Molecule

Description

Cancer

Description

Rucaparib (AG-014699, PF-01367338) is an inhibitor of PARP with K_i of 1.4 nM.

Targets

IC₅₀

In Vitro

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]

In Vivo

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]

Clinical Trials

Rucaparib is currently in Phase II clinical trials for locally advanced/metastatic breast or advanced ovarian cancer.

Features

Rucaparib is the phosphate salt of AG-14447 and the first PARP inhibitor to be used in clinical trials, combined with temozolomide.

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. K_i 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]

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.

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.