cancer prevention: translational research in colon cancer matthew r. young gene regulation section...
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Cancer Prevention: Translational Research in Colon CancerMatthew R. YoungGene Regulation Section Laboratory of Cancer PreventionCCR, NCI
Translational Cancer Prevention: How do we approach Translational Research in Cancer Prevention?
Colon Cancer:Anatomy of Colon Cancer Risk factors for Colon Carcinogenesis
Colon Cancer Prevention: Polyp Prevention Trial (PPT)Mouse Metabolomics
Cancer Stem Cells
Nutrition
Molecular Targeted Carcinogenesis Prevention: Benefits at any stage. Cancer prevention prolongs the natural lifetime due to reduced death from cancer
Cancer progression:1. Initiation can be a single mutagenic event.
2. Promotion results from chronic exposure to tumor promoters e.r. TPA, EGF, UV radiation, TNFά or stress
lead to benign tumors.3. Progression results when benign tumors progress
to carcinoma.Receptors activation increases protein kinase activity,
resulting in and increase in transcription factors. Translation factors lead to mis-regulation of target
proteins.
TRANSLATING PREVENTIONHow do we approach Translational Research in Cancer Prevention? Behavior modification in the general population:
Smoking cessation, Weight reduction Diet modificationExercise
Drugs in high risk groups: Tamoxifen: to prevent breast cancerDFMO + Sulindac: to prevent colon cancerAspirin: breast and colonNSAIDS (Celecoxib) Adenoma Prevention TrailDiet supplements
Vaccines in the general population:HVP vaccine: to prevent cervical cancer.HBV vaccine: to prevent liver cancer
Antibiotics in high risk groups: Block H-pylori induce gastric and esophageal cancer
Colon Cancer is the third most common cause of cancer-related death
Risk factors Associated with Colon Cancer
African-American race..Sedentary Life styleAgeDiabetesA personal history of colorectal cancer or polyps. SmokingObesityInflammatory intestinal conditions. RadiationAlcoholInherited syndromes that increase colon cancer risk.Family history of colon cancer and colon polyps.
Low-fiber, high-fat diet.
Trends in overweight prevalence
Stem cell niche
Transit- amplifying
cells
Differentiated cells
The Anatomy of the Colon
Normal organization of the intestinal crypt
Loss of wild-type APC or β-catenin mutationTransformation of healthy crypts towards an adenoma
Accumulation of other genetic lesions, RAS and PTEN,Progression towards an invasive growing CRC
Myofibroblast
Activated Myofibroblast
HGF
Myeloid cellsIL-6 TNFa
ProgressionInitiation Promotion
Tumor Promotion in the Colon
Stages of colon carcinogenesis ~50% of US population have adenoma(s) by age 70 years
CancerPrevention
TRANSLATING PREVENTION. Basic research uses molecular processes, molecular target identification and targeted
drug discovery. Preclinical research uses target validation and target discovery as well as response biomarkers and molecular targets as
endpoints. Clinical research used drug-based and dietary interventions as well as response biomarfer and molecular target
identification.
