|Year : 2018 | Volume
| Issue : 1 | Page : 74-77
Comparison between antitumor activity of live-attenuated measles virus and cisplatin on Ki-67 expression of colon cancer cell line (SW-480) In vitro
Rasha Fadhel Obaid1, Younis Abdul-Redha K Al-Khafaji2, Hamid Naji Obied3, Salman Azeez AL- Jibouri1
1 Department of Microbiology, College of Medicine, Kufa University, Najaf, Iraq
2 Department of Microbiology, College of Dentistry, Babylon University, Hilla, Iraq
3 Department of Pharmacology, College of Medicine, Babylon University, Hilla, Iraq
|Date of Web Publication||19-Mar-2018|
Hamid Naji Obied
Department of Pharmacology, College of Medicine, University of Babylon, Hilla
Source of Support: None, Conflict of Interest: None
Background: Previous studies declare that Ki-67 protein (Ki-67P) expression was shown to be correlated with cell proliferation and was considered as a prognostic marker for different cancer diseases. Objective: The aim of this study was to investigate the relation between Ki-67 and the cytotoxic effect of MV and cisplatin chemotherapy in inhibition of the proliferation of human colon cancer cell line. Materials and Methods: Colon cancer cell line (SW-480) was cultured in microtiter plates incubated in the presence of different titers of measles virus and different concentrations of cisplatin. The expression titer of Human Ki-67 protein was determined in cell culture supernatant by quantitative ELISA. The optical density (OD) was measured with spectrophotometry at a wavelength of 450 nm. The OD value is proportional to the concentration of human Ki-67P. The concentration of human Ki-67P in samples was calculated by comparing the OD of the samples with the standard curve. Results: The expression of Ki-67P was decreased in relation to the virus titer and concentration of cisplatin in treated cell groups. The results showed significant differences in the level of cellular expression of protein (Ki-67) between the virus-infected cells, cisplatin-treated, and the l untreated control cells group (P ≤ 0.001). The lowest Ki-67P expression was recorded after treatment of SW-480 colon tumor cell with high concentration of measles virus. Conclusion: Both live-attenuated MV (Edmonton strain) and cisplatin could reduce Ki-67P expression in tumor cell line when treated with either one of them. Therefore, they have beneficial effects in reducing the resistance to chemotherapy and radiotherapy.
Keywords: Cisplatin, Ki-67, measles virus, SW-480
|How to cite this article:|
Obaid RF, Al-Khafaji YRK, Obied HN, AL- Jibouri SA. Comparison between antitumor activity of live-attenuated measles virus and cisplatin on Ki-67 expression of colon cancer cell line (SW-480) In vitro. Med J Babylon 2018;15:74-7
|How to cite this URL:|
Obaid RF, Al-Khafaji YRK, Obied HN, AL- Jibouri SA. Comparison between antitumor activity of live-attenuated measles virus and cisplatin on Ki-67 expression of colon cancer cell line (SW-480) In vitro. Med J Babylon [serial online] 2018 [cited 2019 Jul 22];15:74-7. Available from: http://www.medjbabylon.org/text.asp?2018/15/1/74/227816
| Introduction|| |
Cancer is one of the worldwide leading causes of death. Although significant advances were made in cancer treatment, the mortality rates for most malignancies remain alarmingly high. Colorectal cancer is the third common cancer after breast cancer in female and bronchial cancer in male and the second leading cause of death among cancers in western world., Cancer chemotherapies were primarily selected through a panel for agents that killed the rapidly dividing cells. The cytotoxic drugs are still the cornerstone of current treatment, but the benefit was limited due to the narrow therapeutic index, harmful toxicities, and frequently arising resistance. The treatment of cancers with oncolytic viruses offers a very promising approach. These are “living” agents which specifically infect and kill tumor cells as part of the virus replication process. Huge numbers of progeny virions are released, which in turn attack further tumor cells.,
