Analysis of the molecular alterations in cancer cells following nanotechnology-assisted targeted radiotherapy using Raman spectroscopy.

The study unveiled an innovative strategy for precise radiation targeting in cancer treatment, along with the monitoring of molecular changes induced by this therapeutic approach. In this research, we explored the impact of administering anti-HER2-AgNPs nanoconjugates either individually or in conjunction with gamma irradiation on the viability of SKBR3 breast cancer cells. The utilization of nanoconjugates resulted in an enhancement of cellular sensitivity toward radiation. The viability of the cells exhibited a decline as the dose of irradiation increased, and this decrease was further exacerbated by the passage of time following irradiation. The analysis of RS revealed distinct cellular responses in varying conditions. The observed increase in SERS intensity, resulting from the increment in dose from 0 to 2 Gy, can be attributed to the probable upregulation of HER2 expression induced by irradiation. The observed decrease in SERS intensity at doses of 4 and 6 Gy can be attributed to the likely reduction in HER2 expression. It was illustrated that the analysis of Raman spectroscopy data can aid in the identification of radiation-induced biochemical alterations in cancer cells during the application of nanoconjugates-based radiotherapy. The findings revealed that nanoconjugates have the potential to enhance cellular sensitivity to radiation along with facilitating the detection of radiation-induced biochemical alterations within cancer cells.

The Application of ATR Kinase Inhibitor AZD6738 in Combination with Radiotherapy for the Treatment of Melanoma.

Background: Melanoma is categorized as one of the most malignant, severe, and lethal cancers of the skin. Regarding the lack of efficiency of conventional therapies for most patients, novel therapeutic strategies are strongly required. Objective: The current study aimed to assess the impact of AZD6738- an ATR kinase inhibitor- in combination with 6 MV X-ray on the human melanoma cell line (A375). Material and Methods: In this experimental study, cells were treated with different concentrations of AZD6738 for 24 and 48 h in the presence and absence of radiation (2 Gy, 4 Gy, and 6 Gy). The cell viability and cell proliferation assay were examined in both experimental and control groups by MTT and colony formation techniques, respectively. Results: The results indicated that by increasing the concentration of AZD6738, the cell viability was markedly diminished in all treatment groups. As expected, the cell viability of the cells treated with AZD6738 and radiation was significantly lower than the group treated with AZD6738 alone. Besides, the combinatory treatment significantly decreased cell proliferation in the melanoma cell line. The combination of AZD6738 with radiation resulted in a significant increase in cytotoxicity by a 50% increase in cell death when used at concentrations of 0.3 µM, 1 µM, 1.51 µM, and 1.61 µM, respectively. Conclusion: The combination of AZD6738 with radiation possesses a synergistic effect on the reduction of the cell viability and proliferation of melanoma cells. This present study provides insight into the impact of Ataxia Telangiectasia and Rad3-related kinase (ATR) inhibition on the potential role of this kinase in the suppression of melanoma cell proliferation.

The effect of iodine-131 beta-particles in combination with A-966492 and Topotecan on radio-sensitization of glioblastoma: An in-vitro study.

Glioblastoma tumors are resistant to radiotherapy, and the need for drugs to induce radio-sensitization in tumor cells has always been a challenge. Besides, radiotherapy using targeted radionuclide would be effective even for resistant tumors. It has been shown topoisomerase I and poly (ADP-ribose) polymerase (PARP) enzymes have critical roles in the repair process of DNA injury in cells. Therefore, the inhibition of the activity of these enzymes can halt this process and result in the accumulation of damaged DNA in cells and the induction of cell death in tumors. In the present research, the impact of beta-particles of iodine-131 in combination with Topotecan (TPT), as the inhibitor of topoisomerase I, and A-966492, as the inhibitor of the PARP enzyme on the possible increase of radio-sensitivity of glioblastoma cells was assessed. For this purpose, a human glioblastoma cell line, U87MG, was cultured in flasks coated with Poly-Hema to achieve 300 µm-diameter spheroids. Then, nontoxic concentrations of A-966492 and TPT were applied in the cell culture media. The viability of the cells treated with iodine131 in combination with A-966492 and TPT was determined by the clonogenic assay. The expression rate of gamma-H2AX, as a biomarker of DNA double-strand breaks, was analyzed using immunofluorescence microscopy to unravel the effect of TPT, A-966492 (1 µM), and radiation on the cell death induction. The combination of each TPT or A-966492 with radiation resulted in the increased rate of cell death, and the ratios of sensitizer enhancement at 50% survival (SER50) were elevated by 1.45 and 1.25, respectively. Chemo- and radio-sensitization were promoted when iodine-131 was combined with A-966492 and TPT, with the SER50 of 1.68. Also, the expression of γ-H2AX was significantly increased in cells treated with A-966492 and TPT combined with radiation. The results demonstrated that iodine-131, in combination with A-966492 and TPT, had marked effects on radio-sensitizing and can be used as a targeted radionuclide for targeting radiotherapy in combination with topoisomerase I and PARP inhibitors to enhance radiotherapy in clinics.

