Experiment Consulting and Design Service

Background:

A pharmaceutical client required a comprehensive strategy to produce and characterize a multi-pass transmembrane receptor for antibody screening. The target receptor featured seven transmembrane helices, multiple extracellular loops with complex disulfide patterns, and an intracellular signaling domain—a structural architecture notoriously challenging for recombinant production. Previous attempts using conventional bacterial expression systems yielded exclusively insoluble aggregates in inclusion bodies, and mammalian transient expression suffered from low yields and batch-to-batch inconsistency. With the project timeline rapidly approaching critical milestones, an integrated, risk-mitigated approach was urgently needed.

Our Solution:

Profacgen's consulting team conducted a systematic feasibility assessment, evaluating multiple expression hosts (E. coli, insect cells, and mammalian systems), solubilization strategies (including mild detergents, mixed micelles, and styrene-maleic acid copolymers), and a detergent screening matrix of over 60 unique formulations to identify conditions optimal for both extraction and downstream stability. We then designed a phased experimental plan beginning with BacMam-mediated transient transduction in mammalian cells to ensure proper post-translational processing, followed by membrane extraction, detergent exchange, and reconstitution into lipid nanodiscs for stability and functional integrity. Comprehensive biophysical characterization—including size-exclusion chromatography (SEC), thermal shift assays, and surface plasmon resonance (SPR) for ligand-binding activity—was incorporated at each step as go/no-go checkpoints.

Final Results:

The designed workflow yielded 8 mg/L of correctly folded receptor from shaker flask culture, with native glycosylation patterns confirmed by enzymatic deglycosylation and mass spectrometry. Nanodisc-reconstituted protein demonstrated expected ligand-binding affinity (K_D ≈ 15 nM) and conformational stability suitable for downstream applications. The purified material enabled successful antibody panning campaigns, yielding multiple lead candidates that progressed through affinity maturation. The client advanced to preclinical candidate selection six months ahead of the original schedule, significantly reducing R&D costs and accelerating their therapeutic program timeline.

Background:

A biotech startup sought to initiate a targeted protein degradation program against an oncogenic transcription factor that had long been considered "undruggable" due to its intrinsically disordered domain and lack of enzymatic pockets suitable for conventional inhibitor development. The client possessed strong biological validation of the target's role in tumor maintenance but lacked internal expertise in E3 ligase selection, degrader design principles, and the complex biochemistry of ternary complex formation—representing substantial barriers to program initiation.

Our Solution:

We performed comprehensive computational profiling of the target's degron landscape, analyzing sequence features, structural predictions, and previously reported degradation signals to identify permissive surface regions amenable to ubiquitin ligase recruitment. Concurrently, we profiled E3 ligase tissue expression across healthy and tumor tissues to prioritize ligases with favorable specificity profiles and minimal off-tumor liability. To guide molecular design, we performed predictive modeling of ternary complex geometry between the target, candidate ligands, and selected E3 ligases, assessing potential interface complementarity and steric feasibility. Based on these insights, we designed a comprehensive experimental roadmap encompassing in silico screening of ligand libraries, in vitro ubiquitination assays using purified E2/E3 enzymatic systems, and cellular degradation assessments with quantitative Western blot and proteomic readouts.

Final Results:

The integrated strategy identified a potent molecular glue within 12 weeks—a timeline substantially shorter than typical degrader discovery campaigns. The lead compound demonstrated confirmed target engagement (EC₅₀ ≈ 50 nM in cellular NanoBRET assays) and dose-dependent degradation (DC₅₀ ≈ 120 nM) with minimal effects on closely related family members, indicating target selectivity. Based on these compelling proof-of-concept data, the client secured Series A funding from multiple investors, positioning them for rapid progression toward lead optimization and in vivo efficacy studies.