1. From the Division of Hematology, Brigham and Women’s Hospital, Harvard Medical School, Boston; and the Department of Internal Medicine III, University Hospital of Ulm, Ulm, Germany NEJM Aug 14, 2008 p.722-34 R4. 한재준 1

2. Contents 1. Introduction 2. Causes of chromosomal abnormalities 3. Chromosomal rearrangements Chimeric fusion genes Chimeric fusion genes Deregulation of expression of normal genes Deregulation of expression of normal genes 4. Chromosomal imbalances Genomic gains Genomic gains Genomic losses Genomic losses 5. Summary 2

3. Introduction Chromosomes are altered in most types of cancer cells. Chromosomes are altered in most types of cancer cells. 1. In 1950s Theodore Hauschka Theodore Hauschka Albert Levan Albert Levan Saijaro Makino Saijaro Makino Report that tumor cells have multiple chromosome aberrations. 2. To date, clonal chromosome aberrations have been found in all major tumor types. Their identification continues as a result of technical improvements in conventional and molecular cytogenetics. Their identification continues as a result of technical improvements in conventional and molecular cytogenetics. 3

4. Introduction 1. The WHO classification of tumors Uses them to define specific disease entities. Uses them to define specific disease entities. 2. Prognostic and predictive markers 3. Provide insights into the mechanisms of tumorigenesis. 4. In a few instances, led to treatment that targets a specific genetic abnormality. 4

5. Causes of Chromosomal Abnormalities What causes chromosomal abnormalities ? What causes chromosomal abnormalities ? environmental exposures environmental exposures occupational exposures occupational exposures therapy with cytotoxic drugs therapy with cytotoxic drugs Example) Myelodysplastic syndrome or AML Example) Myelodysplastic syndrome or AML after treatment with alkylating agents: deletion or loss of chromosome 5 or 7 (or both) after treatment with alkylating agents: deletion or loss of chromosome 5 or 7 (or both) after treatment with topoisomerase II inhibitors: translocations involving the MLL gene on chromosome band 11q23 after treatment with topoisomerase II inhibitors: translocations involving the MLL gene on chromosome band 11q23 5

6. Causes of Chromosomal Abnormalities Molecular mechanisms underlying the abnormalities Molecular mechanisms underlying the abnormalities Example) Cases of the MDS or AML arising in patients with Fanconi’s anemia Example) Cases of the MDS or AML arising in patients with Fanconi’s anemia Isolated focal gains or cryptic rearrangements of chromosome band 3q26 that cause overexpression of the EVI1 gene. Isolated focal gains or cryptic rearrangements of chromosome band 3q26 that cause overexpression of the EVI1 gene. Complex unbalanced chromosomal abnormalities result from inactivation of the Fanconi’s anemia pathway that regulates the recognition and repair of damaged DNA. Complex unbalanced chromosomal abnormalities result from inactivation of the Fanconi’s anemia pathway that regulates the recognition and repair of damaged DNA. 6

7. Chromosomal Rearrangements reciprocal translocations inversions insertions Functionally 1. Result in the formation of a chimeric fusion gene with new or altered activity 2. Deregulated expression of a structurally normal gene 7

8. Chimeric fusion genes Tyrosine Kinase Genes Tyrosine Kinase Genes The Philadelphia chromosome The Philadelphia chromosome Sequences of the BCR gene on band 22q11.23 are joined to portions of the gene encoding the cytoplasmic ABL1 tyrosine kinase on band 9q34.1 Sequences of the BCR gene on band 22q11.23 are joined to portions of the gene encoding the cytoplasmic ABL1 tyrosine kinase on band 9q34.1 8

9. The Philadelphia chromosome 1. Evidence that human cancer can arise from acquired genetic alterations in somatic cells. 2. Use of selective tyrosine kinase inhibitor, imatinib mesylate, to treat disease. 3. Imatinib-resistant kinase domain mutations have been identified as a major cause of relapse during imatinib therapy, led to second generation BCR-ABL1 inhibitors, such as dasatinib and nilotinib. 9

10. Tyrosine kinase T-cell acute lymphoblastic leukemia: 5% T-cell acute lymphoblastic leukemia: 5% episome(9q34.1) episome(9q34.1) Imatinib-sensitive fusion of ABL1 to the NUP214 gene on band 9q34.1 Imatinib-sensitive fusion of ABL1 to the NUP214 gene on band 9q34.1 Non-small cell lung cancer: 6.7% Non-small cell lung cancer: 6.7% Inv(2)(p22-p21p23) Inv(2)(p22-p21p23) Portions of EML4 and the gene encoding the ALK receptor tyrosine kinase Portions of EML4 and the gene encoding the ALK receptor tyrosine kinase 10