The Use of ß-Elemene to Enhance Radio Sensitization of A375 Human Melanoma Cells.

OBJECTIVE: Melanoma is the most malignant and severe type of skin cancer. It is a tumor with a high risk of metastasis and resistant to conventional treatment methods (surgery, radiotherapy, and chemotherapy). ß-elemene is the most active constituent of Curcuma wenyujin which is a non-cytotoxic antitumor drug, proved to be effective in different types of cancers. The study aimed to investigate the therapeutic effects of ß-elemene in combination with radiotherapy on A375 human melanoma. MATERIALS AND METHODS: In this experimental study, human melanoma cells were grown in the monolayer culture model. The procedure of the treatment was performed by the addition of different concentrations of ß-elemene to the cells. Then, the cells were exposed to 2 and 4 Gy X-ray in different incubation times (24, 48, and 72 hours). The MTT assay was used for the determination of the cell viability. To study the rate of apoptosis response to treatments, the Annexin V/PI assay was carried out. RESULTS: The results of the MTT assay showed ß-elemene reduced the cell proliferation in dose- and time-dependent manners in cells exposed to radiation. Flow cytometry analysis indicated that ß-elemene was effective in the induction of apoptosis. Furthermore, the combination treatment with radiation remarkably decreased the cells proliferation ability and also enhanced apoptosis. For example, cell viability in a group exposed to 40 µg/ml of ß-elemene was 80%, but combination treatment with 6 MV X beam at a dose of 2 Gy reduced the viability to 61%. CONCLUSION: Our results showed that ß-elemene reduced the proliferation of human melanoma cancer cell through apoptosis. Also, the results demonstrated that the radio sensitivity of A375 cell line was significantly enhanced by ß-elemene. The findings of this study indicated the efficiency of ß-elemene in treating melanoma cells and the necessity for further research in this field.

Investigation of combined photodynamic and radiotherapy effects of gallium phthalocyanine chloride on MCF-7 breast cancer cells.

In this study, we evaluated the effect of gallium phthalocyanine chloride (GaPcCl) as a radio- and photosensitizer on MCF-7 breast cancer cell line. We incubated cells with GaPcCl in different concentrations (from 3.125 to 100 µg/ml). Then cells in separate groups were exposed to different light doses (1.8 and 2.8 J/cm2) at wavelength of 660 nm and 2-Gy X-ray ionizing radiation, alone and in combination. Finally, cell survival and apoptosis were determined by MTT assay and flow cytometry, respectively. The results showed that the deactivated GaPcCl at concentration of 100 µg/ml reduces the cell viability up to 15%. While, photoactivated GaPcCl (100 µg/ml) at light dose of 2.8 J/cm2 significantly decreases cell viability up to 55.3%. Although MTT assay demonstrated that GaPcCl is not act as a radiosensitizer, flow cytometry showed significant increase in cell apoptosis when GaPcCl was exposed to 2 Gy X-ray. Using of GaPcCl-PDT (photodynamic therapy) integration with X-ray substantially increased cell death in comparison to the absence of X-ray. Furthermore, flow cytometry displayed a significant increase in apoptosis cells (especially late apoptosis) in this combination therapy. Our result proved that GaPcCl is an effective photosensitizer in MCF-7 human breast cancer cell line. The combination of GaPcCl-PDT and radiotherapy can be an efficient treatment against cancer. This approach needs further investigations on animal models for human purposes.Graphic abstract.

