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A Comprehensive Genomic Approach to the Identification of Predictive Markers using DNA and Tissue A Comprehensive Genomic Approach to the Identification of Predictive Markers using DNA and Tissue Repair Gene Polymorphisms in Radiation Induced Late Toxicity in Prostate Cancer Patients Repair Gene Polymorphisms in Radiation Induced Late Toxicity in Prostate Cancer Patients

Treated with Conformal RadiotherapyTreated with Conformal Radiotherapy

Damaraju S 1,2, Murray D2, Dufour J1, Carandang D1, Myrehaug S2,3, Fallone G2,3, Field C2,3, Greiner R1,3, Hanson J2, Cass CE1,2 and Parliament M1,2

1Alberta Cancer Board PolyomX Program at Cross Cancer Institute, 2Department of Oncology, 3Division of Medical Physics, 4Department of Computing Science, University of Alberta, Edmonton.

A radio-genomic approach to explore the possible relationship A radio-genomic approach to explore the possible relationship between single nucleotide polymorphisms (SNPs) in clinical between single nucleotide polymorphisms (SNPs) in clinical radiation toxicity in a retrospective cohort of prostate cancer radiation toxicity in a retrospective cohort of prostate cancer patients previously treated with conformal radiotherapy patients previously treated with conformal radiotherapy (3DCRT) is attempted.(3DCRT) is attempted.

We selected 279 SNPs in candidate genes encoding DNA We selected 279 SNPs in candidate genes encoding DNA damage recognition/ repair/ response, steroid metabolism, damage recognition/ repair/ response, steroid metabolism, cytokine and tissue repair proteins for this studycytokine and tissue repair proteins for this study

We report initial analysis of association for late tissue toxicity in We report initial analysis of association for late tissue toxicity in 83 patients in a retroactive cohort using 128 SNPs. 83 patients in a retroactive cohort using 128 SNPs.

MethodsMethodsThis study was approved by the institutional ethics board. This study was approved by the institutional ethics board. Informed consent was obtained from eighty three patientsInformed consent was obtained from eighty three patients who who receivedreceived radiation therapy for prostate cancer. These patients radiation therapy for prostate cancer. These patients experienced late tissue (GI) toxicity. We grouped patients with experienced late tissue (GI) toxicity. We grouped patients with RTOG scores of 0-2 (n=55) and 2+ and above (n=28) for RTOG scores of 0-2 (n=55) and 2+ and above (n=28) for analysis, and genotype calls for individual SNPs were also analysis, and genotype calls for individual SNPs were also grouped dichotomously to maximize the differences in time to grouped dichotomously to maximize the differences in time to toxicity. Analysis also included radiation dosimetric variables toxicity. Analysis also included radiation dosimetric variables and age covariates. Cumulative risk of late toxicity curves were and age covariates. Cumulative risk of late toxicity curves were derived according to genotype class for each SNP using Kaplan-derived according to genotype class for each SNP using Kaplan-Meier method and log-rank tests were performed to obtain Meier method and log-rank tests were performed to obtain univariate significance. Exploratory multivariate analysis was univariate significance. Exploratory multivariate analysis was performed using the SAS score method to determine the best performed using the SAS score method to determine the best overall model fit.overall model fit.

Results and Conclusions Results and Conclusions Significant univariate associations with late rectal or bladder Significant univariate associations with late rectal or bladder toxicity (gr. 2+) were found for toxicity (gr. 2+) were found for XRCC3XRCC3 (A>G 5' UTR NT 4541), (A>G 5' UTR NT 4541), LIGIVLIGIV (T>C Asp568Asp), (T>C Asp568Asp), MSH6 (MSH6 (T>C, D180D), T>C, D180D), MLH1MLH1 (C>T (C>T V2191), V2191), CYP2D6*4CYP2D6*4 (G>A splicing defect), mean rectal and (G>A splicing defect), mean rectal and bladder dose, dose to 30% of rectum or bladder, and age <60 bladder dose, dose to 30% of rectum or bladder, and age <60 years. Significant associations with freedom from toxicity were years. Significant associations with freedom from toxicity were found for found for LIGIVLIGIV (T>C Asp568Asp), (T>C Asp568Asp), ERCC2ERCC2 (G>A, D711D), (G>A, D711D), CYP2D6*4CYP2D6*4 (G>A, splicing defect), mean bladder dose >60 (G>A, splicing defect), mean bladder dose >60 Gray, and dose to 30% of rectal volume >75 Gray on Cox Gray, and dose to 30% of rectal volume >75 Gray on Cox multivariate analysis. A study with larger sample size to validate multivariate analysis. A study with larger sample size to validate these findings is warranted to justify radio-genomic approaches these findings is warranted to justify radio-genomic approaches in the clinic. in the clinic.

