Gilbert S. Omenn, M.D., Ph.D.

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Ecogenetics, Evolutionary Biology, Genomics, and
Medicine
Gilbert S. Omenn, M.D., Ph.D. University of
Michigan Conference Darwin y Medicina Mexico
City, UNAM, 25 April, 2008
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15-16 February, 2001
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Its a New World

• New Biology---New Technology
• Genome Expression Microarrays
• Comparative Genomics
• Proteomics
• Bioinformatics, Computational Medicine, and
  Evolutionary Biology

Path to predictive, personalized,
preventive (P3) healthcare
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Break- through of the Year, 2005
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Medicine Needs EvolutionNesse, Stearns, Omenn
Science 24 Feb 2006

• Evolution a vibrant foundation for all biology
• Evolutionary thinking can help researchers and
  clinicians answer common questions
• o Persistence of genes predisposing to
  depression, high blood pressure, diabetes, and
  other diseases
• o Usefulness of cough, fever, diarrhea

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Multi- and Interdisciplinary Research will be
Required to Solve the Puzzle of Complex
Diseases and Conditions
Genes Behavior Diet/Nutrition Infectious
agents Environment Society ??? evolution
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Eco-Genetics

• Known inherited predispositions to
• Foods
• Food additives
• Infectious agents
• Drug chemicals
• Pesticides
• Inhaled pollutants
• Sensory stimuli
• Allergic sensitizing agents

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Eco-Genetics of Food, Nutrition, and Cultural
Sensitivity

• Anthropology and biology of lactase persistence
• Everybody needs milk
• Got Milk?

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Digestion of Milk from the Diet

• Nearly all mammals digest the sugars in Mothers
  milk until weaning.
• The enzyme lactase in the intestine does it.
• Lactase production usually drops 90 during the
  first four years of life in humans, though the
  rate varies widely.
• Certain human populations have a mutation (on
  chromosome 2) which results in persistence of
  lactase activity (gain of function).

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Lactose Intolerance by Group
Human Groups Percent Intolerant Allele Frequency
Swedish 2 0.14
Europeans in Australia 4 0.20
Swiss 10 0.32
American Caucasians 12 0.35
Finns 18 0.42
African Tutsi 20 0.45
African Fulani 23 0.48
American Blacks 75 0.87
Australian Aborigines 85 0.92
African Bantu 89 0.94
Chinese 93 0.96
Thais 98 0.99
American Indians 100 1.00
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Consequences

• Persistence of lactase in the intestine permits
  the unnatural continuation of milk ingestion
  throughout life, enhancing growth and
  development. One benefit is higher calcium
  intake, especially in northern climes where sun
  is insufficient to activate vitamin D.
• People without this mutation shut off lactase
  production and become lactose malabsorbers
  they may become lactose intolerant with
  symptoms of excess gas (hydrogen) and diarrhea
  upon eating lactose-containing dairy products.
• The Navajo used powdered milk supplies to paint
  their adobe homes.

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Why Does Loss of Lactase Lead to Symptoms?

• The lactose in milk remains undigested and
  unabsorbed through the intestinal wall into the
  bloodstream.
• Gut bacteria adapt to the abundance of lactose
  (compared with glucose, etc) and metabolize the
  lactose, generating lots of gas by fermentation.
  
• Gas can cause stomach cramps, bloating, diarrhea
  the osmotic pressure rises, reducing resorption
  of water from the colon a laxative effect.

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Are Lactose Maldigestion and Lactose Intolerance
Important in Mexico?

• Rosado et al, J Nutrition 1241052, 1994
• Children and adults (N926) from Sonora,
  Mexico, and Yucatan
• Milk consumption (in 1982) was 219/-22,
  128/-25, and 19/-1 gm/day
• Whole milk vs lactose-hydrolyzed milk
  major intolerance symptoms uncommon (0-11 of
  children, 7-17 of 13-60 y/o) maldigestion by
  hydrogen breath test, 2-43, highest in Yucatan.

