2013; 145: 1215-1229. (PubMed Link) (IDH1, IDH2 mutations in Figure 2)
¶ Saborowski A, Saborowski M, Davare MA, Druker BJ, Klimstra DS, Lowe SW. Mouse model of intrahepatic cholangiocarcinoma validates FIG-ROS as a potent fusion oncogene and therapeutic target. Proc Natl Acad Sci USA 2013; 110: 19513-19518. (PubMed Link)
¶ Sia D, Tovar V, Moeini A, Llovet JM. Intrahepatic cholangiocarcinoma: pathogenesis and rationale for molecular therapies. Oncogene 2013; 32: 4861-4870. (PubMed Link)
¶ Subbiah IM, Subbiah V, Tsimberidou AM, Naing A, Kaseb AO, Javle M et al. Targeted Therapy of Advanced Gallbladder Cancer and Cholangiocarcinoma with Aggressive Biology: Eliciting Early Response Signals from Phase 1 trials. Oncotarget 2013; 4: 153-162. (PubMed Link)
¶ Jiao Y, Pawlik TM, Anders RA, Selaru FM, Streppel MM, et al. (2013) Exome sequencing identifies frequent inactivating mutations in BAP1, ARID1A and PBRM1 in intrahepatic cholangiocarcinomas. Nat Genet 45: 1470-1473. (PubMed Link)
¶ Grassian AR, Pagliarini R, Chiang DY. Mutations of isocitrate dehydrogenase 1 and 2 in intrahepatic cholangiocarcinoma. Curr Opin Gastroenterol 2014. (PubMed Link)
¶ Razumilava N, Gores GJ (2014) Cholangiocarcinoma. Lancet 10.1016/s0140-6736(13)61903-0. (PubMed Link)
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#6由 sufang 在 五, 10/17/2014 – 09:41 發表。
Fusion transcript discovery in FFPE
Su-Fang Lin <sflin1@gmail.com> Oct 13 to to 黃道揚, 陳所長, FGFR3 Gene Fusions in Taiwan Oral Cancer
Dear Daw-Yang,
早上看到一篇 kinase gene fusion的文章，又燃起我想screen一下台灣口腔癌檢體中究竟FGFR3-TACC3 gene fusion的比例有多高的實驗!
Nat Commun 2014-Sep The landscape of kinase fusions in cancer
不知道以前有沒有跟你提過，當初請Dan/Yi-Mi 幫我們做20例breast cancer RNA-seq的同時，我們也送四株台灣oral cancer cell line的RNA請他們一併分析口腔癌細胞中gene fusion情形。結果其中一株細胞發現有high read的FGFR3-TACC3 fusion. 之後我們以PCR或是RT-PCR檢查成大醫院那邊約65例的臨床檢體，但是很可惜PCR不是contaminated with positive control, 就是all negative…後來因學生畢業了，就停着沒有再往下做。
我綜合Dan/Yi-Mi他們2013年mining TCGA結果(284例HNSC)與現在這篇Nat Commun mining 7,000例 (411例HNSC) 的結果，FGFR3-TACC3在TCGA中HNSC cohort positive rate約僅0.5-1%，很低。但是一想到當初我送四株細胞中，就有一株是FR3-TA3 positive，又覺得應該要好好看ㄧ下多一點的檢體、說不定嚼檳榔的口腔癌組織中，該類fusion會比白人之HNSC多。
想請教你的是，倘使用你的方法，但只focus在FGFR3的gene fusion，FFPE抽出來的RNA有沒有辦法做?
我原本的策略是先以FGFR3之IHC screen臨床切片，若是有強的染色反應，我們再做RT-PCR confirm. 可是如果FFPE也可行，那可以用的檢體數量應該可以多很多! 也不必浪費寶貴的frozen tissues.
另附上一篇 tyrosine kinase gene rearrangement 的review (好像上次你還在美國時給過一次…sorry for my 囉唆)
Looking forward to our first collaboration!!
Su-Fang
Daw-yang Hwang 黃道揚 Oct 13 to me, 陳所長
Dear Su-Fang,
FFPE samples can be analyzed, depends on the RNA quality you can get. If you are going to look at the FGFR3-TACC3 only, it will be even easier (without using NGS) than FGFR3-unknown gene experiment. Just give me a call at any time!! or I can call you when you are free.
Best,
Daw-yang
Su-Fang Lin <sflin1@gmail.com> Oct 13 to Daw-yang Hwang., 陳所長
Thank you very much, Daw-yang, I will contact the doctors first, see what kind of materials/quality I can get. Will come back to you soon.
-Su-Fang
(ps. yes, I only care about FGFR3-TACC3, yet in contrast to FGFR3 whose breakpoint is quite conserved, multiple breakpoints for TACC3 were found, I will give you a map before we start)
Daw-yang Hwang 黃道揚 Oct 13 to me
I tried FFPE sample kits from Epicentre (easy protocol but OD is low), Promega, and Abnova (P and A have similar OD ratio). I wonder what brand you use before?
Sincerely,
Daw-yang
PLoS One. 2014 Apr 11;9(4):e94202. doi: 10.1371/journal.pone.0094202. eCollection 2014.
Fusion transcript discovery in formalin-fixed paraffin-embedded human breast cancer tissues reveals a link to tumor progression.
Ma Y, Ambannavar R, Stephans J, Jeong J, Dei Rossi A, Liu ML, Friedman AJ, Londry JJ, Abramson R, Beasley EM, Baker J, Levy S, Qu K.
Author information
Abstract
The identification of gene fusions promises to play an important role in personalized cancer treatment decisions. Many rare gene fusion events have been identified in fresh frozen solid tumors from common cancers employing next-generation sequencing technology. However the ability to detect transcripts from gene fusions in RNA isolated from formalin-fixed paraffin-embedded (FFPE) tumor tissues, which exist in very large sample repositories for which disease outcome is known, is still limited due to the low complexity of FFPE libraries and the lack of appropriate bioinformatics methods. We sought to develop a bioinformatics method, named gFuse, to detect fusion transcripts in FFPE tumor tissues. An integrated, cohort based strategy has been used in gFuse to examine single-end 50 base pair (bp) reads generated from FFPE RNA-Sequencing (RNA-Seq) datasets employing two breast cancer cohorts of 136 and 76 patients. In total, 118 fusion events were detected transcriptome-wide at base-pair resolution across the 212 samples. We selected 77 candidate fusions based on their biological relevance to cancer and supported 61% of these using TaqMan assays. Direct sequencing of 19 of the fusion sequences identified by TaqMan confirmed them. Three unique fused gene pairs were recurrent across the 212 patients with 6, 3, 2 individuals harboring these fusions respectively. We show here that a high frequency of fusion transcripts detected at the whole transcriptome level correlates with poor outcome (P<0.0005) in human breast cancer patients. This study demonstrates the ability to detect fusion transcripts as biomarkers from archival FFPE tissues, and the potential prognostic value of the fusion transcripts detected. PMID: 24727804