Mizaj as an Index in Persian Traditional Medicine Index Could Associate with Sensitivity to the Radiation

Background: The sensitivity to the radiation among human population depends on various parameters. This variation could lead to dissimilar outcome of radiotherapy in similar situations. Mizaj is a well-known term in Persian medicine that present an individualized medicine viewpoint. All of the people will be categorized in cold, moderate, and warm Mizaj. In this study, we aimed to evaluate the possible association between Mizaj and radiosensitivity by comet assay. Materials and Methods: Peripheral blood sample of 30 healthy volunteers (10 cold, 11 moderate and nine warm Mizaj) were taken and divided into two identical parts. The first part was exposed to 4 Gy x-rays, and the second part was regarded as the sham control. Then, DNA damages of samples were evaluated by the neutral comet assay. Results: The results showed that the mean percentage of damaged cells, in all of the irradiated groups including A (warm), B (moderate) and C (cold) was significantly higher than the controls (P<0.001). Moreover, DNA damage rate in the irradiated warm Mizaj group was higher than both cold and moderate irradiated groups, but the difference between moderate and cold irradiated groups was not statistically significant. Conclusion: The results are indicating that warm Mizaj persons could be more radiosensitive than other groups, which their importance in radiotherapy individualization should be evaluated in more extensive studies. [GMJ.2020;9:e1705]

Restraining Akt1 Phosphorylation Attenuates the Repair of Radiation-Induced DNA Double-Strand Breaks and Reduces the Survival of Irradiated Cancer Cells

The survival kinase protein kinase B (Akt) participates in the regulation of essential subcellular processes, e.g., proliferation, growth, survival, and apoptosis, and has a documented role in promoting resistance against genotoxic stress including radiotherapy, presumably by influencing the DNA damage response and DNA double-strand break (DSB) repair. However, its exact role in DSB repair requires further elucidation. We used a genetic approach to explore the consequences of impaired phosphorylation of Akt1 at one or both of its key phosphorylation sites, Threonine 308 (T308) or Serine 473 (S473), on DSB repair and radiosensitivity to killing. Therefore, we overexpressed either the respective single or the double phosphorylation-deficient mutants (Akt1-T308A, Akt1-S473A, or Akt1-T308A/S473A) in TRAMPC1 murine prostate cancer cells (TrC1) and measured the DSB repair kinetics and clonogenic cell survival upon irradiation. Only the expression of the Akt1-T308A/S473A induced a significant delay in the kinetics of DSB repair in irradiated TrC1 as determined by the γH2A.X (H2A histone family, member X) assay and the neutral comet assay, respectively. Moreover, Akt1-T308A/S473A-expressing cells were characterized by increased radiosensitivity compared to Akt1-WT (wild type)-expressing cells in long-term colony formation assays. Our data reveal that Akt1’s activation state is important for the cellular radiation response, presumably by modulating the phosphorylation of effector proteins involved in the regulation of DSB repair.

PKR Inhibits the DNA Damage Response and Correlates with Poor Clinical Outcomes in AML

Introduction: Dysregulation of the double-stranded RNA-activated protein kinase (PKR) disturbs cellular signaling pathways that influence proliferation, differentiation, inflammation and apoptosis. Given the multifaceted functions of PKR in cellular stress signaling and inflammation, how PKR may play a role in tumorigenesis is a significant but underexplored area. Using a proteomic approach, we stratified AML patients according to PKR expression level in CD34+ leukemic cells and find that patients with high PKR expression have inferior clinical outcomes. Significantly, results from both acute leukemia derived cell lines and primary AML BM samples reveal that increased PKR expression leads to inhibition of DNA damage response (DDR) and DNA double strand break (DSB) repair. Thus, high PKR expression/activity in leukemic cells may retard the DNA damage response, enhance genomic instability and facilitate leukemia progression. Method: Peripheral blood and bone marrow specimens were collected from 414 patients with newly diagnosed AML and evaluated for outcomes at The University of Texas M.D. Anderson Cancer Center (MDACC) between September 1999 and March 2007. Samples were acquired during routine diagnostic assessments in accordance with the regulations and protocols (Lab 01-473) approved by the Investigational Review Board (IRB) of MDACC and analyzed under IRB-approved laboratory protocol (Lab 05-0654). Transgenic PKR (TgPKR) and dominant-negative PKR (DNPKR) mice were previously developed in our lab. PKR knockout mice (PKRKO) were provided by Dr. Robert Silverman (Lerner Research Institute, Cleveland Clinic, Cleveland, OH). The study was approved by UF IACUC protocol #201102224. Sample preparation and RPPA processing was done as described (Kornblau et al., 2011). Proteomic profiling for PKR was performed on AML patient samples by RPPA using the PKR monoclonal M02 antibody (Abnova). Neutral Comet assay and Olive tail moment calculations were performed by Wimasis Image analysis (Munich, Germany). γ-H2AX, p-ATM, p-NBS-1 levels were evaluated by both western blot and flow cytometry. Results: High level of PKR expression in CD34+ blast cells from 414 newly diagnosed AML patients correlates with reduced remission duration and poor survival. Cells with decreased PKR expression or treated with the PKR inhibitor (PKRI) have a significantly more rapid induction of γ-H2AX formation, increased p-ATM and p-NBS1 following ionizing irradiation (IR). On the other hand, bone marrow hematopoietic stem progenitor cells (HSPCs) isolated from transgenic mice overexpressing human PKR (TgPKR) display a decreased level of p-ATm and p-NBS1 indicating a depressed DDR following IR. Significantly in human CD34+ leukemia blasts an inverse relationship exists between PKR expression and γ-H2AX or p-ATM. Results from the Neutral comet assay shows that REH cells with decreased PKR expression or HSPCs from PKRKO mice are able to repair DSBs at a significantly faster rate than HSPCs from WT mice, which repair DSB at faster rate than those from TgPKR mice. In addition treatment of TgPKR cells or CD34+ AML blasts with PKRI can restore the efficiency of DSB repair. Conclusions: Our findings demonstrate that PKR inhibits/retards the DDR and DNA DSB repair following genotoxic stress, the consequence of which results in genomic instability. This finding may explain, at least in part, our observation that high PKR expression in primary AML blast cells is associated with poor survival of patients with AML. Therefore pharmacological inhibition of PKR may represent a novel therapeutic strategy that targets and reduces genomic instability in these cells and may improve the clinical outcomes in such AML patients. Disclosures No relevant conflicts of interest to declare.