The molecular cues for the biological effects of ionizing radiation dose and post-irradiation time on human breast cancer SKBR3 cell line: A Raman spectroscopy study.

Radiotherapy is one of the main modalities of cancer treatment. The utility of Raman spectroscopy (RS) for detecting the distinct radiobiological responses in human cancer cells is currently under investigation. RS holds great promises to provide good opportunities for personalizing radiotherapy treatments. Here, we report the effects of the radiation dose and post-irradiation time on the molecular changes in the human breast cancer SKBR3 cells, using RS. The SKBR3 cells were irradiated by gamma radiation with different doses of 0, 1, 2, 4, and 6 Gy. The Raman signals were acquired 24 and 48 h after the gamma radiation. The collected Raman spectra were analyzed by different statistical methods such as principal component analysis, linear discriminant analysis, and genetic algorithm. A thorough analysis of the obtained Raman signals revealed that 2 Gy of gamma radiation induces remarkable molecular and structural changes in the SKBR3 cells. We found that the wavenumbers in the range of 1000-1400 cm-1 in Raman spectra are selective for discriminating between the effects of the different doses of irradiation. The results also revealed that longer post-irradiation time leads to the relaxation of the cells to their initial state. The molecular changes that occurred in the 2Gy samples were mostly reversible. On the other hand, the exposure to doses higher than 4Gy induced serious irreversible changes, mainly seen in 2700-2800 cm-1 in Raman spectra. The classification models developed in this study would help to predict the radiation-based molecular changes induced in the cancer cells by only using RS. Also, this designed framework may facilitate the process of biodosimetry.

Evaluation of the electromagnetic field intensity in operating rooms and estimation of occupational exposures of personnel.

INTRODUCTION: Operating rooms in hospitals are facilitated with different types of electronic systems, which produce electromagnetic waves. High intensities of magnetic waves may have harmful effects on biological environments. This study aims to evaluate the electromagnetic field intensity at different parts of operating rooms at the first stage and estimate the occupational exposure to operating room personnel at the next phase. MATERIALS AND METHODS: At this cross-sectional study, the magnetic field intensity was evaluated using teslameter at several parts of operating rooms, during operating procedures, while electrical instruments were working. Background electromagnetic field intensity was measured when all the electrical systems were idle. Statistical analysis was performed using SPSS software. The results were compared with ICNIRP standards. RESULTS: The maximum intensity of magnetic field was measured around high-voltage systems at the distance of 50 cm in the personnel's standing area at DCR and PCNL operating procedures were 5.9 and 5.6, respectively. The number of on-mode electrical systems was inconsistent with the intensity of electromagnetic fields at the standing area of operating room personnel's. The intensity of magnetic fields around high-voltage systems, which was about 46.75 mG at the distance of 10 cm, was the highest among measured electromagnetic fields. CONCLUSIONS: The highest magnetic field intensity measured in this study was related to high-voltage systems and is lower than advised intensity by ICNIRP for occupational exposure. Based on this study, it can be concluded that there are no considerable risks of electromagnetic exposure for operating room personnels.

The combination of A-966492 and Topotecan for effective radiosensitization on glioblastoma spheroids.

Radiotherapy is one of the modalities in the treatment of glioblastoma patients, but glioma tumors are resistant to radiation and also chemotherapy drugs. Thus, researchers are investigating drugs which have radiosensitization capabilities in order to improve radiotherapy. PARP enzymes and topoisomerase I enzymes have a critical role in repairing DNA damage in tumor cells. Thus, inhibiting activity of these enzymes helps stop DNA damage repair and increase DSB lethal damages. In the current study, we investigated the combination of TPT as a topoisomerase I inhibitor, and A-966492 as a novel PARP inhibitor for further radiosensitization. U87MG cells (a human glioblastoma cell line) were cultured in Poly-Hema coated flasks to reach 300 µm-diameter spheroids. Treatments were accomplished by using non-toxic concentrations of A-966492 and Topotecan. The surviving fraction of treated cells was determined by clonogenic assay after treatment with drugs and 6 MV X-ray. The γ-H2AX expression was measured by an immunofluorescence staining method to examine the influence of A-966492, TPT and radiation on the induction of double stranded DNA breaks. Treatments using the A-966492 drug were conducted in concentration of 1 µM. Combining A-966492 and TPT with radiation yielded enhanced cell killing, as demonstrated by a sensitizer enhancement ratio at 50% survival (SER50) 1.39 and 1.16 respectively. Radio- and chemo-sensitization was further enhanced when A-966492 was combined with both X-ray and TPT, with SER50 of 1.53. Also γ-H2AX expression was higher in the group treated with a combination of drugs and radiation. A-966492 is an effective PARP inhibitor and has significant radio-sensitivity on U87MG spheroids. By accumulating cells in the S phase and by inhibiting the DNA damage repair, TPT enhanced radio-sensitivity. A-966492 combined with TPT as a topoisomerase I inhibitor had additive radio-sensitizing effects. As a result, applying PARP and topoisomerase I inhibitors can be a suitable strategy for improving radiotherapy in clinics.

