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Evaluation of In Vitro Activity of the Class I PI3K Inhibitor Buparlisib (BKM120) in Pediatric Bone and Soft Tissue Sarcomas.

Anderson JL, Park A, Akiyama R, Tap WD, Denny CT, Federman N - PLoS ONE (2015)

Bottom Line: Anti-proliferative effects were observed in both anchorage dependent and independent conditions and apoptosis was induced within 24 hours of drug treatment.Buparlisib demonstrated cytotoxicity as a single agent, but was found to be more effective when used in combination.The addition of NVP-AEW541 also further reduced phospho-Akt levels and more potently induced apoptosis compared to buparlisib treatment alone.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, Division of Hematology/Oncology, Gwynne Hazen Cherry Memorial Laboratories, University of California, Los Angeles, Los Angeles, California, United States of America.

ABSTRACT
Pediatric bone and soft tissue sarcomas often display increased Akt phosphorylation through up regulation of insulin-like growth factor (IGF1) signaling. Additionally, Akt signaling has been linked to resistance to IGF1 receptor (IGF1R) and mTOR (mammalian target of rapamycin) inhibitors in sarcoma, further demonstrating the role of Akt in tumor survival. This suggests targeting components of the PI3K/Akt pathway may be an effective therapeutic strategy. Here, we investigated the in vitro activity of the pan-class I PI3K inhibitor buparlisib (BKM120) in pediatric bone and soft tissue sarcomas. Buparlisib inhibited activation of Akt and signaling molecules downstream of mTORC1 (mTOR complex 1) in Ewing sarcoma, osteosarcoma, and rhabdomyosarcoma cell lines. Anti-proliferative effects were observed in both anchorage dependent and independent conditions and apoptosis was induced within 24 hours of drug treatment. Buparlisib demonstrated cytotoxicity as a single agent, but was found to be more effective when used in combination. Synergy was observed when buparlisib was combined with the IGF1R inhibitor NVP-AEW541 and the mTORC1 inhibitor rapamycin. The addition of NVP-AEW541 also further reduced phospho-Akt levels and more potently induced apoptosis compared to buparlisib treatment alone. Additionally, the combination of buparlisib with the MEK1/2 inhibitor trametinib resulted in synergy in sarcoma cell lines possessing MAPK pathway mutations. Taken together, these data indicate buparlisib could be a novel therapy for the treatment of pediatric bone and soft tissue sarcomas.

No MeSH data available.


Related in: MedlinePlus

Buparlisib synergizes with NVP-AEW541 to further reduce Akt phosphorylation and augment apoptosis.(A) Cells were exposed to a series of 1.5-fold dilutions of buparlisib and NVP-AEW541 alone or in combination at a constant ratio of 1:3 (TC-32, A4573, RD) or 1:1 (SJCRH30) for 72 hours, then cell viability was determined by MTT assay. Columns represent the average of three independent experiments, error bars represent standard deviation. Combination index values greater than 1, equal to 1, or less than one indicate antagonism, additivity, or synergy. (B) Isobologram plot of the effect of buparlisib combined with NVP-AEW541. The effective doses of NVP-AEW541 and buparlisib are plotted on the x- and y-axis with lines of linear additivity connecting the ED50, ED75, and E90 for individual treatments. Points for combination treatment above, on, or below the lines indicate antagonism, additivity, or synergy, respectively. (C) Immunoblot analysis of phospho-Akt (S473), total Akt, phospho-Erk1/2 (T202/204 on Erk1, T185/187 on Erk2), and total Erk1/2 in A4573 cells after 1 hour of treatment with buparlisib, NVP-AEW541, or both. (D) Immunoblot analysis of phospho-Akt (S473), total Akt, phospho-Erk1/2 (T202/204 on Erk1, T185/187 on Erk2), total Erk1/2, cleaved PARP, and actin in A4573 cells after 24 hours of treatment with buparlisib, NVP-AEW541, or both.
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pone.0133610.g006: Buparlisib synergizes with NVP-AEW541 to further reduce Akt phosphorylation and augment apoptosis.(A) Cells were exposed to a series of 1.5-fold dilutions of buparlisib and NVP-AEW541 alone or in combination at a constant ratio of 1:3 (TC-32, A4573, RD) or 1:1 (SJCRH30) for 72 hours, then cell viability was determined by MTT assay. Columns represent the average of three independent experiments, error bars represent standard deviation. Combination index values greater than 1, equal to 1, or less than one indicate antagonism, additivity, or synergy. (B) Isobologram plot of the effect of buparlisib combined with NVP-AEW541. The effective doses of NVP-AEW541 and buparlisib are plotted on the x- and y-axis with lines of linear additivity connecting the ED50, ED75, and E90 for individual treatments. Points for combination treatment above, on, or below the lines indicate antagonism, additivity, or synergy, respectively. (C) Immunoblot analysis of phospho-Akt (S473), total Akt, phospho-Erk1/2 (T202/204 on Erk1, T185/187 on Erk2), and total Erk1/2 in A4573 cells after 1 hour of treatment with buparlisib, NVP-AEW541, or both. (D) Immunoblot analysis of phospho-Akt (S473), total Akt, phospho-Erk1/2 (T202/204 on Erk1, T185/187 on Erk2), total Erk1/2, cleaved PARP, and actin in A4573 cells after 24 hours of treatment with buparlisib, NVP-AEW541, or both.

