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Identification and Preclinical Pharmacology of the γ-Secretase Modulator BMS-869780.

Toyn JH, Thompson LA, Lentz KA, Meredith JE, Burton CR, Sankaranararyanan S, Guss V, Hall T, Iben LG, Krause CM, Krause R, Lin XA, Pierdomenico M, Polson C, Robertson AS, Denton RR, Grace JE, Morrison J, Raybon J, Zhuo X, Snow K, Padmanabha R, Agler M, Esposito K, Harden D, Prack M, Varma S, Wong V, Zhu Y, Zvyaga T, Gerritz S, Marcin LR, Higgins MA, Shi J, Wei C, Cantone JL, Drexler DM, Macor JE, Olson RE, Ahlijanian MK, Albright CF - Int J Alzheimers Dis (2014)

Bottom Line: Off-target and safety margins were then based on comparisons to the predicted exposure required for robust Aβ1-42 lowering.Because of insufficient safety predictions and the relatively high predicted human daily dose of 700 mg, further evaluation of BMS-869780 as a potential clinical candidate was discontinued.Nevertheless, BMS-869780 demonstrates the potential of the GSM approach for robust lowering of brain Aβ1-42 without Notch-related side effects.

View Article: PubMed Central - PubMed

Affiliation: Exploratory Biology and Genomics, Bristol-Myers Squibb Research and Development, 5 Research Parkway, Wallingford, CT 06492, USA.

ABSTRACT
Alzheimer's disease is the most prevalent cause of dementia and is associated with accumulation of amyloid-β peptide (Aβ), particularly the 42-amino acid Aβ1-42, in the brain. Aβ1-42 levels can be decreased by γ-secretase modulators (GSM), which are small molecules that modulate γ-secretase, an enzyme essential for Aβ production. BMS-869780 is a potent GSM that decreased Aβ1-42 and Aβ1-40 and increased Aβ1-37 and Aβ1-38, without inhibiting overall levels of Aβ peptides or other APP processing intermediates. BMS-869780 also did not inhibit Notch processing by γ-secretase and lowered brain Aβ1-42 without evidence of Notch-related side effects in rats. Human pharmacokinetic (PK) parameters were predicted through allometric scaling of PK in rat, dog, and monkey and were combined with the rat pharmacodynamic (PD) parameters to predict the relationship between BMS-869780 dose, exposure and Aβ1-42 levels in human. Off-target and safety margins were then based on comparisons to the predicted exposure required for robust Aβ1-42 lowering. Because of insufficient safety predictions and the relatively high predicted human daily dose of 700 mg, further evaluation of BMS-869780 as a potential clinical candidate was discontinued. Nevertheless, BMS-869780 demonstrates the potential of the GSM approach for robust lowering of brain Aβ1-42 without Notch-related side effects.

No MeSH data available.


