**9. References**


Inflammatory ROS in Fanconi Anemia Hematopoiesis and Leukemogenesis 53

Cumming, R.C., Lightfoot, J., Beard, K., Youssoufian, H., O'Brien, P.J. & Buchwald, M.

hematopoietic cells through redox regulation of GSTP1. *Nat Med* 7: 814–820.

D'Andrea, A.D. & Grompe, M. (2003) The Fanconi anaemia/BRCA pathway. *Nat Rev Cancer*

D'Andrea, A.D. (2010) Susceptibility pathways in Fanconi's anemia and breast cancer. *N* 

de Cremoux, P., Gluckman, E., Podgorniak, M.P., Menier, C., Thierry, D., Calvo, F. & Socie,

group G gene FANCG is identical with XRCC9. *Nat Gene* 20: 281–283. de Winter, J.P., Leveille, F., van Berkel, C.G., Rooimans, M.A., can Der, W.L., Steltenpool, J.,

anemia complementation Group E gene. *Am Hum Gene* 67:1306–1308. de Winter, J.P., Rooimans, M.A., van Der Weel, L., van Berkel, C.G., Alon, N., Bosnoyan-

Dhar SK, Lynn BC, Daosukho C, St Clair DK. Identification of nucleophosmin as an NF-

Doneshbod–Skibba, G., Martin, J. & Shahidi, N. (1980) Myeloid and erythroid colony growth in non-anemic patients with Fanconi's anemia. *Br J Haematol* 44: 33–38. Du, W., Adam, Z., Rani, R., Zhang, X. & Pang, Q. (2008) Oxidative stress in Fanconi anemia

Dufour, C., Corcione, A., Svahn, J., Haupt, R., Poggi, V., Beka'ssy, A.N., Scime, R., Pistorio,

Essers, M.A., Weijzen, S., de Vries-Smits, A.M., Saarloos, I., de Ruiter, N.D., Bos, J.L. &

Fagerlie, S., Lensch, M.W., Pang, Q. & Bagby, G.C. Jr. (2001) The Fanconi anemia group C

Fiers, W., Beyaert, R., Declercq, W. & Vandenabeele, P. (1999) More than one way to die: Apoptosis, necrosis and reactive oxygen damage. *Oncogene* 18: 7719–7730.

mediated by the small GTPase Ral and JNK. *EMBO J.* 23(24):4802-1812. Fagerlie, S.R., Diaz, J., Christianson, T.A., McCartan, K., Keeble, W., Faulkner, G.R. &

expression of interferon –inducible genes. *Blood* 97: 3017–3024.

novel protein with homology to ROM. *Nat Gene* 24: 15–16.

Coussens, L.M. & Werb, Z. (2002) Inflammation and cancer. *Nature* 420: 860–867.

3: 23–34.

*Engl J Med.* 362(20):1909-1919.

Jul 2;279(27):28209-28219.

vitro. *Blood* 102: 2053–2059.

1921.

(2001) Fanconi anemia group C protein prevents prevents apoptosis in

G. (1996) Decreased IL-1 beta and TNF alpha secretion in long-term bone marrow culture supernatant from Fanconi's anaemia patients. *Eur J Haematol* 57: 202–207. de Winter, J.P., Waisfisz, Q., Rooimans, M.A., van Berkel, C.G., Bosnoyan–Collins, L., Alon,

N., Carreau, M., Bender, O., Demuth, I., Schindler, D., Pronk, J.C., Arwert, F., Hoehn, H., Digweed, M., Buchwald, M. & Joenje, H. (1998) The Fanconi anaemia

Demuth, I., Morgan, N.V., Alon, N., Bosnoyan–Collins, L., Lightfoot, J., Leegwater, P.A., Waisfisz, Q., Komatsu, K., Arwert, F., Pronk, J.C., Mathew, C.G., Digweed, M., Buchwald, M. & Joenje, H. (2000) Isolation of a cDNA representing the Fanconi

Collins, L., de Groot, J., Zhi, Y., Waisfisz, Q., Pronk, J.C., Arwert, F., Mathew, C.G., Scheper, R.J., Hoatlin, M.E., Buchwald, M. & Joenje. H. (2000) FANCF encodes a

kappaB co-activator for the induction of the human SOD2 gene. J Biol Chem. 2004

hematopoiesis and disease progression. *Antioxidants & Redox Signaling* 10:1909-

A. & Pistoia, V. (2003) TNF-alpha and IFNgamma are overexpressed in the bone marrow of Fanconi anemia patients and TNF-alpha suppresses erythropoiesis in

Burgering, B.M. (2004) FOXO transcription factor activation by oxidative stress

Bagby, G.C. (2001) Functional correction of FA-C cells with FANCC suppresses the

gene product: signaling functions in hematopoietic cells. *Exp Hematol* 29: 1371–1381.


Bhatia, R., McGlave, P.B., Dewald, G.W., Blazar, B.R. & Verfaillie, C.M. (1995) Abnormal

Balkwill, F. & Mantovani, A. (2001) Inflammation and cancer: back to Virchow? *Lancet* 357:

Bogliolo, M., Cabré, O., Callén, E., Castillo, V., Creus, A., Marcos, R., & Surrallés, J. (2002)

Brunet, A., Sweeney, L.B., Sturgill, J.F., Chua, K.F., Greer, P.L., Lin, Y., Tran, H., Ross, S.E.,

Cadet, J., Berger, M., Douki, T. & Ravanat, J. L. (2006) *Rev. Physiol. Biochem. Pharmacol.* 131: 1-

Chen, M., Tomkins, D.J., Auerbach, W., McKerlie, C., Youssoufian, H., Liu, L., Gan, O.,

Chen, Q.M. (2000) Replicative senescence and oxidant-induced premature senescence. Beyond the control of cell cycle checkpoints. *Ann. N. Y. Acad. Sci.* 908, 111–125. Cheng, N.C., van de Vrugt, H.J., van der Valk, M.A., Oostra, A.B., Krimpenfort, P., de Vries,

Ciccia, A., Ling, C., Coulthard, R., Yan, Z., Xue, Y., Meetei, A.R., Laghmani, el. H., Joenje, H.,

Cohen–Haguenauer, O., Pult, B., Bauche, C., Daniel, M.I., Casalb, I., Levy, V., Dausset, J.,

Collins, N. & Kupfer, G.M. (2005) Molecular pathogenesis of Fanconi anemia. *Int J Hemato*

Collis, S.J., Ciccia, A., Deans, A.J., Horejsi, Z., Martin, J.S., Maslen, S.L., Skehel, J.M., Elledge,

Cumming, R.C., Liu, J.M., Youssoufian, H. & Buchwald, M. (1996) Suppression of apoptosis

Fanconi anemia homolog Fanca. *Hum. Mol. Genet.* 9, 1805–1811

Carreau, M., Auerbach, A., Groves, T., Guidos, C.J., Freedman, M.H., Cross, J., Percy, D.H., Dick, J.E., Joyner, A.L. & Buchwald, M. (1996) Inactivation of Fac in mice produces inducible chromosomal instability and reduced fertility reminiscent

Y., Joenje, H., Berns, A. & Arwert, F. (2000) Mice with a targeted disruption of the

McDonald, N., de Winter, J.P., Wang, W. & West, S.C. (2007) Identification of FAAP24, a Fanconi anemia core complex protein that interacts with FANCM. *Mol* 

Boiron, M., Auclair, C., Gluckman, E. & Marty, M. (2006) In vivo repopulation ability of genetically corrected bone marrow cells from Fanconi anemia patients.