Mentions: Buparlisib was combined with chemotherapeutic and targeted agents to test for synergy. Sequential treatment of doxorubicin, a chemotherapeutic agent commonly used for the treatment of pediatric sarcomas, and buparlisib displayed additive results in ES and OS cell lines (S4 Fig). We also evaluated buparlisib in combination with the IGF1R inhibitor NVP-AEW541 based on the involvement of Akt signaling in resistance to anti-IGF1R therapy. Synergy was observed in ES and RMS cell lines (Fig 6A and 6B), while OS cell lines displayed additive effects (S5 Fig). The greatest synergistic effects were observed in the ES cell line A4573 and the RMS cell line SJCRH30 (Fig 6A and 6B), which display or low PTEN expression (Fig 1A). We also measured the effects of combining buparlisib and NVP-AEW541 on Akt phosphorylation and induction of apoptosis (Fig 6C and 6D). The drug combination resulted in reduced phospho-Akt levels after one hour of treatment compared to each drug alone (Fig 6C). Furthermore, this response persisted up to 24 hours, indicating IGF1R inhibition can prevent Akt reactivation that was observed with buparlisib treatment alone (Fig 6D). Additionally, the drug combination resulted in increased apoptosis as measured by PARP cleavage at all dose levels after 24 hours of treatment (Fig 6D).


Evaluation of In Vitro Activity of the Class I PI3K Inhibitor Buparlisib (BKM120) in Pediatric Bone and Soft Tissue Sarcomas.

Anderson JL, Park A, Akiyama R, Tap WD, Denny CT, Federman N - PLoS ONE (2015)

