Rethinkng the causes and prevention of breast cancer: a cross-disciplinary evolutionary framework

Rethinkng the causes and prevention of breast cancer: a cross-disciplinary evolutionary framework Paul W. Ewald Holly A Swain Ewald Department of Biology Program in Disease Evolution University of Louisville

DISCLOSURES I do not have anything to disclose.

Categories of disease causation genetic noninfectious environmental (diet, lifestyle, chemicals, radiation; mainly by inducing mutations) infectious

Cancers accepted as caused by pathogens during the last 3 decades Cancer Pathogen acceptance Affected populations Endemic Burkitts Lymphoma adult T-cell leukemia cervical Epstein Barr virus + Plasmodium falciparum Human T Lymphotropic Virus I Human Papilloma virus (HPV) 1980 New Guinea, tropical Africa and So.America 1980 Africa, Japan, Caribbean, So. America 1985 worldwide nasopharyngeal Epstein Barr virus (EBV) 1990 China, Inuit liver Hepatitis B & C viruses 1995 worldwide post-transplant EBV 1995 worldwide proliferative diseas Kaposi s sarcoma human herpesvirus 8 2000 Africa, Mediterranean, MSM MALT & gastric Helicobacter pylori 2000 worldwide stomach oropharyngeal HPV 2005 worldwide

Cancers accepted as caused by pathogens during the last 3 decades Cancer Pathogen acceptance Affected populations Merkel cell Merkel cell 2010 global polyomavirus Gastric EBV 2010 global Sporadic and AIDS- EBV 2015 global associated Burkitt s lymphoma Hodgkin s lymphoma EBV 2015 global Glioblastoma brain cancer CMV 2015-2020? global

Evolutionary processes central to oncogenesis Evolution of normal cells into cancer cells through oncogenic selection Evolution of cellular barriers to and restraints on cancer through natural selection Evolution of pathogen characteristics compromise cellular barriers through natural selection Evolutionary adaptation to ancient environments may make people vulnerable to oncogenesis in present environments

A generally accepted evolutionary explanation of breast cancer Focus: the high rates of breast cancer in economically Prosperous countries relative to hunter gatherers

A generally accepted evolutionary explanation of breast cancer Focus: the high rates of breast cancer in economically prosperous countries relative to hunter gatherers Argument: Higher breast cancer prevalence arises from a greater lifetime number of menstrual cycles in economically prosperous countries

A generally accepted evolutionary explanation of breast cancer Focus: the high rates of breast cancer in economically prosperous countries relative to hunter gatherers Argument: Higher breast cancer prevalence arises from a greater lifetime number of menstrual cycles in economically prosperous countries Logic: proliferation of cells during menstrual cycles - nudges cells toward uncontrolled proliferation - leads to accumulation of mutations

The hormone proliferation hypothesis of breast cancer Hunter-gatherers Americans Relative cancer risk by age 60 1 117 Age at menarche 16.1 12.5 Age at first birth 19.5 24 Live births/female 5.9 1.8 Age at menopause 47 50.5 ovulations per lifetime 160 450 Eaton et al. 1994. Quart. Rev. Biol. 69:353-367

The infection/immune suppression hypothesis Relative breast cancer risk by age 60 Proportional increase in cancer risk 117 : 1 Age at menarche 1.2 : 1 Age at first birth 1.3 : 1 Number of births 1.2 : 1 Age at menopause 1.5 : 1

Evolutionary processes central to oncogenesis Evolution of normal cells into cancer cells Evolution of cellular barriers to and restraints on cancer barrier: a process that blocks oncogenesis; e.g., cell cycle arrest restraint: a biochemical process that inhibits but does not prevent cancer; e.g., DNA proof reading Evolution of pathogen characteristics that compromise cellular barriers

Alternative hypotheses for carcinogenesis Conventional explanation genetic predisposition Infectious exacerbation genetic predisposition Infection as a primary cause genetic predisposition & infection mutation infection (sometimes) mutation mutation metastatic cancer metastatic cancer metastatic cancer

Evolutionary processes central to oncogenesis Evolution of normal cells into cancer cells Evolution of cellular barriers to and restraints on cancer Evolution of pathogen characteristics that compromise cellular barriers