Related in: MedlinePlus

BMS-869780 dose response and evaluation of residual Aβ levels in rat brain. (a) Groups of rats received intraperitoneal (IP) injections of vehicle or BMS-869780 at doses of 100, 30, 10, 3, 1, and 0.3 mg/kg. Additional rats were dosed with GSI BMS-698861 at 30 mg/kg as a positive control for Aβ-lowering. Group sizes were seven rats for each dose of BMS-869780 and 14 rats for vehicle and GSI dose groups. Brain and plasma were harvested 5 hours after dosing. Brain Aβ1-42 (green), brain Aβ1-40 (blue), brain Aβ1-x (black), and plasma Aβ1-40 (red) were determined. Values are expressed as % relative to vehicle group mean. Whiskers represent standard error. ⊗Aβ1-x was not determined in the groups dosed at 3, 1, and 0.3 mg/kg. (b) Brain Aβ1-40 was plotted against brain Aβ1-42 for each rat dosed with BMS-869780 (black ▼) and for each rat dosed with vehicle (grey ●). Values are expressed as % relative to vehicle group mean. Whiskers represent standard error. (c) Brain Aβ1-42 (●), brain Aβ1-40 (■), and plasma Aβ1-40 (▲) were plotted against plasma concentration of BMS-869780 and the data were evaluated by fit to a four-parameter dose response curve. The top of the dose response curve was defined by vehicle group mean (100%), and the apparent IC50 values in terms of the plasma BMS-869780 concentration obtained for brain Aβ1-42, brain Aβ1-40, and plasma Aβ1-40 were 807 nM, 943 nM, and 84 nM, respectively. The respective 95% confidence intervals were 618–1053 nM, 704–1264 nM, and 44–158 nM. (d) Rats were dosed once daily with BMS-869780 for 4 days at 10 and 100 mg/kg or vehicle, plasma, and brain samples were taken 5 hours after the last dose, and immunodepletion of brain extracts was carried out prior to Aβ1-42 ELISA assays. Specific monoclonals used were 565 (Aβ1-42 selective), TSD (Aβ1-40 selective), 4G8 (binds both Aβ1-42 and Aβ1-40), and 6E10 (does not bind rat Aβ). After immunodepletion, Aβ1-42 was assayed by ELISA. (e) Same as described in (d), except that Aβ1-40 ELISA was carried out following the immunodepletion.
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fig7: BMS-869780 dose response and evaluation of residual Aβ levels in rat brain. (a) Groups of rats received intraperitoneal (IP) injections of vehicle or BMS-869780 at doses of 100, 30, 10, 3, 1, and 0.3 mg/kg. Additional rats were dosed with GSI BMS-698861 at 30 mg/kg as a positive control for Aβ-lowering. Group sizes were seven rats for each dose of BMS-869780 and 14 rats for vehicle and GSI dose groups. Brain and plasma were harvested 5 hours after dosing. Brain Aβ1-42 (green), brain Aβ1-40 (blue), brain Aβ1-x (black), and plasma Aβ1-40 (red) were determined. Values are expressed as % relative to vehicle group mean. Whiskers represent standard error. ⊗Aβ1-x was not determined in the groups dosed at 3, 1, and 0.3 mg/kg. (b) Brain Aβ1-40 was plotted against brain Aβ1-42 for each rat dosed with BMS-869780 (black ▼) and for each rat dosed with vehicle (grey ●). Values are expressed as % relative to vehicle group mean. Whiskers represent standard error. (c) Brain Aβ1-42 (●), brain Aβ1-40 (■), and plasma Aβ1-40 (▲) were plotted against plasma concentration of BMS-869780 and the data were evaluated by fit to a four-parameter dose response curve. The top of the dose response curve was defined by vehicle group mean (100%), and the apparent IC50 values in terms of the plasma BMS-869780 concentration obtained for brain Aβ1-42, brain Aβ1-40, and plasma Aβ1-40 were 807 nM, 943 nM, and 84 nM, respectively. The respective 95% confidence intervals were 618–1053 nM, 704–1264 nM, and 44–158 nM. (d) Rats were dosed once daily with BMS-869780 for 4 days at 10 and 100 mg/kg or vehicle, plasma, and brain samples were taken 5 hours after the last dose, and immunodepletion of brain extracts was carried out prior to Aβ1-42 ELISA assays. Specific monoclonals used were 565 (Aβ1-42 selective), TSD (Aβ1-40 selective), 4G8 (binds both Aβ1-42 and Aβ1-40), and 6E10 (does not bind rat Aβ). After immunodepletion, Aβ1-42 was assayed by ELISA. (e) Same as described in (d), except that Aβ1-40 ELISA was carried out following the immunodepletion.