S.J., West, S.C. & Boulton, S.J. (2008) FANCM FAAP24 function in ATR-mediated checkpoint signaling independently of the Fanconi anemia core complex. *Mol Cell.*

in hematopoietic factor-dependent progenitor cell lines by expression of the FAC

processing of DNA damage? *Mutat Res* 408: 75–90.

Cadet, J., Douki, T. & Ravanat, J.L. (2010) *Free Rad. Biol. Med.* 49: 9-21.

of Fanconi anaemia*. Nat. Genet.* 12, 448–451.

*Proc Natl Acad Sci USA* 103:2340–2345.

Leukemia: Role of Malignant Stromal Macrophages. *Blood* 85: 3636-3645 Bagby, G.C. Jr. (2003) Genetic basis of Fanconi anemia. *Curr Opin Hematol* 10: 68–76

539–545.

87.

*Cell.* 25: 331–343.

82: 176–183.

32:313–324.

gene. *Blood* 88: 4558–4567.

*Mutagenesis* 17: 529–538.

Function of the Bone Marrow Microenvironment in Chronic Myelogenous

The Fanconi anaemia genome stability and tumour suppressor network.

Mostoslavsky, R., Cohen, H.Y., Hu, L.S., Cheng, H.L., Jedrychowski, M.P., Gygi, S.P., Sinclair, D.A., Alt, F.W. & Greenberg, M.E. (2004) Stress-dependent regulation of FOXO transcription factors by the SIRT1 deacetylase. *Science* 303: 2011-2015. Buchwald, M. & Moustacchi, E. (1998) Is Fanconi anemia caused by a defect in the


Inflammatory ROS in Fanconi Anemia Hematopoiesis and Leukemogenesis 55

Ibáñez, A., Río, P., Casado, J.A., Bueren, J.A., Fernández-Luna, J.L., Pipaón, C. (2009)

Ichijo, H., Nishida, E., Irie, K., ten Dijke, P., Saitoh, M., Moriguchi, T., Takagi, M.,

Joenje, H., Arwert, F., Eriksson, A.W., de Koning, H. & Oostra, A.B. (1987) Oxygen-

Joenje, H., Levitus, M., Waisfisz, Q., D' Andrea, A.D., Garcia–Higuera, I., Pearson, T., van

Jonkers, J., Meuwissen, R., van der Gulden, H., Peterse, H., van der Valk, M. & Bern, A.

Kops, G.J., Dansen, T.B., Polderman, P.E., Saarloos, I., Wirtz, K.W., Coffer, P.J., Huang, T.T.,

Kennedy, R.D. & D'Andrea, A.D. (2005) The Fanconi Anemia/BRCA pathway: new faces in

Koh, P.S., Hughes, G.C., Faulkner, G.R., Keeble, W.W. & Bagby, G.C. (1999) The Fanconi

Konopleva, M. & Andreeff, M. (2007) Targeting the Leukemia Microenvironment. *Current* 

Kruyt, F.A., Hoshino, T., Liu, J.M., Joseph, P., Jaiswal, A.K. & Youssoufian, H. (1998)

Kuper, H., Adami, H.O. & Trichopoulos, D. (2000) Infections as a major preventable cause

Kutler, D.I., Wreesmann, V.B., Goberdhan, A., Ben-Prat, L., Satagopan, J., Ngai, I., Huvos,

Lensch, M.W., Rathbun, R.K., Olson, S.B., Jones, G.R. & Bagby, G.C. Jr. (1999) Selective

view from the window of Fanconi anemia. *Leukemia* 13: 1784–1789.

Fas ligand–induced neutrophil apoptosis. *Nat Med.* 11(6):666-671.

necrosis factor receptor superfamily. *Exp Hematol* 27: 1–8.

Kryston, T. B., Georgiev, A. & Georgakilas, A. G. (2011) *Mutat. Res.* 711, 193-201.

promoting tumour cell proliferation. *Biochem J.* 29; 422(1):161-170.

patient to group A. *Am J Hum Gene* 67: 759–762.

the crowd. *Genes Dev* 19: 2925–2940.

*Drug Targets,* 2007*, 8,* 685-701.

*Cancer Inst* 95: 1718–1721.

of human cancer. *J Intern Med* 248:171–183.

3050–3056.

mouse model for breast cancer. *Nat Genet* 29: 418–425.

*Science* 275: 90–94.

Elevated levels of IL-1beta in Fanconi anaemia group A patients due to a constitutively active phosphoinositide 3-kinase-Akt pathway are capable of

Matsumoto, K., Miyazono, K. & Gotoh, Y. (1997) Induction of apoptosis by ASK1, a mammalian MAPKKK that activates SAPK/JNK and p38 signaling pathways.

dependence of chromosomal aberrations in Fanconi's anaemia. *Nature* 290: 142–143.

Berkel, C.G., Rooimans, M.A., Morgan, N., Mathew, C.G. & Arwert, F. (2000) Complementation analysis in Fanconi anemia: Assignment of the reference. FA-H

(2001) Synergistic tumor suppressor activity of BRCA2 and p53 in a conditional

Bos, J.L., Medema, R.H., Burgering, B.M. (2002) Forkhead transcription factor FOXO3a protects quiescent cells from oxidative stress. *Nature* 419(6904):316-321. Jonsson, H., Allen, P., Peng, S.L. (2005) Inflammatory arthritis requires Foxo3a to prevent

anemia group C gene product modulates apoptotic responses to tumor necrosis factor- and Fas ligand but does not suppress expression of receptors of the tumor

Abnormal microsomal detoxification implicated in Fanconi anemia group C by interaction of the FAC protein with NADPH cytochrome P450 reductase. *Blood* 92:

A.G., Giampietro, P., Levran, O., Pujara, K., Diotti, R., Carlson, D., Huryn, L.A., Auerbach, A.D. & Singh, B. (2003) Human papillomavirus DNA and p53 polymorphisms in squamous cell carcinomas from Fanconi anemia patients. *J Natl* 

pressure as an essential force in molecular evolution of myeloid leukemic clones: a


Freie, B., Li, X., Ciccone, S.L., Nawa, K., Cooper, S., Vogelweid, C., Schantz, L., Haneline,

Fridman, J.S. & Lowe, S.W. (2003) Control of apoptosis by p53. *Oncogene* 22: 9030–9040. Futaki, M., Igarashi, T., Watanabe, S., Kajigaya, S., Tatsuguchi, A., Wang, J. & Liu, J.M.

protection against oxidative DNA damage. *Carcinogenesis* 23: 67–72. Giaccia, A.J. & Kastan, M.B. (1998) The complexity of p53 modulation: emerging patterns

Gluckman, E., Broxmeyer, H., Auerbach, A., Friedman, H., Douglas, G., Devergie, A.,

Green, A.M. & Kupfer, G.M. (2009) Fanconi anemia. *Hematol Oncol Clin North Am*. 23:193-

Hadjur, S., Ung, K., Wadsworth, L., Dimmick, J., Rajcan–Separovic, E., Scott, R.W.,

Hakim, J. (1993) Reactive oxygen species and inflammation *C R Seances Soc Biol Fil.* 

Hammond, E. M., Dorie, M. J. & Giaccia, A. J. (2003) ATR/ATM targets are phosphorylated

Haneline, L.S., Broxmeyer, H.E., Cooper, S., Hangoc, G., Carreau, M., Buchwald, M. &

Haneline, L.S., Gobbett, T.A., Ramani, R., Carreau, M., Buchwald, M., Yoder, M.C. & Clapp,

Haneline, L.S., Li, X., Ciccone, S.L., Hong, P., Yang, Y., Broxmeyer, H.E., Lee, S.H., Orazi, A.,

Horejsi, Z., Collis, S.J., Boulton, S.J. (2009) FANCM-FAAP24 and HCLK2: roles in ATR

Howlett, N.G., Taniguchi, T., Olson, S., Cox, B., Waisfisz, Q., De Die–Smulders, C., Persky,

Biallelic inactivation of BRCA2 in Fanconi anemia. *Science* 297: 606–609. Huang, H., Tindall, D.J. (2007) Dynamic FoxO transcription factors. *J Cell Sci.* 120:2479-2487.

signalling and the Fanconi anemia pathway. *Cell Cycle* 8:1133–1137.

decreases the risk of clonal evolution. *Blood* 101: 1299–1307.

from divergent signals. *Genes* 12: 2973–2983.

superoxide dismutase. *Blood* 98: 1003–1011.

