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Targeted Knockdown of the Kinetochore Protein D40/Knl-1 Inhibits Human Cancer in a p53 Status-Independent Manner.

Urata YN, Takeshita F, Tanaka H, Ochiya T, Takimoto M - Sci Rep (2015)

Bottom Line: The D40 gene encodes a kinetochore protein that plays an essential role in kinetochore formation during mitosis.These results indicated that D40 siRNA induced apoptotic cell death in human cancer cell lines, and inhibited their growth in vitro and in vivo regardless of p53 status.Therefore, D40 siRNA is a potential candidate anti-cancer reagent.

View Article: PubMed Central - PubMed

Affiliation: Division of Cancer Gene Regulation, Institute for Genetic Medicine, Hokkaido University, Sapporo, Hokkaido, Japan.

ABSTRACT
The D40 gene encodes a kinetochore protein that plays an essential role in kinetochore formation during mitosis. Short inhibitory RNA against D40, D40 siRNA, has been shown to deplete the D40 protein in the human cancer cell line HeLa, which harbors wild-type p53, and this activity was followed by the significant inhibition of cell growth and induction of apoptotic cell death. The p53- cancer cell line, PC-3M-luc, is also sensitive to the significant growth inhibition and cell death induced by D40 siRNA. The growth of PC-3M-luc tumors transplanted into nude mice was inhibited by the systemic administration of D40 siRNA and the atelocollagen complex. Furthermore, D40 siRNA significantly inhibited growth and induced apoptotic cell death in a cell line with a gain-of-function (GOF) mutation in p53, MDA-MB231-luc, and also inhibited the growth of tumors transplanted into mice when administered as a D40 siRNA/atelocollagen complex. These results indicated that D40 siRNA induced apoptotic cell death in human cancer cell lines, and inhibited their growth in vitro and in vivo regardless of p53 status. Therefore, D40 siRNA is a potential candidate anti-cancer reagent.

No MeSH data available.


Related in: MedlinePlus

In vivo Growth Inhibition of p53 mutant cancer cell lines by D40 siRNA.(a) D40 siRNA inhibited the metastatic growth of the PC-3M-luc p53- cell line after transplantation into nude mice. Two million PC-3M-luc cells were transplanted into the left cardiac ventricle of nude mice. The siRNA and atelocollagen complex was injected into the tail veins of mice 4, 7, 13, and 16 days after transplantation. On day 29, the bioluminescence from the tumors was quantitatively detected. Left: Representative bioluminescent images of mice treated with the D40 siRNA/atelocollagen complex (upper) or the control (lower). Right: Statistical analysis of bioluminescent signals from the tumors in mice treated with the D40 siRNA/atelocollagen complex versus the control. Data represent the mean ± sd. (b) D40 siRNA inhibited the growth of the MDA-MB231-luc GOF mutant of p53 cell line after transplantation into scid mice. One million MDA-MB231-luc cells were transplanted into the mammary glands of scid mice. The siRNA and atelocollagen complex was injected into the tumors on mice 4, 6, 8, 11, 13, 15, and 18 days after transplantation. On day 18, the bioluminescence from the tumors was quantitatively detected. Left: Representative bioluminescent images of mice treated with D40 siRNA (upper) or control siRNA (lower). Right: Statistical analysis of the bioluminescent signals from the tumors in mice treated with the D40 siRNA/atelocollagen complex versus the control. Data represent the mean ± se.
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f5: In vivo Growth Inhibition of p53 mutant cancer cell lines by D40 siRNA.(a) D40 siRNA inhibited the metastatic growth of the PC-3M-luc p53- cell line after transplantation into nude mice. Two million PC-3M-luc cells were transplanted into the left cardiac ventricle of nude mice. The siRNA and atelocollagen complex was injected into the tail veins of mice 4, 7, 13, and 16 days after transplantation. On day 29, the bioluminescence from the tumors was quantitatively detected. Left: Representative bioluminescent images of mice treated with the D40 siRNA/atelocollagen complex (upper) or the control (lower). Right: Statistical analysis of bioluminescent signals from the tumors in mice treated with the D40 siRNA/atelocollagen complex versus the control. Data represent the mean ± sd. (b) D40 siRNA inhibited the growth of the MDA-MB231-luc GOF mutant of p53 cell line after transplantation into scid mice. One million MDA-MB231-luc cells were transplanted into the mammary glands of scid mice. The siRNA and atelocollagen complex was injected into the tumors on mice 4, 6, 8, 11, 13, 15, and 18 days after transplantation. On day 18, the bioluminescence from the tumors was quantitatively detected. Left: Representative bioluminescent images of mice treated with D40 siRNA (upper) or control siRNA (lower). Right: Statistical analysis of the bioluminescent signals from the tumors in mice treated with the D40 siRNA/atelocollagen complex versus the control. Data represent the mean ± se.