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Lactose Malabsorption in Mexican-Americans

• Wotecki et al, Amer J Clin Nutr 30470,1977
• Prevalence 53 of Mexican-Americans and 15 of
  Anglo-Americans after 50 gm lactose load (glucose
  increase lt25 mg/dl).
• Mean consumption of milk and dairy products was
  same in lactose absorbers and malabsorbers in
  both groups, though GI symptoms after milk was
  60 vs 24.
• Some reduced milk-drinking due to symptoms
  others continued for laxative benefit!

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What is the Diagnosis? What is Normal?

• In Europe and North America, lactose intolerance
  is regarded as a clinical problem, a medical
  diagnosis. Those who have symptoms may not know
  the cause lactose intolerance actually accounts
  for many cases of irritable bowel syndrome.
• In populations where most people are lactose
  malabsorbers, cultural nutritional practices will
  dominate.

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Prevention of Symptoms

• Avoid milk
• Drink lactose-free milk (enzyme-treated)
• Lactose-reduced (L. acidophilus) milk, as in
  yogurt and in hard cheeses
• Lactase pills
• Gene therapy??
• Note Milk sold for pet cats remarkably,
  European cat breeds have a mutation similar to
  the human European mutation, whereas Asian breeds
  are particularly sensitive!

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When Did This Mutation Arise?

• Most likely
• 1. Separate events in Sweden and in the
  Arabian Peninsula about 4000 BC, converging as
  they spread, or
• 2. Single event in the Middle East about
  4500 BC, with radiation from there.
• 3. Plus a more recent mutation in the East
  African Tutsi.
• A small survival advantage over the 6000-8000
  years since goats, sheep, and cattle were
  domesticated can account for the population
  differences now observed.

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An Associated Polymorphism of Apolipoprotein
A-IV-2 (His360Gln)

• Apo A-IV is a 46 kD plasma glycoprotein
  synthesized in the intestine.
• His/Gln polymorphism in Caucasians at codon 360
  (chromosome 11) modifies milkfat absorption and
  chylomicron clearance.
• Worldwide distribution resembles lactase
  persistence across 29 population groups,
• r 0.94 (plt0.000001). Highest prevalence in
  Iceland probably carried into Europe by
  Vikings.
• Weinberg, Genetic Epi 17285, 1999

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Eco-Genetics

• Known inherited predispositions to
• Drug chemicals
• Pesticides
• Inhaled pollutants
• Foods
• Food additives
• Sensory stimuli
• Allergic sensitizing agents
• Infectious agents

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Variation in Susceptibility to Infections

• Malaria (P. falciparum type) children with
  sickle hemoglobin trait have greater survival.
• Malaria (P. vivax type) people lacking the
  Duffy (Fy) red blood cell antigen are resistant
  to this parasite, which enters the cell through
  this surface antigen.
• People lacking CCR5 receptors on lymphocytes are
  resistant to infection by HIV.

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Distinguishing between Host Defenses and
Infectious Agent Offenses

• Symptoms of host defenses include pain, nausea,
  vomiting, diarrhea, fatigue, anxiety
• Treatment of these symptoms may suppress
  defenses, promoting pathogen transmission and
  harming patients
• Shigella diarrhea faster recovery and fewer
  complications without anti-diarrheal meds.
• Cough suppressants raise risk of pneumonia
• Acute phase response to infection, trauma, or
  cancers both behavioral (fever) and physiological
  (sequestration of Fe, Zn). Counterpart in insects.

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Evolution and Persistence of Virulence

• Host-parasite relationships are greatly
  influenced by the rapidity of transmission.
• When transmission is rapid, pathogens can
  disregard many host variables.
• When transmission is slow, pathogens benefit by
  establishing a long-term balanced relationship
  with the host.
• Modeling shows vaccines which prevent infection
  constrain prevalence and virulence vaccines that
  offer only anti-toxin immunity have opposite
  effects.

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Emergence of Meta-Genomics

• Microbes run the worldconverting key elements
  of life (C,N,O,S) into accessible forms
  performing photosynthesis living in ecological
  and human host niches, making nutrients, metals,
  and vitamins available to the hosts, digesting
  foods, breaking down toxins, fighting off
  disease-causing microbes.
• Microbial communities have adapted through
  billions of years of environmental changes. New
  molecular methods permit analysis of whole
  communities, not just individual species or
  isolates.