50986–50993.

Med 321: 1174–1178.

187(3):286-295.

*Chem.* 278, 12207–12213.

repopulating ability. *Blood* 94: 1–8.

214.

4098.

L.S., Orazi, A., Broxmeyer, H.E., Lee, S.H. & Clapp, D.W. (2003) Fanconi anemia type C and p53 cooperate in apoptosis and tumorigenesis. *Blood* 102: 4146–4152. Freie, B.W., Ciccone, S.L., Li, X., Plett, P.A., Orschell, C.M., Srour, E.F., Hanenberg, H.,

Schindler, D., Lee, S.H. & Clapp, D.W. (2004) A role for the Fanconi anemia C protein in maintaining the DNA damage-induced G2 checkpoint. *J Biol Chem* 279:

(2002) The FANCG Fanconi anemia protein interacts with CYP2E1: possible role in

Esperou, H., Thierry, D., Socie, G., Lehn, P., Cooper, S., English, D., Kurtzberg, J., Bard, J. & Boyse, E. (1989) Hematopoietic reconstitution in a patient with Fanconi's anemia by means of umbilical-cord blood from an HLA identical sibling. *N Engl J* 

Buchwald, M. & Jirik, F.R. (2001) Defective hematopoiesis and hepatic steatosis in mice with combined deficiencies of the genes encoding Fancc and Cu/Zn

by ATR in response to hypoxia and ATM in response to reoxygenation. *J. Biol.* 

Clapp, D.W. (1998) Multiple inhibitory cytokines induce deregulated progenitor growth and apoptosis in hematopoietic cells from FAC-/- mice. *Blood* 91: 4092–

D.W. (1999) Loss of Fancc function results in decreased hematopoietic stem cell

Srour, E.F. & Clapp, D.W. (2003) Retroviralmediated expression of recombinant Fancc enhances the repopulating ability of Fancc-/- hematopoietic stem cells and

N., Grompe, M., Joenje, H., Pals, G., Ikeda, H., Fox, E.A. & D'Andrea, A.D. (2002)


Inflammatory ROS in Fanconi Anemia Hematopoiesis and Leukemogenesis 57

Mackay, I.R. & Rose, N.R. (2001) Autoimmunity and lymphoma: tribulations of B cells. *Nat* 

Mantovani, G., Macciò, A., Pisano, M., Versace, R., Lai, P., Esu, S., Massa, E., Ghiani, M.,

Mantovani, G., Macciò, A., Madeddu, C., Mura, L., Gramigano, G., Lusso, M.R., Mulas, C.,

Marx, J. (2004) Cancer research. Inflammation and cancer: the link grows stronger. *Science*

Martin, O. A., Redon, C., Nakamura, A. J., Dickey, J. S., Georgakilas, A. G. & Bonner, W. M.

Meetei, A.R., de Winter, J.P., Medhurst, A.L., Wallisch, M., Waisfisz, Q., van de Vrugt, H.J.,

Meetei, A.R., Levitus, M., Xue, Y., Medhurst, A.L., Zwaan, M., Ling, C., Rooimans, M.A.,

Meindl, A., Hellebrand, H., Wiek, C., Erven, V., Wappenschmidt, B., Niederacher, D.,

RAD51C as a human cancer susceptibility gene. *Nat Genet.* 42 (5): 410-414. Miyamoto, K., Araki, K.Y., Naka, K., Arai, F., Takubo, K., Yamazaki, S., Matsuoka, S.,

Mukhopadhyay, S.S., Leung, K.S., Hicks, M.J., Hastings, P.J., Youssoufian, H. & Plon, S.E.

Nakamura, H., Nakamura, K. & Yodoi, J. (1997) Redox regulation of cellular activation.

Nakata, S., Matsumura, I., Tanaka, H., Ezoe, S., Satoh, Y., Ishikawa, J., Era, T. & Kanakura, Y.

anemia complementation group M *Nat Genet* 37: 958–963.

stress in Fanconi anemia *J Cell Biol* 175: 225–235.

Oostra, A.B., Yan, Z., Ling, C., Bishop, C.E., Hoatlin, M.E., Joenje, H. & Wang, W. (2003) A novel ubiquitin ligase is deficient in Fanconi anemia. *Nat Genet* 35: 165–

Bier, P., Hoatlin, M., Pals, G., de Winter, J.P., Wang, W. & Joenje, H. (2004) X-linked inheritance of Fanconi anemia complementation group B. *Nat Gene* 36: 1219–1224. Meetei, A.R., Medhurst, A.L., Ling, C., Xue, Y., Singh, T.R., Bier, P., Steltenpool, J., Stone, S.,

Dokal, I., Mathew, C.G,, Hoatlin, M., Joenje ,H., de Winter, J.P, & Wang, W. (2005) A human ortholog of archael DNA repair protein HEF is defective in Fanconi

Freund, M., Lichtner, P., Hartmann, L., Schaal, H., Ramser, J., Honisch, E., Kubisch, C., Wichmann, H.E., Kast, K., Deissler, H., Engel, C., Müller-Myhsok, B., Neveling, K., Kiechle, M., Mathew, C.G., Schindler, D., Schmutzler, R.K., Hanenberg, H. (2010) Germline mutations in breast and ovarian cancer pedigrees establish

Miyamoto, T., Ito, K., Ohmura, M., Chen, C., Hosokawa, K., Nakauchi, H., Nakayama, K., Nakayama, K.I., Harada, M., Motoyama, N., Suda, T. & Hirao, A. (2007) Foxo3a is essential for maintenance of the hematopoietic stem cell pool. *Cell* 

(2006) Defective mitochondrial peroxiredoxin-3 results in sensitivity to oxidative

(2004) NF-kappa B family proteins proteins participate in multiple steps of

Dessi, D., Melis, G.B. & Del Giacco, D.S. (1997) Tumor-associated lymphomonocytes from neoplastic effusions are immunologically defective in comparison with patient autologous PBMCs but are capable of releasing high amounts of

Mudu, M.C., Murgia, V., Camboni, P., Massa, E., Ferreli, L., Contu, P., Rinaldi, A., Sanjust, E., Atzei, D. & Elsener, B. (2002) Quantitative evaluation of oxidative stress, chronic inflammatory indices and leptin in cancer patients: correlation with stage

*Immunol* 2: 793–795.

306: 966–968.

170.

(2011) *Cancer Res*. 71: 1-5.

*Stem Cell.* 1(1):101-112.

*Annu Rev Immunol* 15: 351–369.

various cytokines. *Int J Cancer* 71: 724–731.

and performance status. *Int J Cancer* 98: 84–91.