Photothermal therapy using folate conjugated gold nanoparticles enhances the effects of 6MV X-ray on mouth epidermal carcinoma cells.

The aim of this study was to develop an optimized method for preparation of folate conjugated gold nanoparticles (F-AuNPs) and to investigate its cytotoxic effects and cell apoptosis in combination with photothermal therapy (PTT) and radiotherapy (RT) for the treatment of mouth epidermal carcinoma cells KB. For this purpose, cells were treated with synthesized F-AuNPs at different concentrations for 6h and then irradiated them with laser beam (532nm, 0.5W/cm2, 15min). After photothermal therapy, the cells were exposed to 6MV X-ray with a single dose of 2Gy. MTT assay were performed to evaluate the cell survival rate and apoptosis was determined by flow cytometry using an annexin V-fluorescein isothiocyanate/propidium iodide apoptosis detection kit. No significant cell damage or cell apoptosis from the individual treatment of laser light or F-AuNPs was observed, while viability of cells incubated with F-AuNPs and then exposed to the laser was significantly decreased. Additionally, our results demonstrated that F-AuNPs is good radiosensitizers even at a low concentration such as 20µM when megavoltage X-ray is used. Also, when KB cells were treated with F-AuNPs under both laser and X-ray irradiation, the cell viability substantially decreased more than F-AuNPs-enhanced PTT alone or F-AuNPs-enhanced RT alone. Flow cytometry assay clearly indicated that F-AuNPs-mediated photo-thermo-radio therapy significantly induced apoptosis. These results confirm that F-AuNPs is a promising and research-worthy nanoconjugate in the field of targeted photo-thermo-radiotherapy of cancer.

Synergistic Effects of Arsenic Trioxide and Radiation: Triggering the Intrinsic Pathway of Apoptosis

Background: Arsenic trioxide (ATO) has been reported as an effective anti-cancer and a US Food and Drug Administration (FDA) approved drug for treatment of some cancers. The aim of this study was to determine the underlying apoptosis molecular and cellular mechanisms of ATO in the presence or absence of ionizing radiation (IR) in vitro in the glioblastoma multiforme (GBM) cell line, U87MG. Methods: Cells were treated by different concentrations of ATO either in presence or absence of IR. Viability and apoptosis pathway of both treated and control groups were evaluated using MTT assay and the expression analysis of Bax, Bcl-2, and caspase-3 genes, respectively. All treatments were performed on 100-µm diameter spheroids. Results: Results showed a significant reduction in the survival of the cells in all treated groups. As expected, cell survival was much less in combination treatment than treatment with only ATO. Moreover, combination therapy made Bax and caspase-3 up-regulated and Bcl-2 down-regulated. Conclusion: ATO and radiation had a synergistic apoptotic effect on GBM cells by up-regulation of caspase-3 and alteration of the Bax-Bcl-2 balance; therefore, ATO may act as a potential anti-cancer agent against GBM cells through triggering the mitochondrial pathway of apoptosis.

Evaluation of the cytotoxic effects of hyperthermia and 5-fluorouracil-loaded magnetic nanoparticles on human colon cancer cell line HT-29.

