Characteristics of sphingomyelin metabolism in the MCF7 and BT474 radiotherapy­resistant HER2­positive breast cancer cell lines.

Breast cancer is the most common cancer globally in terms of incidence. This cancer is classified into subtypes based on histological or immunological characteristics. HER2-positive cases account for 15-25% of breast cancer cases, and one of the first events in breast carcinogenesis is HER2 upregulation. Furthermore, HER2 expression increases the detection rate of metastatic or recurrent breast cancers by 50-80%. The epidermal growth factor receptor family includes HER2, which is a transmembrane receptor protein. In our previous case report, patients who were resistant to anti-HER2 monoclonal antibody therapy, chemotherapy and radiotherapy had higher concentrations of phospholipid metabolites such as phosphatidylcholine and sphingomyelin (SM), which was associated with cancer recurrence progression. To better understand the relationship between radiotherapy resistance and SM expression, breast cancer cell lines with and without HER2 expression (MCF7 and BT474) after exposure to ionizing radiation (IR) were examined. In the cell culture supernatant, similar levels of SM in MCF7 cells were identified after 1-4 Gy exposure. However, SM levels in BT474 cells were upregulated compared with those of in the control group. Intracellular SM levels were upregulated in BT474 cells exposed to 1 and 4 Gy compared with the non-irradiated control group. Furthermore, significantly increased mRNA expression levels of sphingomyelin synthase 2 (SGMS2) in BT474 cells exposed to IR were observed compared with those in nonirradiated cells; however, the SGMS2 levels in MCF7 cells did not differ significantly among the 0, 2 and 4 Gy groups. These findings suggested that a higher dose of IR induced the secretion of SM and its associated gene expression in HER2-positive breast cancer cells.

Additive antitumor effect of arsenic trioxide with exposure to ionizing radiation to human acute promyelocytic leukemia HL­60 cells.

Arsenic trioxide (ATO) is expected to be a chemical drug with antitumor activity against acute promyelocytic leukemia (APL), a type of acute myeloid leukemia. In Japan, its antitumor effects were confirmed in clinical trials for APL, and it has been approved in various countries around the world. However, there have been no reports on ATO's antitumor effects on radioresistant leukemia cells, which can be developed during radiotherapy and in combination with therapeutic radiation beams. The present study sought to clarify the antitumor effect of ATO on APL cells with radiation resistance and determine its efficacy when combined with ionizing radiation (IR). The radiation­resistant HL60 (Res­HL60) cell line was generated by subjecting the native cells to 4­Gy irradiation every week for 4 weeks. The half­maximal inhibitory concentration (IC50) for cell proliferation by ATO on native cell was 0.87 µM (R2=0.67), while the IC50 for cell proliferation by ATO on Res­HL60 was 2.24 µM (R2=0.91). IR exposure increased the sub­G1 and G2/M phase ratios in both cell lines. The addition of ATO resulted in a higher population of G2/M after 24 h rather than 48 h. When the rate of change in the sub­G1 phase was examined in greater detail, the sub­G1 phase in both control cells without ATO significantly increased by exposure to IR at 24 h, but only under the condition of 2 Gy irradiation, it had continued to increase at 48 h. Res­HL60 supplemented with ATO showed a higher rate of sub­G1 change at 24 h; however, 2 Gy irradiation resulted in a decrease compared with the control. There was a significant increase in the ratio of the G2/M phase in cells after incubation with ATO for 24 h, and exposure to 2 Gy irradiation caused an even greater increase. To determine whether the inhibition of cell proliferation and cell cycle disruptions is related to reactive oxygen species (ROS) activity, intracellular ROS levels were measured with a flow cytometric assay. Although the ROS levels of Res­HL60 were higher than those of native cells in the absence of irradiation, they did not change after 0.5 or 2 Gy irradiation. Furthermore, adding ATO to Res­HL60 reduced intracellular ROS levels. These findings provide important information that radioresistant leukemia cells respond differently to the antitumor effect of ATO and the combined effect of IR.

Exploring predictive molecules of acute adverse events in response to volumetric­modulated arc therapy for prostate cancer using urinary metabolites.

Volumetric-modulated arc therapy (VMAT) is a radiotherapy technique used to treat patients with localized prostate cancer, which is frequently associated with acute adverse events (AEs) that can affect subsequent treatment. Notably, the radiation dose of VMAT can be tailored to each patient. In the present study, a retrospective analysis was performed to predict acute AEs in response to a therapeutic high radiation dose rate based on urinary metabolomic molecules, which are easily collected as noninvasive biosamples. Urine samples from 11 patients with prostate cancer who were treated with VMAT (76 Gy/38 fractions) were collected. The study found that seven patients (~64%) exhibited genitourinary toxicity (Grade 1) and four patients had no AEs. A total of 630 urinary metabolites were then analyzed using a mass spectrometer (QTRAP6500+; AB SCIEX), and 234 relevant molecules for biological and clinical applications were extracted from the absolute quantified metabolite values using the MetaboINDICATOR tool. In the Grade 1 acute AE group, there was a significant negative correlation (rs=-0.297, P<0.05) between the number of VMAT fractions and total phospholipase A2 activity in the urine. Additionally, patients with Grade 1 AEs exhibited a decrease in PC aa C40:1, a phospholipid. These findings suggested that specific lipids found in urinary metabolites may serve as predictive biomarkers for acute AEs in response to external radiotherapy.