During the past decade, measles virus (MV) vaccine strains have emerged oncolytic platform virus, that may not be compromised by preexisting MV immunity. MV vaccine Edmonston or Schwarz strains use mainly CD46 molecules to infect cells. During cancer development, tumor cells often express high levels of CD46 molecules, which inhibit the complement system. This CD46 overexpression makes the tumor cells less sensitive to lysis by the complement but highly sensitive to MV vaccine infection. These viruses, either naturally occurring or genetically engineered, are replication-competent viruses that are able to selectively infect and destroy cancer cells either intratumoral or systemic administration. Oncolytic viruses display an anticancer mechanism in comparison to ordinary treatments (chemotherapy and radiotherapy), permitting the opportunity for synergistic effect in cancer treatments. Slowing the growth and progression of cancer cells is one of the major tasks that faced by modern medicine. Ki-67 as a nuclear protein expressed in dividing mammalian cells is widely used in cancer biology as prognostic marker of cancer and has been widely investigated. Nishimura et al. suggest that the Ki-67 titer before neoadjuvant chemotherapy is a sold diagnostic parameter for evaluation of the usefulness of the chemotherapies, also they found that the pathological documented response was significantly associated with Ki-67 values using different analysis. Patients with higher cell proliferation (Ki-67 >25%) may be considered better candidates for neoadjuvant chemotherapy. After neoadjuvant chemotherapy, lower Ki-67 values indicate a low chance for pathological complete response but a better prognosis. The aim of this study was to investigate the relation between Ki-67 and the cytotoxic effect of MV and cisplatin chemotherapy in inhibition of the proliferation of human colon cancer cell line.
| Materials and Methods|| |
Live-attenuated MV (Edmonston) was obtained from vaccine and sera institute in Baghdad. Each vial vaccine contains 10 doses of lyophilized hyperattenuated MV (Edmonton strain); each dose contains at least 1000 CCID50. Virus was prepared as recommended by manufacturing company (serum institute of India Ltd (siil) ) by adding 5 ml of sterile DDW to the lyophilized powder with continuous stirring. Then stored at -80o C till use.
The human colon cancer cell line (SW-480) was purchased from National Cell Bank of Iran, Pasteur Institute. They were cultured in medium 1640 (RPMI-1640, Gibco-BRL), with 10% heat-inactivated fetal bovine serum (Gibco) and penicillin/streptomycin (100 U/mL) (Sigma). Cell line was grown at 37°C incubation. Cells were plated in 25 cm 2 tissue culture flasks for the subsequent experiments. Information was recorded on data sheet and flask checked daily for observation of any changes in media and/or monolayer formation.
Colon cancer cell line (SW-480) was cultured in four microtiter plates of 96-well flat-bottomed and incubated in the presence of different titers of MV and different concentrations of cisplatin. The plated cells were divided into three groups: Group 1 was the untreated control group; Group 2 was MV-treated group; and Group 3 cell line that treated with cisplatin. Three replicates were used for each dilution as well as the control wells which were treated with serum-free medium only. The cells were incubated at 37°C in CO2 incubator for 24, 48, 72, and 96 h; then, the supernatants were collected, and the cells were detached for the estimation of Ki-67 protein (Ki-67P).
Estimation of human Ki-67 protein
Ki-67P concentrations were determined in cell culture supernatant by quantitative ELISA Kit (Abcam). The optical density (OD) could be measured with spectrophotometry at a wavelength of 450 nm ± 2 nm. The OD value is proportional to the concentration of human Ki-67P. The concentration of human Ki-67P in samples was calculated by comparing the OD of the samples with the standard curve.
Statistical analysis was done using SPSS (statistical package for social sciences) version 20 in which we used mean and standard deviation as descriptive statistics and analysis of variance with least significant difference for comparison between groups. P ≤ 0.05 was considered statistically significant.
| Results|| |
Ki-67 profile was measured by ELISA test in supernatant of human colon cell line that was treated with different titer of MV (10−1, 10−2, 10−3, 10−4, and 10−5), different concentration of cisplatin (3.125, 6.25, 12.5, 25, and 50 μg/ml), and non-treated cell group for 24, 48,72, and 96 h.