Evolutionary focus: hosts evolve barriers to cancer viruses compromise barriers to persist viruses evolve to exploit hosts

Fundamental Barriers 1 to Cancer Barrier cell cycle arrest Apoptosis (=cell suicide) regulation of telomerase cell adhesion Effect blocks cellular proliferation destroys precancerous cells caps # of cell divisions inhibits metastasis 1 barriers must be compromised (or missing) for metastatic cancer to occur

Barriers to Cancer & Viral Persistence Barrier to cancer to persistence cell cycle arrest Apoptosis (=cell suicide) regulation of telomerase blocks cellular proliferation destroys precancerous cells caps # of cell divisions prevents vertical viral proliferation prevents viral survival caps total amount of vertical viral proliferation cell adhesion inhibits metastasis inhibits spread of viruses within body

Disruption of cellular barriers to cancer Disruption Pathogen Cancer Stimulation of proliferation Immortalization Apoptosis Epstein Barr virus Burkitt s; Nasopharyngeal; post-transplant; EBNA3C* inhibits and degrades prb; Hodgkin s; breast? EBNA=Epstein Barr Nuclear Antigen

Disruption of cellular barriers to cancer Disruption Pathogen Cancer Stimulation of proliferation Immortalization Apoptosis Epstein Barr virus Burkitt s, Nasopharyngeal post-transplant, Hodgkin s, breast? EBNA3C* inhibits and degrades prb; BRAF0 ups HER2/HER3 signaling cascades; LMP2 ups ras EBNA=Epstein Barr Nuclear Antigen BRAF0= an EBV latency protein HER=human epidermal growth factor receptor LMP=Latent Membrane Protein of Epstein Barr virus ras=protein that can enhance replication

Disruption of cellular barriers to cancer Disruption Pathogen Cancer Stimulation of proliferation Immortalization Apoptosis Epstein Barr virus Burkitt s, Nasopharyngeal post-transplant, Hodgkin s breast? EBNA3C inhibits and degrades prb; LMP2 ups ras & leads to methylation of PTEN promoter EBNA2 & LMP1 induce TERT1 expression and telomerase activation LMP2 ups antiapoptotic ras effects & Bcl; LMP2 leads to methylation of PTEN promoter; EBNA3C inhibits p53 activity EBNA=Epstein Barr Nuclear Antigen prb=retinoblastoma protein LMP=Latent Membrane Protein of Epstein Barr virus ras=protein that can enhance replication and enhance or suppress apoptosis TERT=telomerase reverse transcriptase (=active subunit of telomerase) PTEN=Phosphatase and TENsin homolog: restricts cell division & fosters apoptosis

Disruption of cellular barriers to cancer Pathogen Cancer Disruption cell cycle arrest Immortalization: telomerase rgltn Apoptosis Cell adhesion Epstein Barr virus Burkitt s, nasopharyngeal yes Human papilloma virus cervical, head & neck HTLV adult T-cell leukemia/ lymphoma Human herpes virus 8 Kaposi's sarcoma Hepatitis B virus liver

Disruption of cellular barriers to cancer Disruption Pathogen Cancer Stimulation of proliferation Immortalization Apoptosis Epstein Barr virus Burkitt s, Nasopharyngeal post-transplant, EBNA3C inhibits and degrades prb; BRAF0 ups HER2/HER3 LMP1 induces TERT expression and telomerase activation Hodgkin s, LMP2 ups Ras breast? EBNA=Epstein Barr Nuclear Antigen prb=retinoblastoma protein LMP=Latent Membrane Protein of Epstein Barr virus ras=protein that can enhance replication and apoptosis TERT=telomerase reverse transcriptase (=active subunit of telomerase)

Disruption of cellular barriers to cancer Disruption Pathogen Cancer Stimulation of proliferation Immortalization Apoptosis Epstein Barr virus Burkitt s, Nasopharyngeal post-transplant, Hodgkin s, breast? EBNA3C inhibits and degrades prb; BRAF0 ups HER2/HER3 signaling cascades LMP2 ups Ras LMP1 induces TERT1 expression and telomerase activation LMP2 ups antiapoptotoic Ras effects & Bcl EBNA=Epstein Barr Nuclear Antigen prb=retinoblastoma protein LMP=Latent Membrane Protein of Epstein Barr virus ras=protein that can enhance replication TERT=telomerase reverse transcriptase (=active subunit of telomerase) Bcl=inhibits apoptosis by preventing activation of caspases

