Nirogacestat (PF-03084014)

Name: Nirogacestat (PF-03084014)
Brand: Selleck Chemicals
SKU: S8018
Rating: 5 (7 reviews)

For research use only. Not for use in humans.

Catalog No.S8018 Synonyms: PF-3084014

7 publications

Nirogacestat (PF-03084014) Chemical Structure

Molecular Weight(MW): 489.64

Nirogacestat (PF-03084014, PF-3084014) is a selective gamma-secretase inhibitor with IC50 of 6.2 nM in a cell-free assay. Phase 2.

Selleck's Nirogacestat (PF-03084014) has been cited by 7 publications

Mol Ther Oncolytics, 2019, 13:58-66

PubMed: 31016228 ( click the link to review the publication )

Front Immunol, 2019, 10:162

PubMed: 30792717 ( click the link to review the publication )

J Cell Physiol, 2019, 234(5):5940-5952

PubMed: 30515785 ( click the link to review the publication )

J Biol Chem, 2019, 294(21):8543-8554

PubMed: 30940724 ( click the link to review the publication )

Exp Cell Res, 2019, 10.1016/j.yexcr.2019.06.015

PubMed: 31211955 ( click the link to review the publication )

Arch Oral Biol, 2019, 100:93-99

PubMed: 30822705 ( click the link to review the publication )

Cell Prolif, 2018, 51(3):e12424

PubMed: 29232766 ( click the link to review the publication )

2 Customer Reviews

The expression of p-EGFR and NICD was assessed by western blot after treatment for 3 d.

Cell Prolif, 2018, 51(3):e12424. Nirogacestat (PF-03084014) purchased from Selleck.

(F) Tumour tissues were further analysed to detect the efficacy of the inhibitors by IHC staining using p-EGFR and Notch1. (4Fa) Notch1 staining was mostly confined on the cell membrane. (4Fc) After Erlotinib treatment, the Notch1 protein was cleaved and translocated to the nucleus. (4Fb) p-EGFR was overexpressed in the control group. (4Fd) Erlotinib treatment inhibited p-EGFR expression. (4Fe-f) PF-03084014 treatment reduced both Notch1 and p-EGFR expression. (4Fg-h) Synthetic therapy reduced the activation of Notch1 and EGFR pathway. Scale bar = 50 μm. *P < .05, **P < .01, ***P < .001

Cell Prolif, 2017, doi: 10.1111/cpr.12424. Nirogacestat (PF-03084014) purchased from Selleck.

Purity & Quality Control

Choose Selective Gamma-secretase Inhibitors

Biological Activity

Description

Nirogacestat (PF-03084014, PF-3084014) is a selective gamma-secretase inhibitor with IC50 of 6.2 nM in a cell-free assay. Phase 2.

Targets

γ-secretase ^[1]
(cell-free assay)

6.2 nM

In vitro

PF-03084014 inhibits Notch receptor cleavage in cellular assays using HPB-ALL cells that harbor mutations in both the heterodimerization and PEST domains in Notch1with IC50 of 13.3 nM. PF-03084014 downregulates Notch target genes Hes-1, and cMyc expression in HPB-ALL cells with IC50 of <1 nM and 10 nM, respectively. PF-03084014 inhibits cell growth of a subset of human T-ALL cell lines (HPB-ALL, DND-41, TALL-1,and Sup-T1) through induction of cell cycle arrest and apoptosis with IC50s of 30-100 nM. ^[1] PF-03084014 reduces proliferation of HUVECs with IC50 of 0.5 μM, and decreases the lumen formation with an IC50 value of 50 nM. PF-03084014 (1 μM) has no antiproliferative effect in MX1 cells; however, it inhibits migration by 95%. ^[2]

Assay

Methods	Test Index	PMID
Western blot	N1ICD / Hes-1 / Hey-1 / p-MEK / MEK / c-PARP ; PubMed: 23402814 Cancer Lett Immunobloting of lysates from Panc265 orthotopic experiment demonstrating that PF-03084014 and combination of GEM plus PF-03084014 strongly inhibit N1ICD and the expression of Notch targets Hes-1 and Hey-1. Cleaved-PARP (c-PARP) levels were elevated in GEM and GEM plus PF-03084014 treatment, indicating higher apoptosis compared to Control or PF-03084014 treatment groups. While the treatments could not modulate MEK1/2 protein expression, PF-03084014 and combination of GEM plus PF-03084014 treatment inhibit MEK1/2 phosphorylation. Numbers on the left side of the blots indicate the molecular weight of the proteins in kilodaltons.	23402814

In vivo

PF-03084014 orally administrated in a single dose of 200 mg/kg, causes maximal NICD inhibition for ∼80% in xenograft HPB-ALL tumors. PF-03084014 shows robust antitumor activity in this mode with a maximal tumor growth inhibition of ∼ 92% at dose of 150 mg/kg, accompanied by a significant reduction of NICD/Notch1, tumor mitotic index (Ki67), and apoptosis (activated caspase-3) staining. ^[1] PF-03084014 (120 mg/kg) induces apoptosis, antiproliferation, reduces tumor cell self-renewal ability, impaires tumor vasculature, and decreases metastasis activity in breast cancer HCC1599 tumor-bearing mice. PF-03084014 treatment displays significant antitumor activity in various types of the breast xenograft models with TGI value of at least 50%. ^[2]