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Metagenomics, A New Science Revealing the
Secrets of our Microbial Planet National
Research Council U.S., 2007

• Combines the power of genomics, bioinformatics,
  and systems biology
• Involves study of sequences and functions of
  genomes of many organisms simultaneously,
  including a great many not culturableas
  presented by Dr. Francino today
• Libraries already prepared from oceans, coral
  reefs, soils, whale carcasses, thermal vents, and
  hot springs
• Expect quite different microbial communities in
  oral cavity, respiratory passages, gut,
  genito-urinary, and skin environments of people
  with different genotypes and different
  environmental circumstances, including diets,
  diseases, infections, and therapies.

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Framework for Regulatory Decision-Making for
Chemicals

• Epidemiology
• Hazard Identification Lifetime rodent
  bioassays
• Short-term, in vitro/in vivo tests
• Structure / activity
• Potency (dose/response)
• Risk Characterization Exposure analysis
• Variation in susceptibility
• Information
• Risk Reduction Substitution
• Regulation / Prohibition

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Impetus for Susceptibility Analysis

• Clinical Occupational Medicine - Why me,
  Doc?
• OSH Act requires that standards be set so
  that no
• workershall suffer adverse effects.
• Clean Air Act requires standards set to
  protect most
• susceptible subgroups.
• Food Quality Protection Act requires
  protection of
• children and other vulnerable
  subpopulations.
• To discover mechanisms or markers for diseases
  that
• may appear only years later -- and
  could be prevented
• getting inside the black box

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Eco-Genetic Polymorphisms

• Tissue susceptibility
• G6PD deficiency
• Alpha1-anti-trypsin deficiency
• Biotransformation enzymes
• Acetylator phenotype
• Debrisoquine hydroxylation
• (Carbon oxidation - P450)
• Paraoxonase activity

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Occupational Bladder Cancer(Huddersfield, U.K.
Data)

• Slow Acetylators
• Controls 57
• Bladder CA 67
• Dye workers (naphthylamines) 96
• Clerical workers 40
• Machine oils workers 44

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Acetylator Phenotype bimodal distribution of
plasma level of active drug (INH) _at_ 6 hours after
standard dose

• Genetics Single gene
• Enzyme N-acetyl-transferase in liver
• Other drugs that require acetylation
• hydralazine dapsone other sulfas
• phenalzine cyclophosphamide
• What about occupational or environmental
  chemicals that are detoxified by acetylation?
• Industrial anti-oxidants
• Naphthylamines
• Benzidines, biphenyls

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Susceptibility to Chronic Beryllium Disease

• An immune-mediated pulmonary granulomatous
• disease among exposed workers
• HLA-DP gene contains a Glu lysine mutation
  
• at position 69 (Glu-69) predisposes to
  CBD
• a genetic biomarker of susceptibility
• Can be combined with lymphocyte proliferation
• test for evidence of sensitization to
  beryllium
• Value-of-information from mandatory and
• voluntary screening programs has been
• estimated for risk reduction
  possibilities
• social, legal, and ethical issues are
  crucial

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Molecular Signatures

• Numerous studies underway using gene expression
  microarrays and proteomics to identify and
  utilize chemical-specific molecular signatures
  for exposures, early effects, and evidence of
  variation in susceptibility.
• Model chemical acetaminophen--toxigenomics and
  toxicoproteomics of liver damage.

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The Thrifty Genotype and Maladaptation to
Modern Environments

• Humans adapted to the Paleolithic environment of
  hunters and gatherers are maladapted to modern
  lifestyles.
• Phenotypes are obesity, high BP, metabolic
  syndrome, and adult-onset diabetes.
  
  
  
  
  
• Most humans consume much more fat, sugars, and
  salt, but less fiber and phytochemicals. Not
  simply digestive and cardiovascular systems, but
  how our brains perceive and process stimuli from
  food sources.
• The evolution from quadripedal to bipedal gait
  puts pressure on the lower spine, with low back
  pain, complications of childbirth, and tendency
  to fainting and circulating problems.