Mentions: IC50 determinations for Aβ1-42 and Aβ1-40 in H4-APPsw cultures in 384-well format were determined using homogeneous time-resolved fluorescence immunoassays as previously described [59]. The principle of these assays is illustrated in Figures 1(c) and 1(d). In other experiments using H4-APPsw cultures, Aβ was quantified by ELISA; for Aβ1-42 the combination of monoclonal antibodies was 26D6 and 565-HRP; for Aβ1-40 it was TSD and 26D6-HRP, and for Aβ1-x it was 4G8 and 26D6-HRP. In some experiments a novel 4-plex Aβ electrochemiluminescence immunoassay was used (Mesoscale Discovery catalog number N45ZA-1). Briefly, the 4-plex was carried out in 96-well format, with 4 separate spots of capture antibodies in each well. The 96-well plates were prepared by the manufacturer, with spots of monoclonal antibodies for Aβ1-42, Aβ1-40, and Aβ1-38, and an additional fourth spot of streptavidin in each well. Plates were initially incubated with blocking buffer (5% BSA in phosphate buffered saline, 200 μL per well) for 2 hours at ambient temperature and then with D2A6H-biotin conjugate (50 ng/mL in 1% BSA, phosphate buffered saline, 25 μL per well) for 1 hour. Plates were then rinsed with phosphate buffered saline before addition of experimental samples for determination of Aβ1-42, Aβ1-40, Aβ1-38, and Aβ1-37 levels, following the manufacturer's instructions as for the Aβ 3-plex kit (catalog number K15148E-1). Rat brain extracts for use in the 4-plex assay were made in 0.2% diethylamine, as previously described [62]. For detection of rat Aβ peptides in the 4-plex (Figure 5), 252Q6-sulfo-tag conjugate was used, and for detection of human Aβ peptides from cell cultures 6E10-sulfo-tag conjugate was used. For Aβ in transiently transfected PS1/PS2 dKO fibroblasts, Aβ1-42 was quantified using an ELISA kit (WACO), and Aβ1-40 was quantified by ELISA as described above for H4-APPsw cultures. For triple transgenic mice (3xTg; [63]), human transgenic Aβ1-42 was assayed in brain homogenates using an ELISA kit (WACO). For the rat and mouse experiments illustrated in Figures 7 and 8, brain samples were prepared by solid phase extraction [64], and endogenous rat brain Aβ1-42 and Aβ1-40 were quantified by ELISA as previously described for wild type mice [65]. For brain extracts made using solid phase extraction, calibration of Aβ was relative, based on the approximately linear response of the assay in the range tested. For brain extracts made in 0.2% diethylamine, Aβ1-42, Aβ1-40, Aβ1-38, and Aβ1-37 concentrations in brain and cell culture samples were determined by fitting the results of immunoassays against calibration curves derived from a range of dilutions of the corresponding synthetic peptides on each assay plate using a quadratic curve fit (Graphpad Prizm 5.0). Aβ1-x was calibrated in the same way against synthetic Aβ1-40 peptide. Results were expressed in units of pM, corrected for sample dilution.


Identification and Preclinical Pharmacology of the γ-Secretase Modulator BMS-869780.

Toyn JH, Thompson LA, Lentz KA, Meredith JE, Burton CR, Sankaranararyanan S, Guss V, Hall T, Iben LG, Krause CM, Krause R, Lin XA, Pierdomenico M, Polson C, Robertson AS, Denton RR, Grace JE, Morrison J, Raybon J, Zhuo X, Snow K, Padmanabha R, Agler M, Esposito K, Harden D, Prack M, Varma S, Wong V, Zhu Y, Zvyaga T, Gerritz S, Marcin LR, Higgins MA, Shi J, Wei C, Cantone JL, Drexler DM, Macor JE, Olson RE, Ahlijanian MK, Albright CF - Int J Alzheimers Dis (2014)

BMS-869780 dose response and evaluation of residual Aβ levels in rat brain. (a) Groups of rats received intraperitoneal (IP) injections of vehicle or BMS-869780 at doses of 100, 30, 10, 3, 1, and 0.3 mg/kg. Additional rats were dosed with GSI BMS-698861 at 30 mg/kg as a positive control for Aβ-lowering. Group sizes were seven rats for each dose of BMS-869780 and 14 rats for vehicle and GSI dose groups. Brain and plasma were harvested 5 hours after dosing. Brain Aβ1-42 (green), brain Aβ1-40 (blue), brain Aβ1-x (black), and plasma Aβ1-40 (red) were determined. Values are expressed as % relative to vehicle group mean. Whiskers represent standard error. ⊗Aβ1-x was not determined in the groups dosed at 3, 1, and 0.3 mg/kg. (b) Brain Aβ1-40 was plotted against brain Aβ1-42 for each rat dosed with BMS-869780 (black ▼) and for each rat dosed with vehicle (grey ●). Values are expressed as % relative to vehicle group mean. Whiskers represent standard error. (c) Brain Aβ1-42 (●), brain Aβ1-40 (■), and plasma Aβ1-40 (▲) were plotted against plasma concentration of BMS-869780 and the data were evaluated by fit to a four-parameter dose response curve. The top of the dose response curve was defined by vehicle group mean (100%), and the apparent IC50 values in terms of the plasma BMS-869780 concentration obtained for brain Aβ1-42, brain Aβ1-40, and plasma Aβ1-40 were 807 nM, 943 nM, and 84 nM, respectively. The respective 95% confidence intervals were 618–1053 nM, 704–1264 nM, and 44–158 nM. (d) Rats were dosed once daily with BMS-869780 for 4 days at 10 and 100 mg/kg or vehicle, plasma, and brain samples were taken 5 hours after the last dose, and immunodepletion of brain extracts was carried out prior to Aβ1-42 ELISA assays. Specific monoclonals used were 565 (Aβ1-42 selective), TSD (Aβ1-40 selective), 4G8 (binds both Aβ1-42 and Aβ1-40), and 6E10 (does not bind rat Aβ). After immunodepletion, Aβ1-42 was assayed by ELISA. (e) Same as described in (d), except that Aβ1-40 ELISA was carried out following the immunodepletion.
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Related In: Results  -  Collection