Levitus, M., Rooimans, M.A., Steltenpool, J., Cool, N.F., Oostra, A.B., Mathew, C.G., Hoatlin,

Li, J., Sejas, D.P., Zhang, X., Qiu, Y., Nattamai, K.J., Rani, R., Rathbun, K.R., Geiger, H.,

Li, J., Du, W., Maynard, S., Andreassen, P.R. & Pang, Q. (2010) Oxidative stress-specific interaction between FANCD2 and FOXO3a. *Blood.* 115(8):1545-1548. Li, X., Yang, Y., Yuan, J., Hong, P., Freie, B., Orazi, A., Haneline, L.S. & Clapp, D.W. (2004)

Li, Y. & Youssoufian, H. (1997) MxA overexpression reveals a common genetic link in four Fanconi anemia complementation groups. *J Clin Invest* 100: 2873–2880. Liu, J. (2000) Fanconi's anemia. In: Young NS, ed. *Bone Marrow Failure Syndromes*. 47–68. Liu, T.X., Howlett, N.G., Deng, M., Langenau, D.M., Hsu, K., Rhodes, J., Kanki, J.P.,

Lo Ten Foe, J.R., Rooimans, M.A., Bosnoyan–Collins, L., Alon, N., Wijker, M., Parker, L.,

Luna–Fineman, S., Shannon, K.M. & Lange, B.J. (1995) Childhood monosomy 7: epidemiology, biology, and mechanistic implications. *Blood* 85: 1985–1989. Luke-Glaser, S., Luke, B., Grossi, S. & Constantinou, A. (2010) FANCM regulates DNA chain elongation, is stabilized by S-phase checkpoint signalling. *EMBO J.* 29: 795–805. Ma, D.J., Li, S.J., Wang, L.S., Dai, J., Zhao, S.L & Zeng, R. (2009) Temporal and spatial

Macciò, A., Lai, P., Santona, M.C., Pagliara, L., Melis, G.B. & Mantovani, G. (1998) High

Macdougall, I.C. & Cooper, A.C. (2002) Erythropoitin resistence: the role of inflammation

Maciejewski, J.P., Selleri, C., Sato, T., Anderson, S. & Young, N.S. (1995) Increased

and pro-inflammatory cytokines. *Nephrol Dial Transplant* 17: 39–43.

anemia. *Nat Genet* 37: 931–933.

type cells in Fancc-/- mice. *Blood* 104: 1204– 1209.

members. *Cell Death Differ* 6: 1162– 1168.

gene, FAA. *Nature Gene* 14: 320–323.

cancer. *Gynecol Oncol* 69: 248–252.

anaemia. *Br J Haematol* 91: 245–252.

*Res.* 19(5):651-664.

3295.

M.E., Waisfisz, Q., Arwert, F., De Winter, J.P. & Joenje, H. (2004) Heterogeneity in Fanconi anemia: evidence for two new genetic subtypes. *Blood* 103(7): 2498–2503. Levran, O., Attwooll, C., Henry, R.T., Milton, K.L., Neveling, K., Rio, P., Batish, S.D., Kalb,

R., Velleuer, E., Barral, S., Ott, J., Petrini, J., Schindler, D., Hanenberg, H. & Auerbach, A.D. (2005) The BRCA1-interacting helicase BRIP1 is deficient in Fanconi

Williams, D.A. & Bagby, G.C. & Pang, Q. (2007) TNF- induces leukemic clonal evolution ex vivo in Fanconi anemia group C stem cells. *J Clin Invest* 117: 3283–

Continuous in vivo infusion of interferon- gamma (IFN-gamma) preferentially reduces myeloid progenitor numbers and enhances engraftment of syngeneic wild-

D'Andrea, A.D. & Look, A.T. (2003) Knockdown of zebrafish Fancd2 causes developmental abnormalities via p53-dependent apoptosis. *Dev Cell* 5: 903–914. Lohrum, M.A. & Vousden, K.H. (1999) Regulation and activation of p53 and its family

Lightfoot, J., Carreau, M., Callen, D.F., Savoia, A., Cheng, N.C., van Berkel, C.G., Strunk, M.H., Gille, J.J., Pals, G., Kruyt, F.A., Pronk, J.C., Arwert, F., Buchwald, M. & Joenje, H. (1996) Expression cloning of a cDNA for the major Fanconi anaemia

profiling of nuclei-associated proteins upon TNF-alpha/NF-kappaB signaling. *Cell* 

serum levels of soluble IL-2 receptor, cytokines, and C-reactive protein correlate with impairment of T cell response in patients with advanced epithelial ovarian

expression of Fas antigen on bone marrow CD34\_ cells of patients with aplastic


Inflammatory ROS in Fanconi Anemia Hematopoiesis and Leukemogenesis 59

Rossi, D.J., Bryder, D., Seita, J., Nussenzweig, A., Hoeijmakers, J., Weissman, I.L.(2007)

Rosselli, F., Sanceau, J., Wietzerbin, J. & Moustacchi, E. (1992) Abnormal lymphokine

Rosselli, F., Sanceau, J., Gluckman, E., Wietzerbin, J. & Moustacchi, E. (1994) Abnormal

Rubin, C.M,, Arthur, D.C., Woods, W.G., Lange, B.J., Nowell, P.C., Rowley, J.D., Nachman,

redox- regulated apoptotic pathway. *J Biol Chem* 279: 16805–16812.

ambient oxygen. *Am J Hum Genet* 43(4): 429–435.

in Fancc-deficient mice. *J Immunol* 178: 5277–5287.

in Fanca-/- and Fancg-/- mice. *Blood* 108: 4283–4287.

anemia pathway via FANCM. *Mol Cell.* 37:879–886.

anemia. *Am J Hematol* 42:196–201.

W. M.(2010) *Mutat. Res.* 704, 152-159.

Scheller, J., Chalaris, A., Schmidt-Arras, D. & Rose-John, S. (2011) The pro- and anti-

Schindler, D. & Hoehn, H. (1988) Fanconi anemia mutation causes cellular susceptibility to

Schultz, J.C. & Shahidi, N.T. (1993) Tumor necrosis factor-alpha overproduction in Fanconi's

Schwab, R.A., Blackford, A.N. & Niedzwiedz, W. (2010) ATR activation, replication fork restart are defective in FANCM-deficient cells. *EMBO J.* 29: 806–818. Sedelnikova, O. A., Redon, C. E., Dickey, J. S., Nakamura, A. J., Georgakilas, A. G. & Bonner,

Sejas, D.P., Rani, R., Qiu, Y., Zhang, X., Fagerlie, S.R., Nakano, H., Williams, D.A. & Pang, Q.

Si, Y., Ciccone, S., Yang, F.C., Yuan, J., Zeng, D., Chen, S., van de Vrugt, H., Critser, J.,

Singh, T.R., Saro, D., Ali, A.M., Zheng, X.F., Du, C.H., Killen, M.W., Sachpatzidis, A.,

Smogorzewska, A., Matsuoka, S., Vinciguerra, P., McDonald, E.R. 3rd, Hurov, K.E., Luo, J.,

(2007) Inflammatory reactive oxygen species-mediated hematopoietic suppression

Arwert, F., Haneline, L.S. & Clapp, D.W. (2006) Continuous in vivo infusion of interferon-gamma (IFNgamma) enhances engraftment of syngeneic wild-type cells

Wahengbam, K., Pierce, A.J., Xiong, Y., Sung, P. & Meetei. A.R. (2010) MHF1- MHF2, a histone-fold-containing protein complex, participates in the Fanconi

Ballif, B.A., Gygi, S.P., Hofmann, K., D'Andrea, A.D. & Elledge, S.J. (2007)

to childhood cancer. *Nat Genet* 39: 162–164.

with age. *Nature.* 447(7145):725-729.

interleukin- 6. *Hum Genet* 89: 42–48.

*Blood* 83: 1216–1225.

1813(5):878-888.