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Evolution and High Blood PressureA. B. Weder,
Hypertension 2007 49260

• Sodium Hypothesis to explain higher incidence of
  high blood pressure in African-Americans
  thrifty genotypes to maintain sodium levels,
  prevent excretion in kidneys, in times of poor
  diets, including slave ships.
• When BP increases, kidney output of Na and water
  are increased, and BP falls feedback.
• Hypertension is viewed as an emergent property
  of the entire cardiovascular system with feedback
  loops.
• Epi studies 30-50 of HBP risk due to heritable
  factors.

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Genes/Alleles Predisposing to High BP

• Angiotensinogen, AGT -6A
• G-Protein, beta-3 subunit, GNB3, 825T
• These two are in genomic regions of broad linkage
  disequilibrium and low haplotype diversityhence
  arose recently, were exposed to positive
  selection
• Beta-2 adrenergic receptor, 47A and 79C
• Epithelial sodium channel alpha-subunit (--946G)
• Cytochrome P450, CYP3A51
• All 5 sodium-conserving ancestral variants are
  much more prevalent in African (equatorial)
  populations.

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Growth Development and Evolution of High Blood
Pressure (HBP)

• Fetus is programmed to survive nutritional
  scarcity in the mother if born into an
  environment of excess sodium and calories, leads
  to metabolic syndrome, obesity, and high BP.
• Fetal growth retardation with decreased
  glomerular filtration capacity set for higher
  range of BP
• Thus, good maternal nutrition could prevent or
  delay onset of some HBP cases.

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Allometry and Kidney Growth

• Kidney weight is proportional to body weight with
  an allometric constant of 0.8, while blood volume
  scales with constant 1.0.
• Kidney growth ceases at sexual maturity smaller
  as menarche and andrarche move to younger ages.
• Adaptation sets blood pressure and sodium
  excretion higher.

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Uricase and Uric Acid

• Serum uric acid levels are higher in humans that
  most other mammals (except guinea pigs and
  Dalmatian dogs).
• Humans lack hepatic uricase.
• Uric acid can help maintain BP via
  vasoconstriction in low sodium environments
• In high-sodium societies, high uric acid may
  cause subtle renal injury, induce chronic salt
  sensitivity, and lead to high BP.

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Dopamine-4 Receptor Repeat Polymorphism and High
Blood Pressure

• Dopamine release in the kidney increases renal
  sodium excretion. In essential hypertension, DA
  effect on sodium excretion is blunted.
• DA-4 receptors are expressed in juxtaglomerular
  and cortical collecting cells associations
  between long allele in exon 3 and slight
  increases in systolic BP, diastolic BP, and
  history of alcohol use.
• DA-1 receptor (A48G) and GPCR kinase-4 gene
  (GRK3gA142V) polymorphisms also associated with
  high BP.

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Metabolic Syndrome in Mexico and in Mexican
AmericansBastarrachea et al, Human Biology
79121, 2007

• GEMM Family Study so far, 375 individuals in 21
  extended families.
• All metabolic and clinical endpoints
  significantly heritable.
• Previous U.S. study Mexican-Americans have
  highest prevalence (32) of metabolic syndrome
  (ATP-III definition) non-Hispanic whites 24,
  African Americans 22.

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Population Genetics From Evolutionary History to
Genetic Medicine Jorde et al, Human Mol
Genetics 2001

• The effective human population size is
  surprisingly small, about 10,000, far less than
  the census size, based on mitochondrial DNA, Y
  chromosome variation, and autosomal
  polymorphisms.
• Human population probably expanded rapidly in
  late Pleistocenetell-tale rare variants
• Genetic diversity between the major continental
  populations is only 10-15 vs 85-90 within each
  continent.
• Much evidence supports African origin of modern
  humans, with subset colonizing Asia and Europe
  about 100,000 years ago. Most persuasive
  haplotypes found outside Africa are usually
  subsets of those found within Africa.

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Genome-wide Detection and Characterization of
Positive Selection in Human Populations (1)

• Dense maps of human genetic variation 3 million
  polymorphisms (HapMap2).
• Long-range haplotype methods identify alleles
  segregating in a population undergoing recent
  selection.
• Cross-population comparisons help discover
  alleles swept to near-fixation in a population.
• Found gt300 strong candidate regions focused on
  22 strongest regions.
• .

