Curcumin combined with Glu-GNPs enhancing radiosensitivity of transplanted tumor of MDA-MB-231-luc cells in nude mice

Breast cancer is the first cause of cancer death in women all over the world. And the morbidity of breast cancer has been increasing in recent years. Nowadays, the treatment of breast cancer is still a problem. Radiotherapy resistance is one of the important factors leading to poor prognosis of clinical treatment. Curcumin, a plant polyphenol from Curcuma longa, has antitumor activity and inhibition of tumor recurrence and metastasis. However, the influence of curcumin on radiosensitivity of breast carcinoma xenografts, the synergistic effect and possible mechanism of curcumin and Glucose-Gold nanoparticles (Glu-GNPs) in nude mice in vivo need to be further explored. The model of transplanted tumor in nude mice was established and sensitized by intravenous injection of curcumin and Glu-GNPs. The results showed that curcumin can significantly inhibit the growth of tumor, and has no obvious effect on the body weight of nude mice, and Curcumin, Glu-GNPs and X-ray irradiation treatment alone or in combination could reduce the expression of VEGF and HSP90 mRNA and protein compared with model group in tumor tissue. The inhibition of irradiation resistance may be related to inhibiting the synthesis of VEGF and HSP90. This study suggested that curcumin and Glu-GNPs have the radiosensitization effect in vivo, which provides an experimental basis for the clinical development of green radiosensitizers.

is no effective strategies to deal with breast cancer metastasis. Establishing a nude 66 mouse model of breast cancer xenograft and studying the important related genes in 67 breast cancer growth and development, is an important breast cancer research method, 68 and plays a key role in studying the etiology, pathogenesis, effective prevention, 69 diagnosis and treatment, drug screening and rehabilitation of breast cancer [5] . In 70 recent years, bioluminescence imaging system has become a sensitive research 71 method in animal model. It can monitor the growth and metastasis of tumor in real 72 time. In addition, it can be accurately detected in vivo without killing mice [6] . Zhang 73 et al [7] found that the results of in vivo imaging and pathological detection are 74 consistent, but compared with HE staining, the growth and metastasis of breast cancer 75 can be more directly and clearly observed using in vivo imaging system. In vivo 76 imaging method is applied to study transplanted tumor in mice. Compared with 77 traditional measurement method, in vivo imaging shows scientific methods to 78 measure tumor growth more accurately, which greatly reduces the error of artificial 79 measurement.

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The development of Gold nanoparticles (GNPs) can be traced back to the 16th 81 century. GNPs, a type of nanomaterial approved for clinical trials by FDA in the US, 82 had become a hot area in scientific research [8] . It was reported that when GNPs 83 received X-ray irradiation, the ionizing ability of the radiation to tumor cells was 84 enhanced, which promoted tumor cell damage and apoptosis [9][10][11][12] . Our previous study 85 suggested that Glu-GNPs assimilated in MCF-7 adherent cells and THP-1 suspension 86 cells were easier than GNPs and enhanced the cancer killing of THP-1 cells 20% 87 more than GNPs [13] . It pointed that glucose tagging may be a correct way to 88 promoting the uptake of GNPs in tumor cells, as tumor cells can take up more glucose 89 than normal cells. In addition, it was previously demonstrated that these nanometal 90 particles enhanced killing effects on tumor cells without increasing damage to the 91 surrounding normal tissues in a mouse model, thereby reducing the adverse effects of 92 radiotherapy [14][15] .

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In recent years, scientific research has found that traditional Chinese materia 94 medica has unique advantages and efficacy in treating tumors. Curcumin, a plant 95 polyphenol extracted from the rhizome of Curcuma longa, is the main ingredient of 96 curry, with the characteristics of high efficiency, safety and low toxicity [16] . It was 97 reported that curcumin has killing and therapeutic effects on many kinds of cancer 98 including lung cancer [17] , gastric cancer [18] and breast cancer [19] in vitro and in vivo. 99 Hu Shan, et al [20] found that curcumin can inhibit the proliferation of various breast 100 cancer cells such as MDA-MB-231, MCF-7, MDA-MB-468 and T47D, which also 101 induced cell apoptosis. Palange's study showed that curcumin treatment can limit the 5 102 metastatic potential of breast cancer cell lines, possibly by altering the expression of 103 related adhesion molecules [21] . These findings suggested that curcumin may have 104 important research significance in the treatment of breast cancer. However, it reminds 105 to be studied whether curcumin enhances radiosensitivity effect and curcumin has 106 synergistic effect with Glu-GNPs. And the mechanism of curcumin on breast cancer 107 is not clear.     lying on their sides, and the tumor was exposed. The fluorescence intensity in all 9 183 groups of mice was detected and recorded 10 minutes after the injection by using 184 in-vivo bioluminescence imaging system. 186 Tumor tissue samples were fixed in 4% paraformaldehyde at 4℃ for 48 hours.

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After that, the samples were dehydrated, embedded, paraffin sectioned and stained  increased correspondingly (Fig 1A and B). It indicated that MDA-MB-231-luc cells 224 could express luciferase stably. Three to seven days after inoculated, nude mice formed tumors gradually and the 241 tumor formation rate was 100% except for control group (Fig 1C). The shape of the from that of the model group (Fig 2C and D). and Cur+Glu-GNPs+IR group decreased significantly (Fig 3). It also showed that  The results showed that breast carcinoma xenograft was invasive ductal carcinoma 296 with high malignancy. (Fig 4A).  tumor volume and the body weight of mice, the curative effect of drugs was 348 preliminarily evaluated. We found that cisplatin was effective in the treatment 349 process, but the body weight loss of mice in cisplatin group was obvious. X-ray has synergistic therapeutic effect.

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Studies have shown that its radiotherapy resistance mechanism is that blood 362 vessels are irregularly distributed in tumor tissue, blood circulation is blocked, 363 hypoxic areas exist in tumor tissue, hypoxic cells have high radiation tolerance and 364 DNA damage of tumor stem cells [23] . Therefore, the combination of drugs and 365 irradiation may be sensitive to tumor cells. In this study, we found that curcumin   Hu. 19