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Home > Cellular Therapy > Sourcing Blood Cells Abstract

Sourcing Blood Cells Abstract

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Human-Derived Raw Materials: Controlled, Consistent Collections Enable Successful Manufacturing of Cell-Based Regenerative Medicine Products

 

 

 

Thomas Ramos, MS, MBA1, Wenshi Wang, PhD1, Jacob Okhovat1, Scott Burger, MD2

 

1HemaCare Corporation 15350 Sherman Way, Suite 350, Van Nuys, CA 91406

(877) 397-3087 • tramos@hemacare.com

 

2Advanced Cell & Gene Therapy, LLC, 105 Highgrove Drive, Chapel Hill, NC 27516

(919) 969-1103 | celltherapy@ac-gt.com

 

 

Abstract

Human cells are critical raw materials for manufacturing cell therapy products, but often introduce significant variability. Rigorous operational controls and quality systems, however, enable optimal collection of high-quality, consistent cellular material. HemaCare, a long-standing supplier of human-derived blood components, controls apheresis procedures and collection sites under a formal quality system, with GMP-compliant, validated procedures and equipment, and GTP-compliant donor screening and tracking. HemaCare performed 86,799 cellular apheresis collections in the last seven years (year ending July 31, 2013), including patient and normal-donor peripheral blood mononuclear cells (MNCs), and plateletpheresis products for research, clinical trials, and commercial products. HemaCare’s unmobilized apheresis products showed consistently high MNC purity, with 93.8% of products containing ≥75% MNC, and an average of 85.66% MNC ± 7.1% (mean ± 1 SD). Red blood cell contamination was low, with hematocrit averaging 1.78% ± 0.7%. Approximately 85% of HemaCare donors have donated apheresis products 5 or more times; this repeat-donor pool also contributes to product consistency. HemaCare’s laboratory is equipped with Miltenyi Biotec technology for isolation of cellular raw material into purified cellular subpopulations. The consistency and viability of the purified end products are measured with flow cytometry. Using BioLife Solutions’ serum-free and protein-free fully-defined cGMP CryoStor™ cryopreservation media with purified cells, post-thaw recovery rates of cell fractions have been above 95%, based on 7AAD staining. Dendritic cell and macrophage have demonstrated post-thaw recovery rates of ≥90%. CryoStor™ cryopreservation medium, in combination with freezing in the BioCision CoolCell™ freezing container, has enabled HemaCare to standardize the cryopreservation process, reducing variability while optimizing post-thaw viable cell recovery of its research products.

 

 

Cell subpopulations - CD45

About HemaCare

HemaCare is a leading provider of apheresis products, human blood cells, apheresis collection services, and therapeutic apheresis services

  • Apheresis collections and blood-derived products for preclinical research, clinical studies from Phase I to Phase IV, and commercial applications
  • Supports applications in immunotherapy, cell therapy, assay development, and medical devices
  • Apheresis PBMC
  • G-CSF-mobilized PBSC
  • Bone marrow
  • Cord blood
  • Peripheral blood
  • Plasma, serum
  • Cell subpopulations
    • CD34+
    • CD3+, CD4+, CD8+
    • CD19+, CD56+, others
  • Healthy-donor and disease-state products
  • Fresh and cryopreserved products

 

The HemaCare Advantage

HemaCare is committed to providing our customers with experienced, personalized, responsive, cost effective, and value added services.

 

Research Products and Cellular Therapy Services

  • Donor pool is already pedigreed and will continue to be expanded
  • Extensive donor registry with ability to request repeat donor collections
  • Predictable, reliable, and validated collection procedures
  • Optimized Standard Operating Procedures leading to high degree of standardization and control
  • Ability to collect based on specific, customizable protocols
  • High-yield, consistent cell collections
  • Validated, automated cell counts and five-part WBC differentials
  • Established distribution redundancies leading to the ability to ship via FedEx, UPS, World Courier, and various local couriers
  • Access to our scientific/technical support 24/7/365

Human Cells: Standardizing Living Biological Raw Material Through Quality Processes

 

Human Cells
  • The quality and consistency of Human blood-derived cells are critical to enable research for cell therapy, tissue-engineered products, and ex vivo gene therapy products.
  • Controlling cell collections minimizes variability and provides an optimal product for use in research and/or manufacturing.
  • Training is rigorous and all employees are signed off on critical SOPs for collection.
  • Experienced staff lay the foundation of the collections and are critical for success.
  • Quality Systems standardize and guide operations aimed to yield optimal, consistent products.

 

Apheresis Expertise in a
Quality Systems Framework

 

Optimize cell collection by controlling and qualifying the apheresis processes based on cGMP and cGTP requirements

  • Qualification of donors
  • Qualification of vendors, equipment, supplies and sites
  • Monitoring equipment, supplies and environment
  • Development and maintenance of procedures
  • Comprehensive staff training and qualification
  • Documentation and records management
  • Management of exceptions and deviations
  • Monitoring quality indicators for process improvement
  • Internal and external inspections

 

Donors – The Critical Source

  • All donors are qualified per regulations and protocol requirements, with IRB-approved informed consent
  • Pedigreed, well-characterized apheresis donor population
    • 85% of HemaCare donors have donated ≥ 5 times/year
    • Facilitates recruitment of donors with specific characteristics required by investigator
      • Medical history, HLA type, other laboratory test results, age, gender, ethnicity…
    • Repeat donors further minimize variability
The Critical Source

% MNC of a donor sample pool

  • Mean 3.5% CV for products per donor
  • Mean 7.7% CV for all products
  • n=21 donors, 3-5 products/donor

