Presentation on theme: "Cebulska-Wasilewska A. 1,2, Miszczyk J. 1, Dobrowolska B. 3, Dobrowolski Z. 3 1 Environmental and Radiation Biology Department, The H. Niewodniczański."— Presentation transcript:
Cebulska-Wasilewska A. 1,2, Miszczyk J. 1, Dobrowolska B. 3, Dobrowolski Z. 3 1 Environmental and Radiation Biology Department, The H. Niewodniczański Institute of Nuclear Physics PAN, Cracow, Poland, 2 Epidemiology and Preventive Medicine Departmen CM UJ, Poland, 3 Urology Department and Clinic CM UJ, Poland Detection of Chromosomal Translocation in Prostate Cancer and Benign Prostatic Hyperplasia by Fluorescence in situ Hybridization (FISH)
Prostate cancer epidemiology Benign Prostatic Hyperplasia [BPH] and Prostate Cancer [PC] are the most common males diseases. In Poland prostate cancer is the third most common malignant cancer in males. Prostate cancer incidence varies widely between: ethnic populations, countries and increases sharply with older age.
Prostate cancer risk is strongly influenced by: GENETIC FACTORS > mutation > genetic susceptibility FAMILIAL HISTORY EPIGENETIC FACTORS > dietary factors (vitamin D, fats,) > androgens > ethnic origin > lifestyle, smoking > genes involved in familial prostate cancer (for example: HPC1 on chromosome 1) > gene expression
> genetic alteration on multiple chromosomes including especially chromosome 1, > many susceptibility loci have been reported at this chromosome, > many types of cancers are associated with specific types of chromosomal aberrations Becouse...
Aim of study Compare the vulnerability to the induction in chromosome 1 translocation in lymphocytes from prostate cancer with that from benign prostatic hyperplasia.
Materials and methods Investigated groups [PCP] – 30 prostate cancer patients. (average age 62.4±5.3) [BPH] – 27 persons from the control group with benign prostatic hyperplasia. (average age 68.9 ± 8.3) Challenging dose In the laboratory tubes with blood were irradited with X-rays doses of 2 Gy. Fluorescent in situ hybridisation (FISH) Biotin-labeled whole chromosome probes specific to chromosome 1 (Star Fish Cambio, UK). Donors peripheral blood samples Fixation methanol/acetic acid (3:1) KC L 80% RPMI 1640, 20% fetal calf serum antibiotics, PHA Incubation at 37ºCfor 72 hours colcemid X-rays irradiation Standard cytogenetic procedureFig. 1. Culture were set up according to standard cytogetic procedure, then were harvested and followed by fixation procedure.
Materials and methods The slides were examined at 1000x magnification of the epifluoescence microscope (Nicon Eclipse E400). Donors were examined for presence in their in peripheral blood lymphocytes of chromosome translocations according to the criteria of Protocol for Aberration Identification and Nomenclature-PAINT . Labeled probe Place probe on slide Hybridization 37ºC, 72 hours detection General FISH protocolFig. 2.  Tucker J.D. et al. A proposed system for scoring structural aberrations detected by chromosome painting. Cytog. Cell Genet. (1995), 211-221. Fig. 3. Fig. 4. Fig. 5.
Materials and methods 2 types of parameters were used to describe the extent of chromosomal damage t – frequency of chromosome 1 translocation F G /100 – genomic frequency of chromosome 1 translocation F G =Fg/2.05fp(1-fp) . F G - the total genomic translation frequency Fg – the translocation frequency measured by FISH after painting Fp – the fraction of th genome represented the painted chromosome, for chromosome 1 = 0.084 fraction of the genome 8.4%)  Lucas J.N., Sachs R.K. Using three-color chromosomepainting to test chromome aberration models. Proc. Natl. Sci90, 1484-14
Results t Number of translocations/1000 cells (t) was significantly higher in patients with prostate cancer (14.60±0.91) than in the control group (10.24 ±1.10; p<0.01). Sig. 1. X-rays effect on frequency of chromosome 1 translocation determined by FISH in peripheral lymphocytes in patients with prostate cancer [PCP] and benign prostate hyperplasia [BPH]. T – number of translocations/1000 cells
Results F G /100 Percentage of F G /100 was significantly higher in patients with prostate cancer (0.55±0.03) that obtained for the reference group (0.38 ±0.04, p<0.01). F G /100 - genomic frequency of translocation Sig. 2. Genomic frequency of chromosome 1 translocation for patients with prostate cancer [PCP] and benign prostatic hyperplasia [BPH].
We want to study correlation between occur cancer in family and frequency of chromosome 1 translocation.
Results t AgeR0.50 p<.001 CiFR0.11 p<.001 Tab. 1. Correlations for patients with prostate cancer between age of donors, existing cancer in the closely related members of family and t. CiF – reported cancer in the immediate family R – correlation coefficient t – number of translocations/1000 cells High and significant correlation between age of donors and frequency of chromosome 1 translocation was observed (0.50; p<0.001). Furthermore, there was also correlation between frequency of chromosome 1 translocation observed in patients who had reported other cancers in family.
Conclusions 1. These studies, although preliminary, are suggesting that frequency of translocation detected in the response to challenging treatment might be used as predictor of susceptibility for prostate cancer patients. However, more studies are necessary of other factors which could affect genomic frequency of translocations such as: life style, diet or genetic polymorphism. 2. Our results might confirm hypothesis that exist an association between predisposition to genetic instability chromosome 1 and hereditary or familial conditioning of prostate cancer.
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