Study Design and Scope

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DNA Repair Pathways

•Base Excision Repair- Oxidized bases, spontaneous damage (ADPRT, APE1, Lig, Pol B, XRCC1 etc)

•Nucleotide Excision Repair- UV and Chemicals (ERCC, Lig, Pol D1, XPA, XPC, RAD23A, and RAD23B)

•Double Strand Break/Recombination Repair- Ionizing Radiation and radiometric drugs (Lig, NBS1, XRCC, RAD50, RAD51)

•Damage Recognition and Cell Cycle Check Points- Indirect Role in Repair (CDK4, CDKN2A, RAD52, RAD54, XRCC9, MLH, MSH)

•Mismatch Repair- Replication errors (MSH)

Supported by ACF and ACB

ACumulative risk of grade 2+ Toxicity,

all cases+/- 95% CI, N = 83

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BRisk of Gr. 2+ Toxicity:

ERCC2 D711D p = 0.078

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CRisk of Gr. 2+ Toxicity:

Lig IV D568D p = 0.014

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DRisk of Gr. 2+ Toxicity:

MSH6 D180D p = 0.053

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ERisk of Gr. 2+ Toxicity:

CYP2D6*4 Splicing defectp = 0.013

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FRisk of Gr. 2+ Toxicity:

Dose to 30% of rectal volume p = 0.0013

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HRisk of Gr. 2+ Toxicity:

Mean bladder dose p = 0.0036

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>60

GRisk of Gr. 2+ Toxicity:

Age at diagnosis >60 yearsp = 0.0055

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Fig 1: Kaplan-Meier graphs of radiation induced toxicity in prostate cancer patients and significance levels for the variables in univariate analysis

                 

Table 4:Multivariate analysis of factors associated with RTOG grade 2+ chronic toxicity  

 

Best Subset of 4 predictors Score Chi-Square = 36.6, p < 0.0001

Second Best Subset of 4 predictors Score Chi-Square = 34.9, p < 0.0001

 

  Variable p-value Hazard 95% CI p-value Hazard 95% CI  

  LIG IV T>C, Asp568Asp 0.0004 4.86 2.04 - 11.56 0.0034 3.56 1.52 - 8.31  

  CYP2D6*4 G>A, Splicing defect - - - 0.0110 2.85 1.27 - 6.38  

  Age at diagnosis 0.0108 0.24 0.08 - 0.72 0.0002 0.13 0.04 - 0.38  

  Mean bladder dose 0.0111 3.02 1.29 - 7.08 - - -  

  Dose to 30% of rectal volume 0.0037 3.27 1.47 - 7.29 0.0005 4.14 1.87 - 9.17  

Best Subset of 5 predictors Score Chi-Square = 42.0, p < 0.0001

Second Best Subset of 5 predictors Score Chi-Square = 41.7, p < 0.0001

  Variable p-value Hazard 95% CI p-value Hazard 95% CI  

  LIG IV T>C, Asp568Asp 0.0002 5.37 2.22 - 12.97 0.0003 5.22 2.14 - 12.73  

  ERCC2 G>A, D711D 0.0207 3.54 1.21 - 10.35 - - -  

  MSH6 T>C, D180D - - - 0.0074 3.08 1.35 - 7.01  

  Age at diagnosis 0.0036 0.19 0.06 - 0.58 0.0057 0.22 0.07 - 0.64  

  Mean bladder dose 0.0086 3.19 1.34 - 7.59 0.0029 3.90 1.60 - 9.55  

  Dose to 30% of rectal volume 0.0105 2.89 1.28 - 6.53 0.0012 3.81 1.70 - 8.55