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Heat shock protein 90 inhibitors repress latent membrane protein 1 (LMP1) expression and proliferation of Epstein-Barr virus-positive natural killer cell lymphoma.

Murata T, Iwata S, Siddiquey MN, Kanazawa T, Goshima F, Kawashima D, Kimura H, Tsurumi T - PLoS ONE (2013)

Bottom Line: We here screened small molecule inhibitors and isolated HSP90 inhibitors, Radicicol and 17-AAG, as candidates that suppress LMP1 expression and cell proliferation not only in EBV-positive SNK6 Natural Killer (NK) cell lymphoma cells, but also in B and T cells.Tumor formation in immuno-defficient NOD/Shi-scid/IL-2Rγ() (NOG) mice was also retarded.These results suggest that HSP90 inhibitors can be alternative treatments for patients with EBV-positive malignancies.

View Article: PubMed Central - PubMed

Affiliation: Division of Virology, Aichi Cancer Center Research Institute, Nagoya, Aichi, Japan.

ABSTRACT
Epstein-Barr virus (EBV) LMP1 is a major oncoprotein expressed in latent infection. It functions as a TNFR family member and constitutively activates cellular signals, such as NFκB, MAPK, JAK/STAT and AKT. We here screened small molecule inhibitors and isolated HSP90 inhibitors, Radicicol and 17-AAG, as candidates that suppress LMP1 expression and cell proliferation not only in EBV-positive SNK6 Natural Killer (NK) cell lymphoma cells, but also in B and T cells. Tumor formation in immuno-defficient NOD/Shi-scid/IL-2Rγ() (NOG) mice was also retarded. These results suggest that HSP90 inhibitors can be alternative treatments for patients with EBV-positive malignancies.

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Effects of HSP90 inhibitors in vivo.HSP90 inhibitors repress EBV copy numbers in the whole blood (A) and tumor size (B) in mice implanted with EBV-positive NK cell lymphoma. Twenty-four NOG mice were subcutaneously implanted with 5×106 of SNK6 cells per mouse on day 0. From day 14, DMSO (vehicle) or 17-AAG (6 times in two weeks, 50 mg/kg in total) was injected to twelve mice each, intra-peritoneally. Peripheral blood was collected from the tail veins once in two weeks, and levels of EBV genomic DNA in whole blood were measured by Real-Time PCR (A). Subcutaneous tumor masses (B) and LMP1 mRNA levels in the tumors (C) were also measured. *p<0.05 (Mann-Whitney U test).
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pone-0063566-g004: Effects of HSP90 inhibitors in vivo.HSP90 inhibitors repress EBV copy numbers in the whole blood (A) and tumor size (B) in mice implanted with EBV-positive NK cell lymphoma. Twenty-four NOG mice were subcutaneously implanted with 5×106 of SNK6 cells per mouse on day 0. From day 14, DMSO (vehicle) or 17-AAG (6 times in two weeks, 50 mg/kg in total) was injected to twelve mice each, intra-peritoneally. Peripheral blood was collected from the tail veins once in two weeks, and levels of EBV genomic DNA in whole blood were measured by Real-Time PCR (A). Subcutaneous tumor masses (B) and LMP1 mRNA levels in the tumors (C) were also measured. *p<0.05 (Mann-Whitney U test).

Mentions: It has been already reported that HSP90 inhibitors block proliferation of EBV-positive malignancies, including NK/T lymphomas [27], [28], although the earlier studies did not test LMP1 levels. For examination of effects in vivo, we here adopted a novel mouse xenograft model using severely immuno-deficient mice of the NOG strain [29], allowing assessment of whether the HSP90 inhibitors might repress EBV-positive malignant cells without seriously affecting other normal part of tissues and organs. The NOG mice were injected subcutaneously with 5×106 of SNK6 cells at day 0, and low doses of 17-AAG (50 mg/kg in total) or DMSO were administered into the abdominal cavity for 6 times between days 14 and 25. EBV genomic DNA titers in whole blood (Fig. 4A) and tumor sizes (Fig. 4B) were measured. As shown in Fig. 4, 17-AAG markedly reduced the viral titer in blood and growth of the NK lymphoma cells in NOG mice. Because engraftment of NK lymphomas in mice is not very efficient [29], only 50% of the mice developed subcutaneous tumor masses and others failed to nurture the lymphoma cells even in the DMSO treatment group. Due to this low efficiency of transplantation, statistical significance could be exhibited only once for EBV DNA load (Fig. 4A, 14 weeks), but we gained the clear impression that 17-AAG worked more efficiently. This is an important difference when compared to LCLs, with which 100% of injected mice develop tumors, and thereby the efficacy of 17-AAG was exhibited more significantly [28]. We then compared the LMP1 expression levels in the tumor developed in the absence or presence of 17-AAG (Fig. 4C). LMP1 transcript was lower in the tumor treated with 17-AAG, indicating that the inhibitor reduced LMP1 transctiption in vivo, too. To summarize, our results imply high potential of HSP90 inhibitors for treating EBV-positive cancers, including EBV-positive NK lymphomas.