The purpose of this study was to evaluate the combined effects of heat and polylactic-co-glycolic acid (PLGA) nanoparticles, as 5-fluorouracil carriers with/without iron oxide core, on the viability and proliferation capacity of human colon cancer cell line HT-29 in the spheroid model. HT-29 spheroid cells were treated with different concentrations of 5-FU or 5-FU-loaded into both nanoparticles for 74 h. Hyperthermia was then performed at 43 °C for 60 min. Finally, the effects of the mentioned treatments on cell viability and proliferation capacity were evaluated using the trypan blue dye exclusion test and colony formation assay, respectively. Our results showed that hyperthermia, in combination with 5-FU or PLGA nanoparticles as 5-FU carriers, significantly enhanced the cytotoxic effects as compared to the control group. Considering that nanoparticles could increase the intracellular concentration of drugs in cancer cells, the extent of cytotoxic effects following treatment with 5-FU-loaded into both nanoparticles was significantly higher than that with free 5-FU. In addition, the presence of iron oxide cores in nanoparticles during hyperthermia enhanced the cytotoxic effects of hyperthermia compared with nanoparticles without iron oxide core. Based on this study, hyperthermia in combination with 5-FU-loaded PLGA nanoparticles with iron oxide core drastically reduced the proliferation capacity of HT-29 cells; therefore, it may be considered as a new direction in the treatment of colon cancer.

Enhancement of radiosensitivity of melanoma cells by pegylated gold nanoparticles under irradiation of megavoltage electrons.

PURPOSE: Gold nanoparticles (GNP) have significant potential as radiosensitizer agents due to their distinctive properties. Several studies have shown that the surface modification of nanoparticles with methyl polyethylene glycol (mPEG) can increase their biocompatibility. However, the present study investigated the radiosensitization effects of mPEG-coated GNP (mPEG-GNP) in B16F10 murine melanoma cells under irradiation of 6 MeV Electron beam. MATERIALS AND METHODS: The synthesized GNP were characterized by UV-Visible spectroscopy, dynamic light scattering, transmission electron microscopy, and zeta potential. Enhancement of radiosensitization was evaluated by the clonogenic assay at different radiation doses of megavoltage electron beams. RESULTS: It was observed that mPEG-GNP with a hydrodynamic size of approximately 50 nm are almost spherical and cellular uptake occurred at all concentrations. Both proliferation efficiency and survival fraction decreased with increasing mPEG-GNP concentration. Furthermore, significant GNP sensitization occurred with a maximum dose enhancement factor of 1.22 at a concentration of 30 µM. CONCLUSIONS: Pegylated-GNP are taken up by B16F10 cancer cells and cause radiosensitization in the presence of 6 MeV electrons. The radiosensitization effects of GNP may probably be due to biological processes. Therefore, the underlying biological mechanisms beyond the physical dose enhancement need to be further clarified.

Role of iron oxide core of polymeric nanoparticles in the thermosensitivity of colon cancer cell line HT-29.

PURPOSE: In this study the effect of PLGA polymeric nanoparticles as a 5-fluorouracil (5-FU) carrier with and without iron oxide core and hyperthermia were investigated on the level of DNA damage in a spheroid culture model of HT-29 colon cancer cell lines by alkaline comet assay. MATERIALS AND METHODS: First, HT-29 colon cancer cells were cultured in vitro as spheroids with a mean diameter of 100 µm. The spheroids were then treated with different concentrations of 5-FU or nanoparticles as 5-FU carriers with and without an iron oxide core for one volume-doubling time of the spheroids (71 h) and hyperthermia at 43 °C for 1 h. Finally, the effect of treatment on viability and level of DNA damage was examined using trypan blue dye exclusion assay and alkaline comet assay, respectively. RESULTS: Results showed that hyperthermia in combination with 5-FU or nanoparticles as 5-FU carriers significantly induced the most DNA damage as compared with the control group. The extent of DNA damage following treatment with 5-FU-loaded nanoparticles combined with hyperthermia was significantly more than for 5-FU combined with hyperthermia. In comparison to the effect of 5-FU-loaded nanoparticles with the iron oxide core and 5-FU-loaded nanoparticle without the iron oxide core, the nanoparticles with the iron oxide core combined with hyperthermia induced more DNA damage than the nanoparticles without the iron oxide core. CONCLUSIONS: According to this study, hyperthermia is a harmful agent and nanoparticles are effective delivery vehicles for drugs into colon cancer cells. The iron oxide filled nanoparticles increased the effect of the hyperthermia. All these factors have a significant role in the treatment of colorectal cancer cells.