11. Chimeric fusion genes Transcription Factor Genes Transcription Factor Genes enhanced transcriptional activity enhanced transcriptional activity Ewing’s sarcoma Ewing’s sarcoma t(11;22)(q24.1-q24.3;q12.2) t(11;22)(q24.1-q24.3;q12.2) t(21;22)(q22.3;q12.2) t(21;22)(q22.3;q12.2) Fuse the EWSR1 gene on band 22q12.2 to a gene encoding a member of the ETS family of transcription factors, most frequently FLI1 on band 11q24.1-q24.3) and ERG on band 21q22.3 Fuse the EWSR1 gene on band 22q12.2 to a gene encoding a member of the ETS family of transcription factors, most frequently FLI1 on band 11q24.1-q24.3) and ERG on band 21q22.3 11

12. Chimeric fusion genes Transcription Factor Genes Transcription Factor Genes aberrant transcriptional repression aberrant transcriptional repression Acute myeloid leukemia Acute myeloid leukemia PML-RARA PML-RARA RUNX1-RUNX1T1 RUNX1-RUNX1T1 CBFB-MYH11 CBFB-MYH11 Causes aberrant transcriptional repression, include genes required for normal myeloid differentiation, thereby contributing to the accumulation of immature myeloid cells in AML. Causes aberrant transcriptional repression, include genes required for normal myeloid differentiation, thereby contributing to the accumulation of immature myeloid cells in AML. 12

13. Deregulation of expression of normal genes Chromosomal rearrangements that juxtapose tissue-specific regulatory elements, such as gene promotors or enhancer sequences, to the coding sequence of a proto-oncogene deregulate expression of the proto- oncogene. Chromosomal rearrangements that juxtapose tissue-specific regulatory elements, such as gene promotors or enhancer sequences, to the coding sequence of a proto-oncogene deregulate expression of the proto- oncogene. 13

14. Deregulation of expression of normal genes Burkitt’s lymphoma Burkitt’s lymphoma Enhancer of an immunoglobulin gene IGHG1 drives the constitutive expression of the gene encoding the MYC transcription factor on band 8q24.21 Enhancer of an immunoglobulin gene IGHG1 drives the constitutive expression of the gene encoding the MYC transcription factor on band 8q24.21 14

15. Deregulation of expression of normal genes Prostate cancer Prostate cancer Fuse all coding regions of the ERG gene to androgen- regulated sequences in the promotor of the prostate- specific TMPRSS2 gene; these sequences mediate the aberrant expression of ERG in prostate tissue. Fuse all coding regions of the ERG gene to androgen- regulated sequences in the promotor of the prostate- specific TMPRSS2 gene; these sequences mediate the aberrant expression of ERG in prostate tissue. 15

16. Chromosomal Imbalances Alterations spanning entire chromosomes to intragenic duplications or deletions. Alterations spanning entire chromosomes to intragenic duplications or deletions. Most chromosomal imbalances have functional consequences that are unknown, because most imbalances affect large genomic regions containing multiple genes, and many tumors have numerous unbalanced chromosomal abnormalities. Most chromosomal imbalances have functional consequences that are unknown, because most imbalances affect large genomic regions containing multiple genes, and many tumors have numerous unbalanced chromosomal abnormalities. 16

17. Genomic gains Most recurrent genomic gains probably contribute to tumorigenesis by enhancing the activity of specific genes in the affected chromosomal regions. Most recurrent genomic gains probably contribute to tumorigenesis by enhancing the activity of specific genes in the affected chromosomal regions. Example) ~30% women with breast cancer Example) ~30% women with breast cancer Amplification of the gene on band 17q21.1 that encodes the ERBB2 receptor tyrosine kinase. Amplification of the gene on band 17q21.1 that encodes the ERBB2 receptor tyrosine kinase. The resulting overexpression of ERBB2 represents a target for the monoclonal antibody trastuzumab. The resulting overexpression of ERBB2 represents a target for the monoclonal antibody trastuzumab. 17

18. Genomic gains Large-scale Genomic Gains Large-scale Genomic Gains Commonly arise from chromosomal nondisjunction or unbalanced translocations, which cause complete or partial chromosomal trisomies, or from amplification events affecting DNA segments of different size. Commonly arise from chromosomal nondisjunction or unbalanced translocations, which cause complete or partial chromosomal trisomies, or from amplification events affecting DNA segments of different size. Malignant melanoma Malignant melanoma MITF on 3p14.2-p14.1 MITF on 3p14.2-p14.1 NEDD9 on 6p25-p24 NEDD9 on 6p25-p24 Hepatocellular carcinoma Hepatocellular carcinoma YAP1 on 11q13 YAP1 on 11q13 BIRC2 on 11q22 BIRC2 on 11q22 18

19. Genomic gains Focal Genomic Gains Focal Genomic Gains Gains affecting small genomic regions or even single genes have been described less frequently than large gains. Gains affecting small genomic regions or even single genes have been described less frequently than large gains. New high-resolution methods New high-resolution methods Comparative genomic hybridization Comparative genomic hybridization Single-nucleotide polymorphism genotyping Single-nucleotide polymorphism genotyping Breast cancer Breast cancer Amplification of small segment of band 6q25.1 containing the gene encoding estrogen receptor 1 (ESR1) Amplification of small segment of band 6q25.1 containing the gene encoding estrogen receptor 1 (ESR1) Increased ESR1 protein levels and increased sensitivity to tamoxifen. Increased ESR1 protein levels and increased sensitivity to tamoxifen. Non-small cell lung cancer Non-small cell lung cancer Amplification of 480-kb interval on band 14q13 Amplification of 480-kb interval on band 14q13 The NKX2-1 gene encodes a lung specific transcription factor The NKX2-1 gene encodes a lung specific transcription factor 19