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Genome-wide Detection and Characterization of
Positive Selection in Human Populations (2)

• Identified 26 non-synonymous, coding, SNPs
  showing regional evidence of positive selection.
• Found 2 genes in a common biological process with
  positive selection in the same population
• LARGE and DMD in West Africa, related to
  infection by Lassa virus
• SLC24A5 and SLC45A2 in Europe, related to skin
  pigmentation
• EDAR and EDA2R in Asia, involved in hair follicle
  development.
• Sabeti et al. Nature
  2007449913-919

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Genome-wide Detection and Characterization of
Positive Selection in Human Populations (3)

• The most differentiated genes among 3 million
  markers studied globally are those determining
  skin pigmentation.
• May have been linked to risk of ricketts (failure
  of bone production) if there is not sufficient
  vitamin D and activation of vitamin D by
  sunlight, as in northern European populations.

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A Genome-Wide Association Study of Global Gene
Expression (1)Dixon et al. Nature Genetics
2007391202-1207

• Aim build a database to systematically examine
  potential effects of disease-associated variants
  on transcript expression.
• Created a global map of effects of polymorphisms
  on gene expression in 400 children from 206
  British families in asthma study. Looking for
  regulatory effects (non-coding SNPs).
• Genotyped 408,000 SNPs measured gene expression
  of 54,000 transcripts from 20,599 genes in
  lymphoblastoid cell lines.
• 28 of transcripts, representing 6600 genes, had
  narrow heritability H2 gt0.3.

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A Genome-Wide Association Study of Global Gene
Expression (2)

• Highly heritable expression levels are best
  evidence that trait is genetically regulated.
• LCL cells provide general indicator of gene
  expression.
• Use Affy chips for gene expression, Illumina
  beads to genotype
• Identified 88 genes separately associated with 3
  or more SNPs (cis and trans).
• Only 3 SNPs assd with gt5 transcriptsno evidence
  for master regulators

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A Genome-Wide Association Study of Global Gene
Expression (3)

• Used Gene Ontology to identify genes enriched
  among highly heritable traits
• Greatest enrichment response to unfolded
  proteins--chaperonins and heat shock proteins
• May represent an evolutionary response to
  cellular stress unfolded proteins key in prion
  disease, Alzheimer disease, diabetic pancreatic
  beta cells
• Cell cycle, RNA processing, DNA repair, and
  immune response genes also highly heritable.

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Genome-Wide Association Study (4) Applied to
Childhood AsthmaMoffatt et al, Nature 2007
448 470-47

• New susceptibility locus non-coding SNPs
  residing within a 206-kb segment on chromosome
  17q23.
• Of 19 genes in this region, the expression
  database pointed to ORMDL3 (plt10-22), correlated
  (r0.67) with asthma. Codes a transmembrane
  protein anchored in endoplasmic reticulum.
• Disease-associated alleles accounted for 30 of
  variance of expression.
• Can expose LCL cells to asthma triggers.

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Other Disease-Associated Genome-Wide Studies (5)

• Crohns disease PTGER4, chromosome 5
• Type 2 adult-onset diabetes PHACS on chr 11
• Fetal hemoglobin expression in adults markers
  upstream of HBS1L on chr 6q23.
• LDL and HDL cholesterol levels 11 genes
  confirmed, 7 novel genes identified

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Genome-Wide Disease Association Studies Willer
et al, Nature Genetics 200840161-169

• Study of 20,000 people revealed 18 genes which
  influence cholesterol levels, of which 7 are
  newly identified.
• Those increasing total and LDL-cholesterol match
  those predisposing to coronary heart disease.
• However, no protective variants were found for
  action via HDL-C. Maybe this explains recent
  failures of estrogen replacement in Womens
  Health Initiative trials and of Pfizer
  HDL-C-enhancing drug in coronary disease patients.

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Genomic Evidence for Evolution of the Lung
Torday Rehan, Amer J Resp Cell Mol Biol 318,
2004

• Parathyroid hormone-related protein (PTHrP) is a
  highly conserved, stretch-regulated gene
  necessary for embryonic transition from branching
  morphogenesis to alveolization of the lung,
  beginning with the fish swim bladder.
• Growth and stretching up-regulate, while
  microgravity and overdistension down-regulate,
  PTHrP and PTHrP receptor mRNA in alveolar type II
  cells.
• Both surfactant homeostasis and alveolar
  capillary perfusion are under PTHrP control.
• Phylogeny across vertebrate species reveals
  evolutionary amplification of PTHrP signaling
  mediating development, modeling, and remodeling
  of the alveolar wall.

