PBMNC Cryopreservation in Platelet Plasma for Ischemia

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Solution Overview

Problem

Current methods for preparing autologous peripheral blood mononuclear cell (PBMNC) concentrates for treating critical limb ischemia suffer from high neutrophil contamination, red blood cell contamination, and qualitative/quantitative composition issues, leading to reduced efficacy and limited storage and administration time.

Innovation Solution

An autologous preparation comprising PBMNCs suspended in autologous platelet-enriched plasma or platelet concentrate, processed under controlled conditions to minimize red blood cells and neutrophils, with DMSO added for cryopreservation, and diluted with autologous plasma upon thawing to enhance angiogenic signals.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Quantity of substance

If conventional apheresis systems and centrifugation systems are used to concentrate PBMNCs, then cell concentration is achieved, but high neutrophil contamination and red blood cell contamination occur, reducing therapeutic efficacy

Engineering Contradiction:

ImprovePBMNC concentrationVSAvoidneutrophil contamination and red blood cell contamination

Core Design Contradiction:

Quantity of substanceVSObject-affected harmful factors

Solution Approach 1:

The patent applies extraction by removing harmful components (neutrophils and red blood cells) from the PBMNC concentrate through specific filtration steps. The filtration system selectively extracts contaminating cells while retaining the therapeutic PBMNCs, thereby improving purity without sacrificing concentration.

Inventive Principle:

Principle #2Taking out (Extraction)

Solution Approach 2:

The patent implements local quality by creating distinct functional zones in the filtration system with different pore sizes and selectivity characteristics. The first filter removes larger contaminants (red blood cells) while the second filter removes smaller contaminants (neutrophils), with each filter optimized for its specific function to achieve progressive purification.

Inventive Principle:

Principle #3Local quality

2Duration of action of stationary object

If PBMNC concentrate is prepared and stored for later administration, then treatment flexibility is improved, but cellular viability and efficacy decrease over time

Engineering Contradiction:

Improvestorage timeVSAvoidcellular viability and efficacy

Core Design Contradiction:

Duration of action of stationary objectVSReliability

Solution Approach 1:

The patent applies parameter changes by optimizing the composition of the suspension medium with specific concentrations of antioxidants, growth factors, and protective agents. These compositional parameters are tuned to create a microenvironment that maintains cellular viability during storage, extending the usable storage period while preserving therapeutic efficacy.

Inventive Principle:

Principle #35Parameter changes

3Reliability

If platelet-rich plasma is used as suspension medium, then angiogenic activity is enhanced, but preparation complexity increases

Engineering Contradiction:

Improveangiogenic activityVSAvoidpreparation process complexity

Core Design Contradiction:

ReliabilityVSDevice complexity

Solution Approach 1:

The patent applies merging by combining multiple beneficial components (PBMNCs, platelet-rich plasma, growth factors, and protective agents) into a single integrated suspension medium. This unified formulation provides synergistic effects that enhance angiogenic activity while the standardized protocol simplifies the overall preparation process despite the multiple components involved.

Inventive Principle:

Principle #5Merging (Combining)

Applied Scientific Principles

This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.

Function Achieved in This Case

The solution results in a standardized, high-quality product with extended storage and administration flexibility, improved angiogenic activity, and enhanced therapeutic efficacy by maintaining cellular viability and platelet growth factor expression.

Implementation Method 1

Freezing the autologous concentrate and the autologous platelet-enriched plasma or platelet concentrate; thawing, upon use, the thawed autologous concentrate and the thawed autologous platelet-enriched plasma or platelet concentrate

Methodology Applied

Scientific EffectCryopreservation: Freezing

Implementation Method 2

treating a blood sample taken from the patient to obtain autologous PBMNCs and autologous platelet-enriched plasma or platelet concentrate

Methodology Applied

Scientific EffectCentrifugal separation: Centrifugal Separation

Implementation Method 3

processed under controlled conditions to minimize red blood cells and neutrophils

Methodology Applied

Scientific EffectFiltration: Filter (physical)

Data Source

PatentEP3342856B1Peripheral blood mononucleate cells formulations subjected to hypoxia

Publication Date: 2022.05.04 QUANTIX SWISS SA

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AI summary

The object of the present invention is an autologous preparation (A) substantially free of red blood cells and neutrophil-depleted, comprising peripheral blood mononuclear cells (PBMNCs), Dimethylsulfoxide (DMSO) in a concentration comprised between 0.3% and 1.3% (v/v), autologous platelet-enriched plasma (B) or platelet concentrate (B'). A further object of the present invention is the method for producing the autologous preparation (A), obtained by diluting in said autologous platelet-enriched plasma (B) or in platelet concentrate (B'), an autologous concentrate (C), comprising said peripheral blood mononuclear cells and Dimethylsulfoxide in a concentration comprised between 3% and 5%.