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Based in Scotland - UK Tools for cell biology and biomanufacturing Scottish Life-Sciences cluster (at BioCity) Technology based on world-class science 12 staff
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Physiological cell death:
viable cell dying cell (apoptotic) dead cell (necrotic) sub-cellular debris apoptosis phagocyte
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Cell death in vitro: phagocytes absent
viable cells dying cell (apoptotic) dead cell (necrotic) phagocytes absent cell corpses persist and accumulate inhibitory cascade initiated… - build-up of cell debris - inappropriate signalling mechanisms - slowed growth (and secretory activity) sub-cellular debris
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Consequences of cell death in vitro:
dead cell (necrotic) dying cell (apoptotic) sub-cellular debris decreased purity loss of reproducibility slower growth/yield
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ClioCell - GMP quality removal of dead cells:
Product comprises: nanoparticles (250 nm) YES medium* magnet Key attributes: Cost-effective Simple to use Efficient Disposable Scalable GMP compatible *yield enhancing separation
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ClioCell = rapid removal of dead cells:
Input cells & debris ClioCell nanoparticles Output cells Debris Simple Magnetic Separation Viable Cell High Viability Non-Viable Cell Dead cells & debris retained
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Benefits of ClioCell to cell-based systems:
Input Output Extended growth Antibody Concentration (ng/Cell) Dead-cell-depleted Culture Untreated Culture Extended productivity ES Cells Antibody Production* *Landes Bioscience - Volume 1, Issue 4 July/August 2009
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ClioCell – history of use:
CELL TYPE: SPECIES: Cardiomyocytes Human NSO myeloma Mouse Hybridomas (n=6) Burkitt lymphomas Lambda-MYC lymphomas Neutrophil Thymocyte Macrophage THP-1 monocyte/macrophage RAW-264 monocyte/macrophage J774 monocyte/macrophage Bladder carcinoma Foetal liver A549 lung carcinoma Human & Human Embryonic stem cells Very small embryonic-like stem cells Burkitt lymphoma xenograft Human in Mouse CHO Hamster U937 monocytic leukaemia L929 fibroblasts Dendritic cells HL60 leukaemia Monomac 6 leukaemia H345 lung carcinoma Cytotoxic T lymphocytes PER.C6
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ClioCell – research applications:
Stem cell research: rescue of ‘inhibited’ population (from Roslin Cells, UK) cryo-damaged E14-IV embryonic stem cells Untreated Single separation
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ClioCell – research applications:
Drug screening (chemotaxis): neutrophil chemotaxis towards fMLP (5 µm Boyden chamber) migration inhibited by apoptotic cells removal of apoptotic cells increased assay sensitivity by 170% IN OUT ClioCell
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‘Pinch-points’ in the cell culture cycle:
point of care
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BL2 frozen for 24 h then cultured for up to 5 h in BL2 medium
Optimizing cell growth (manufacturing): Removal of dead cells BL2 frozen for 24 h then cultured for up to 5 h in BL2 medium 20 40 60 80 100 0 min 15 min 30 min 45 min 1 h 2 h 3 h 4 h 5 h BL2 cells % of ungated cells
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Reduction of dead cell number speeds proliferation
Optimizing cell growth (manufacturing): Reduction of dead cell number speeds proliferation 20 40 60 80 100 0 min 15 min 30 min 45 min % of ungated cells 1 h 2 h 3 h 4 h 5 h BL2 cells
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Clinical application of ClioCell:
Cell therapy: Inconsistent supply of product due to dead/dying cells reduction in manufacturing cycle (lower CoGs) enhanced purity and potency (regulatory and quality) increased shelf-life GREATER EFFICACY Treat Discard
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Therapeutic use of ClioCell:
EBV-specific cytotoxic T lymphocytes* Cells thawed, incubated at 37oC for 2 hours Viability counts performed using trypan blue and FACS Two ClioCell separations performed… Untreated After ClioCell Untreated st nd ClioCell *Collaboration with University of Edinburgh
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Further uses of ClioCell:
Bioprocessing: rescue of ‘inhibited’ populations increased (per-cell) yield of protein less requirement to scale-up reactor BENEFITS TO CUSTOMER: … ‘more product for less cost’ IgM produced per cell (pg) Untreated ClioCell® 5-Day Yield… 50 100 150 200 250
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ClioCell – USPs: Cost-effective - 25 uses for (approx) $425
CoGs = $40 for nanoparticles, YES medium & magnet* Simple to use – one formulation ‘fits all’ Applicable to all cell biology sectors No cell loss GMP compatible 12 month shelf-life (potentially 24 months) Regulatory requirement (quality, efficacy) * Assumes batch size of 250 or more
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ClioCell – target market:
Cell biologists… - research (academia) - product development - cell therapy Simple message – ‘negative influence of dead cells’ Bespoke applications (in-house technical support) - debris removal prior to FACS - drug screening, in vitro toxicity - RNA purification - biomanufacturing - rescue of senescent populations Requirement to educate and advise
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ClioCell – SUMMARY: Simple to use Cost effective
Numerous applications include: rescue of ‘dead-cell inhibited’ population recovery of viability following cold-shock removal of inhibitory components from media maintenance of (therapeutic) phenotype GMP compliant Patent protected