Ariffin, H., Tischkowitz, M., Mathew, C.G., Auerbach, A.D. & Rahman, N. (2006) Biallelic mutations in PALB2 cause Fanconi anemia subtype FA-N and predispose

Deficiencies in DNA damage repair limit the function of haematopoietic stem cells

production: a novel feature of the genetic disease Fanconi anemia. I. Involvement of

lymphokine production: a novel feature of the genetic disease Fanconi anemia. II. In vitro and in vivo spontaneous overproduction of tumor necrosis factor alpha.

J., Bostrom, B., Baum, E.S., Suarez, C.R., Shah, N.R., Morgan, E., Mauer, H.S., McKenzie, S.E., Larson, R.A. & Le Beau, M.M. (1991) Therapy-related myelodysplastic syndrome and acute myeloid leukemia in children: correlation between chromosomal abnormalities and prior therapy. *Blood* 78: 2982–2988. Saadatzadeh, M.R., Bijangi–Vishehsaraei, K., Hong, P., Bergmann, H. & Haneline, L.S. (2004)

Oxidant hypersensitivity of Fanconi anemia type C-deficient cells is dependent on a

inflammatory properties of the cytokine interleukin-6. *Biochim Biophys Acta.*

hematopoiesis through elimination of reactive oxygen species*. J Biol Chem* 279: 55578–55586.


Nijnik, A., Woodbine, L., Marchetti, C., Dawson, S., Lambe, T., Liu, C., Rodrigues, N.P.,

Otsuki, T., Nagakura, S., Wang, J., Bloom, M., Grompe, M. & Liu, J.M. (1999) Tumor necrosis

Pagano, G., Degan, P., d'Ischia, M., Kelly, F. J., Nobili, B., Pallardó, F.V., Youssoufian, H. &

Pang, Q. Fagerlie, S., Christianson, T.A., Keeble, W., Faulkner, G., Diaz, J., Rathbun, R.K. &

Pang, Q., Keeble, W., Christianson, T.A., Faulkner, G.R. & Bagby, G.C. (2001) FANCC

Pang, Q., Keeble, W., Diaz, J., Christianson, T.A., Fagerlie, S., Rathbun, K., Faulkner, G.R.,

Pang, Q., Christianson, T.A., Keeble, W., Koretsky, T. & Bagby, G,C. (2002) The anti-

Park, S.J., Ciccone, S.L., Beck, B.D., Hwang, B., Freie, B., Clapp, D.W. & Lee, S.H. (2004)

Potente, M., Urbich, C., Sasaki, K., Hofmann, W.K., Heeschen, C., Aicher, A., Kollipara, R.,

Rathbun, R.K., Faulkner, G.R., Ostroski, M.H., Christianson, T.A., Hughes, G., Jones, G.,

Rathbun, R.K., Christianson, T.A., Faulkner, G.R., Jone, G., Keeble, W., O'Dwyer, M. &

Reid, S., Schindler, D., Hanenberg, H., Barker, K., Hanks, S., Kalb, R., Neveling, K., Kelly, P.,

haematopoietic stem cells during ageing. *Nature.* 447(7145):686-690.

knockout mice. *J Cell Physiol* 179: 79–86.

*Cell. Biol.* 20, 4724–4735.

2392.

*Blood* 90: 974–985.

cytotoxicity. *EMBO J* 20: 4478–4489.

stranded RNA. *Blood.* 2001 97(6):1644-1652.

anemia proteins. *J Biol Chem 279*: 30053–30059.

caspase 3 family members. *Blood* 96: 4204–4211.

anemia protein, FANCC. *J Biol Chem.* 277(51):49638-49643.

clinical phenotype. *Eur J Haematol* 75: 93–100.

55578–55586.

hematopoiesis through elimination of reactive oxygen species*. J Biol Chem* 279:

Crockford, T.L., Cabuy, E., Vindigni, A., Enver, T., Bell, J.I., Slijepcevic, P., Goodnow, C.C., Jeggo, P.A. & Cornall, R.J. (2007) DNA repair is limiting for

factor- and CD95 ligation suppress erythropoiesis in Fanconi anemia C gene

Zatterale, A. (2005) Oxidative stress as a multiple effector in Fanconi anaemia

Bagby, G.C. (2000) The Fanconi anemia protein FANCC binds to and facilitates the activation of STAT1 by gamma interferon and hematopoietic growth factors. *Mol.* 

interacts with Hsp70 to protect hematopoietic cells from IFN-/TNF--mediated

O'Dwyer, M. & Bagby, G.C. Jr. (2001) Role of double-stranded RNA-dependent protein kinase in mediating hypersensitivity of Fanconi anemia complementation group C cells to interferon gamma, tumor necrosis factor-alpha, and double-

apoptotic function of Hsp70 in the interferon-inducible double-stranded RNAdependent protein kinase-mediated death signaling pathway requires the Fanconi

Oxidative stress/damage induces multimerization and interaction of Fanconi

DePinho, R.A., Zeiher, A.M., Dimmeler, S. (2005) Involvement of Foxo transcription factors in angiogenesis and postnatal neovascularization *J Clin Invest.* 115(9):2382-

Cahn, R., Maziarz, R., Royle, G., Keeble, W., Heinrich, M.C/, Grompe, M., Tower, P.A. & Bagby, G.C. (1997) Inactivation of the Fanconi anemia group C gene augments interferon-gamma-induced apoptotic responses in hematopoietic cells.

Bagby, G.C. (2000) Interferon—induced apoptotic responses of Fanconi anemia group C hematopoietic progenitor cells involve caspase 8-dependent activation of

Seal, S., Freund, M., Wurm, M., Batish, S.D., Lach, F.P., Yetgin, S., Neitzel, H.,

Ariffin, H., Tischkowitz, M., Mathew, C.G., Auerbach, A.D. & Rahman, N. (2006) Biallelic mutations in PALB2 cause Fanconi anemia subtype FA-N and predispose to childhood cancer. *Nat Genet* 39: 162–164.


Inflammatory ROS in Fanconi Anemia Hematopoiesis and Leukemogenesis 61

Wajant, H., Pfizenmaier, K. & Scheurich, P. (2003) Tumor necrosis factor signaling. *Cell* 

Walsh, C.E., Nienhuis, A.W., Samulski, R.J., Brown, M.G., Miller, J.L., Young, N.S. & Liu,

West, R.R., Stafford, D.A., White, A.D., Bowen, D.T. & Padua, R.A. (2000) Cytogenetic

Whitney, M.A., Royle, G., Low, M.J., Kelly, M.A., Axthelm, M.K., Reifsteck, C., Olsen, S.,

Wilson, A., Laurenti, E., Oser, G., van der Wath, R.C., Blanco-Bose, W., Jaworski, M., Offner,

Wong, J.C. & Buchwald, M. (2002) Disease model: Fanconi anemia. *Trends Mol Med.* 8(3):139-

Xia, B., Dorsman, J.C., Ameziane, N., de Vries, Y., Rooimans, M.A., Sheng, Q., Pals, G.,

Yamamoto, K.N., Kobayashi, S., Tsuda, M., Kurumizaka, H., Takata, M., Kono, K., Jiricny, J.,

Yang, Y., Kuang, Y., Montes De Oca, R., Hays, T., Moreau, L., Lu, N., Seed, B. & D'Andrea,

Young, N.S. & Maciejewski, J. (1997) The pathophysiology of acquired aplastic anemia. *N.* 

Zhang, X., Li, J., Sejas, D.P., Rathbun, K.R., Bagby, G.C. & Pang, Q. (2004) The Fanconi

Zhang, X., Li, J., Sejas, D.P. & Pang, Q. (2005) Hypoxia-reoxygenationinduces premature senescence in FA bone marrow hematopoietic cells. *Blood* 106: 75–85. Zhang, X., Li, J., Sejas, D.P. & Pang, Q. (2005) The ATM/p53/p21 pathway influences cell

Zhang, X., Sejas, D.P., Qiu, Y., Williams, D.A. & Pang, Q. (2007) Inflammatory ROS promote

targeted disruption of the Fanconi anemia C gene. *Blood* 88: 49–58.

during homeostasis and repair. *Cell.* 135(6):1118-1129.

