Reverse Phase Protein Microarray Reveals a Correlation Between Akt Activation and Altered...
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This study was designed to assay the DNA ploidy using fresh biopsy tissues by flow cytometry (FCM), and investigate the prog- nostic significance of DNA ploidy in NPC. Materials/Methods: Between January 1999 and February 2000, the DNA ploidy of fresh samples in the 53 untreated NPC patients with poor differential squamous cell carcinomas were analyzed by flow cytometry (FCM). Twenty-one patients were treated with one course of cisplatin plus 5-Fluorouracil chemotherapy at the end of 4 th week in addition to radiotherapy. Results: Of the 53 patients, 32 (60.4%) had DNA diploid and 21 (39.6%) had DNA heteroploid. The differences of age, sex, clin- ical stage, N stage and chemotherapy were not significant between diploid group and heteroploid group (p= 0.695, 0.657, 0.088, 0.972 and 0.335). From end of radiotherapy to February 2006, the median follow-up was 73 months (range, 12–84 months). The 5- year overall survival rate was 65.61%, and it was 80.92% in DNA diploid group and 42.86% in heteroploid group (p= 0.002), respectively. The 5-year distant metastasis-free survival rates of DNA diploid and DNA heteroploid patients were 84.26%. and 44.53% (p= 0.003). The 5-year local progression-free survival rates of DNA diploid and DNA heteroploid patients were 92.59% and 72.65% (p= 0.118). By Cox regression analysis, DNA ploidy and clinical stage were the correlative factors for the overall survival rate and distant metastasis-free survival rate (p= 0.020, 0.017 and 0.007, 0.011) (see Tables). Conclusions: DNA ploidy and clinical stage are independent prognostic indices of NPC. The NPC patients with DNA heteroploid were more easily emerged distant metastases than those with DNA diploid. Table: Multivariate analysis of distant metastases-free survival for 53 NPC patients Variate b SE Wald p OR 95% CI Ploidy 1.319 0.554 5.676 0.017 3.740 1.264–11.069 Stage 1.020 0.403 6.416 0.011 2.775 1.260–6.111 Table: Multivariate analysis of overall survival for 53 NPC patients Variate b SE Wald p OR 95% CI Ploidy 1.147 0.493 5.402 0.020 3.148 1.197–8.821 Stage 0.971 0.357 7.401 0.007 2.641 1.312–5.315 Author Disclosure: F. Han, None; H. Wang, None; Y. Xia, None; M. Liu, None; C. Zhao, None; T. Lu, None. 2765 Reverse Phase Protein Microarray Reveals a Correlation Between Akt Activation and Altered Insulin-Like Growth Factor Receptor Expression Following Exposure to Ionizing Radiation B. P. Soule 1 , V. Espina 2 , A. VanMeter 2 , C. Council 1 , L. A. Liotta 2 , J. B. Mitchell 1 , N. L. Simone 3 1 Radiation Biology Branch, National Cancer Institute, Bethesda, MD, 2 Center for Applied Proteomics & Molecular Medicine, George Mason University, Manassas, VA, 3 Radiation Oncology Branch, National Cancer Institute, Bethesda, MD Purpose/Objective(s): The role of Akt (protein kinase B) in cell survival, proliferation and carcinogenesis has been well estab- lished. Akt is activated by stressors such as UV radiation, ionizing radiation and free radicals. This activation is promoted by insulin like growth factor (IGF). In this study, we demonstrate that ionizing radiation-induced Akt activation by phosphorylation of serine- 473 (Akt Ser473 ), is a time and dose dependent phenomenon that correlates with alterations in IGF-receptor 1 expression. Materials/Methods: Murine melanoma cells in culture were irradiated to 10 Gy or 0.5 Gy using a 60 Co sealed source irradiator. Cell lysates were prepared and analyzed using a reverse phase protein microarray probed with antibodies against components of several cell cycle, apoptosis and survival pathways and detected using a colorimetric assay. Altered protein expression was con- firmed by western blotting. Results: Thirty minutes after exposure to 0.5 Gy, Akt Ser473 expression increased 3.4-fold above baseline levels, then returned to baseline 6 hours post-irradiation. This correlated with IGF-receptor expression which also increased at 30 min post-irradiation to 1.7 times basal levels, returned to baseline by 1 hour post-irradiation, then decreased to nearly 2 1 ˇ / 2 times below baseline levels by 72 hours post-irradiation. Conversely, after exposure to 10 Gy, Akt Ser473 levels increased linearly to nearly 3 times basal levels by 72 hours. IGF-receptor levels remained unchanged from basal levels following 10 Gy exposure. Conclusions: These data suggest that Akt activation by ionizing radiation is a dose and time dependent phenomenon that may be mediated by radiation-induced alterations in IGF-receptor expression. It is logical, therefore, that therapies aimed at altering IGF and IGF-receptor expression may potentiate the effects of radiotherapy. Author Disclosure: B.P. Soule, None; V. Espina, None; A. VanMeter, None; C. Council, None; L.A. Liotta, None; J.B. Mitchell, None; N.L. Simone, None. 2766 Analysis of Biologically Equivalent Lung Tissue Dose for 3-D Conformal Partial Breast Irradiation and Whole Breast Irradiation A. K. Jain, L. Vallow, A. Gale Mayo Clinic, Jacksonville, FL Purpose/Objective(s): To examine radiation dose to lung tissue for 3 dimensional (3-D) conformal external beam partial breast irradiation (PBI) and standard whole breast irradiation (WBI). Biologically equivalent dose (BED) was used to account for the altered dose and fractionation of PBI. S622 I. J. Radiation Oncology d Biology d Physics Volume 69, Number 3, Supplement, 2007
Transcript of Reverse Phase Protein Microarray Reveals a Correlation Between Akt Activation and Altered...
