Acute Myelogenous Leukemia and Myelodysplasia Secondary to Breast Cancer Treatment: Case Studies and Literature Review

CLINICAL INVESTIGATION

Acute Myelogenous Leukemia and Myelodysplasia Secondary to Breast Cancer Treatment: Case Studies and Literature Review Marion Cole, MD and Roger Strair, MD, PhD

Abstract: Background and Purpose: Chemotherapy and radiation therapy for breast cancer are known to increase the risk of developing a myelodysplastic syndrome (MDS) and/or acute myelogenous leukemia (AML). Alkylating agents and topoisomerase II inhibitors, fundamental to the treatment of breast cancer, are the most likely contributors to this increase in risk. Radiation therapy adds to the risk, and there is speculation that granulocyte colony-stimulating factor (G-CSF) may also predispose to leukemia. The purpose of this systemic review is to bring to the attention of family physicians the unintended consequence of leukemia secondary to aggressively treated breast cancer. Methods: The medical records of several patients from Robert Wood Johnson University Hospital, with previously treated breast cancer admitted for therapy for AML or myelodysplasia, were reviewed. In addition, the recent literature on this topic was reviewed. Results: Cases of patients whose AML was likely secondary to their treatment for breast cancer were used to illustrate the role of chemotherapy, radiation therapy, and perhaps G-CSF in the development of leukemia. Conclusions: Chemotherapy and radiation therapy administered for breast cancer predispose patients to the development of MDS or AML. We hypothesize that the breast cancer (BRCA) gene mutations might add to the risk and that primary care physicians must be aware of the long-term risks of cytotoxic therapy, including the development of MDS or AML. Key Indexing Terms: Breast cancer; AML; MDS. [Am J Med Sci 2010;339(1):36–40.]

W

omen treated for breast cancer have a 3.5-fold increased risk of developing a myelodysplastic syndrome (MDS) or acute myelogenous leukemia (AML).1 Symptoms of AML or MDS (a common predecessor to AML) often include fatigue or easy bruising caused by anemia along with thrombocytopenia. Diagnosis of AML or MDS is determined through examination of peripheral blood and the bone marrow. When abnormal white blood cells with the characteristic appearance of myeloblasts (or blasts, immature white blood cells) predominate the blood smear, the diagnosis of AML is made readily. However, in many cases, review of the blood will only reveal cytopenias (anemia and thrombocytopenia); rare blasts; or more differentiated white blood cells with features of abnormal maturation such as a lack of granules. In these cases, review of a bone marrow aspirate and biopsy will be necessary to make the diagnosis of MDS or AML. Abnormalities of white blood cell, red blood cell, and platelet formation are more apparent in review of the bone marrow and are used to characterize the From the University of Medicine and Dentistry of New Jersey, New Brunswick, New Jersey. Submitted June 6, 2009; accepted in revised form August 26, 2009. Presented at Resident/Fellow Research Day on May 7, 2008 at Robert Wood Johnson University Hospital, New Brunswick, NJ. Correspondence: Marion Cole, MD, Internal Medicine Residency Program, Department of Medicine, One Robert Wood Johnson Place, MEB 486, PO Box 19, New Brunswick, NJ 08903-0019 (E-mail: marionwomack@ hotmail.com).