 

Equipment Management

Equipment Management

  • Equipment management processes ensure apheresis equipment operates efficiently and within process specifications throughout equipment lifecycle
    • Equipment selection
    • Equipment validation – IQ, OQ, PQ
    • Preventive maintenance
    • Corrective maintenance
    • Equipment QC and performance tracking
  • Apheresis platforms
    • TRIMA Accel® Automated Collection System – Platelets
    • COBE® Spectra Apheresis System - PBMCs, Stem Cells, TA

 

Controlled Procedures and Documentation

  • All activities governed by written procedures and policies
    • Developed using a standardized process
    • Reviewed and approved by subject matter experts and QA
    • Validated prior to release
    • Incorporate regulations, manufacturer’s requirements, research protocols and IRB requirements
  • Controlled: Computerized document management system
  • Proposed variances approved through Exception Management
  • Documentation and record retention requirements met
  • Traceability of each product collected

Controlled Procedures and Documentation

Staff Qualification and Training

 

  • Recruitment, training, and proficiency are critical to a consistent product output
  • Job descriptions outline licensure, qualifications, educational requirements, work experience
  • Training program covers standardized procedures and equipment, and “soft skills”

Staff Qualification and Training

  • Training includes 6 months with preceptor and multiple competency assessments prior to release
  • Quality indicators monitor staff performance

 

Deviation Management

 

  • Deviation: A variation from SOPs, cGMP, cGTP standards or specifications; may affect product quality or donor safety
  • Capturing deviations
    • All staff report deviations when noted
    • Immediate actions are performed
  • Corrective and Preventive Action (CAPA)
    • Root cause analysis
    • Develop and carry out corrective/preventive actions
  • Track/trend deviations for process improvement
  • Managing and preventing deviations maintains product consistency and reduces final product variability

 

Quality Indicators

Internal Quality Indicators
  • Tracking and trending
    • Donor reactions, deviations, exceptions, suppliers, risks, equipment performance, etc…
  • Product QC analysis
    • Automated cell counter/analyzer, 5-part WBC differential (Horiba Pentra analyzer)
      • Nucleated cell (WBC) content and subpopulations, % MNC, HCT, product volume, etc…
    • Immunophenotyping (MACS Quant flow cytometer)
    • Functional Assays Internal Quality Indicators
  • Donor testing
    • Screening, infectious disease testing, CBC with 5-part WBC differential
  • Internal and external audits

 

Selected Apheresis Product Quality Indicators

 

Mononuclear Cells

 

 

MNC Purity

85.66% ± 7.1% (mean ± 1 SD)
93.8% of products ≥ 75% MNC

 

MNC Content

11.5 ± 4.0 x 109 MNC (mean ± 1 SD)

 

Volume

360.62 ± 82.71 ml (mean ± 1 SD)

 

RBC Contamination

Hematocrit 1.78% ± 0.7% (mean ± 1 SD) 91.3% of products ≤ 2.5% hematocrit

 

Internal Quality Indicators

Internal Quality Indicators

  • Deviation management summary reports
  • Equipment performance monitoring and tracking
  • Staff competency assessment
  • Product characteristics, statistical evaluation
    • Nucleated cell (WBC) content and subpopulations
    • Mononuclear cell % and total content
    • Immunophenotype, if needed
    • Volume, hematocrit, other

 

External Quality Indicators

External Quality Indicators

External Quality Indicators

External Quality Indicators

 

High level of satisfaction led to request for expanded services at additional sites

Best In Class Partners

 

A major strategy to maintain HemaCare’s quality of cells has been to partner with companies which have high quality standards of their own.

  • Cold (4oC) shipment in HypoThermosol® (BioLife Solutions) has been shown to increase stability of a variety of cell types, and could extend shelf-life of apheresis PBMCs and other cell types.
  • Cell processing procedures are performed with isolation technology from Miltenyi Biotec. Large scale isolations are done on the Prodigy® and CliniMACS® for an output of billions of cells. Smaller focused isolations are done on the AutoMACS® and manual columns. Both systems were optimized to achieve over 90% purities and over 90% viabilities.
  • Cryopreservation was implemented with the use of the CoolCell® manual freezing container (BioCision). The containers contain no liquid component and ensure a consistent cooling rate of 1°C/min each use. This in combination with CryoStor™ cryopreservation medium (BioLife Solutions) allows for highly efficient procedures during cryopreservation.

 

Non-Cryopreserved PBMC Storage
Plot Data

Non-Cryopreserved PBMC Storag

Post Thaw QC Data

 

Upon thawing cryopreserved cells, HemaCare’s recovery has been consistently stable with respect to purity and viability.

Post Thaw QC Data

Functional Assay

 

Neutrophils

Functional Assay

Summary

  • Collecting blood-based cellular products in a manner that minimizes variability brings a higher degree of reproducibility to the research project or manufacturing effort
  • Quality-based controls such as standardized SOPs, staff training and competency assessments, equipment management, and monitoring of quality indicators reduce this variability
  • Availability of repeat donors from a pedigreed donor base enhances the quality and value of this critical, living biological material
  • Relationships with other best in class organizations allows for consistent product development and collaborations focused in improving the cell manufacturing process.
  • Use of cGMP, serum-free, protein-free biopreservation media such as HypoThermosol and CryoStor shows great promise to enable worldwide shipment of fresh or frozen cellular products isolated from apheresis collection, extending shelf-life of cell therapy products.
  • Development of functional assays are established to monitor the health and functionality of a variety of cells after processing. These assays are applied to both fresh and cryopreserved cells.