Protocol

Kinase Assay:^[3]	- Collapse γ-secretase assay: A DNA fragment encoding amino acids 596 - 695 of the 695-aa isoform of APP (APP695) and the Flag sequence (DYKDDDDK) at the C terminus is generated by PCR amplification with suitably designed oligonucleotides and the APP695 cDNA. The Met that serves as the translation start site is residue 596 of APP695 (the P1 residue with respect to theβ-secretase cleavage site). This DNA fragment is inserted into the prokaryotic expression vector pET2-21b. The recombinant protein, C100Flag, is overproduced in Escherichia coli [strain BL21(DE3)] and purified by Mono-Q column chromatography. C100Flag (1.7 μM) is incubated with cell membranes (0.5 mg/mL) in the presence of CHAPSO, CHAPS (3-[(3-cholamidopropyl)dim-ethylammonio]-1-propanesulfonate), or Triton X-100 (0, 0.125, 0.25, 0.5, or 1%) in buffer B (50 mM Pipes, pH 7.0y 5mM MgCl₂/5 mM CaCl₂/150 mM KCl) at 37°C. The reactions are stopped by adding RIPA (150 mM NaCl/1.0% NP-40/0.5% sodium deoxycholatey 0.1% SDS/50 mM Tris HCl, pH 8.0) and boiling for 5 min. The samples ae centrifuged and the supernatant solutions are assayed for the Aβ peptides by ECL. The Aβ40- and Aβ42-related products from γ-secretase-mediated processing of C100Flag possess a Met at the N terminus and are thus defined as M-Aβ40 and M-Aβ42, respectively. Likewise, supernatant solution (0.125 mg/mL) from CHAPSO-extracted HeLa cell membranes (solubilized γ-secretase) is incubated with C100Flag (1.7 μM) in buffer B containing 0.25% CHAPSO and subsequently assayed for M-Aβ40 and M-Aβ42 by using ECL.
Cell Research:^[1]	- Collapse Cell lines: Human T-ALL cell lines HPB-ALL Concentrations: ~1 μM Incubation Time: 7 days Method: Cells are seeded in 96-well plates at 10,000 cells/well in growth media supplemented with 10% fetal bovine serum. Serial dilutions of PF-03084014 are done in DMSO, appropriate controls or designated concentrations of PF-03084014 are added to each well, and cells are incubated at 37℃ for 7 days (final DMSO content 0.1%). Resazurin at a final concentration of 0.1 mg/mL is added to the cells and plates are incubated for 2 to 4 hours. Fluorescent signals are read as emission at 590 nm after excitation at 560 nm. (Only for Reference)
Animal Research:^[1]	- Collapse Animal Models: Human T-cell acute lymphoblastic leukemia xenografts HPB-ALL Formulation: 0.5% methylcellulose Dosages: 150 mg/kg Administration: p.o. twice daily (Only for Reference)

References

Solubility (25°C)

In vitro	DMSO	97 mg/mL warmed (198.1 mM)
	Ethanol	97 mg/mL warmed (198.1 mM)
	Water	Insoluble

* Please note that Selleck tests the solubility of all compounds in-house, and the actual solubility may differ slightly from published values. This is normal and is due to slight batch-to-batch variations.

Chemical Information Download Nirogacestat (PF-03084014) SDF

Molecular Weight	489.64
Formula	C₂₇H₄₁F₂N₅O
CAS No.	1290543-63-3
Storage	3 years -20°C powder
Storage	2 years -80°C in solvent
Synonyms	PF-3084014
Smiles	CCCC(NC1CCC2=CC(=CC(=C2C1)F)F)C(=O)NC3=C[N](C=N3)C(C)(C)CNCC(C)(C)C

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Clinical Trial Information (data from https://clinicaltrials.gov, updated on 2019-10-14)

NCT Number	Recruitment	interventions	Conditions	Sponsor/Collaborators	Start Date	Phases
NCT02338531	Withdrawn	Drug: PF-03084014\|Procedure: Breast cancer surgery	Breast Cancer	Jules Bordet Institute	June 2015	Phase 2
NCT02299635	Terminated	Drug: PF-03084014	Triple Negative Breast Neoplasms	Pfizer	February 3 2015	Phase 2
NCT02109445	Terminated	Drug: PF-03084014\|Drug: Gemcitabine\|Drug: Nab-paclitaxel	Metastatic Cancer Pancreas	Pfizer\|Academic GI Cancer Consortium (AGICC)	September 3 2014	Phase 2
NCT01876251	Terminated	Drug: PF-03084014\|Drug: Docetaxel	Breast Cancer Metastatic	Pfizer	November 4 2013	Phase 1
NCT01981551	Active not recruiting	Drug: PF-03084014	Desmoid Tumors\|Aggressive Fibromatosis	National Cancer Institute (NCI)\|National Institutes of Health Clinical Center (CC)	October 31 2013	Phase 2

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Gamma-secretase Signaling Pathway Map

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Nirogacestat (PF-03084014)