The study has shown that the Ki-67P expression in colon (SW-480) cell line that had treated with high dosage of MV was 63.529 ± 3.116 after 24 h of inoculation and then started to decrease gradually with increased postinoculation time intervals and reach its minimum value of Ki-67 expression at 96 h postinoculation which was 22.530 ± 1.167 in colonic tumor cells which was more lower than control nontreated cell group.
Whereas Ki-67P expression level after treatment with high concentration of cisplatin was 79.136 ± 0.568 at 24 h post treatment and then gradually decrease to reach its lowest mean value of Ki-67 (38.233 ± 0.651) at 96 h after treatment. [Table 1] and [Figure 1] and [Figure 2] reflected significant differences in the level of KI-67P expression between cell that was treated with (measles virus or cisplatin ) and untreated control group.
|Table 1: Comparison between antitumor activity of live-attenuated measles virus and cisplatin on Ki-67 expression of colon cancer cell lines|
Click here to view
|Figure 1: Influence of cisplatin on Ki-67 protein expression in SW-480 colon cancer cells. Data are represented as mean ± standard deviation (n = 3) (P < 0.001)|
Click here to view
|Figure 2: Influence of attenuated measles virus (Edmonton strain) on Ki-67 protein expression in SW-480 colon cancer cells. Data are represented as mean ± standard deviation (n = 3) (P < 0.001)|
Click here to view
| Discussion|| |
Beside that, Ki-67P expression titer reflects the biological activity of tumor cells; also, it clinically has clinical correlation with metastasis and stage of tumors. In addition, it is significantly higher in malignant tissues with poorly differentiated tumor cells in comparison with normal tissue. It was used as a prognostic and predictive marker for cancer diagnosis and treatment, increasing the evidence which may indicate that Ki-67P may be an effective cellular target in cancer therapy. Ki-67P expression in proliferating cancer is a prognostic marker for various cancers.
In the present study, the association between the Ki-67 expression and the therapeutic effects of MV and cisplatin was investigated in colon cancer cell line that treated with different titer of MV and different concentration of cisplatin at different periods of time. The results revealed a reduction in expression of Ki-67P proliferation marker in cancer cell line that was treated with MV and cisplatin in comparison with control group. Ki-67 expression was decreased with increasing titer of MV and concentration of cisplatin.
This result is consistent with other studies which were documented a reduced expression of Ki-67P proliferation markers in cancer cell line that treated with MV or cisplatin. Shi and Tian  reported that cisplatin can inhibit the growth of HeLa cell and decreased expression of Ki-67Ps during treatment with different concentrations of cisplatin. Zhao et al. showed that Ki-67 expression was declined within the tumors which treated with 106 CCID50/ml MV, compared with the control group which treated with phosphate buffer saline. Furthermore, the result of this study is also consistent with previous studies which were documented that oncolytic virus inhibits Ki-67 expression during growth leading to inhibition of cancer cell line. Li et al. found that adenovirus could inhibit Ki-67 expression and cellular proliferation beside induce apoptosis of prostate carcinoma in vivo, which is consistent with previous studies in vitro experiments. These observations indicated that Ki-67 may be an effective target in cancer therapy. The results documented that the inhibition of cell growth was associated with a decrement in the expression of Ki-67 protein and increased in the ratio apoptosis compared with control group. And the inhibition of cell growth an important mechanism of anti-tumor activity of measles virus and cisplatin.
| Conclusions|| |
Both live-attenuated MV (Edmonton strain) and cisplatin could reduce Ki-67P expression in tumor cell line when treated with either one of them. Therefore, they have beneficial effects in reducing the resistance to chemotherapy and radiotherapy.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Singh PK, Doley J, Kumar GR, Sahoo AP, Tiwari AK. Oncolytic viruses & their specific targeting to tumour cells. Indian J Med Res 2012;136:571-84.