The Polyp Prevention Trial (PPT)
Multicenter randomized controlled trial examining the effect of a
low-fat (20% of total energy intake), high-fiber (18 g/1000 kcal), high-vegetable and -fruit (5-8 daily servings)
dietary pattern on the recurrence of adenomatous polyps of
the large bowel,
Eligibility one or more adenomas removed within 6
monthscomplete nonsurgical polyp removal complete colonic examination age 35 years or older; no history of colorectal cancer, inflammatory
bowel disease, or large bowel resection; satisfactory completion of a food frequency
questionnaire and 4-day food record
P-trend: 0.001A
dvan
ced
Ade
nom
a R
ecur
renc
e O
R (
95%
CI)
Q2 Q3 Q43.4 12.0 41.5
∆ Dry Bean Intake (T(1,2,3)-T0; in g/d)
Dry Bean Intake Inversely Associatedwith Advanced Adenoma Recurrence
Ob/Ob Obese MiceSingle mutation within the Ob (leptin) geneDevelop obesity, hyperphagia, hyperinsulinemia, and hyperglycemiaInjected with colon carcinogen azoxymethane (AOM) to induce colon cancer Placed on diets after final AOM( injection for 40 weeks
1) Control diet (modified AIN-93G)2) Cooked Whole navy bean diet3) Bean Residue fraction diet 4) Bean ethanol Extract fraction diet
Navy Beans and their Fractions Decrease Colon Lesion Incidence* in AOM-Induced Obese Mice
Biomarkers that predict Colon Cancer and Efficacy of interventions in mice and humans IL-6 a response biomarker for dietary prevention of colonCarcinogenesis in Ob mice,Mentor-Marcel Can Prev Res ,2009
Decrease in serum levels of IL-6 anindicator of efficacious response to bean diet
0.05.0
10.015.020.025.030.035.040.045.050.055.060.065.070.075.080.0
IL-6
Ser
um
IL
-6 (
% d
etec
tab
le)
No Bean
WH Bean
RES Bean
EXT Beanp=.151
p=.0009
p=.018
p=.0035
0.00
5.00
10.00
15.00
20.00
25.00
30.00
Fo
ld c
ha
ng
e r
ela
tiv
e t
o N
o A
OM
No
Be
an
s
IL-6 Tnfrsf8 Stat 4 Sftpd
AOMAOM + Bean extract
**
* *
Bean diet attenuates colon gene expression changes induced by AOM
in ob/ob mice
Human Relevance of IL-6 as a Biomarker of Response to Dietary Intervention
Interleukin-6 as a Potential Indicator for Dietary PreventionOf High Risk Adenoma Recurrence in the Polyp Prevention Trial, Bobe G et al, Cancer Prevention Research, 2010
Colon Carcinogenesis stages in the mouse
Day 0 7 14 21 28 35 42 49 56 70
1-2% DSS in dist. water
AOM 10 mg/kg BWMice 6 wks of age
First tumors appearedStart Diet
Two-Stage Colon Carcinogenesis Model AOM/DSS
Mice develop ACFs, dysplastic lesions, adenomas and
adenocarcinomas.Lesions have elevated b-catenin, cyclooxygenase-2 (COX-2)
and inducible nitric oxide synthase (iNOS) activity
PolypsDetected
Initial polypsEnlarge
Tumor burdenUnhealthy
Normal
Untreated 29 50 60Days after AOM injection
MRI is useful for monitoring efficacy of dietary and/or pharmacological interventions in colon carcinogenesis
MRI is useful for monitoring efficacy of dietary and/or pharmacological interventions in colon carcinogenesis
Bean Extracts and Isorhamnetin diets inhibit inflammation induced colon cancer. Isorhamnetin, Kaempferol and Bean Extracts decreased tumor burden
Isorhamnetin and the Bean Extracts decreased morbidity associated with AOM/DSS treatment
Pilot study to assess efficacy of lifestyle alteration.
Legume Inflammation Feeding Experiment (LIFE). The effects of a high legume (dry bean) diet on markers of insulin resistance
(IR) and inflammation in patients at high risk for CRC.
Feces (F)
American Diet American Diet
Legume DietLegume Diet
Blood (B) B B B B B
F F F
Legume DietAd libitum
Weight change
Legume DietAd Libitum
Weight change
weeks
Legume Inflammation Feeding ExperimentElaine Lanza, Cytonix; Terry Hartman, PSU; Robb
Chapkin Tx A & M
Candidate molecular biomarkers identified from exfoliated colonocytes.
Two- and three-gene combinations provide robust classifiers with potential to noninvasively identify discriminative
molecular signatures for differential diagnostic purposes.