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disorder (see below). Of the many chemotherapeutic agents now used to treat breast cancer, agents known to predispose to MDS or AML include alkylating agents, such as ifosfamide, melphalan, and cyclophosphamide; topoisomerase II inhibitors, such as anthracyclines (including doxorubicin and mitoxantrone); and epipodophyllotoxins, such as etoposide. Some studies have also shown that radiotherapy can also be a significant risk factor for leukemia. Other treatments such as granulocyte colony-stimulating factor (G-CSF) may also predispose to leukemia, although the evidence to date is not conclusive. Case Summaries Six patients with AML or MDS after therapy for breast cancer were recently admitted to the Leukemia Service at Robert Wood Johnson University Hospital. The cases are summarized in Table 1. As can be seen, chemotherapy was administered before the development of AML in all of the cases. Six patients received adjuvant chemotherapy for breast cancer and 2 were treated for ovarian cancer previously. The time from chemotherapy to development of MDS or AML ranged from 1 to 7 years. All patients presented to their primary care physicians with symptoms such as fatigue, bruising, or fever and were found to have abnormal hemograms that prompted evaluation by their oncologist. A subsequent bone marrow aspirate and biopsy diagnosed MDS or AML, and the patient was referred for management. Three patients had BRCA mutations (Table 1). Chemotherapy and Radiation Therapy Adjuvant chemotherapy and radiotherapy with or without hormonal treatment extend the disease-free and overall survival of patients with breast cancer.2 Alkylating agents (eg, cyclophosphamide) and topoisomerase II inhibitors (eg, doxorubicin, mitoxantrone, or etoposide) are some of the oldest and most potent chemotherapeutic agents used to treat patients with cancer and were used to treat all the patients listed in Table 1 who were treated with adjuvant therapy. Alkylating agents such as cyclophosphamide produce covalent bonding of alkyl groups to biological molecules. When alkylating agents bind DNA, they cause strand breaking and cross-linking. Topoisomerase II inhibitors such as doxorubicin or mitoxantrone prevent enzyme-mediated unwinding of DNA, resulting in the accumulation of DNA that has been cleaved but not religated. They also form free radicals, which may result in further DNA strand breaks. Radiation therapy also alters DNA (and other biological molecules) with directed ionizing energy, resulting in DNA breaks and other cellular damage. Although the treatmentinduced damage results in cancer cytotoxicity, the damage may also result in genetic alterations that have the unintended effect of predisposing patients to MDS and AML. Myelodysplasia and AML MDSs are heterogeneous hematopoietic stem cell disorders that are characterized by ineffective hematopoiesis, vari-

The American Journal of the Medical Sciences • Volume 339, Number 1, January 2010

AML, MDS, and Breast Cancer Treatment

TABLE 1. Patients With t-MDS or t-AML After Treatment for Breast Cancer Age

Stage of Breast Cancer

Initial Treatment

37/F

Stage III

Mastectomy

65/F

Stage II

Lumpectomy and radiation

70/F

Stage II

Lumpectomy and radiation

55/Fa

Stage IVb

Bilateral mastectomy

54/Fa

Stage III

Bilateral mastectomy

78/Fa

Stage I DCISb

Mastectomy

Adjuvant Treatment

Time From Chemotherapy to AML

Type of MDS-AML

Cyclophosphamide Doxorubicin Paclitaxel Capecitabine Cyclophosphamide Doxorubicin Paclitaxel Cyclophosphamide Doxorubicin Paclitaxel Letrozole Tamoxifen Cyclophosphamide Methotrexate Fluorouracil Doxorubicin Cyclophosphamide Doxorubicin Paclitaxel Herceptin None

2 yr

FAB M4—multilineage dysplasia—normal cytogenetics

1 yr

FAB M4 with eosinophilia— inversion chromosome 16

2 yr

FAB M4—normal cytogenetics

7 yr

MDS RAEB 2—loss of chromosomes 5, 7, 9, and 21⫹ trisomy 8

2 yr

FAB M4 with eosinophilia— inversion of chromosome 16

1 yr

FAB M4—multilineage dysplasia with additions of chromosomes 5, 6, and 11 and deletion of chromosome 12

a

Patients with BRCA gene mutations. Patients also treated with paclitaxel and carboplatin for ovarian cancer. MDS, myelodysplastic syndrome; AML, acute myeloid leukemia; FAB, French-American-British; RAEB, refractory anemia with excess blasts.

