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Genome biology and gynecology: the application of oligonucleotide microarrays to leiomyomata
FERTILITY AND STERILITY威 VOL. 80, NO. 2, AUGUST 2003 Copyright ©2003 American Society for Reproductive Medicine Published by Elsevier Inc. Printed on acid-free paper in U.S.A.
Genome biology and gynecology: the application of oligonucleotide microarrays to leiomyomata William A. Freije, M.D. Department of Obstetrics and Gynecology, Department of Human Genetics, David Geffen School of Medicine, University of California, Los Angeles, California
The technological developments of the prior decades have led to the sequencing of multiple organisms, most notably the human species itself as reported in the preliminary draft of the human genome in February 2001 (1, 2). The automated sequencer as developed by Leroy Hood at Caltech in 1986 (3) has made the Human Genome Project a reality, and the benefit of this research effort has now led to a variety of commercially available, relatively inexpensive microarray platforms that researchers may use to examine genome biology. The term genome refers to the ability to simultaneously examine all of the approximately 30,000 human genes in a single experiment. This technology has already been applied to ovarian cancer (4) as well as to benign uterine tumors such as leiomyoma as reported in this (5) and previous issues (6) of Fertility and Sterility.
Received November 25, 2002; revised and accepted March 4, 2003. Reprint requests: William A. Freije, M.D., Department of Obstetrics and Gynecology, David Geffen School of Medicine, University of California, Los Angeles, 10833 LeConte Avenue, Room 27-139 CHS, Los Angeles, CA 90095-1740 (FAX: 310-794-5446; E-mail: [email protected]). 0015-0282/03/$30.00 doi:10.1016/S0015-0282(03) 00729-5
The ability to evaluate thousands of genes in a single hybridization allows researchers to approach problem solving not at an individual gene level; rather, researchers can take a holistic approach that evaluates the full breadth of cellular activity. The papers submitted by Tsibris and colleagues in the July 2002 Fertility and Sterility issue and Wang et al. in this issue are the first steps toward the ultimate goal of systems biologic analysis, as propounded by Dr. Hood, in a gynecologic field. In this latest article of Wang et al., entitled “Distinctive Proliferative Phase Differences in Gene Expression in Human Myometrium and Leiomyomata,” the investigators report on 68 genes that are significantly different in leiomyomata as compared with normal adjacent myometrium from a screen of 6,800 genes. The categories of genes involved include signal transduction, growth factors, hor-
mones/receptors, transcription factors, extracellular matrix proteins, and prostaglandin-related genes. When these 68 genes were employed to discriminate without a priori identity of the tissue samples, discrimination between the normal myometrium and leiomyomatous tissue was easily apparent (see Fig. 2 of Wang et al. (5), dendogram of tissue samples). In addition, the investigators have extended their microarray results with real time PCR, as well as with estrogen receptor immunohistochemistry. An examination of both studies reveals a handful of genes that are consistently differentially regulated in leiomyomata as compared with normal myometrium. Significant up-regulated genes that are shared in both studies include IGF-II, HBGR2, CRABP-II, MEST, and thymidylate synthase. Significant down-regulated genes that are shared in both studies include ADH, SLC, PGD2, PGE3R, MAPKKK5, and dermatopontin. It is encouraging that different researchers have identified similar genes thought to be important in the development of leiomyomata. However, the list of differentially regulated genes identified in both studies is not identical in either number or content, despite the similar platform of Affymetrix oligonucleotide match/mismatch microarrays. This highlights the importance of tissue related issues, such as patient selection and timing of tissue procurement during either the proliferative or follicular phases, and normalization of the technique, as well as microarray related issues such as the platform used and the normalization technique employed. Therefore, the future holds the promise of the application of these new technologies, including proteomic analysis, to dissect the pathophysiology of gynecologic disorders. The application of 277
this technology will undoubtedly transform the practice of gynecology: novel small molecules will be developed that may molecularly abrogate the abnormal physiology of leiomyomata and allow new treatment regimens. Tumor subclass identification will aid both diagnosis and molecular treatment. It is an exciting time in medicine, most particularly in the field of human genetics, where discovery of functional pathways will lead to improved clinical treatment methods. References 1. Lander ES, Linton LM, Birren B, Nusbaum C, Zody MC, Baldwin J, et al. Initial sequencing and analysis of the human genome. Nature 2001; 409:860 –921.
278
Freije
Microarray analysis of leiomyomata
2. Venter JC, Adams MD, Myers EW, Li PW, Mural RJ, Sutton GG, et al. The sequence of the human genome. Science 2001;291:1304 –51. 3. Smith LM, Sanders JZ, Kaiser RJ, Hughes P, Dodd C, Connell CR, et al. Fluorescence detection in automated DNA sequence analysis. Nature 1986;321:674 –9. 4. Welsh JB, Zarrinkar PP, Sapinoso LM, Kern SG, Behling CA, Monk BJ, et al. Analysis of gene expression profiles in normal and neoplastic ovarian tissue samples identifies candidate molecular markers of epithelial ovarian cancer. Proc Natl Acad Sci USA 2001;98:1176 –81. 5. Wang H, Mahadevappa M, Yamamoto K, Wen Y, Chen B, Warrington JA, et al. Distinctive proliferative phase differences in gene expression in human myometrium and leiomyomata. Fertil Steril 2003;80:266 –76. 6. Tsibris JC, Segars J, Coppola D, Mane S, Wilbanks GD, O’Brien WF, et al. Insights from gene arrays on the development and growth regulation of uterine leiomyomata. Fertil Steril 2002;78:114 –21.
