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Research Area
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Description
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Cancer |
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Biological Activity
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Description
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Rucaparib (AG-014699, PF-01367338) is an inhibitor of PARP with Ki of 1.4 nM. |
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Targets
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PARP |
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IC50 |
1.4 nM (Ki) [1] |
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In Vitro
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Rucaparib is a potent inhibitor of purified full-length human PARP-1 and shows higher inhibition of cellular PARP in LoVo and SW620 cells. [1] The radio-sensitization by Rucaparib is due to downstream inhibition of activation of NF-κB, and is independent of SSB repair inhibition. Rucaparib could target NF-κB activated by DNA damage and overcome toxicity observed with classical NF-κB inhibitors without compromising other vital inflammatory functions. [2] Rucaparib inhibits PARP-1 activity by 97.1% at a concentration of 1 μM in permeabilised D283Med cells. [3] |
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In Vivo
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Rucaparib is not toxic but significantly enhances temozolomide-induced TGD in the DNA repair protein-competent D384Med xenografts. Pharmacokinetics studies also show that Rucaparib is detected in the brain tissue, which indicates that Rucaparib has potential in intra-cranial malignancy therapy. [3] Rucaparib significantly potentiates the cytotoxicity of topotecan and temozolomide in NB-1691, SH-SY-5Y, and SKNBE (2c) cells. Rucaparib enhances the antitumor activity of temozolomide and indicates complete and sustained tumor regression in NB1691 and SHSY5Y xenografts. [4] |
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Clinical Trials
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Rucaparib is currently in Phase II clinical trials for locally advanced/metastatic breast or advanced ovarian cancer. |
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Features
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Rucaparib is the phosphate salt of AG-14447 and the first PARP inhibitor to be used in clinical trials, combined with temozolomide. |
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Protocol
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Kinase Assay
[1]
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| Ki Determination |
Inhibition of human full-length recombinant PARP-1 by [32P]NAD+ incorporation is measured. The [32P]ADP-ribose incorporated into acid insoluble material is quantified using a PhosphorImager. Ki is calculated by nonlinear regression analysis. |
| Inhibition of PARP activity |
Inhibition of PARP activity in 5 × 103 D283Med cells is measured using various concentrations of Rucaparib (0-1 μM), compared with DMSO-only. Maximally stimulated PARP activity is measured in samples of permeabilised cells by immunological detection of the amount of poly(ADP-ribose) (PAR) formed, using the 10H anti-PAR antibody, during a 6-min incubation with NAD+ and oligonucleotide (substrate and activator). A PAR standard curve using a GCLP-validated assay is used as reference for the assay. [3] |
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Cell Assay
[3]
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| Cell Lines |
D425Med, D283Med and D384Med cells |
| Concentrations |
0.4 μM |
| Incubation Time |
3 or 5 days |
| Methods |
Medulloblastoma cell lines are seeded in 96-well plates at a density of 1 × 103, 3 × 103 and 3 × 103, respectively. At 24 hours (D384Med) or 48 hours (D283Med and D425Med) after seeding, the cells are exposed to various concentrations of temozolomide in the presence or absence of 0.4 μM Rucaparib. After 3 days (D425Med and D384Med) or 5 days (D283Med) of culture, cell viability is evaluated by a XTT cell proliferation kit assay. Cell growth is expressed as a percentage in relation to DMSO or 0.4 μM Rucaparib-alone controls. The concentration of temozolomide, alone or in combination with Rucaparib that inhibited growth by 50% (GI50) is calculated. The potentiation factor 50 (PF50) is defined as the ratio of the GI50 of temozolomide in the presence of Rucaparib to the GI50 of temozolomide alone. |
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Animal Study
[3]
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| Animal Models |
CD-1 nude mice bearing established D283Med xenografts |
| Formulation |
Normal saline |
| Doses |
1 mg/kg |
| Administration |
One or four daily by i.p. |
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References |
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[1] Thomas HD, et al. Mol Cancer Ther, 2007, 6(3), 945-956.
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[2] Hunter JE, et al. Oncogene, 2012, 31(2), 251-264.
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[3] Daniel RA, et al. Br J Cancer, 2010, 103(10), 1588-1596.
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[4] Daniel RA, et al. Clin Cancer Res, 2009, 15(4), 1241-1249.
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