Benchmark Degraders and Chemical Probes

A cellular degradation readout reports the outcome of a long chain of events, since the compound must cross the membrane, form a ternary complex, drive ubiquitination of the target and deliver it to the proteasome within the time window of the experiment. A compound whose behavior in that chain is already established therefore carries a large share of the interpretive weight in any degradation assay, whether it serves as the positive control that qualifies the cell system, the benchmark against which an internal series is ranked, or the tool that tests which ligase an observed degradation event depends on. Amerigo Scientific provides published degraders covering a range of targets and both canonical E3 ligases, Homo-PROTAC tools that degrade the ligase itself, cereblon-binding molecular glues, and a high-affinity VHL pathway probe.

Benchmark Degraders

Degradation assays are compared through their response shape rather than a single number, since a degrader is described by the concentration producing half-maximal degradation, the maximum extent of degradation reached, the time course over which the target disappears, and the concentration above which binary complexes begin to compete with the productive ternary complex. Establishing that shape in a given cell line and readout requires a compound that has already been characterized elsewhere.

We provide published degraders as reference compounds for assay development and compound series benchmarking, covering targets across bromodomain, kinase, nuclear receptor and other target families, and recruiting either cereblon or VHL.

Applications

• Positive control in cellular degradation assays
• Assay development and validation
• Benchmarking of internal compound series
• Time course and hook effect characterization
• Target biology and pathway studies

Homo-PROTAC E3 Ligase Degradation Tools

A Homo-PROTAC carries two copies of the same E3 ligase ligand, so it recruits the ligase to itself and drives its degradation. This converts an E3 ligase into a chemically depletable component of the experiment, which offers a route to test whether a degradation event depends on the ligase that a compound was designed to recruit.

Amerigo Scientific offers Homo-PROTAC tools directed at the two ligases most widely used in degrader design, VHL and cereblon.

Applications

• Chemical depletion of an E3 ligase
• Confirmation of ligase dependency for a degradation event
• Studies of E3 ligase turnover and stability
• Control compound alongside ligase-recruiting degraders

Cereblon-Binding Molecular Glues

Cereblon-binding immunomodulatory compounds reshape the substrate preference of the CRL4^CRBN ligase, which brings proteins to the ligase that it would otherwise leave alone. These compounds established the mechanism that molecular glue programs now pursue deliberately, and they remain the reference points for cereblon-directed work.

We provide lenalidomide, a thalidomide analog and cereblon binding compound that induces ubiquitination and degradation of casein kinase 1α by the CRL4^CRBN ligase, and that also acts as an inhibitor of TNF-α and of angiogenesis. We also provide pomalidomide, a thalidomide derivative that binds cereblon and inhibits its ubiquitination, a potent TNF-α inhibitor with an IC50 of 13 nM, with reported antiproliferative activity in the Namalwa lymphoma cell line and antiangiogenic properties. Both compounds also serve as the cereblon ligand starting point for PROTAC synthesis.

Applications

• Reference compounds for molecular glue mechanisms
• Cereblon binding and substrate recruitment studies
• Casein kinase 1α degradation studies
• Immunomodulatory and antiangiogenic activity studies
• Cereblon ligand starting point for PROTAC synthesis

E3 Ligase Pathway Probes

Occupying the substrate binding site of an E3 ligase with a high-affinity ligand blocks the recruitment of its natural substrates, which reveals the downstream biology of the ligase and provides a competition control for degraders that recruit the same ligase.

VH298 is a cell permeable high-affinity inhibitor of the VHL E3 ubiquitin ligase with a Kd of 80 to 90 nM. It blocks the interaction between VHL and HIF-α downstream of HIF-α hydroxylation and initiates a hypoxic response, producing time- and concentration-dependent accumulation of hydroxylated HIF-α and upregulating the mRNA and protein levels of HIF target genes.

Applications

• VHL pathway investigation
• Hypoxic response induction
• HIF-α stabilization studies