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NPOT®
NPOT® (Nematic Protein Organization Technique) is a biophysical platform that maps how drugs interact with proteins in their native, unaltered state. By analysing tissues, cell lines, or patient biopsies directly, NPOT® identifies drug targets, interaction networks, and biomarkers – without disrupting the natural biological environment. Compatible with small molecules, antibodies, peptides, and large compounds, NPOT® accelerates drug discovery by revealing how therapies truly work in disease contexts.
Leveraging Kirkwood-Buff molecular crowding and aggregation theory, NPOT® harnesses the power of any chemical entity—be it a small molecule, protein, or high molecular weight compound—to shift the equilibrium of cellular proteins in a pH gradient, reflecting different cell compartments. This shift triggers the formation of a heteroassembly, which is then isolated and analyzed via LC-MS/MS.
Next, we use InOPERA®, Inoviem Scientific proprietary database, to distinguish nonspecific proteins (which display a high frequency of appearance within database integrated biosamples), from specific proteins, which include the target candidate.
NPOT® is a game-changer, offering an unbiased approach that not only identifies targets but also reveals the early stages of the mode of action (MOA). Proven effective across nearly all therapeutic areas, NPOT® pinpoints direct drug targets (ON- and OFF-targets) and indirect effectors, uncovering pathways and MOAs. Each identified protein undergoes thorough multidirectional analysis, including functional studies, physico-chemical property assessments, and UniProt annotation.
To ensure precision, NPOT® includes a parallel analysis with a negative control compound, such as an inactive enantiomer or a structurally similar compound with minimal pharmacological activity. This critical step guarantees the identification of selective and specific proteins within the interactome, setting a new standard for drug discovery and development.
NPOT® combines biophysical principles with advanced proteomics to study drug effects in native tissues:
Works in human tissues, including biopsies—no artificial models.
Uses inactive compounds as controls to filter out false positives.
Go from sample to actionable insights in days, accelerating translational research.
Following LC-MS/MS sequencing, Inoviem’s proprietary software (InOPERA®) rigorously mines the data to reveal true interactome members. Our InOPERA® database cross-references each protein against more than 300 NPOT® studies, using frequency analysis to exclude nonspecific ontaminants (proteins that appear in over 25% of studies, such as actin).
In parallel, we conduct detailed physicochemical profiling – comparing
hydrophobicity (GRAVY index), net charge, and interaction potential (Boman index) between test compounds and controls – to prioritize selective interactors. Combined with functional annotations from resources like UniProt, this comprehensive workflow provides an unbiased, complementary approach for pinpointing the primary targets of drug candidates.
InOPERA® application user interface. A Comparison filter. Selection of imported dataset(s) and/or selection of biosample(s) to be compared in the InoPERA® database. B. Selection of the reference filter(s) to define the subset of the database used for frequency calculation. Different reference filter strategies can be combined, based on the cell types, organs, therapeutic areas, species, and protocols.
The proteins enriched by NPOT® are further interrogated using multiple functional enrichment analysis tools. These tools integrate data across various species and validation methodologies, associating the identified proteins with biological functions and pathways. Complementary network analyses further enhance our understanding by visualizing the functional networks and elucidating the interplay among interactome components:
Post-analysis, Inoviem validates both target candidates and the identified protein-protein interactions using Surface Plasmon Resonance (SPR). This validation step is applicable to both membrane-bound and cytosolic proteins, allowing for precise measurements of binding specificity, active concentration, and the kinetic and thermodynamic parameters of the interactions.
By preserving the native state of biological systems, NPOT® ensures that insights into drug action and target engagement are as close to the in vivo reality as possible, thus driving precision medicine forward.
NPOT® bridges the gap between lab experiments and real-world biology. Whether you’re optimizing lead compounds or de-risking clinical candidates, NPOT® delivers mechanistic clarity for smarter decisions.