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fig7: BMS-869780 dose response and evaluation of residual Aβ levels in rat brain. (a) Groups of rats received intraperitoneal (IP) injections of vehicle or BMS-869780 at doses of 100, 30, 10, 3, 1, and 0.3 mg/kg. Additional rats were dosed with GSI BMS-698861 at 30 mg/kg as a positive control for Aβ-lowering. Group sizes were seven rats for each dose of BMS-869780 and 14 rats for vehicle and GSI dose groups. Brain and plasma were harvested 5 hours after dosing. Brain Aβ1-42 (green), brain Aβ1-40 (blue), brain Aβ1-x (black), and plasma Aβ1-40 (red) were determined. Values are expressed as % relative to vehicle group mean. Whiskers represent standard error. ⊗Aβ1-x was not determined in the groups dosed at 3, 1, and 0.3 mg/kg. (b) Brain Aβ1-40 was plotted against brain Aβ1-42 for each rat dosed with BMS-869780 (black ▼) and for each rat dosed with vehicle (grey ●). Values are expressed as % relative to vehicle group mean. Whiskers represent standard error. (c) Brain Aβ1-42 (●), brain Aβ1-40 (■), and plasma Aβ1-40 (▲) were plotted against plasma concentration of BMS-869780 and the data were evaluated by fit to a four-parameter dose response curve. The top of the dose response curve was defined by vehicle group mean (100%), and the apparent IC50 values in terms of the plasma BMS-869780 concentration obtained for brain Aβ1-42, brain Aβ1-40, and plasma Aβ1-40 were 807 nM, 943 nM, and 84 nM, respectively. The respective 95% confidence intervals were 618–1053 nM, 704–1264 nM, and 44–158 nM. (d) Rats were dosed once daily with BMS-869780 for 4 days at 10 and 100 mg/kg or vehicle, plasma, and brain samples were taken 5 hours after the last dose, and immunodepletion of brain extracts was carried out prior to Aβ1-42 ELISA assays. Specific monoclonals used were 565 (Aβ1-42 selective), TSD (Aβ1-40 selective), 4G8 (binds both Aβ1-42 and Aβ1-40), and 6E10 (does not bind rat Aβ). After immunodepletion, Aβ1-42 was assayed by ELISA. (e) Same as described in (d), except that Aβ1-40 ELISA was carried out following the immunodepletion.
Mentions: IC50 determinations for Aβ1-42 and Aβ1-40 in H4-APPsw cultures in 384-well format were determined using homogeneous time-resolved fluorescence immunoassays as previously described [59]. The principle of these assays is illustrated in Figures 1(c) and 1(d). In other experiments using H4-APPsw cultures, Aβ was quantified by ELISA; for Aβ1-42 the combination of monoclonal antibodies was 26D6 and 565-HRP; for Aβ1-40 it was TSD and 26D6-HRP, and for Aβ1-x it was 4G8 and 26D6-HRP. In some experiments a novel 4-plex Aβ electrochemiluminescence immunoassay was used (Mesoscale Discovery catalog number N45ZA-1). Briefly, the 4-plex was carried out in 96-well format, with 4 separate spots of capture antibodies in each well. The 96-well plates were prepared by the manufacturer, with spots of monoclonal antibodies for Aβ1-42, Aβ1-40, and Aβ1-38, and an additional fourth spot of streptavidin in each well. Plates were initially incubated with blocking buffer (5% BSA in phosphate buffered saline, 200 μL per well) for 2 hours at ambient temperature and then with D2A6H-biotin conjugate (50 ng/mL in 1% BSA, phosphate buffered saline, 25 μL per well) for 1 hour. Plates were then rinsed with phosphate buffered saline before addition of experimental samples for determination of Aβ1-42, Aβ1-40, Aβ1-38, and Aβ1-37 levels, following the manufacturer's instructions as for the Aβ 3-plex kit (catalog number K15148E-1). Rat brain extracts for use in the 4-plex assay were made in 0.2% diethylamine, as previously described [62]. For detection of rat Aβ peptides in the 4-plex (Figure 5), 252Q6-sulfo-tag conjugate was used, and for detection of human Aβ peptides from cell cultures 6E10-sulfo-tag conjugate was used. For Aβ in transiently transfected PS1/PS2 dKO fibroblasts, Aβ1-42 was quantified using an ELISA kit (WACO), and Aβ1-40 was quantified by ELISA as described above for H4-APPsw cultures. For triple transgenic mice (3xTg; [63]), human transgenic Aβ1-42 was assayed in brain homogenates using an ELISA kit (WACO). For the rat and mouse experiments illustrated in Figures 7 and 8, brain samples were prepared by solid phase extraction [64], and endogenous rat brain Aβ1-42 and Aβ1-40 were quantified by ELISA as previously described for wild type mice [65]. For brain extracts made using solid phase extraction, calibration of Aβ was relative, based on the approximately linear response of the assay in the range tested. For brain extracts made in 0.2% diethylamine, Aβ1-42, Aβ1-40, Aβ1-38, and Aβ1-37 concentrations in brain and cell culture samples were determined by fitting the results of immunoassays against calibration curves derived from a range of dilutions of the corresponding synthetic peptides on each assay plate using a quadratic curve fit (Graphpad Prizm 5.0). Aβ1-x was calibrated in the same way against synthetic Aβ1-40 peptide. Results were expressed in units of pM, corrected for sample dilution.