D Legume P-value D Healthy American P-valuesTNFR1 - 3.7% 0.005 - 4.4% 0.001CRP -20.2% 0.018 -18.3% 0.007C-peptide - 2.8% 0.407 - 0.1 0.605
American Diet American Diet
Legume Diet Legume Diet
Potential biomarkers associated with consumption of Legume Enriched and Healthy American diets
American Diet American Diet
Legume Diet Legume Diet Legume Diet
Ad libitumWeight change
Potential biomarkers associated with consumption of reduced energy legume enriched diets. Mean body weight - 4.4 P<0.001BMI -4.5% P<0.001
Other markers significantly reduced (P<0.001)
Total Cholesterol, LDL-C, TG, C-peptide, fasting glucose, Leptin
Metabolomics for identification of Biomarkers for Dietary Intervention and Protection.
Metabolomics: The systematic study of all metabolites in an organism and how they change in relation to a biological
perturbation such as diet, disease or intervention
American Diet
American DietLegume Diet
Legume Diet
B B
B B
Metabolomics for identifying biomarkers from the LIFE study. Serum was collected from participants before and after consumption of bean enriched weight maintenance diet. Anticipated results:Identification of biomarkers for complianceDiscovery of biomarkers of efficacy
Metabolite Name
Sca
led
In
ten
sity
Treatment Group
01
23
45
pipecolate
1 2 3 4 1 2 3 4
Baseline End
Median ValueExtreme Mean Value
Data Points
Upper QuartileLower Quartile
“Max” of distribution“Min” of distribution
___
Box and Whiskers Legend
Metabolomics for identifying biomarkers from the LIFE study274 named biochemicals identified; 87 biochemicals were significanly different between pre and post bean dietsPipecolate increased more than 6-fold in post bean diet.
Diet-derived Metabolites Diet-derived metabolites showed significantly different
plasma levels in pre-diet and post-diet samples.0
.00
.51
.01
.5
1,5-anhydroglucitol (1,5-AG)
1 2 3 4 1 2 3 4
Baseline End
01
23
45
trigonelline (N-methylnicotinate)
1 2 3 4 1 2 3 4
Baseline End
Trigonelline (N-methylnicotinate)
Gut Bacterial Metabolites. Several metabolites generated by gut bacterial metabolism showed significantly
different plasma levels in pre-diet and post-diet samples
01
23
45
pipecolate
1 2 3 4 1 2 3 4
Baseline End
01
23
4
indolepropionate
1 2 3 4 1 2 3 4
Baseline End
Potential Markers of Dietary CompliancePotential candidates for markers of compliance to bean diet include the following. Gut bacterial metabolites pipecolate and indole proprionate. Diet derived 1,5-anhydroglucitol (1,5-AG)Modified amino acid N-acetylornithine
01
23
45
pipecolate
1 2 3 4 1 2 3 4
Baseline End
01
23
4
indolepropionate
1 2 3 4 1 2 3 4
Baseline End
0.0
0.5
1.0
1.5
2.0
2.5
N-acetylornithine
1 2 3 4 1 2 3 4
Baseline End
0.0
0.5
1.0
1.5
1,5-anhydroglucitol (1,5-AG)
1 2 3 4 1 2 3 4
Baseline End
AOM 2% DSS Diet serum/feces
0 7 12 14 21 28 35 42 49 56 63 70 Days
Metabolomics for identifying biomarkers from the AOM/DSS induced CRC in mice
Metabolomics from Mice fed Bean Extracts
Correlates with Metabolomics from Human (LIFE) Study
Pipecolate and N-acetyl-ornithine, proposed biomarkers of bean diet
compliance identified elevated in both bean diet plasma groups. Also a similar subtle yet significant decrease in 1,5-anhydroglucitol was observed in both
animal groups on bean diet.