Buparlisib synergizes with NVP-AEW541 to further reduce Akt phosphorylation and augment apoptosis.(A) Cells were exposed to a series of 1.5-fold dilutions of buparlisib and NVP-AEW541 alone or in combination at a constant ratio of 1:3 (TC-32, A4573, RD) or 1:1 (SJCRH30) for 72 hours, then cell viability was determined by MTT assay. Columns represent the average of three independent experiments, error bars represent standard deviation. Combination index values greater than 1, equal to 1, or less than one indicate antagonism, additivity, or synergy. (B) Isobologram plot of the effect of buparlisib combined with NVP-AEW541. The effective doses of NVP-AEW541 and buparlisib are plotted on the x- and y-axis with lines of linear additivity connecting the ED50, ED75, and E90 for individual treatments. Points for combination treatment above, on, or below the lines indicate antagonism, additivity, or synergy, respectively. (C) Immunoblot analysis of phospho-Akt (S473), total Akt, phospho-Erk1/2 (T202/204 on Erk1, T185/187 on Erk2), and total Erk1/2 in A4573 cells after 1 hour of treatment with buparlisib, NVP-AEW541, or both. (D) Immunoblot analysis of phospho-Akt (S473), total Akt, phospho-Erk1/2 (T202/204 on Erk1, T185/187 on Erk2), total Erk1/2, cleaved PARP, and actin in A4573 cells after 24 hours of treatment with buparlisib, NVP-AEW541, or both.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
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pone.0133610.g006: Buparlisib synergizes with NVP-AEW541 to further reduce Akt phosphorylation and augment apoptosis.(A) Cells were exposed to a series of 1.5-fold dilutions of buparlisib and NVP-AEW541 alone or in combination at a constant ratio of 1:3 (TC-32, A4573, RD) or 1:1 (SJCRH30) for 72 hours, then cell viability was determined by MTT assay. Columns represent the average of three independent experiments, error bars represent standard deviation. Combination index values greater than 1, equal to 1, or less than one indicate antagonism, additivity, or synergy. (B) Isobologram plot of the effect of buparlisib combined with NVP-AEW541. The effective doses of NVP-AEW541 and buparlisib are plotted on the x- and y-axis with lines of linear additivity connecting the ED50, ED75, and E90 for individual treatments. Points for combination treatment above, on, or below the lines indicate antagonism, additivity, or synergy, respectively. (C) Immunoblot analysis of phospho-Akt (S473), total Akt, phospho-Erk1/2 (T202/204 on Erk1, T185/187 on Erk2), and total Erk1/2 in A4573 cells after 1 hour of treatment with buparlisib, NVP-AEW541, or both. (D) Immunoblot analysis of phospho-Akt (S473), total Akt, phospho-Erk1/2 (T202/204 on Erk1, T185/187 on Erk2), total Erk1/2, cleaved PARP, and actin in A4573 cells after 24 hours of treatment with buparlisib, NVP-AEW541, or both.
Mentions: Buparlisib was combined with chemotherapeutic and targeted agents to test for synergy. Sequential treatment of doxorubicin, a chemotherapeutic agent commonly used for the treatment of pediatric sarcomas, and buparlisib displayed additive results in ES and OS cell lines (S4 Fig). We also evaluated buparlisib in combination with the IGF1R inhibitor NVP-AEW541 based on the involvement of Akt signaling in resistance to anti-IGF1R therapy. Synergy was observed in ES and RMS cell lines (Fig 6A and 6B), while OS cell lines displayed additive effects (S5 Fig). The greatest synergistic effects were observed in the ES cell line A4573 and the RMS cell line SJCRH30 (Fig 6A and 6B), which display or low PTEN expression (Fig 1A). We also measured the effects of combining buparlisib and NVP-AEW541 on Akt phosphorylation and induction of apoptosis (Fig 6C and 6D). The drug combination resulted in reduced phospho-Akt levels after one hour of treatment compared to each drug alone (Fig 6C). Furthermore, this response persisted up to 24 hours, indicating IGF1R inhibition can prevent Akt reactivation that was observed with buparlisib treatment alone (Fig 6D). Additionally, the drug combination resulted in increased apoptosis as measured by PARP cleavage at all dose levels after 24 hours of treatment (Fig 6D).

Bottom Line: Anti-proliferative effects were observed in both anchorage dependent and independent conditions and apoptosis was induced within 24 hours of drug treatment.Buparlisib demonstrated cytotoxicity as a single agent, but was found to be more effective when used in combination.The addition of NVP-AEW541 also further reduced phospho-Akt levels and more potently induced apoptosis compared to buparlisib treatment alone.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, Division of Hematology/Oncology, Gwynne Hazen Cherry Memorial Laboratories, University of California, Los Angeles, Los Angeles, California, United States of America.

ABSTRACT
Pediatric bone and soft tissue sarcomas often display increased Akt phosphorylation through up regulation of insulin-like growth factor (IGF1) signaling. Additionally, Akt signaling has been linked to resistance to IGF1 receptor (IGF1R) and mTOR (mammalian target of rapamycin) inhibitors in sarcoma, further demonstrating the role of Akt in tumor survival. This suggests targeting components of the PI3K/Akt pathway may be an effective therapeutic strategy. Here, we investigated the in vitro activity of the pan-class I PI3K inhibitor buparlisib (BKM120) in pediatric bone and soft tissue sarcomas. Buparlisib inhibited activation of Akt and signaling molecules downstream of mTORC1 (mTOR complex 1) in Ewing sarcoma, osteosarcoma, and rhabdomyosarcoma cell lines. Anti-proliferative effects were observed in both anchorage dependent and independent conditions and apoptosis was induced within 24 hours of drug treatment. Buparlisib demonstrated cytotoxicity as a single agent, but was found to be more effective when used in combination. Synergy was observed when buparlisib was combined with the IGF1R inhibitor NVP-AEW541 and the mTORC1 inhibitor rapamycin. The addition of NVP-AEW541 also further reduced phospho-Akt levels and more potently induced apoptosis compared to buparlisib treatment alone. Additionally, the combination of buparlisib with the MEK1/2 inhibitor trametinib resulted in synergy in sarcoma cell lines possessing MAPK pathway mutations. Taken together, these data indicate buparlisib could be a novel therapy for the treatment of pediatric bone and soft tissue sarcomas.

No MeSH data available.


Related in: MedlinePlus