20. Genomic gains Non-small cell lung cancer Treatment Non-small cell lung cancer Treatment 1. Point mutations in the catalytic domain of the EGFR receptor tyrosine kinase is associated with responsiveness to the kinase inhibitors gefitinib and erlotinib. 2. Amplification and overexpression of the gene encoding the MET receptor tyrosine kinase can restore aberrant signal transduction downstream of mutant EGFR treated with an EGFR inhibitor. 20

21. Genomic losses Cytogenetically visible alterations, such as complete or partial chromosomal monosomies, to single or intragenic deletions that are detectable only by techniques that provide high spatial resolution. Cytogenetically visible alterations, such as complete or partial chromosomal monosomies, to single or intragenic deletions that are detectable only by techniques that provide high spatial resolution. Most recurrent genomic losses probably contribute to malignant transformation by reducing the function of specific genes in the affected chromosomal regions. Most recurrent genomic losses probably contribute to malignant transformation by reducing the function of specific genes in the affected chromosomal regions. 21

22. Genomic losses Glioblastoma, prostate cancer, endometrial cancer Glioblastoma, prostate cancer, endometrial cancer Inactivation of the PTEN tumor-suppressor gene increases signaling through the phosphoinositise-3-kinase-AKT- mammalian target of rapamycin (PI3K-AKT-mTOR) pathway and promotes tumor cell proliferation and survival. Inactivation of the PTEN tumor-suppressor gene increases signaling through the phosphoinositise-3-kinase-AKT- mammalian target of rapamycin (PI3K-AKT-mTOR) pathway and promotes tumor cell proliferation and survival. 22

23. Genomic losses Large-Scale Genomic Losses Large-Scale Genomic Losses 1. The classic approach to identifying a tumor-suppressor gene compares multiple tumors with a specific chromosomal deletion to determine the minimal genomic region that is lost in all cases. RB1 (13q14.2),TP53 (17p13.1), APC (5q21-q22), NF1(17q11.2), PTEN (10q23.3), ATM (11q22-q23) RB1 (13q14.2),TP53 (17p13.1), APC (5q21-q22), NF1(17q11.2), PTEN (10q23.3), ATM (11q22-q23) 2. For many recurrent genomic losses, however, the critical genes are unknown. 1p deletions in neuroblastoma, 3p deletions in lung cancer, 7q deletions in myeloid cancers. 1p deletions in neuroblastoma, 3p deletions in lung cancer, 7q deletions in myeloid cancers. 3. Some deletions have proved to be of great value for determining the prognosis and guiding treatment decisions del(5q) in AML, del(11q), del(13q), del(17p) in CLL del(5q) in AML, del(11q), del(13q), del(17p) in CLL 23

24. Genomic losses Genomic losses Resulting in Allelic Insufficiency Genomic losses Resulting in Allelic Insufficiency Genes that contribute to tumorigenesis by inactivation of a single gene. Genes that contribute to tumorigenesis by inactivation of a single gene. 5q minus syndrome of the myelodysplastic syndrome 5q minus syndrome of the myelodysplastic syndrome Patients with the 5q minus syndrome are highly responsive to the thalidomide derivative lenalidomide. Patients with the 5q minus syndrome are highly responsive to the thalidomide derivative lenalidomide. 24

25. Genomic losses Genomic losses Affecting Noncoding Genes Genomic losses Affecting Noncoding Genes Inactivation of genes that do not encode proteins. Inactivation of genes that do not encode proteins. microRNA genes microRNA genes Small RNA involved in posttranscriptional regulation of gene expression Small RNA involved in posttranscriptional regulation of gene expression MIRN15A and MIRN16-1 in CLL MIRN15A and MIRN16-1 in CLL Negatively regulate the expression of antiapoptotic protein BCL2 Negatively regulate the expression of antiapoptotic protein BCL2 25

26. Summary Cancer is caused by genetic alterations that disrupt the normal balance among cell proliferation, survival, and differentiation. Cancer is caused by genetic alterations that disrupt the normal balance among cell proliferation, survival, and differentiation. For treatment of cancer many of the most specific drug targets undergo genetic changes that conventional cytogenetic methods can detect. For treatment of cancer many of the most specific drug targets undergo genetic changes that conventional cytogenetic methods can detect. The analysis of chromosomal abnormalities can be used to identify the subpopulation of patients who are most likely to benefit from a particular drug treatment The analysis of chromosomal abnormalities can be used to identify the subpopulation of patients who are most likely to benefit from a particular drug treatment Continued improvements in genetic techniques will lead to identification of additional genetic changes that can be exploited to design better therapeutic strategies. Continued improvements in genetic techniques will lead to identification of additional genetic changes that can be exploited to design better therapeutic strategies. 26