b

ous degrees of cytopenia, and dysplastic features in peripheral blood and marrow cells.3 There is often a high level of apoptosis in bone marrow cells of patients with MDS, contributing to the ineffective hematopoiesis and resulting cytopenias. They are clonal disorders, although several clones may coexist in a given patient. Given the clonality, abnormal cellular differentiation and tendency for evolution, including the development of AML, they are considered malignant myeloid disorders. The modern classification of MDS incorporates morphologic, cytogenetic, and laboratory parameters. The French-American-British classification defined 5 different subsets: refractory anemia (RA), RA with ringed sideroblasts, RA with excess blasts (RAEB), RAEB in transformation (RAEBt), and chronic myelomonocytic leukemia.4 Patients whose blood and bone marrow findings meet the French-American-British criteria of RAEB (bone marrow blasts 5%–20%) or RAEBt (bone marrow blasts 20%–29%) have a high frequency of developing AML (50%–100%). A more recent classification by the World Health Organization (WHO) dropped RAEBt as a distinct subset because it did not offer extra diagnostic or prognostic information, often behaving similar to AML.5 Patients with ⬎20% blasts are now considered to have AML. The WHO classification of myeloid neoplasms also recognized therapy-related acute myeloid leukemias (t-AML) and MDSs (t-MDS) as a distinct subclass of myeloid neoplasms.5 Furthermore, the WHO subdivided these treatmentrelated myeloid malignancies based on the presumed causative therapy: alkylating agent-radiation type and topoisomerase II inhibitor-related type. These 2 subtypes are recognized because © 2010 Lippincott Williams & Wilkins

of distinct biological and clinical features. Alkylating agentradiation-related t-MDS/t-AML usually is diagnosed 3 to 7 years after the causative treatment.6,7 Most patients present with either MDS or AML with multilineage dysplasia and abnormalities of chromosomes 5 and/or 7 are often detected on cytogenetic analysis. In contrast, topoisomerase II inhibitorrelated myeloid malignancies have a shorter latency period and generally present as AML without an antecedent MDS phase.8 The leukemic cells from these patients often have abnormalities/translocations of chromosome 11q23, 21q22, or others that are sometimes seen in de novo AML. These mutations seem to modulate the transcription of key genes that regulate myeloid differentiation. The specific genetic alterations often result in fusion transcription factors or altered protein complexes that modify the state of chromatin, resulting in suppressed expression of key genes. Rarely, acute lymphocytic leukemia with abnormalities of chromosome 11q23 may arise secondary to treatment with topoisomerase II inhibitors. The presence of abnormalities of chromosome 11q23 in patients with congenital leukemia has led to conjecture concerning possible in utero exposure to topoisomerase II inhibitor-like agents as an etiologic factor for infant leukemia.9 t-MDS/t-AML is the first secondary malignancy recognized as a long-term complication of primary cancer therapy. It was initially described in survivors of Hodgkin lymphoma who were treated with alkylating agents with or without radiation therapy. In several retrospective studies, the incidence of t-MDS/t-AML was 0.3% to 10% in patients who were treated for Hodgkin disease with an alkylating agent-based regimen