Disruption of cellular barriers to cancer Pathogen Cancer Disruption cell cycle arrest Immortalization Apoptosis Cell adhesion Epstein Barr virus Human papilloma virus HTLV Human herpes virus 8 Hepatitis B virus Burkitt s, nasopharyngeal cervical, head & neck adult T-cell leukemia/ lymphoma Kaposi's sarcoma liver yes yes yes yes

Disruption of cellular barriers to cancer Pathogen Cancer Disruption cell cycle arrest Immortalization Apoptosis Cell adhesion Epstein Barr virus Human papilloma virus HTLV Human herpes virus 8 Burkitt s, nasopharyngeal cervical, head & neck adult T-cell leukemia/ lymphoma Kaposi's sarcoma yes yes yes yes yes yes yes yes yes yes yes yes yes yes yes yes Hepatitis B virus liver yes yes yes yes

Infectious agents and breast cancer Mouse Mammary Tumor Virus (MMTV) -Causes mammary tumors in Mus domesticus -In 30-45% of human breast cancer & < 5% normal tissue within M. domesticus s range (USA, Australia) [Wang et al 1995, Liu et al 2001, Etkind et al. 2000, Ford et al 2003, Levine et al 2004, Lawson et al 2006]

MMTV in Breast cancer Country Within M. domesticus s range? % of breast cancer positive for MMTV Tunisia yes 74% United States yes 36% Italy yes 38% Australia yes 42% Argentina yes 31% Vietnam no 1% Reference: Levine PH et al. Increasing evidence for a human breast carcinoma virus with geographic differences Cancer. 2004. 101:721-6

Infectious agents and breast cancer Epstein Barr Virus -US: EBV in 51% of BC; < 10% of normal adjacent tissue [Bonnet et al 1999] -France, Denmark, the Netherlands, France, Algiers & Tunisia: EBV in BC 30-40% > in adjacent tissues [Fina et al. 2001] -More strongly associated with breast cancer when florid EBV infections occur during adolescence [Yasui et al 2001]

Infectious agents and breast cancer Human papillomavirus -Oncogenic HPV in 15.9% of 107 breast cancers [Kroupis et al 2006] & 24.8% of 101 breast cancers [Damin et al 2004] -2008-2010: Confirmations in Japan, Mexico, Korea, Hungary, Syria & Australia -HPV (serotype 16) in breast cancer concurrent with cervical cancer --46% of 41 cases of when cervical cancer started first --0% of 9 cases when breast cancer started first [Hennig et al 1999] -Transmission route to breast is uncertain --self contamination from genital area? --genital to oral to breast sexual contact? --via blood or lymph?

Infectious agents and breast cancer Cytomegalovirus US: --CMV in 97% of BC (ductal carcinoma in situe and infiltrating ductal carcinoma); --CMV in 63% of normal breast tissue [Harkins et al. 2010]

Infectious agents and breast cancer Mouse Mammary Tumor Virus (MMTV) -Causes mammary tumors in Mus domesticus -In 30-45% of human breast cancer & < 5% normal tissue within M. domesticus s range (USA, Australia) [Wang et al 1995, Liu et al 2001, Etkind et al. 2000, Ford et al 2003, Levine et al 2004, Lawson et al 2006] -Breast cancer rates reduced by ~1/3 outside of M. domesticus s range (eastern Europe, northern & eastern Asia) [Stewart et al. 2000]

Breast cancer & rodent host range for MMTV Country Within Mus domesticus s range? Incidence Breast Cancer* Deaths China Viet Nam Japan Italy Switzerland Australia Canada Netherlands UK Sweden Denmark France United States no no no yes yes yes yes yes yes yes yes yes yes 18.7 15.6 32.7 74.4 81.7 83.2 84.3 86.7 87.2 87.8 88.7 91.9 101.1 5.5 8.3 18.9 19.8 18.4 21.1 27.5 24.3 17.3 27.8 21.5 19.0 *per 100,000 population; from Ferlay et al. GLOBOCAN 2002. Cancer Incidence, Mortality and Prevalence Worldwide. IARC CancerBase No. 5, version 2.0. IARCPress, Lyon, 2004

