Page 1 of 6

In this issue bull PI Highlight Brian C Betts MD

bull Process Development ofLarge-Scale Mesenchymal StemCell Derived ExosomeManufacturing

bull Upcoming Scientific Meetings

bull Upcoming Web Seminars

bull PACT Program Announcement

bull PACT General Facility SOPs

bull PACT Cell Processing Facilities

bull Products amp Services

Contact PACTPlease contact the PACT Coordinating Center if you have any questions regarding the PACT program Visit our website at

wwwpactgroupnet

ContactLani Ibenana and Ashraf El Fiky

AddressThe Emmes Company LLC401 North Washington StreetSuite 700Rockville MD 20850Telephone (301) 251-1161Email pactinfopactgroupnet

Brian C Betts MDAssociate Professor of MedicineDivision of Hematology Oncology and TransplantationUniversity of Minnesota

Q Please describe your clinical and research interestsA I am a physician-scientist with expertise in allogeneic hematopoietic cell transplantation and cell therapy In the clinic I care for patients with blood cancers provide stem cell transplants and manage potentialcomplications like relapse or graft-versus-host disease (GVHD) Theexperiences in the clinic directly guide the work in my lab where we are focused on finding better ways to prevent GVHD Mechanistically GVHD occurs when the donors immune system fails to recognize the patientrsquos body as ldquoselfrdquo and can cause substantial damage to the skin gastrointestinal tract and liver While we have medications to prevent and treat GVHD they carry their own risks and toxicities Primarilystandard GVHD prophylaxis can increase the risk for opportunistic infections and even increase the risk for cancer relapse after trans-plant Moreover the medications are not always successful in prevent-ing GVHD Today clinically relevant acute GVHD effects about 30-40 of patients receiving a stem cell transplant from ahealthy donor When GVHD does not respond to upfront therapy the risk of dying exceeds 90 Thus this represents a critical unmet need in the field

Beyond standard and often broadly suppressive pharmacologic immune suppression my lab has engineered a highly potent celltherapy product to prevent GVHD Specifically we generate regula-tory T cells (Treg) that have the capacity to selectively dampen thedonorrsquos immune response against the patientrsquos tissues In collabora-tion with experts from the University of Minnesota Medical College of Virginia Moffitt Cancer Center and the National Institutes of Health we purify donor T cells and direct them to become Tregs by blocking a proinflammatory cell signal called STAT3 Since these cells do not begin life as a regulatory T cell we refer to them as ldquoinduciblerdquo Tregs These STAT3-inhibited inducible Tregs exhibit significantly greater suppression over dangerous donor T cells yet still maintain beneficial anti-leukemia effects of the transplant Additionally we discovered that blocking STAT3 in these cells unexpectedly offsets their metabolism which could be detrimental over time Importantly we found a creative solution to this problem using a drug you can easily find in a vitamin or supplement store By adding this vitamin to STAT3-inhibited inducible Tregs their metabolic profile is restored and they acquire enhanced suppressive activity We have demonstrated this in rigorous humanized mouse models and we are now focused on bringing these novel cells to the clinic

April 2020 - Volume 15 No2

Page 2 of 6

PI Highlight (continued)

Q How has PACT been a resource for you in pursuing a clinical trial

A PACT is absolutely vital to the success of this innovative therapy While my lab can readily demonstrate the efficacy of STAT3-inhibited inducible Tregs on a small scale true translation of this concept requires a substantial increase in scale For example we typically treat a 30 gram mouse with 100000 Tregs For a typical adult patient we expect we will need 225x109 STAT3-inhibited inducible Tregs This is based on the need to match the ratio of Tregs to circulating alloreactive T cells one to one Additionally these cells need to be generated under specialized and stringent conditions Thankfully PACT and Molecular and Cellular Therapeutics at the University of Minnesota led by David McKenna MD has the exper-tise technology and infrastructure to design and execute large-scale expansion of these specialized Tregs In fact PACT is supporting the pathway for regulatory approval of the STAT3-inhibited inducible Tregs as well as clinical scale-up and translation As a team we look forward to testing these innovative Tregs in a phase I GVHD prevention study