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with or without radiation therapy.10,11 In addition to Hodgkin lymphoma and breast cancer, t-MDS/t-AML has also been reported with the treatment of a wide spectrum of other diseases, including solid and hematologic malignancies, myeloproliferative disorders (eg, polycythemia vera and essential thrombocytosis); and nonmalignant conditions, including rheumatoid arthritis, ankylosing spondylitis, psoriasis, and others. A common feature is therapy with alkylating agents, topoisomerase II inhibitors, or radiation. MDS and AML After Chemotherapy and Radiation Therapy There are many examples of t-MDS/t-AML after defined courses of chemotherapy. A recent follow-up of patients treated for early-stage breast cancer on National Surgical Adjuvant Breast and Bowel Project clinical trials with doxorubicin and cyclophosphamide confirmed an increased incidence of acute leukemia.12 In these women, the latency period from the beginning of the adjuvant therapy to the development of MDS/ AML was 10 to 125 months. More intense regimens of adjuvant chemotherapy were associated with a higher incidence of MDS/AML. Patients treated with the highest doses of cyclophosphamide (in combination with doxorubicin) had an MDS/ AML incidence of ⬃1%, which further increased in patients receiving radiation therapy. Patients receiving standard dose cyclophosphamide (in combination with doxorubicin) had an incidence of ⬃0.2%. Of note, patients treated with the highest doses of cyclophosphamide also received G-CSF, resulting in speculation that the cytokine may contribute to development of t-MDS/t-AML. As opposed to potential carcinogenic mechanisms associated with the cytotoxicity and mutagenicity of alkylating agents and topoisomerase inhibitors, G-CSF, a cytokine used to diminish the intensity and duration of neutropenia, is linked to the Jak/STAT, p21Ras/MAP kinase, and PI-3K/PKB pathways by way of the G-CSF receptor. Parts of these pathways have all been associated with hematopoietic malignancies, and stimulation of mutated cells may foster selection of damaged cells. Thus, G-CSF has been studied as a potential risk factor for secondary AML.13,14 Some studies have suggested that the risk for MDS/AML was increased in women 65 years or older treated with adjuvant chemotherapy and growth factor (versus adjuvant therapy without growth factor) for stage I–III breast cancer.15 Of the 906 patients reported who had stage I–III breast cancer and received at least one dose of G-CSF, 16 (1.77%) developed secondary AML, whereas 48 (1.04%) of the 4604 patients with stage I–III breast cancer who did not receive G-CSF developed secondary AML.15 Results of this study might have been influenced by the fact that in older women, there could be more susceptibility to t-MDS/t-AML than in younger women. Furthermore, indications of use of G-CSF, the cumulative dose and duration of G-CSF therapy, and the dose of chemotherapy received by the patients were not reported. Mechanisms Underlying the Development of MDS and AML Induction of mutations by chemotherapeutic agents or radiation has been demonstrated and presumably underlies the development of t-MDS/t-AML. Some studies associate polymorphisms in drug metabolism or cellular repair mechanisms with the development of or protection from t-MDS/t-AML.16 Of note, 3 of the patients reported in Table 1 had germline mutations in BRCA 1 or BRCA 2, genes that encode proteins that are essential components of a large pathway involved in error-free repair of DNA double-strand breaks. Thus, BRCA