J.M. (1994) Phenotypic correction of Fanconi anemia in human hematopoietic cells with a recombinant adeno-associated virus vector. *J Clin Invest* 94: 1440–1448. Wang, J., Otsuki, T., Youssoufian, H., Foe, J.L., Kim, S., Devetten, M., Yu, J., Li, Y., Dunn, D.

& Liu, J.M. (1998) Overexpression of the Fanconi anemia group C gene (FAC) protects hematopoietic progenitors from death induced by Fas-mediated apoptosis.

abnormalities in the myelodysplastic syndromes and occupational or

Braun, R.E., Heinrich, M.C., Rathbun, R.K., Bagby, G.C. & Grompe, M. (1996) Germ cell defects and hematopoietic hypersensitivity to -interferon in mice with a

S., Dunant, C.F., Eshkind, L., Bockamp, E., Lió, P., Macdonald, H.R. & Trumpp, A. (2008) Hematopoietic stem cells reversibly switch from dormancy to self-renewal

Errami, A., Gluckman, E., Llera, J., Wang, W., Livingston, D.M., Joenje, H. & de Winter, J.P. (2006) Fanconi anemia is associated with a defect in the BRCA2 partner

Takeda, S. & Hirota, K. (2011) Involvement of SLX4 in interstrand cross-link repair is regulated by the Fanconi anemia pathway. *Proc Natl Acad Sci U S A.* 108(16):6492-

A.D. (2001) Targeted disruption of the murine Fanconi anemia gene, Fancg/Xrcc9.

anemia proteins functionally interact with the protein kinase regulated by RNA

fate decision between apoptosis and senescence in reoxygenated hematopoietic

and cooperate with Fanconi anemia mutation for hematopoietic senescence. *J Cell* 

*Death Differ* 10: 45–65.

*Cancer Res* 58: 3538–3541.

PALB2. *Nat Genet* 39: 159–161.

142.

6496.

*Blood* 98, 3435–3440

*Engl. J. Med.* 336: 1365–1372.

*Science* 120: 1572–1583.

(PKR). *J Biol Chem.* 279(42):43910-43919.

progenitor cells. *J Biol Chem 280*: 19635–19640.

environmental exposure. *Blood* 95: 2093–2097.

Identification of the FANCI Protein, a Monoubiquitinated FANCD2 Paralog Required for DNA Repair. *Cell* 129: 1–13.


Somyajit, K., Subramanya, S., Nagaraju, G. (2010) RAD51C: a novel cancer susceptibility

Stark, R., Andre, C., Thierry, D., Cherel, M., Galibert, F. & Gluckman, E. (1993) The

Stoepker, C., Hain, K., Schuster, B., Hilhorst-Hofstee, Y., Rooimans, M.A., Steltenpool, J.,

Strathdee, C.A., Gavish, H., Shannon, W.R. & Buchwald, M. (1992) Cloning of cDNAs for Fanconi's anaemia by functional complementation. *Nature* 356: 763–767. Suematsu, N., Tsutsui, H., Wen, J., Kang, D., Ikeuchi, M., Ide, T., Hayashidani, S., Shiomi, T.,

Timmers, C., Taniguchi, T., Hejna, J., Reifsteck, C., Locas, L., Bruun, D., Thayer, M., Cox, B.,

Tischkowitz, M. & Dokal, I. (2004) Fanconi anaemia and leukaemia—clinical and molecular

Tothova, Z. & Gilliland, D.G. (2002) FoxO Transcription Factors and Stem Cell Homeostasis:

Tothova, Z., Kollipara, R., Huntly, B.J., Lee, B.H., Castrillon, D.H., Cullen, D.E., McDowell,

Tsai, W.B., Chung, Y.M., Takahashi, Y., Xu, Z. & Hu, M.C. (2008) Functional interaction

Turrel-Davin, F., Tournadre, A., Pachot, A., Arnaud, B., Cazalis, M.A,, Mougin, B. &

Umeda, T. & Hino, O. (2002) Molecular aspects of human hepatocarcinogenesis mediated by

Ventura, J.J., Cogswell, P., Flavell, R.A., Baldwin, A.S. Jr & Davis, R.J. (2004) JNK potentiates

Insights from the Hematopoietic System *Stem Cells*. 20(5):438-447.

Required for DNA Repair. *Cell* 129: 1–13.

*Natl Acad Sci U S A.* 108(16):6492-6496.

cardiac myocytes. *Circulation* 107:1418–1423.

aspects. *Br J Haematol* 126(2): 176–191.

novel Fanconi anemia gene, FANCD2. *Mol Cell* 7: 241–248. Tischkowitz, M.D. & Hodgson, S.V. (2003) Fanconi anaemia*. J Med Genet* 40: 1–10.

2038.

566.

339.

10(4):460-467

*Oncology* 62: 38–42.

species. *Genes Dev* 18: 2905–2915.

Identification of the FANCI Protein, a Monoubiquitinated FANCD2 Paralog

gene is linked to Fanconi anemia and breast cancer. *Carcinogenesis.* 31(12):2031-

expression of cytokine and cytokine receptor genes in long-term bone marrow culture in congenital and acquired bone marrow hypoplasias. *Br J Haematol* 83: 560–

Oostra, A.B., Eirich, K., Korthof, E.T., Nieuwint, A.W., Jaspers, N.G., Bettecken, T., Joenje ,H., Schindler, D., Rouse, J. & de Winter, J.P. (2011) SLX4, a coordinator of structure-specific endonucleases, is mutated in a new Fanconi anemia subtype. *Proc* 

Kubota, T., Hamasaki, N. & Takeshita, A. (2003) Oxidative stress mediates tumor necrosis factor-alpha-induced mitochondrial DNA damage and dysfunction in

Olson, S., D'Andrea, A.D., Moses, R. & Grompe, M. (2001) Positional cloning of a

E.P., Lazo-Kallanian, S., Williams. I.R., Sears, C., Armstrong, S.A., Passegué, E., DePinho, R.A. & Gilliland, D.G. (2002) FoxOs are critical mediators of hematopoietic stem cell resistance to physiologic oxidative stress. *Cell.* 128(2):325-

between FOXO3a and ATM regulates DNA damage response. *Nat Cell Biol.* 

Miossec, P. (2010) FoxO3a involved in neutrophil and T cell survival is overexpressed in rheumatoid blood and synovial tissue. *Ann Rheum Dis* 69:755-760.

inflammation: from hypercarcinogenic state to normo- or hypocarcinogenic state.

TNF-stimulated necrosis by increasing the production of cytotoxic reactive oxygen


*University of Alabama at Birmingham* 

*USA* 

Somaira Nowsheen, Alexandros G. Georgakilas and Eddy S. Yang

Despite decades of research, cancer continues to affect millions of people each year. However, the more we discover about cancer, the more we realize that no single therapeutic strategy can effectively treat it. As we learn about the aberrant signals and pathways which lead to cancer, prevention may be a more feasible strategy. Vaccines, chemo preventive compounds, and healthier lifestyle choices are our arms in the battle against this deadly disease. In this chapter we discuss the importance of cancer prevention, how chemoprevention can be our first line of defense, and consider the role of small molecules and vaccines in cancer prevention and therapy. Deciphering the role of early disease detection and understanding how biomarkers and epigenetics can be a tool against cancer is

vital. Finally, tackling the causes of cancer is critical for eradicating this malignancy.