] [Full text]
George EL, Suresh HM. Multistage carcinogenesis and the incidence of colorectal cancer. Proc Natl Acad Sci 2002;99:20-2.
Siegel RL, Miller KD, Jemal A. Cancer statistics, 2016. CA Cancer J Clin 2016;66:7-30.
Jemal A, Murray T, Ward E, Samuels A, Tiwari RC, Ghafoor A, et al.
Cancer statistics, 2005. CA Cancer J Clin 2005;55:10-30.
Hartkopf AD, Fehm T, Wallwiener D, Lauer U. Oncolytic virotherapy of gynecologic malignancies. Gynecol Oncol 2011;120:302-10.
Hartkopf AD, Fehm T, Wallwiener M, Lauer U. Oncolytic viruses to treat ovarian cancer patients – A review of results from clinical trials. Tumor Diagn Ther 2012;33:279-83.
Huang F, Wang BR, Wu YQ, Wang FC, Zhang J, Wang YG, et al.
Oncolytic viruses against cancer stem cells: A promising approach for gastrointestinal cancer. World J Gastroenterol 2016;22:7999-8009.
Knuchel MC, Marty RR, Morin TN, Ilter O, Zuniga A, Naim HY, et al.
Relevance of a pre-existing measles immunity prior immunization with a recombinant measles virus vector. Hum Vaccin Immunother 2013;9:599-606.
Fishelson Z, Donin N, Zell S, Schultz S, Kirschfink M. Obstacles to cancer immunotherapy: Expression of membrane complement regulatory proteins (mCRPs) in tumors. Mol Immunol 2003;40:109-23.
Guillerme JB, Boisgerault N, Roulois D, Ménager J, Combredet C, Tangy F, et al.
Measles virus vaccine-infected tumor cells induce tumor antigen cross-presentation by human plasmacytoid dendritic cells. Clin Cancer Res 2013;19:1147-58.
Zamarin D, Pesonen S. Replication-competent viruses as cancer immunotherapeutics: Emerging clinical data. Hum Gene Ther 2015;26:538-49.
Nishimura R, Osako T, Okumura Y, Hayashi M, Arima N. Clinical significance of ki-67 in neoadjuvant chemotherapy for primary breast cancer as a predictor for chemosensitivity and for prognosis. Breast Cancer 2010;17:269-75.
Jonat W, Arnold N. Is the ki-67 labelling index ready for clinical use? Ann Oncol 2011;22:500-2.
Ghaleb RA. The effect of diode laser 635nm on mitochondrial membrane potential and apoptosis induction of CHO47 cells lin. Med J Babylon 2016;13:7-16.
Li LT, Jiang G, Chen Q, Zheng JN. Ki67 is a promising molecular target in the diagnosis of cancer (review). Mol Med Rep 2015;11:1566-72.
Cidado J, Wong HY, Rosen DM, Cimino-Mathews A, Garay JP, Fessler AG, et al.
Ki-67 is required for maintenance of cancer stem cells but not cell proliferation. Oncotarget 2016;7:6281-93.
Shi ZP, Tian XZ. Effect of cisplatin on apoptosis of cervical cancer cell line HeLa and the expression of FHIT, Ki67 and Bax proteins. J Shanxi Med Univ 2008;10:012.
Zhao D, Chen P, Yang H, Wu Y, Zeng X, Zhao Y, et al.
Live attenuated measles virus vaccine induces apoptosis and promotes tumor regression in lung cancer. Oncol Rep 2013;29:199-204.
Li J, Luo J, Gu D, Jie F, Pei N, Li A, et al.
Adenovirus-mediated angiotensin II type 2 receptor overexpression inhibits tumor growth of prostate cancer in vivo
. J Cancer 2016;7:184-91.
[Figure 1], [Figure 2]