Q How important is it to have PACT support for cellular therapy initiatives

A Though cell therapy is a familiar treatment option in hematologic malignancies due to stem cell transplantation many other medical disciplines are now designing cutting-edge therapies based in unique cell products For example the emer-gence of chimeric antigen receptor (CAR) T cells has had substantial impact in the treatment of refractory lymphoma and leukemia and this is likely only the beginning For instance my lab has collaborated with the Davila Lab at Moffitt Cancer Center to design CAR T cells to prevent and treat GVHD Other effector cells like NK cells and expanded tumor-infiltrating lymphocytes are also actively being investigated at Centers worldwide Additionally Treg therapy is being tested in non-on-cology applications such as autoimmune syndromes Therefore PACT is truly at the leading-edge of medical innovation and their commitment to science is accelerating access to important therapies for patients in need

Process Development of Large-Scale Mesenchymal Stem Cell Derived Exosome Manufacturing Michael Bellio Yee-Shaun Lee Joshua Hare Aisha Khan

Exosomes have recently gained interest as a potentially new therapeutic agent as an alternative or combined therapy to stem cell and other cell-based therapy Exosomes are small secreted vesicles (typically about 30-150 nm) that contain an array of nucleic acid protein or other biomolecules These cargos may be important for cell-cell inter-communication and creating a paracrine effect when using as a therapeutic agent As a mechanism of cell-cell communication exosomes regulate many pathways including but not limited to angiogenesis inflammation and migration1-3 As animal models pro-duce promising results researchers have begun working towards phase I clinical trials and the need to scale up exosome production as critically important4-6 However the main challenges of translating exosome products into the clinics are the large-scale production of cell generated conditioned medium and precipitation of exosomes from this medium

At Interdisciplinary Stem Cell Institute (ISCI) we have completed process development assessments of large-scale exo-some production using the Quantum Bioreactor (Terumo) for scale- bone marrow Mesenchymal Stem cell (MSC) derived exosome products This process development was initiated in response to the growing need of exosome products for Inves-tigational New Drug (IND) enabling studies and rapid clinical translation Our group was able to complete this development work in part through PACT sponsored research with Drs Peiman Hematti and John Kink (University of Wisconsin) in which we aimed to produce large scale exosome batches for rapid and repeated testing in small animal models

To address the large-scale challenges associated with exosome production we utilized Quantum Bioreactors to collect exosome-rich MSC conditioned medium to yield large amounts of exosomes effectively Quantum Bioreactor is an auto-matic close system using a 3-dimensional hollow-fiber core as the culture vessel The core (cell culture vessel) contains approximately 11500 hollow fibers that provides approximately 21 m2 of surface area for cell growth which is roughly equivalent to 120 T175-flasks that would require two regular size incubators A typical T-175 flask is cultured with 25 mL of medium Implementing the Quantum Bioreactor has proven to be a cost effective and efficient tool for achieving high cell yields within minimal technical work hours required If a Quantum Bioreactor is roughly equivalent to 120 T-175 flasks the handling volume of conditioned medium is approximately 3000L as compared to 200-500 mL from one Quantum Bioreactor collecting for one day Quantum Bioreactor is superior in the volume handling of the collected culture medium used for exo-some harvest Large volume handling is time consuming and often limited by the capacity of the equipment that are used for exosome processing

Our exosome production begins with an initial MSC expansion phase in conventional culture flasks for both passage 0 and 1 At P1 (passage 1) harvest the cells were cryopreserved in aliquots of 30x106 cells per 20 mL of cryopreservation reagentin our developmental studies These cryopreserved aliquots are used to build a P1 cell stock that can be directly loadedonto the bioreactor when needed If cryopreservation is not needed freshly harvest P1 cells can be loaded directly into theQuantum Bioreactor

Page 2 of 6

Page 3 of 6

Process Development of Large-Scale Mesenchymal Stem Cell Derived Exosome Manufacturing (continued)

