5 research outputs found
Automated Affinity Capture and On-Tip Digestion to Accurately Quantitate <i>in Vivo</i> Deamidation of Therapeutic Antibodies
Deamidation
of therapeutic antibodies may result in decreased drug
activity and undesirable changes in pharmacokinetics and immunogenicity.
Therefore, it is necessary to monitor the deamidation levels [during
storage] and after <i>in vivo</i> administration. Because
of the complexity of <i>in vivo</i> samples, immuno-affinity
capture is widely used for specific enrichment of the target antibody
prior to LC–MS. However, the conventional use of bead-based
methods requires large sample volumes and extensive processing steps.
Furthermore, with automation difficulties and extended sample preparation
time, bead-based approaches may increase artificial deamidation. To
overcome these challenges, we developed an automated platform to perform
tip-based affinity capture of antibodies from complex matrixes with
rapid digestion and peptide elution into 96-well microtiter plates
followed by LC–MS analysis. Detailed analyses showed that the
new method presents high repeatability and reproducibility with both
intra and inter assay CVs < 8%. Using the automated platform, we
successfully quantified the levels of deamidation of a humanized monoclonal
antibody in cynomolgus monkeys over a time period of 12 weeks after
administration. Moreover, we found that deamidation kinetics between <i>in vivo</i> samples and samples stressed <i>in vitro</i> at neutral pH were consistent, suggesting that the <i>in vitro</i> stress test may be used as a method to predict the liability to
deamidation of therapeutic antibodies <i>in vivo</i>
Discovery of GDC-0853: A Potent, Selective, and Noncovalent Bruton’s Tyrosine Kinase Inhibitor in Early Clinical Development
Bruton’s
tyrosine kinase (Btk) is a nonreceptor cytoplasmic
tyrosine kinase involved in B-cell and myeloid cell activation, downstream
of B-cell and Fcγ receptors, respectively. Preclinical studies
have indicated that inhibition of Btk activity might offer a potential
therapy in autoimmune diseases such as rheumatoid arthritis and systemic
lupus erythematosus. Here we disclose the discovery and preclinical
characterization of a potent, selective, and noncovalent Btk inhibitor
currently in clinical development. GDC-0853 (<b>29</b>) suppresses
B cell- and myeloid cell-mediated components of disease and demonstrates
dose-dependent activity in an <i>in vivo</i> rat model of
inflammatory arthritis. It demonstrates highly favorable safety, pharmacokinetic
(PK), and pharmacodynamic (PD) profiles in preclinical and Phase 2
studies ongoing in patients with rheumatoid arthritis, lupus, and
chronic spontaneous urticaria. On the basis of its potency, selectivity,
long target residence time, and noncovalent mode of inhibition, <b>29</b> has the potential to be a best-in-class Btk inhibitor for
a wide range of immunological indications
Discovery of GDC-0853: A Potent, Selective, and Noncovalent Bruton’s Tyrosine Kinase Inhibitor in Early Clinical Development
Bruton’s
tyrosine kinase (Btk) is a nonreceptor cytoplasmic
tyrosine kinase involved in B-cell and myeloid cell activation, downstream
of B-cell and Fcγ receptors, respectively. Preclinical studies
have indicated that inhibition of Btk activity might offer a potential
therapy in autoimmune diseases such as rheumatoid arthritis and systemic
lupus erythematosus. Here we disclose the discovery and preclinical
characterization of a potent, selective, and noncovalent Btk inhibitor
currently in clinical development. GDC-0853 (<b>29</b>) suppresses
B cell- and myeloid cell-mediated components of disease and demonstrates
dose-dependent activity in an <i>in vivo</i> rat model of
inflammatory arthritis. It demonstrates highly favorable safety, pharmacokinetic
(PK), and pharmacodynamic (PD) profiles in preclinical and Phase 2
studies ongoing in patients with rheumatoid arthritis, lupus, and
chronic spontaneous urticaria. On the basis of its potency, selectivity,
long target residence time, and noncovalent mode of inhibition, <b>29</b> has the potential to be a best-in-class Btk inhibitor for
a wide range of immunological indications
Discovery of Potent and Selective Tricyclic Inhibitors of Bruton’s Tyrosine Kinase with Improved Druglike Properties
In our continued effort to discover
and develop best-in-class Bruton’s tyrosine kinase (Btk) inhibitors
for the treatment of B-cell lymphomas, rheumatoid arthritis, and systemic
lupus erythematosus, we devised a series of novel tricyclic compounds
that improved upon the druglike properties of our previous chemical
matter. Compounds exemplified by <b>G-744</b> are highly potent,
selective for Btk, metabolically stable, well tolerated, and efficacious
in an animal model of arthritis
Discovery of Novel PI3-Kinase δ Specific Inhibitors for the Treatment of Rheumatoid Arthritis: Taming CYP3A4 Time-Dependent Inhibition
PI3Kδ is a lipid kinase and a member of a larger
family of enzymes, PI3K class IAÂ(α, β, δ) and IB
(γ), which catalyze the phosphorylation of PIP2 to PIP3. PI3Kδ
is mainly expressed in leukocytes, where it plays a critical, nonredundant
role in B cell receptor mediated signaling and provides an attractive
opportunity to treat diseases where B cell activity is essential,
e.g., rheumatoid arthritis. We report the discovery of novel, potent,
and selective PI3Kδ inhibitors and describe a structural hypothesis
for isoform (α, β, γ) selectivity gained from interactions
in the affinity pocket. The critical component of our initial pharmacophore
for isoform selectivity was strongly associated with CYP3A4 time-dependent
inhibition (TDI). We describe a variety of strategies and methods
for monitoring and attenuating TDI. Ultimately, a structure-based
design approach was employed to identify a suitable structural replacement
for further optimization