The process of carcinogenesis is extremely slow, offering ample opportunity for intervention and prevention. A mutation in the genome may lead to transformation to a precancerous lesion and eventually to cancer with unchecked cell growth. An untold number of genetic changes can trigger cells to become cancerous. Predicting the changes and the susceptible

Mutations, whether acquired or inherited and caused by endogenous and exogenous agents, result in oncogenic transformation. In the human genome, there are many different types of genes that control cell growth in a very systematic, precise way. Error in these genes leads to further alterations or mutations. An accumulation of many mutations in different genes occurring in a specific group of cells over time is required to cause malignancy. In general, mutations in two classes of genes, proto-oncogenes and tumor suppressor genes, lead to cancer.

Proto-oncogenes are typically responsible for promoting cell growth but alterations lead to transformation into oncogenes and promotion of tumor growth. Mutations in these genes are typically dominant in nature. Often, proto-oncogenes encode proteins that function to stimulate cell division, inhibit cell differentiation, and halt cell death. Oncogenes, however, typically exhibit increased production of these proteins, thus leading to increased cell division, decreased cell differentiation, and inhibition of cell death. These phenotypes typify cancer cells. Underlying genetic mechanisms associated with oncogene activation include point mutations, deletions, or insertions that lead to a hyperactive gene product. Other examples include alterations in the promoter region of a proto-oncogene that lead to

**1. Introduction** 

population is even more daunting.

**1.1 Proto-oncogenes** 

Ziech, D., Franco, R., Georgakilas, A. G., Georgakila, S., Malamou-Mitsi, V., Schoneveld, O., Pappa, A. & Panayiotidis, M. I. (2010) Chem. *Biol. Interact.* 188: 334-339. **3** 

Somaira Nowsheen, Alexandros G. Georgakilas and Eddy S. Yang *University of Alabama at Birmingham USA* 

### **1. Introduction**

62 Cancer Prevention – From Mechanisms to Translational Benefits

Ziech, D., Franco, R., Georgakilas, A. G., Georgakila, S., Malamou-Mitsi, V., Schoneveld, O.,

Despite decades of research, cancer continues to affect millions of people each year. However, the more we discover about cancer, the more we realize that no single therapeutic strategy can effectively treat it. As we learn about the aberrant signals and pathways which lead to cancer, prevention may be a more feasible strategy. Vaccines, chemo preventive compounds, and healthier lifestyle choices are our arms in the battle against this deadly disease. In this chapter we discuss the importance of cancer prevention, how chemoprevention can be our first line of defense, and consider the role of small molecules and vaccines in cancer prevention and therapy. Deciphering the role of early disease detection and understanding how biomarkers and epigenetics can be a tool against cancer is vital. Finally, tackling the causes of cancer is critical for eradicating this malignancy.

The process of carcinogenesis is extremely slow, offering ample opportunity for intervention and prevention. A mutation in the genome may lead to transformation to a precancerous lesion and eventually to cancer with unchecked cell growth. An untold number of genetic changes can trigger cells to become cancerous. Predicting the changes and the susceptible population is even more daunting.

Mutations, whether acquired or inherited and caused by endogenous and exogenous agents, result in oncogenic transformation. In the human genome, there are many different types of genes that control cell growth in a very systematic, precise way. Error in these genes leads to further alterations or mutations. An accumulation of many mutations in different genes occurring in a specific group of cells over time is required to cause malignancy. In general, mutations in two classes of genes, proto-oncogenes and tumor suppressor genes, lead to cancer.

#### **1.1 Proto-oncogenes**

Proto-oncogenes are typically responsible for promoting cell growth but alterations lead to transformation into oncogenes and promotion of tumor growth. Mutations in these genes are typically dominant in nature. Often, proto-oncogenes encode proteins that function to stimulate cell division, inhibit cell differentiation, and halt cell death. Oncogenes, however, typically exhibit increased production of these proteins, thus leading to increased cell division, decreased cell differentiation, and inhibition of cell death. These phenotypes typify cancer cells. Underlying genetic mechanisms associated with oncogene activation include point mutations, deletions, or insertions that lead to a hyperactive gene product. Other examples include alterations in the promoter region of a proto-oncogene that lead to

An alternate theory is Dr. Alfred Knudson's two-hit theory of cancer causation. This model accounts for both hereditary and non-hereditary cancer. Normal cells have two undamaged chromosomes, one from each parent, containing thousands of genes. People with a hereditary susceptibility to cancer inherit a damaged gene on one of the chromosomes. Thus, their first hit or mutation occurs at conception. Others receive the first hit in their lifetime. A subsequent damage to the same gene on the second chromosome may lead to cancer. Therefore, people with a hereditary susceptibility to cancer just need one hit during their lifetime to produce cancer. An overview of this model is given in **Figure 2**. This model is applicable for cancer such as retinoblastoma where inheritance of the first hit leads to a far

Fig. 1. The cascade of events that leads to colon carcinogenesis.

greater chance of developing a second cancer causing mutation.

Fig. 2. The two-hit model of carcinogenesis.

increased transcription. Gene amplification events leading to extra chromosomal copies of a proto-oncogene may also lead to oncogenesis. Chromosomal translocation events that relocate a proto-oncogene to a new chromosomal site that leads to higher expression or lead to a fusion between a proto-oncogene and a second gene, which produces a fusion protein with oncogenic activity may lead to cancer as well.

#### **1.2 Tumor suppressors**

Tumor suppressor genes are also present in our cells to control cell growth and apoptosis. Exquisite control over these processes suppresses tumor development. Mutations in tumor suppressors, as mentioned above, can lead to carcinogenesis. Tumor suppressors are in place to oppose threats to the genome. p53, the guardian of the genome, is one of the most commonly mutated tumor suppressor genes in human cancer. p53, a transcription factor, plays a critical role in numerous signaling pathways, from development to maintaining genomic stability and cell death (Brosh and Rotter 2009). Mutant p53 has been shown to exhibit gain–of–function properties that drive tumor progression and metastasis (Brosh and Rotter 2009). p53 is a stress response protein that functions primarily as a tetrameric transcription factor which regulates a large number of genes in response to a variety of cellular insults, including oncogene activation and DNA damage. These signals activate p53 primarily through post-translational modifications that result in augmented p53 protein level and transactivation activity. Activated p53 suppresses cellular transformation mainly by inducing growth arrest, apoptosis, DNA repair and differentiation in damaged cells (Oren 2003). Not surprisingly, p53 function is almost always compromised in tumor cells. Mutations in p53, usually due to somatic mutations, are observed in approximately half of all human cancers and constitute a cornerstone in tumorigenesis (Hollstein et al. 1991, Vogelstein, Lane and Levine 2000).