Heat shock protein 90 inhibitors repress latent membrane protein 1 (LMP1) expression and proliferation of Epstein-Barr virus-positive natural killer cell lymphoma.

Murata T, Iwata S, Siddiquey MN, Kanazawa T, Goshima F, Kawashima D, Kimura H, Tsurumi T - PLoS ONE (2013)

Effects of HSP90 inhibitors in vivo.HSP90 inhibitors repress EBV copy numbers in the whole blood (A) and tumor size (B) in mice implanted with EBV-positive NK cell lymphoma. Twenty-four NOG mice were subcutaneously implanted with 5×106 of SNK6 cells per mouse on day 0. From day 14, DMSO (vehicle) or 17-AAG (6 times in two weeks, 50 mg/kg in total) was injected to twelve mice each, intra-peritoneally. Peripheral blood was collected from the tail veins once in two weeks, and levels of EBV genomic DNA in whole blood were measured by Real-Time PCR (A). Subcutaneous tumor masses (B) and LMP1 mRNA levels in the tumors (C) were also measured. *p<0.05 (Mann-Whitney U test).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3643901&req=5

pone-0063566-g004: Effects of HSP90 inhibitors in vivo.HSP90 inhibitors repress EBV copy numbers in the whole blood (A) and tumor size (B) in mice implanted with EBV-positive NK cell lymphoma. Twenty-four NOG mice were subcutaneously implanted with 5×106 of SNK6 cells per mouse on day 0. From day 14, DMSO (vehicle) or 17-AAG (6 times in two weeks, 50 mg/kg in total) was injected to twelve mice each, intra-peritoneally. Peripheral blood was collected from the tail veins once in two weeks, and levels of EBV genomic DNA in whole blood were measured by Real-Time PCR (A). Subcutaneous tumor masses (B) and LMP1 mRNA levels in the tumors (C) were also measured. *p<0.05 (Mann-Whitney U test).
Mentions: It has been already reported that HSP90 inhibitors block proliferation of EBV-positive malignancies, including NK/T lymphomas [27], [28], although the earlier studies did not test LMP1 levels. For examination of effects in vivo, we here adopted a novel mouse xenograft model using severely immuno-deficient mice of the NOG strain [29], allowing assessment of whether the HSP90 inhibitors might repress EBV-positive malignant cells without seriously affecting other normal part of tissues and organs. The NOG mice were injected subcutaneously with 5×106 of SNK6 cells at day 0, and low doses of 17-AAG (50 mg/kg in total) or DMSO were administered into the abdominal cavity for 6 times between days 14 and 25. EBV genomic DNA titers in whole blood (Fig. 4A) and tumor sizes (Fig. 4B) were measured. As shown in Fig. 4, 17-AAG markedly reduced the viral titer in blood and growth of the NK lymphoma cells in NOG mice. Because engraftment of NK lymphomas in mice is not very efficient [29], only 50% of the mice developed subcutaneous tumor masses and others failed to nurture the lymphoma cells even in the DMSO treatment group. Due to this low efficiency of transplantation, statistical significance could be exhibited only once for EBV DNA load (Fig. 4A, 14 weeks), but we gained the clear impression that 17-AAG worked more efficiently. This is an important difference when compared to LCLs, with which 100% of injected mice develop tumors, and thereby the efficacy of 17-AAG was exhibited more significantly [28]. We then compared the LMP1 expression levels in the tumor developed in the absence or presence of 17-AAG (Fig. 4C). LMP1 transcript was lower in the tumor treated with 17-AAG, indicating that the inhibitor reduced LMP1 transctiption in vivo, too. To summarize, our results imply high potential of HSP90 inhibitors for treating EBV-positive cancers, including EBV-positive NK lymphomas.

Bottom Line: We here screened small molecule inhibitors and isolated HSP90 inhibitors, Radicicol and 17-AAG, as candidates that suppress LMP1 expression and cell proliferation not only in EBV-positive SNK6 Natural Killer (NK) cell lymphoma cells, but also in B and T cells.Tumor formation in immuno-defficient NOD/Shi-scid/IL-2Rγ() (NOG) mice was also retarded.These results suggest that HSP90 inhibitors can be alternative treatments for patients with EBV-positive malignancies.

View Article: PubMed Central - PubMed

Affiliation: Division of Virology, Aichi Cancer Center Research Institute, Nagoya, Aichi, Japan.

ABSTRACT
Epstein-Barr virus (EBV) LMP1 is a major oncoprotein expressed in latent infection. It functions as a TNFR family member and constitutively activates cellular signals, such as NFκB, MAPK, JAK/STAT and AKT. We here screened small molecule inhibitors and isolated HSP90 inhibitors, Radicicol and 17-AAG, as candidates that suppress LMP1 expression and cell proliferation not only in EBV-positive SNK6 Natural Killer (NK) cell lymphoma cells, but also in B and T cells. Tumor formation in immuno-defficient NOD/Shi-scid/IL-2Rγ() (NOG) mice was also retarded. These results suggest that HSP90 inhibitors can be alternative treatments for patients with EBV-positive malignancies.

Show MeSH
Related in: MedlinePlus