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mutations may lead to DNA repair that is less effective and more error prone. With a more error-tolerant pathway, there may be a greater likelihood of developing genetic alterations that result in t-MDS or t-AML. In fact, 32% of primary AML samples and 75% of t-AML samples were reported to have reduced expression of the BRCA 1 gene, with hypermethylation of the promoter detected in some patient samples.17,18 In addition, BRCA 2 is one of the genes mutated in Fanconi anemia, a disease with a marked predisposition to the early development of AML.17 The distribution of drug-induced mutations may be semi-random, and disease induction may occur in only a subset of patients whose susceptible hematopoietic target cells by chance accumulate the aggregate alterations that give rise to the malignancy. Cells that give rise to t-MDS or t-AML must be capable of self-renewal; hence, the target cells have characteristics of hematopoietic stem cells. In some cases, selective pressure imposed by myelosuppressive therapy may also contribute to the expansion of mutated hematopoietic cells that have a mutation-induced survival advantage. This is best demonstrated by the microsatellite instability present in AML cells obtained from 7 of 7 patients with azathioprine-associated secondary AML developing after solid-organ transplantation.19 Defects in DNA mismatch repair diminish thiopurine-induced toxicity and are selected by continuous exposure of cells to thiopurine. The resulting defective DNA mismatch repair results in microsatellite instability, an increased mutation load, and leukemia. Microsatellite instability is only rarely detected in de novo AML.20 Many pathways have been implicated in the development of MDS and AML. Although the pathways are the same in both de novo MDS and AML and secondary MDS and AML, the frequencies with which they are observed in the 2 groups are different.21 Three types of genetic mutations are proposed to be involved in MDS and AML. Class I mutations are those that activate genes in the tyrosine kinase-RAS/BRAF transduction pathway, resulting in increased cell proliferation. Class II mutations are those that inactivate genes encoding hematopoietic transcription factors, thus disturbing cell differentiation. Class III mutations are those that inactivate tumor suppressor gene p53. Fourteen or more different genes have been found to be mutated in secondary MDS and AML, and these are involved in 8 different proposed genetic pathways. The most common genetic defect observed in secondary MDS and AML after therapy containing an alkylating agent includes multiple unbalanced aberrations and loss or gain of chromosomal material. Often, this entails loss of parts of the long arm or all of chromosome 5 or 7 (5q-/-5, 7q-/-7) as well as gain of chromosome 8 (⫹8). These specific defects are in 50% to 70% of secondary MDS cases and 40% to 50% of secondary AML cases and are most likely caused by previous treatment with alkylating agents. The second most common genetic defect observed in secondary AML involved recurrent balanced aberrations in chromosomal rearrangement without any loss or gain of genetic material. Most commonly, this defect involves reciprocal translocations involving rearrangement of transcription factor or chromatin remodeling genes, most notably MLL at 11q23, the AML1 at 21q22, RARA at 17q21, and CBFB at 16q22, leading to a disturbance in cellular differentiation. These specific defects are rarely seen in secondary MDS cases but are seen in 15% to 20% of secondary AML cases and are most likely caused by topoisomerase II inhibitors. Although a significant share of patients with secondary MDS and AML has a normal karyotype, the frequency Volume 339, Number 1, January 2010

AML, MDS, and Breast Cancer Treatment

of patients with a normal karyotype in secondary disease is far less than in de novo disease. Different mechanisms are involved in the development of genetic mutations depending on the karyotypes of patients with secondary AML.22 Those with gain or loss of chromosomal material often have mutations in the tumor suppressor gene p53 and activation of the transcription factor AML1.21 These patients often present with complex chromosomal rearrangements and have a poor prognosis. Other genetic mutations involved with this mechanism include activation of several tyrosine kinases, including FLT3, cKIT, cFMS, or JAK2 as well as genes further downstream in the RAS-BRAF-MEK-ERK signal transduction pathway, resulting in continuous activation of the cell cycle and cell proliferation. In summary, the chemotherapeutic combinations used in these patients react with biological molecules, inducing biochemical alterations of DNA, proteins, membranes, and other cellular components. These agents have little specificity for malignant cells. However, in normal cells, the damage that is done is detected by cellular molecular complexes that signal through p53 to result in cell cycle arrest or apoptosis depending on the nature of the damage. Arrested cells can then repair damage, allowing recovery from the chemotherapy. In contrast, malignant cells often lack the mechanisms to signal via p53associated pathways, resulting in less cell cycle arrest and further damage, including cytotoxicity. In addition, polymorphisms in drug metabolism or DNA may shift the balance and enhance mutagenesis.23 Furthermore, inherited mutations in BRCA genes are hypothesized to enhance mutagenesis by decreasing the fidelity of DNA repair. In some cases, specific bone marrow cells with the potential for self-renewal will accumulate chemotherapy-induced mutations that result in altered cell signaling, enhanced proliferation, diminished apoptosis, or abnormal differentiation. Key sequential mutations may develop and the resultant cell may proliferate abnormally and fail to differentiate, resulting in t-MDS or t-AML.