S622 I. J. Radiation Oncology d Biology d Physics Volume 69, Number 3, Supplement, 2007
This study was designed to assay the DNA ploidy using fresh biopsy tissues by flow cytometry (FCM), and investigate the prog-nostic significance of DNA ploidy in NPC.
Materials/Methods: Between January 1999 and February 2000, the DNA ploidy of fresh samples in the 53 untreated NPC patientswith poor differential squamous cell carcinomas were analyzed by flow cytometry (FCM). Twenty-one patients were treated withone course of cisplatin plus 5-Fluorouracil chemotherapy at the end of 4th week in addition to radiotherapy.
Results: Of the 53 patients, 32 (60.4%) had DNA diploid and 21 (39.6%) had DNA heteroploid. The differences of age, sex, clin-ical stage, N stage and chemotherapy were not significant between diploid group and heteroploid group (p = 0.695, 0.657, 0.088,0.972 and 0.335). From end of radiotherapy to February 2006, the median follow-up was 73 months (range, 12–84 months). The 5-year overall survival rate was 65.61%, and it was 80.92% in DNA diploid group and 42.86% in heteroploid group (p = 0.002),respectively. The 5-year distant metastasis-free survival rates of DNA diploid and DNA heteroploid patients were 84.26%. and44.53% (p = 0.003). The 5-year local progression-free survival rates of DNA diploid and DNA heteroploid patients were92.59% and 72.65% (p = 0.118). By Cox regression analysis, DNA ploidy and clinical stage were the correlative factors forthe overall survival rate and distant metastasis-free survival rate (p = 0.020, 0.017 and 0.007, 0.011) (see Tables).
Conclusions: DNA ploidy and clinical stage are independent prognostic indices of NPC. The NPC patients with DNA heteroploidwere more easily emerged distant metastases than those with DNA diploid.
Table: Multivariate analysis of distant metastases-free survival for 53 NPC patients
Variate b SE Wald p OR 95% CI
Ploidy
1.319 0.554 5.676 0.017 3.740 1.264–11.069Stage
1.020 0.403 6.416 0.011 2.775 1.260–6.111Table: Multivariate analysis of overall survival for 53 NPC patients
Variate b SE Wald p OR 95% CI
Ploidy
1.147 0.493 5.402 0.020 3.148 1.197–8.821Stage
0.971 0.357 7.401 0.007 2.641 1.312–5.315Author Disclosure: F. Han, None; H. Wang, None; Y. Xia, None; M. Liu, None; C. Zhao, None; T. Lu, None.
2765 Reverse Phase Protein Microarray Reveals a Correlation Between Akt Activation and Altered Insulin-Like
Growth Factor Receptor Expression Following Exposure to Ionizing RadiationB. P. Soule1, V. Espina2, A. VanMeter2, C. Council1, L. A. Liotta2, J. B. Mitchell1, N. L. Simone3
1Radiation Biology Branch, National Cancer Institute, Bethesda, MD, 2Center for Applied Proteomics & Molecular Medicine,George Mason University, Manassas, VA, 3Radiation Oncology Branch, National Cancer Institute, Bethesda, MD
Purpose/Objective(s): The role of Akt (protein kinase B) in cell survival, proliferation and carcinogenesis has been well estab-lished. Akt is activated by stressors such as UV radiation, ionizing radiation and free radicals. This activation is promoted by insulinlike growth factor (IGF). In this study, we demonstrate that ionizing radiation-induced Akt activation by phosphorylation of serine-473 (AktSer473), is a time and dose dependent phenomenon that correlates with alterations in IGF-receptor 1 expression.
Materials/Methods: Murine melanoma cells in culture were irradiated to 10 Gy or 0.5 Gy using a 60Co sealed source irradiator.Cell lysates were prepared and analyzed using a reverse phase protein microarray probed with antibodies against components ofseveral cell cycle, apoptosis and survival pathways and detected using a colorimetric assay. Altered protein expression was con-firmed by western blotting.
Results: Thirty minutes after exposure to 0.5 Gy, AktSer473 expression increased 3.4-fold above baseline levels, then returned tobaseline 6 hours post-irradiation. This correlated with IGF-receptor expression which also increased at 30 min post-irradiation to1.7 times basal levels, returned to baseline by 1 hour post-irradiation, then decreased to nearly 2 1̌̌/2 times below baseline levels by72 hours post-irradiation. Conversely, after exposure to 10 Gy, AktSer473 levels increased linearly to nearly 3 times basal levels by72 hours. IGF-receptor levels remained unchanged from basal levels following 10 Gy exposure.
Conclusions: These data suggest that Akt activation by ionizing radiation is a dose and time dependent phenomenon that may bemediated by radiation-induced alterations in IGF-receptor expression. It is logical, therefore, that therapies aimed at altering IGFand IGF-receptor expression may potentiate the effects of radiotherapy.
Author Disclosure: B.P. Soule, None; V. Espina, None; A. VanMeter, None; C. Council, None; L.A. Liotta, None; J.B. Mitchell,None; N.L. Simone, None.
2766 Analysis of Biologically Equivalent Lung Tissue Dose for 3-D Conformal Partial Breast Irradiation
and Whole Breast IrradiationA. K. Jain, L. Vallow, A. Gale
Mayo Clinic, Jacksonville, FL
Purpose/Objective(s): To examine radiation dose to lung tissue for 3 dimensional (3-D) conformal external beam partial breastirradiation (PBI) and standard whole breast irradiation (WBI). Biologically equivalent dose (BED) was used to account for thealtered dose and fractionation of PBI.