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PTHrP Signaling Between Germ Layers

• By simultaneously stimulating activities of PTHrP
  and PTHrP-receptor, alveolar wall distention
  increases surfactant production and alveolar
  capillary blood flow, with ventilation/perfusion
  (V/Q) matching.
• V/Q matching is the evolutionary integration of
  cell/molecular interactions by which the lung and
  vasculature have adapted to the progressive
  increase in metabolic demand for oxygen.
• PTHrP activates cyclic AMP-dependent protein
  kinase A, which drives adipose differentiation
  related protein (ADRP) to promote trafficking of
  lipid substrate for surfactant and leptin to
  stimulate differentiation of alveolar type II
  cells, during thinning of the alveolar wall due
  to inhibition of fibroblasts by PTHrP.

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Evidence from Ontology and Phylogeny

• Three dramatically different types of lung
• Single-chambered lung of fish, frogs, lizards
  100X diameter interstitial muscle dominates
• Bronchoalveolar lung of mammals conducting and
  terminal airways are convoluted, facilitating gas
  exchange increased saturation and greater
  turnover of surfactant protects the smaller
  alveoli
• Cross-current saccular lung of birds the
  exception to prove the rule lack alveoli, lungs
  are stiff and fixed to thorax, yet possess type
  II cells that express PTHrP and surfactant.
  PTHrPs role in bone morphogenesis links hollow
  bones with air sacs of the respiratory system of
  birds.

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Effects of Deletion of PTHrP Clinical Insights
and Advances

• Models of what pathology might result in humans
  with mutations of PTHrP have been produced as
  knock-outs in mice.
• Deletion of PTHrP results in failed alveolization
  and death due to pulmonary insufficiency within
  minutes to hours of birth. The lung develops
  normally up to the stage of canalization,
  coincident with the point in development when
  PTHrP is produced in mice, rats, and humans.
  Functionally, the lung fails to produce
  surfactant due to immaturity of the embryonic
  lung tissue layers.
• These abnormalities could account for various
  types of surfactant-deficient syndromes in human
  newborns.
• PPARgamma agonists can help premature babies with
  bronchopulmonary dysplasia (Torday Rehan, Peds
  Res 2007).

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Admixture Studies in MexicoJimenez-Sanchez,
Science 300295, 2003

• Mexico has a characteristic genetic population
  structure from admixture of 65 native Indian
  groups with those of Spanish ancestry.
• Genomic medicine was designated a priority in
  health care and medical research during the
  tenure of Dr. Julio Frenk as Secretary of Health.
• A 15-year plan for Genomic Medicine began with
  the establishment of INMEGEN, Instituto de
  Medicina Genomica in the M-NIH, a Consortium for
  the INMEGEN, and public debate on ELSI.

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Genome-wide Admixture Panel for Hispanic/Latino
PopulationsMao et al, Amer J Human Genetics
801171, 2007

• Admixture mapping is a powerful method for
  identifying genetic risk factors for complex
  traits and diseases having prevalence differences
  across populations.
• Here a panel of 2120 ancestry-informative
  markers maximizes differences between Native
  American and European populations while
  minimizing differences between Mexican (Maya and
  Nahua) and South American (Aymara and Quechua)
  populations.
• Can reduce by 100X the genotyping required for
  whole-genome association studies in these
  populations.

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Mao, 2007 PCO 3D Plot of Genetic Distances with
Affy 500K Array
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Mao et al, 2007 Principal coordinate analysis
and 3D plot separating European from
MesoAmerican, Maya from others.
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SUMMARY

• Genome-wide studies and databases are directly
  useful for analyses of the risk factors and
  pathways for specific diseases in different
  populations around the world and in diverse
  populations within individual countries such as
  Mexico and the United States.
• Evolutionary biology provides helpful insights
  for a wide array of developmental, metabolic, and
  disease phenotypes in humans.
• As we celebrate 200th anniversary of Darwins
  birth, evolution by natural selection is a
  foundation for all of biology, including medicine.