Bottom Line: Off-target and safety margins were then based on comparisons to the predicted exposure required for robust Aβ1-42 lowering.Because of insufficient safety predictions and the relatively high predicted human daily dose of 700 mg, further evaluation of BMS-869780 as a potential clinical candidate was discontinued.Nevertheless, BMS-869780 demonstrates the potential of the GSM approach for robust lowering of brain Aβ1-42 without Notch-related side effects.

View Article: PubMed Central - PubMed

Affiliation: Exploratory Biology and Genomics, Bristol-Myers Squibb Research and Development, 5 Research Parkway, Wallingford, CT 06492, USA.

ABSTRACT
Alzheimer's disease is the most prevalent cause of dementia and is associated with accumulation of amyloid-β peptide (Aβ), particularly the 42-amino acid Aβ1-42, in the brain. Aβ1-42 levels can be decreased by γ-secretase modulators (GSM), which are small molecules that modulate γ-secretase, an enzyme essential for Aβ production. BMS-869780 is a potent GSM that decreased Aβ1-42 and Aβ1-40 and increased Aβ1-37 and Aβ1-38, without inhibiting overall levels of Aβ peptides or other APP processing intermediates. BMS-869780 also did not inhibit Notch processing by γ-secretase and lowered brain Aβ1-42 without evidence of Notch-related side effects in rats. Human pharmacokinetic (PK) parameters were predicted through allometric scaling of PK in rat, dog, and monkey and were combined with the rat pharmacodynamic (PD) parameters to predict the relationship between BMS-869780 dose, exposure and Aβ1-42 levels in human. Off-target and safety margins were then based on comparisons to the predicted exposure required for robust Aβ1-42 lowering. Because of insufficient safety predictions and the relatively high predicted human daily dose of 700 mg, further evaluation of BMS-869780 as a potential clinical candidate was discontinued. Nevertheless, BMS-869780 demonstrates the potential of the GSM approach for robust lowering of brain Aβ1-42 without Notch-related side effects.

No MeSH data available.


Related in: MedlinePlus