Mentions: In subsequent experiments, we determined whether D40 siRNA could inhibit the growth of tumors transplanted in experimental animals. The p53- PC-3M-luc cells were inoculated into the left cardiac ventricles of nude mice, and the D40 siRNA/atelocollagen complex, in which atelocollagen was used as a carrier for the siRNA, was then injected intravenously to examine the effects of D40 siRNA on the metastatic growth of tumors in nude mice2728. As PC-3M-luc cells were engineered to constitutively express luciferase, the growth of tumors in nude mice was monitored using an extracorporeal device after the injection of a substrate for luciferase. Representative extracorporeal bioluminescent images of tumors in mice are shown in Fig. 5a Left. The bioluminescent signals observed in mice treated with the D40 siRNA/atelocollagen complex (upper) were significantly weaker than those in control mice (lower). Statistical analysis of the quantified bioluminescent signals from mice revealed that metastatic growth of the PC-3M-luc cell line was significantly inhibited by the D40 siRNA/atelocollagen complex compared with control (Fig. 5a Right).


Targeted Knockdown of the Kinetochore Protein D40/Knl-1 Inhibits Human Cancer in a p53 Status-Independent Manner.

Urata YN, Takeshita F, Tanaka H, Ochiya T, Takimoto M - Sci Rep (2015)