Vol. 80, No. 2, August 2003
Genome biology and gynecology: the application of oligonucleotide microarrays to leiomyomata William A. Freije, M.D. Department of Obstetrics and Gynecology, Department of Human Genetics, David Geffen School of Medicine, University of California, Los Angeles, California
The technological developments of the prior decades have led to the sequencing of multiple organisms, most notably the human species itself as reported in the preliminary draft of the human genome in February 2001 (1, 2). The automated sequencer as developed by Leroy Hood at Caltech in 1986 (3) has made the Human Genome Project a reality, and the benefit of this research effort has now led to a variety of commercially available, relatively inexpensive microarray platforms that researchers may use to examine genome biology. The term genome refers to the ability to simultaneously examine all of the approximately 30,000 human genes in a single experiment. This technology has already been applied to ovarian cancer (4) as well as to benign uterine tumors such as leiomyoma as reported in this (5) and previous issues (6) of Fertility and Sterility.
Received November 25, 2002; revised and accepted March 4, 2003. Reprint requests: William A. Freije, M.D., Department of Obstetrics and Gynecology, David Geffen School of Medicine, University of California, Los Angeles, 10833 LeConte Avenue, Room 27-139 CHS, Los Angeles, CA 90095-1740 (FAX: 310-794-5446; E-mail: [email protected]). 0015-0282/03/$30.00 doi:10.1016/S0015-0282(03) 00729-5
The ability to evaluate thousands of genes in a single hybridization allows researchers to approach problem solving not at an individual gene level; rather, researchers can take a holistic approach that evaluates the full breadth of cellular activity. The papers submitted by Tsibris and colleagues in the July 2002 Fertility and Sterility issue and Wang et al. in this issue are the first steps toward the ultimate goal of systems biologic analysis, as propounded by Dr. Hood, in a gynecologic field. In this latest article of Wang et al., entitled “Distinctive Proliferative Phase Differences in Gene Expression in Human Myometrium and Leiomyomata,” the investigators report on 68 genes that are significantly different in leiomyomata as compared with normal adjacent myometrium from a screen of 6,800 genes. The categories of genes involved include signal transduction, growth factors, hor-
mones/receptors, transcription factors, extracellular matrix proteins, and prostaglandin-related genes. When these 68 genes were employed to discriminate without a priori identity of the tissue samples, discrimination between the normal myometrium and leiomyomatous tissue was easily apparent (see Fig. 2 of Wang et al. (5), dendogram of tissue samples). In addition, the investigators have extended their microarray results with real time PCR, as well as with estrogen receptor immunohistochemistry. An examination of both studies reveals a handful of genes that are consistently differentially regulated in leiomyomata as compared with normal myometrium. Significant up-regulated genes that are shared in both studies include IGF-II, HBGR2, CRABP-II, MEST, and thymidylate synthase. Significant down-regulated genes that are shared in both studies include ADH, SLC, PGD2, PGE3R, MAPKKK5, and dermatopontin. It is encouraging that different researchers have identified similar genes thought to be important in the development of leiomyomata. However, the list of differentially regulated genes identified in both studies is not identical in either number or content, despite the similar platform of Affymetrix oligonucleotide match/mismatch microarrays. This highlights the importance of tissue related issues, such as patient selection and timing of tissue procurement during either the proliferative or follicular phases, and normalization of the technique, as well as microarray related issues such as the platform used and the normalization technique employed. Therefore, the future holds the promise of the application of these new technologies, including proteomic analysis, to dissect the pathophysiology of gynecologic disorders. The application of 277
this technology will undoubtedly transform the practice of gynecology: novel small molecules will be developed that may molecularly abrogate the abnormal physiology of leiomyomata and allow new treatment regimens. Tumor subclass identification will aid both diagnosis and molecular treatment. It is an exciting time in medicine, most particularly in the field of human genetics, where discovery of functional pathways will lead to improved clinical treatment methods. References 1. Lander ES, Linton LM, Birren B, Nusbaum C, Zody MC, Baldwin J, et al. Initial sequencing and analysis of the human genome. Nature 2001; 409:860 –921.
278
Freije
Microarray analysis of leiomyomata
2. Venter JC, Adams MD, Myers EW, Li PW, Mural RJ, Sutton GG, et al. The sequence of the human genome. Science 2001;291:1304 –51. 3. Smith LM, Sanders JZ, Kaiser RJ, Hughes P, Dodd C, Connell CR, et al. Fluorescence detection in automated DNA sequence analysis. Nature 1986;321:674 –9. 4. Welsh JB, Zarrinkar PP, Sapinoso LM, Kern SG, Behling CA, Monk BJ, et al. Analysis of gene expression profiles in normal and neoplastic ovarian tissue samples identifies candidate molecular markers of epithelial ovarian cancer. Proc Natl Acad Sci USA 2001;98:1176 –81. 5. Wang H, Mahadevappa M, Yamamoto K, Wen Y, Chen B, Warrington JA, et al. Distinctive proliferative phase differences in gene expression in human myometrium and leiomyomata. Fertil Steril 2003;80:266 –76. 6. Tsibris JC, Segars J, Coppola D, Mane S, Wilbanks GD, O’Brien WF, et al. Insights from gene arrays on the development and growth regulation of uterine leiomyomata. Fertil Steril 2002;78:114 –21.
Vol. 80, No. 2, August 2003