Molecular Glue/PROTAC® Optimization Kit for CDK2/CDK9-Cereblon Binding

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Application: Identify and optimize Molecular Glues/PROTACs targeting CDK2, CDK9, and CRBN.Design novel molecules targeting CDK2, CDK9, and CRBN.Directly compare the activity of different Molecular Glues/PROTACs.

Background: CDKs (cyclin-dependent kinases) are serine/threonine kinases involved in critical cellular functions. CDK2 (Cyclin-Dependent Kinase 2) is a cell cycle regulator specifically involved in the transition from G1 to S phase of the cell cycle. It forms complexes with Cyclin E and Cyclin A to phosphorylate key substrates involved in DNA replication and cell cycle progression. CDK9 (Cyclin-Dependent Kinase 9) primarily regulates transcription. It forms the catalytic core of the positive transcription elongation factor b (P-TEFb) complex and phosphorylates the C-terminal domain (CTD) of RNA polymerase II, playing a crucial role in promoting transcriptional elongation.CDK9 is an emerging target for cancer therapy, especially in hematological malignancies. Since it is also involved in HIV replication, it is a potential target for anti-HIV therapies. CDK9 inhibitors are being investigated for their potential in treating inflammatory diseases and cardiovascular conditions. CDK2 is a well validated target in oncology, particularly in cancers with dysregulated cell cycle control. The protein may also play a role in certain neurodegenerative diseases. In cancer, CDK2 has emerged as a mechanism of resistance acquired after treatment of estrogen receptor-positive breast cancer patients with CDK4/6 inhibitors like Kisqali, Verzenio and Ibrance, a phenomenon driven by upregulation of CCNE1 (cyclin E1) which directly modulates CDK2 activity. These tumors may be activated by the amplification of Cyclin E1 or E2, loss of the AMBRA1 (activating molecule in Beclin-regulated autophagy) gene, or loss of retinoblastoma. Many CDK2 inhibitors are in development, however the protein is relatively difficult to drug, and toxic off-targets effects are an issue. Degraders are envisioned as an alternative to small molecule inhibitors.  Both CDK2 and CDK9 are subjects of ongoing clinical research, with various inhibitors in different stages of clinical trials for various conditions, primarily cancer.

Contraindications: The final concentration of DMSO in the assay should not exceed 1%.Avoid green and blue dyes that absorb light in the AlphaScreen signal emission range (λ=520-620 nm), such as Trypan Blue.Avoid the use of potent singlet oxygen quenchers such as sodium azide (NaN₃) or metal ions (Fe²⁺, Fe³⁺, Cu²⁺, Zn²⁺ and Ni²⁺).The presence of >1% RPMI 1640 culture medium leads to a signal reduction due to the presence of excess biotin and iron in this medium. Media like MEM, which lacks these components, does not affect AlphaScreen assays.

Description: The Molecular Glue/PROTAC® Optimization Kit for CDK2/CDK9-Cereblon Binding is designed for the testing and profiling of Molecular Glues (MG) and PROTACs targeting CDK2 (cyclin dependent kinase 2) or CDK9 (cyclin dependent kinase 9) and Cereblon (CRBN). The Molecular Glue/PROTAC® Optimization Kit for CDK2/CDK9-Cereblon Binding comes in a convenient AlphaLISA® format, with enough PROTAC® buffer, purified CDK2/CyclinA2 and CDK9/CyclinK complexes, and CRBN for 384 reactions. This kit also contains the control PROTAC® (CDK2/9 Degrader) and CDK inhibitor FN-1501.The Molecular Glue/PROTAC® of interest is incubated with Cereblon (CRBN) and CDK2 or CDK9, bringing them into proximity. CRBN contains a FLAG-tag, which is recognized by anti-FLAG AlphaLISA„¢ acceptor bead. CDK contains a GST-tag that binds to the donor bead. Upon excitation of the donor bead, a singlet oxygen is generated by the bead. The singlet oxygen excites the acceptor bead, which emits light proportionally to the level of interaction between CDK2/CDK9 and Cereblon.

Storage Stability: This assay kit will perform optimally for up to 6 months from date of receipt when the materials are stored as directed.

Warnings: Avoid freeze/thaw cycles

Biosafety Level: Not applicable (BSL-1)

References: Gerosa R., et al., 2024 Crit Rev Oncol Hematol. 196: 104324. 
Tadesse S., et al., 2020 Drug Discov Today. 25(2): 406-413. 

• Artikelnummer: BPS82643
• Hersteller: BPS Bioscience
• Hersteller Artikelnummer: 82643
• Verpackungseinheit: 384 reactions
• Mengeneinheit: PAK