In vivo Growth Inhibition of p53 mutant cancer cell lines by D40 siRNA.(a) D40 siRNA inhibited the metastatic growth of the PC-3M-luc p53- cell line after transplantation into nude mice. Two million PC-3M-luc cells were transplanted into the left cardiac ventricle of nude mice. The siRNA and atelocollagen complex was injected into the tail veins of mice 4, 7, 13, and 16 days after transplantation. On day 29, the bioluminescence from the tumors was quantitatively detected. Left: Representative bioluminescent images of mice treated with the D40 siRNA/atelocollagen complex (upper) or the control (lower). Right: Statistical analysis of bioluminescent signals from the tumors in mice treated with the D40 siRNA/atelocollagen complex versus the control. Data represent the mean ± sd. (b) D40 siRNA inhibited the growth of the MDA-MB231-luc GOF mutant of p53 cell line after transplantation into scid mice. One million MDA-MB231-luc cells were transplanted into the mammary glands of scid mice. The siRNA and atelocollagen complex was injected into the tumors on mice 4, 6, 8, 11, 13, 15, and 18 days after transplantation. On day 18, the bioluminescence from the tumors was quantitatively detected. Left: Representative bioluminescent images of mice treated with D40 siRNA (upper) or control siRNA (lower). Right: Statistical analysis of the bioluminescent signals from the tumors in mice treated with the D40 siRNA/atelocollagen complex versus the control. Data represent the mean ± se.
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f5: In vivo Growth Inhibition of p53 mutant cancer cell lines by D40 siRNA.(a) D40 siRNA inhibited the metastatic growth of the PC-3M-luc p53- cell line after transplantation into nude mice. Two million PC-3M-luc cells were transplanted into the left cardiac ventricle of nude mice. The siRNA and atelocollagen complex was injected into the tail veins of mice 4, 7, 13, and 16 days after transplantation. On day 29, the bioluminescence from the tumors was quantitatively detected. Left: Representative bioluminescent images of mice treated with the D40 siRNA/atelocollagen complex (upper) or the control (lower). Right: Statistical analysis of bioluminescent signals from the tumors in mice treated with the D40 siRNA/atelocollagen complex versus the control. Data represent the mean ± sd. (b) D40 siRNA inhibited the growth of the MDA-MB231-luc GOF mutant of p53 cell line after transplantation into scid mice. One million MDA-MB231-luc cells were transplanted into the mammary glands of scid mice. The siRNA and atelocollagen complex was injected into the tumors on mice 4, 6, 8, 11, 13, 15, and 18 days after transplantation. On day 18, the bioluminescence from the tumors was quantitatively detected. Left: Representative bioluminescent images of mice treated with D40 siRNA (upper) or control siRNA (lower). Right: Statistical analysis of the bioluminescent signals from the tumors in mice treated with the D40 siRNA/atelocollagen complex versus the control. Data represent the mean ± se.
Mentions: In subsequent experiments, we determined whether D40 siRNA could inhibit the growth of tumors transplanted in experimental animals. The p53- PC-3M-luc cells were inoculated into the left cardiac ventricles of nude mice, and the D40 siRNA/atelocollagen complex, in which atelocollagen was used as a carrier for the siRNA, was then injected intravenously to examine the effects of D40 siRNA on the metastatic growth of tumors in nude mice2728. As PC-3M-luc cells were engineered to constitutively express luciferase, the growth of tumors in nude mice was monitored using an extracorporeal device after the injection of a substrate for luciferase. Representative extracorporeal bioluminescent images of tumors in mice are shown in Fig. 5a Left. The bioluminescent signals observed in mice treated with the D40 siRNA/atelocollagen complex (upper) were significantly weaker than those in control mice (lower). Statistical analysis of the quantified bioluminescent signals from mice revealed that metastatic growth of the PC-3M-luc cell line was significantly inhibited by the D40 siRNA/atelocollagen complex compared with control (Fig. 5a Right).

Bottom Line: The D40 gene encodes a kinetochore protein that plays an essential role in kinetochore formation during mitosis.These results indicated that D40 siRNA induced apoptotic cell death in human cancer cell lines, and inhibited their growth in vitro and in vivo regardless of p53 status.Therefore, D40 siRNA is a potential candidate anti-cancer reagent.

View Article: PubMed Central - PubMed

Affiliation: Division of Cancer Gene Regulation, Institute for Genetic Medicine, Hokkaido University, Sapporo, Hokkaido, Japan.

ABSTRACT
The D40 gene encodes a kinetochore protein that plays an essential role in kinetochore formation during mitosis. Short inhibitory RNA against D40, D40 siRNA, has been shown to deplete the D40 protein in the human cancer cell line HeLa, which harbors wild-type p53, and this activity was followed by the significant inhibition of cell growth and induction of apoptotic cell death. The p53- cancer cell line, PC-3M-luc, is also sensitive to the significant growth inhibition and cell death induced by D40 siRNA. The growth of PC-3M-luc tumors transplanted into nude mice was inhibited by the systemic administration of D40 siRNA and the atelocollagen complex. Furthermore, D40 siRNA significantly inhibited growth and induced apoptotic cell death in a cell line with a gain-of-function (GOF) mutation in p53, MDA-MB231-luc, and also inhibited the growth of tumors transplanted into mice when administered as a D40 siRNA/atelocollagen complex. These results indicated that D40 siRNA induced apoptotic cell death in human cancer cell lines, and inhibited their growth in vitro and in vivo regardless of p53 status. Therefore, D40 siRNA is a potential candidate anti-cancer reagent.

No MeSH data available.


Related in: MedlinePlus