Metabolomics from Mice fed Bean Extracts Correlates with Metabolomics from Human (LIFE) StudyDecreased lysophospholipids Decreased medium- chain fatty acids Decrease in carnitine/acylcarnitines, No notable change in long-chain FA;
Collectively indicating increased FA metabolism for energy in bean diet-fed animals
Fetal metabolomics from mice fed bean extract diet Increase in Alcohol sugars,
Krebs cycle intermediates (citrate, alpha-ketoglutarate, fumarate and malate) were also significantly elevated in feces of bean extract fed mice.
Fecal nucleotide breakdown products including nitrogenous bases, ribose and 2-deoxyribose, as well as phosphate were substantially increased in bean extract fed mice
The NCI-Translational Research Working Group:Lifestyle Alteration Developmental Pathway LIFE: Short term feeding study to measure the effects of a bean diet on markers of insulin resistance (IR) and inflammation in patients at high risk for CRC.
Develop Biomarkers-Metabolomics: Metabolic biomarkers of compliance identified in human serum
Develop Biomarkers-Metabolomics: Metabolic biomarkers of compliance identified in human serum also detected in mouse serum and feces. Young et al., unpublished
The NCI-Translational Research Working Group:Lifestyle Alteration Developmental Pathway LIFE: Short term feeding study to measure the effects of a bean diet on markers of insulin resistance (IR) and inflammation in patients at high risk for CRC.
Develop Biomarkers-Metabolomics: Metabolic biomarkers of compliance identified in human serum
Develop Biomarkers-Metabolomics: Metabolic biomarkers of compliance identified in human serum also detected in mouse serum and feces. Young et al., unpublished
Metabolomic analysis from the Polyp Prevention Trial: Identification of metabolic biomarkers associated with reduced adenoma recurrence. 3 groups of 125 participants 2 time points, baseline and after 3 years
Control: Participants with no change in tumors
Intervention, bean consumption: participants who consumed high bean diet and showed a reduced recurrence of adenomas
Tumor Positive: Participants with increased recurrence of
adenoma after 3 yr
BIOMARKERS AND MOLECULAR TARGETS OF NON-TOXIC DIETARY INTERVENTIONS FOR CANCER PREVENTION
Laboratory of Cancer Prevention Nancy Colburn, Noriko Yoshikawa, Alyson Baker, Qiou Wei, Glenn Hegamyer, Shakir Saud, Elaine Lanza
LIFE Study, Terryl J. Hartman, Pennsylvania State, Zhiying ZhangRobb Chapkin, Texas A & M
Obese mice, Marcie Bennink, Michigan State University, Kati Barrett Division of Cancer Prevention, John Milner, Young Kim, Gerd Bobe, Prevention Fellow, Roycelynn Mentor-Marcel, Prevention Fellow
Statistician Paul Albert, NCI Small Animal Imaging Program, Pete Choyke, Marcelino Bernardo, Lilia
Ileva, Joe Kalen, Lisa RIffle
AP-1 and NF-kB Matthew Young, Arindam Dhar, Jing Hu, Connie Matthews, Moon-IL Kang, Brett Hollingshead, Qiou Wei, Gerd Bobe,Roycelynn Mentor-Marcel Jim McMahon, MTDP NCI; Curt Henrich, MTDP, Powel Brown, Baylor Univ; Peter Choyke and SAIP, NCI; Elaine Lanza, Cytonix; Terry Hartman, PSU; Rob Chapkin, TX A&M; Gary Stoner, OHU; Michel Toledano, IBITECS, France
Pdcd4Hsin-Sheng Yang, Joan Cmarik, Aaron Jansen, Halina ZakowiczArti Santhanam, Tobias Schmid, Brett Hollingshead, Noriko Yoshikawa, Nahum Sonenberg, McGill Univ.; Myung Cho, Seoul Nat Univ; Alex Wlodawer, Nicole LaRonde, NCI; Michele Pagano, NYU; Heike Allgayer, Klinikum Mannheim; Bruce Shapiro, NCI
LCP 2009Laboratory of Cancer Prevention