Palbociclib Isethionate

Cancer Res,2006 Aug 1;66(15):7661-7.

A novel orally active small molecule potently induces G1 arrest in primary myeloma cells and prevents tumor growth by specific inhibition of cyclin-dependent kinase 4/6.[Pubmed: 16885367]

METHODS AND RESULTS:
Cell cycle deregulation is central to the initiation and fatality of multiple myeloma, the second most common hematopoietic cancer, although impaired apoptosis plays a critical role in the accumulation of myeloma cells in the bone marrow. The mechanism for intermittent, unrestrained proliferation of myeloma cells is unknown, but mutually exclusive activation of cyclin-dependent kinase 4 (Cdk4)-cyclin D1 or Cdk6-cyclin D2 precedes proliferation of bone marrow myeloma cells in vivo. Here, we show that by specific inhibition of Cdk4/6, the orally active small-molecule PD 0332991 potently induces G(1) arrest in primary bone marrow myeloma cells ex vivo and prevents tumor growth in disseminated human myeloma xenografts. PD 0332991 inhibits Cdk4/6 proportional to the cycling status of the cells independent of cellular transformation and acts in concert with the physiologic Cdk4/6 inhibitor p18(INK4c). Inhibition of Cdk4/6 by PD 0332991 is not accompanied by induction of apoptosis. However, when used in combination with a second agent, such as dexamethasone, PD 0332991 markedly enhances the killing of myeloma cells by dexamethasone.
CONCLUSIONS:
PD 0332991, therefore, represents the first promising and specific inhibitor for therapeutic targeting of Cdk4/6 in multiple myeloma and possibly other B-cell cancers.

J Biol Chem,2001 May 18;276(20):16617-23.

Cell cycle and biochemical effects of PD 0183812. A potent inhibitor of the cyclin D-dependent kinases CDK4 and CDK6.[Pubmed: 11278443]

CDK assays for IC50 determinations and kinetic evaluation are performed in 96-well filter plates. All CDK-cyclin kinase complexes are expressed in insect cells through baculovirus infection and purified. The substrate for the assays is a fragment (amino acids 792–928) of pRb fused to GST (GST•RB-Cterm). The total reaction volume is 0.1 mL containing a final concentration of 20 mM Tris-HCl, pH 7.4, 50 mM NaCl, 1 mM dithiothreitol, 10 mM MgCl2 25 μM ATP (for CDK4-cyclin D1, CDK6-cyclin D2, and CDK6-cyclin D3) containing 0.25 μCi of [γ-32P]ATP, 20 ng of enzyme, 1 μg of GST•RB-Cterm, and appropriate dilutions of inhibitor. All components except the [γ-32P]ATP are added to the wells, and placed on a plate mixer for 2 min. The reaction is started by adding the [γ-32P]ATP, and incubated at 25℃ for 15 min. The reaction is terminated by addition of 0.1 mL of 20% trichloroacetic acid, and the plate is kept at 4 ℃ for at least 1 hr to allow the substrate to precipitate. The wells are then washed five times with 0.2 mL of 10% trichloroacetic acid, and radioactive incorporation is determined with a β plate counter.

Breast Cancer Res,2009;11(5):R77.

PD 0332991, a selective cyclin D kinase 4/6 inhibitor, preferentially inhibits proliferation of luminal estrogen receptor-positive human breast cancer cell lines in vitro.[Pubmed: 19874578]

Cell lines: Human breast cancer cells MDA-MB-435
Concentrations:  2 μM
Incubation Time:  6 days
Method:
Cells are seeded in duplicate at 5,000 to 10,000 cells per well in 24-well plates. The day after plating, different concentrations of PD 0332991 are added. Control wells without drug are also seeded. At the end of incubation, cells are trypsinizated and placed in Isotone solution and counted immediately using a Coulter Z2 particle counter.

Mol Cancer Ther,2004 Nov;3(11):1427-38.

Specific inhibition of cyclin-dependent kinase 4/6 by PD 0332991 and associated antitumor activity in human tumor xenografts.[Pubmed: 15542782]

Animal Models:Human colon carcinoma xenografts Colo-205
Dosages:150 mg/kg
Administration: o.p. injection every day