DISCUSSION In the cases presented above, no one agent seems to be solely responsible for causing secondary AML. Most patients received several agents implicated in t-MDS or t-AML. None of the women in the above cases had any other known environmental exposures; however, BRCA mutations were present in 3 of our patients. Although there is little available data, the presence of BRCA mutations in 3 of our patients emphasizes the need for formal studies to determine whether BRCA mutations predispose to t-MDS or t-AML. Hence, as proposed by others, it is conceivable that deficiency in expression of genes for BRCA 1 and 2 leave patients more vulnerable to adverse effects of chemotherapy and therefore at an increased risk for t-AML as a result of breast cancer treatment.17,18,24

CONCLUSION As highlighted by the cases above and what has been learned about the unintended consequences of certain treatments for breast cancer, physicians and patients should be cognizant of the risk for t-MDS and t-AML. Although all the women who received adjuvant chemotherapy for breast cancer had standard indications for treatment, the potential of these agents to result in t-MDS or t-AML must be discussed with the patient. Therapeutic development of newer and more targeted agents for patients with breast cancer might reduce this complication. The influence of BRCA mutations on the development of t-MDS and t-AML should be formally studied to © 2010 Lippincott Williams & Wilkins

determine whether there is incremental risk. If such a risk is detected, subsequent clinical trials will need to determine the optimum adjuvant therapy for such patients. Some promising antitumor results have recently been reported after using olaparib, a polyadenosine diphosphate polymerase inhibitor, in BRCA mutation carriers with breast, ovarian, and prostate cancer.25 Such molecularly targeted therapies for tumors of different origins but containing the same molecular defect may provide new therapeutic options. Other agents such as sulforaphane26 and diindolylmethane,27,28 found in cruciferous vegetables, including broccoli, have been shown to inhibit cancer cells through various mechanisms, including induction of phase II detoxification and histone deacetylase inhibition. Further studies might be useful in determining whether there is a role for these agents in pretreatment of high-risk patients with breast cancer. These have the potential to increase the cancer killing potential of current radiation and chemotherapy while reducing possible side effects, including the risk of a second malignancy. ACKNOWLEDGMENTS All authors had full access to all the data in the study and take responsibility for the integrity of the data and the accuracy of the data analysis. REFERENCES 1. Renella R, Verkooijen HM, Fioretta G, et al. Increased risk of acute myeloid leukaemia after treatment for breast cancer. Breast 2006;15: 614 –19. 2. Bedard P, Cardoso F. Recent advances in adjuvant systemic therapy for early-stage breast cancer. Ann Oncol 2008;19(suppl 5):v122–v127. 3. Nimer SD. Myelodysplastic syndromes. Blood 2008;111:4841–51. 4. List A. New approaches to the treatment of myelodysplasia. Oncologist 2002;7(suppl 1):39 – 49. 5. Harris NL, Jaffe ES, Diebold J, et al. World Health Organization classification of neoplastic diseases of the hematopoietic and lymphoid tissues: report of the Clinical Advisory Committee meeting-Airlie House, Virginia, November 1997. J Clin Oncol 1999;17:3835– 49. 6. Sahin FI, Yilmaz Z, Karakuçs S, et al. t(8;16) AML developed subsequent to breast cancer therapy. Hematology 2006;11:153–5. 7. Hortobagyi G. Adjuvant therapy for breast cancer. Annu Rev Med 2000;51:377–92. 8. Andersen MK, Christiansen DH, Jensen BA, et al. Therapy-related acute lymphoblastic leukaemia with MLL rearrangements following DNA topoisomerase II inhibitors, an increasing problem: report on two new cases and review of the literature since 1992. Br J Haematol 2001;114:539 – 43. 9. Spector LG, Xie Y, Robison LL, et al. Maternal diet and infant leukemia: the DNA topoisomerase II inhibitor hypothesis: a report from the children’s oncology group. Cancer Epidemiol Biomarkers Prev 2005;14:651–5. 10. Leone G, Mele L, Pulsoni A, et al. The incidence of secondary leukemias. Haematologica 1999;84:937– 45. 11. Leone G, Pagano L, Ben-Yehuda D, et al. Therapy-related leukemia and myelodysplasia: susceptibility and incidence. Haematologica 2007; 92:1389 –98. 12. Smith RE, Bryant J, DeCillis A, et al. Acute myeloid leukemia and myelodysplastic syndrome after doxorubicin-cyclophosphamide adjuvant therapy for operable breast cancer: the National Surgical Adjuvant Breast and Bowel Project Experience. J Clin Oncol 2003;21:1195–204. 13. Le Deley MC, Suzan F, Cutuli B, et al. Anthracyclines, mitoxantrone, radiotherapy, and granulocyte colony-stimulating factor: risk factors for leukemia and myelodysplastic syndrome after breast cancer. J Clin Oncol 2007;25:292–300.