Infectious agents and breast cancer Mouse Mammary Tumor Virus (MMTV) -Causes mammary tumors in Mus domesticus -In 30-45% of human breast cancer & < 5% normal tissue within M. domesticus s range (USA, Australia) [Wang et al 1995, Liu et al 2001, Etkind et al. 2000, Ford et al 2003, Levine et al 2004, Lawson et al 2006] -Breast cancer rates reduced by ~1/3 outside of M. domesticus s range (eastern Europe, southern Asia) [Stewart et al. 2000] -Effects of MMTV on barriers to cancer --disregulates cell cycle arrest --inhibits apoptosis --inhibits cell adhesion

Infectious agents and breast cancer Bovine Leukemia Virus (BLVV) [Buehring et al. 2015] -retrovirus related to HTLV and MMTV -causes leukemia in cows -transmitted to humans in dairy milk -positivity in breast cancer patient 30% > in controls (59% vs 29%) -infects about half of the women in the US

Candidate viruses in breast cancer Virus # research groups % of BC explainable Epstein Barr 2 30-50% 1-10% Mouse (Human) Mammary Tumor Human papillomavirus 2 25-40% Virus:cell % several 15-40% 1-10% Bovine leukemia 1 30%?? Human cytomegalovirus 2 36%?

Does a low ratio of viral genome/tumor cell warrant rejection of infectious causation? Cancer Virus Viral/cell % Hodgkin s lymphoma Epstein Barr 1-10% glioblastoma cytomegalovirus 10%

The hormone proliferation hypothesis of breast cancer Hunter-gatherers Americans Relative cancer risk by age 60 1 117 Age at menarche 16.1 12.5 Age at first birth 19.5 24 Live births/female 5.9 1.8 Age at menopause 47 50.5 ovulations per lifetime 160 450 Eaton et al. 1994. Quart. Rev. Biol. 69:353-367

The infection/immune suppression hypothesis Infections are a primary causes of breast cancer Estrogen & progesterone exacerbate infectious causation through immune suppression

Infection/immune suppression hypothesis genetic predisposition & infection Exacerbation of infection Estrogen and progesterone suppress cell-mediated immunity mutation metastatic cancer

The infection/immune suppression hypothesis Relative cancer risk by age 60 Age at menarche Age at first birth Number of births Age at menopause Proportional increase in cancer risk Explanation 117 : 1 Differential exposure to pathogens according to age, geography, population isolation, & social norms 1.2 : 1 Earlier menarche associated with greater exposure to STIs 1.3 : 1 Delayed child bearing associated with greater exposure to STIs 1.2 : 1 Smaller families associated with greater exposure to STIs 1.5 : 1 Later menopause associated with greater exposure to STIs

Infection/mmune suppression hypothesis of breast cancer Association Pregnancy is associated with increased breast cancer Pregnancy is associated with receptor negative breast cancer Early pregnancies most associated with lower postmenopausal breast cancer Nulliparity is associated with increased breast cancer only after menopause Interpretation Estrogen & progesterone suppress immunity to infection during pregnancy Infection rather than hormones directly cause cancer Women who have kids early may have fewer lifetime sexual partners Nulliparous women may have more sexual partners over a lifetime Nuns reportedly have increased rates of breast cancer Hormones elevated during pregnancy cause increased cancer rates postpartum, after hormone levels decline? Increased rates are attributable to misdiagnosis of breast cancer among nuns with tuberculosis of breast Hormones open the door to infections that persist after hormone levels decline

Risk factors and disease causation: breast cancer genetic BRCA alleles noninfectious environmental Mutagens, hormones? breast cancer infectious Human Papillomavirus, Mouse Mammary Tumor Virus, Epstein Barr Virus, Cytomegalovirus

Candidate interventions if viruses are primary causes of breast cancer Virus % of bc explainable Intervention Epstein Barr 30-50% Vaccines? Antivirals? Mouse (Human) Mammary Tumor Human papillomavirus 25-40% Bovine leukemia 30% Human cytomegalovirus 15-40% Vaccine prevention of transmission via mice prevention of transmission via milk 36% Vaccines? Antivirals?