#### **1.3 Models of carcinogenesis**

There are several models of carcinogenesis. One of the models proposed by Dr. Bert Vogelstein proposes the loss of function of tumor suppressors such as p53 which paves the way for genomic instability, changes in metabolism, insensitivity to apoptotic signals, invasiveness and motility. However, the nature of the causal link between early tumorigenic events and the induction of the p53-mediated checkpoints that constitute a barrier to tumor progression remains uncertain. Loss of p53 function occurs during the development of most, if not all, tumor types. The cascade of events starts with a mutation that inactivates tumor suppressor gene leading to hyper-proliferation of epithelial cells. The mutation may also inactivate DNA repair genes while mutation of proto-oncogene creates an oncogene. The same mutation may lead to a cascade of inactivation of several more tumor suppressor genes before resulting in cancer. For example in colon carcinogenesis, loss or mutation of APC gene leads to overexpression on cyclooxygenase (COX) genes, transforming the normal tissue to hyperproliferative epithelium, and resulting in early adenoma. Subsequent DNA hypomethylation leads to mutations such as in k-ras gene, and results in intermediate adenoma. Another mutation following this, such as loss or mutation of DCC or SMAD 4, results in late adenoma. Subsequent mutation in p53 leads to carcinogenesis. Further mutations result in metastasis and greater genomic instability. It is thus quite apparent that the perturbations necessary to form cancer are numerous and complex. **Figure 1** gives an overview of this model of carcinogenesis.

increased transcription. Gene amplification events leading to extra chromosomal copies of a proto-oncogene may also lead to oncogenesis. Chromosomal translocation events that relocate a proto-oncogene to a new chromosomal site that leads to higher expression or lead to a fusion between a proto-oncogene and a second gene, which produces a fusion protein

Tumor suppressor genes are also present in our cells to control cell growth and apoptosis. Exquisite control over these processes suppresses tumor development. Mutations in tumor suppressors, as mentioned above, can lead to carcinogenesis. Tumor suppressors are in place to oppose threats to the genome. p53, the guardian of the genome, is one of the most commonly mutated tumor suppressor genes in human cancer. p53, a transcription factor, plays a critical role in numerous signaling pathways, from development to maintaining genomic stability and cell death (Brosh and Rotter 2009). Mutant p53 has been shown to exhibit gain–of–function properties that drive tumor progression and metastasis (Brosh and Rotter 2009). p53 is a stress response protein that functions primarily as a tetrameric transcription factor which regulates a large number of genes in response to a variety of cellular insults, including oncogene activation and DNA damage. These signals activate p53 primarily through post-translational modifications that result in augmented p53 protein level and transactivation activity. Activated p53 suppresses cellular transformation mainly by inducing growth arrest, apoptosis, DNA repair and differentiation in damaged cells (Oren 2003). Not surprisingly, p53 function is almost always compromised in tumor cells. Mutations in p53, usually due to somatic mutations, are observed in approximately half of all human cancers and constitute a cornerstone in tumorigenesis (Hollstein et al. 1991,

There are several models of carcinogenesis. One of the models proposed by Dr. Bert Vogelstein proposes the loss of function of tumor suppressors such as p53 which paves the way for genomic instability, changes in metabolism, insensitivity to apoptotic signals, invasiveness and motility. However, the nature of the causal link between early tumorigenic events and the induction of the p53-mediated checkpoints that constitute a barrier to tumor progression remains uncertain. Loss of p53 function occurs during the development of most, if not all, tumor types. The cascade of events starts with a mutation that inactivates tumor suppressor gene leading to hyper-proliferation of epithelial cells. The mutation may also inactivate DNA repair genes while mutation of proto-oncogene creates an oncogene. The same mutation may lead to a cascade of inactivation of several more tumor suppressor genes before resulting in cancer. For example in colon carcinogenesis, loss or mutation of APC gene leads to overexpression on cyclooxygenase (COX) genes, transforming the normal tissue to hyperproliferative epithelium, and resulting in early adenoma. Subsequent DNA hypomethylation leads to mutations such as in k-ras gene, and results in intermediate adenoma. Another mutation following this, such as loss or mutation of DCC or SMAD 4, results in late adenoma. Subsequent mutation in p53 leads to carcinogenesis. Further mutations result in metastasis and greater genomic instability. It is thus quite apparent that the perturbations necessary to form cancer are numerous and complex. **Figure 1** gives an

with oncogenic activity may lead to cancer as well.

**1.2 Tumor suppressors** 

Vogelstein, Lane and Levine 2000).

overview of this model of carcinogenesis.

**1.3 Models of carcinogenesis** 

Fig. 1. The cascade of events that leads to colon carcinogenesis.

An alternate theory is Dr. Alfred Knudson's two-hit theory of cancer causation. This model accounts for both hereditary and non-hereditary cancer. Normal cells have two undamaged chromosomes, one from each parent, containing thousands of genes. People with a hereditary susceptibility to cancer inherit a damaged gene on one of the chromosomes. Thus, their first hit or mutation occurs at conception. Others receive the first hit in their lifetime. A subsequent damage to the same gene on the second chromosome may lead to cancer. Therefore, people with a hereditary susceptibility to cancer just need one hit during their lifetime to produce cancer. An overview of this model is given in **Figure 2**. This model is applicable for cancer such as retinoblastoma where inheritance of the first hit leads to a far greater chance of developing a second cancer causing mutation.

Fig. 2. The two-hit model of carcinogenesis.

Whereas some lesions are subject to direct protein-mediated reversal, most are repaired by a cascade of catalytic events mediated by multiple proteins. In MMR-mediated repair, detection of mismatches and insertion/deletion loops triggers a single-strand incision that is then worked upon by nuclease, polymerase and ligase enzymes. In BER-mediated repair, a damaged base is often recognized by a DNA glycosylase enzyme that mediates base removal before nuclease, polymerase and ligase proteins complete the repair in processes overlapping with those used in single strand break repair. In contrast, NER-mediated repair recognizes helix-distorting base lesions. It includes two sub-pathways that differ in the mechanism of lesion recognition: transcription-coupled NER, which specifically targets lesions that block transcription, and global-genome NER. A key aspect of NER is that the damage is excised as a 22–30-base oligonucleotide, producing single-stranded DNA that is

acted upon by DNA polymerases and associated factors before ligation proceeds.

cycle.

2009, Jiang et al. 2011, Wang et al. 2010).

**2.2.1 Cell cycle checkpoints** 

In NHEJ, DSBs are recognized by the Ku protein that then binds and activates the protein kinase DNA-PKcs, leading to recruitment and activation of end-processing enzymes, polymerases and DNA ligase IV. NHEJ repair, predominantly utilized in the repair of radiation induced DNA damage, is a highly efficient but error-prone process that often results in mutations in the repaired DNA. The NHEJ repair process is dependent on the DNA-dependent protein kinase (DNA-PK) catalytic subunit (DNA-PKcs), the Ku70/Ku80 heterodimer, and the XRCC4–ligase IV complex and ultimately rejoins the ends of DSBs with little or no homology. In response to radiation, DNA-PKcs is autophosphorylated at threonine 2609. This is required for the functional activation of the NHEJ repair pathway. Consistent with the role of NHEJ repair in the repair of radiation-induced DSBs, cells deficient in any NHEJ repair protein have been shown to be hypersensitive to radiationmediated cytotoxicity (Iliakis et al. 2004, Yang et al. 2009, van Gent, Hoeijmakers and Kanaar 2001). A less-well-characterized Ku-independent NHEJ pathway, called micro-homologymediated end-joining (MMEJ) or alternative end-joining, results in sequence deletions. Although both NHEJ and MMEJ are error-prone, they can operate in any phase of the cell

In contrast, HR is generally restricted to S and G2 because it uses sister-chromatid sequences as the template to mediate faithful repair. Although there are several HR sub-pathways, HR is always initiated by single strand DNA generation, which is promoted by various proteins including the MRE11–RAD50–NBS1 (MRN) complex. In events catalyzed by RAD51 and the breast-cancer susceptibility proteins BRCA1 and BRCA2, the single strand DNA then invades the undamaged template and, following the actions of polymerases, nucleases, helicases and other components, DNA ligation and substrate resolution occur. HR is also used to restart stalled replication forks and to repair inter-strand DNA crosslinks, the repair of which also involves the Fanconi anaemia protein complex. This high-fidelity, error-free process is also critical in the repair of lesions resulting from replicative stress (Yang et al.

Checkpoints are also put in place throughout the cell cycle that halt further progression of DNA replication and cell division upon detection of damaged DNA. This can arrest the cell either transiently or permanently (senescence), as well as activate specific DNA repair pathways in response to certain types of DNA damage. Some of the proteins in these