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14. Patt DA, Duan Z, Fang S, et al. Acute myeloid leukemia after adjuvant breast cancer therapy in older women: understanding risk. J Clin Oncol 2007;25:3871– 6. 15. Hershman D, Neugut A, Jacobson J, et al. Acute myeloid leukemia or myelodysplastic syndrome following use of granulocyte colonystimulating factors during breast cancer adjuvant chemotherapy. J Natl Cancer Inst 2007;99:196 –205. 16. Valagussa P, Santoro A, Fossati-Bellani F, et al. Second acute leukemia and other malignancies following treatment for Hodgkin’s disease. J Clin Oncol 1986;4:830 –7. 17. Friedenson B. The BRCA 1⁄2 pathway prevents hematologic cancers in addition to breast and ovarian cancers. BMC Cancer 2007;7:152. 18. Scardocci A, Guidi F, D‘Alo’ F, et al. Reduced BRCA1 expression due to promoter hypermethylation in therapy related acute myeloid leukemia. Br J Cancer 2006;95:1108 –13. 19. Tester WJ, Kinsella TJ, Waller B, et al. Second malignant neoplasms complicating Hodgkin’s disease: the National Cancer Institute experience. J Clin Oncol 1984;2:762–9. 20. Lillington DM, Micallef IN, Carpenter E, et al. Detection of chromosomal abnormalities pre-high-dose treatment in patients with leukemia after treatment for non-Hodgkin’s lymphoma. J Clin Oncol 2001; 19:2472– 81. 21. Petersen-Bjergaard J, Andersen MS, Andersen MK. Genetic path-

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ways in the pathogenesis of therapy-related myelodysplasia and acute myeloid leukemia. Hematology 2007;1:392–7. 22. Larson R. Etiology and management of therapy-related myeloid leukemia. Hematology 2007;1:453–9. 23. Weinstein I. Cell culture studies on the mechanism of action of chemical carcinogens and tumor promoters. In: Carcinogenesis: a comprehensive survey: the role of chemicals and radiation in the etiology of cancer, Vol. 10. New York (NY): Raven Press; 1985. 24. Shih HA, Nathanson KL, Seal S, et al. BRCA1 and BRCA2 mutations in breast cancer families with multiple primary cancers. Clin Cancer Res 2000;6:4259 – 64. 25. Fong P, Boss D, Yap T, et al. Inhibition of poly(ADP-ribose) polymerase in tumors from BRCA mutation carriers. N Engl J Med 2009;361:1–12. 26. Gamet-Payrastre L. Signaling pathways and intracellular targets of sulforaphane mediating cell cycle arrest and apoptosis. Curr Cancer Drug Targets 2006;6:135– 45. 27. Wahidur Rahman KM, Ali S, Aboukameel A, et al. Inactivation of NF-kappaB by 3,3⬘-diindolylmethane contributes to increased apoptosis induced by chemotherapeutic agent in breast cancer cells. Mol Cancer Ther 2007;6:2757– 65. 28. Bhatnagar N, Li X, Chen Y, et al. 3,3⬘-diindolylmethane enhances the efficacy of butyrate in colon cancer prevention through down-regulation of survivin. Cancer Prev Res 2009;2:581.

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