To begin cell expansion for exosome production cryopreserved cells were loaded directly onto the Quantum Bioreactor for the P2 expansion to occur over the next seven days For clinical production 30x106 cells were loaded onto the Quantum Bioreactor(s) directly after harvest This critical change eliminated the need for any required testing to establish a cryopre-served master cell bank (MCB) At the completion of expansion medium used for expansion was washed out and replaced by the serum free medium Exosome collection began after this medium replacement Quantum Bioreactorrsquos automated fea-ture allowed fresh medium to flow into the bioreactor core at previously validated inlet rate where the spent medium would be collected as waste During the exosome collection stage the spent conditioned medium that usually would be designated as ldquowasterdquo would therefore be collected as the exosome-rich conditioned medium used for final product collection At an optimized rate approximately 200-500 mL of the exosome-rich conditioned medium would be collected daily This feature allowed for the repetitive collection of exosome-rich conditioned medium over a determined period of time with a simple step of replacing the medium collection bag For our process developmental work exosome-rich conditioned medium was collected up to 96 hours post medium replacement Each day the collected exosome-rich conditioned medium underwent a sequential step of centrifugation and ultracentrifuge precipitation resulting in final purified exosome product The optimal duration for exosome collection is yet to be determined for bone marrow MSC derived exosome as it is closely associated with its temporal exosome potency profile Exosome collected throughout the entire duration will be pooled together as one final exosome product This critical step is to ensure each batch of product has the maximum yield to increase the cost-effec-tiveness associated with required testing Nanosight NS300 nanoparticle tracking analysis was used to provide quantitative and qualitative data on the exosomes collected daily Our developmental exosomes were about 107 nm (Figure 1A) and the daily yield ranged from 1x1012 to over 4x1012 (Figure 1B) particles The presence of exosomal associated markers CD63 CD81 and CD9 was confirmed using flow cytometry (Figure 1C) Importantly the total particle yield collected during each daily time point showed the large-scale capabilities of this system (Figure 1B)

In anticipation of FDArsquos batch release requirements of allogenic more than minimally manipulated exosome products it is critical to harvest a portion of the cells at P3 and final exosome products to satisfy the release requirements These require-ments include testing for fungalbacterialyeast contamination (sterility and endotoxin testing) mycoplasma potency purity (test for residuals and in-vitro adventitious virus testing) and identity (exosome phenotype particle sizeconcentration protein concentration) of the exosome product Future exosome production utilizing the Quantum Bioreactors for clinical application will allow a single yield of large quantity of exosome deeming its economic value as transitional into future com-mercial product A single high yield of exosome product will not only provide enough product to perform the required testing but also reduce the cost to perform repetitive validation testing satisfying the FDA requirements Furthermore production of small batches will require a large percentage of the lot being used only for safety testing resulting in dramatically increased costs and loss of product

In conclusion we have developed a manufacturing process utilizing the Quantum Bioreactor to produce large-scale MSC-de-rived exosomes for use in both animal studies and clinical trials (Figure 2) This methodology has considerable scale up potential compared to standard cell culture flask cultures that require manual tasks Furthermore this system allowed us to move forward with the completion of IND enabling studies and collect relevant data that can satisfy FDA requirements

Figure 1 (A B amp C) Process Validation Data of Large-Scale Exosome Production (A) Nanosight nanoparticle tracking analysis of particle size and concentration distribution in a representative sample (B) The total particle harvest from daily conditioned medium collections and precipitations (C) Flow cytometry analysis of exosome associated markers from a representative sample

Page 3 of 6

Page 4 of 6

Process Development of Large-Scale Mesenchymal Stem Cell Derived Exosome Manufacturing (continued)

Figure 2 Schematic Representation of Manual Cell Culture Flask vs Quantum Bioreactor Methodologies

Acknowledgment We would like to thank Dr Peiman Hematti and Dr John Kink from University of Wisconsin for their support We would also like to thank PACT for proving the funding for this project

Page 5 of 6

Process Development of Large-Scale Mesenchymal Stem Cell Derived Exosome Manufacturing (continued)

References

1 Kishore R and Khan M More Than Tiny Sacks Stem Cell Exosomes as Cell-Free Modality for Cardiac Repair Circ Res 2016118330-43

2 Mathivanan S Ji H and Simpson RJ Exosomes extracellular organelles important in intercellular communication J Proteomics 2010731907-20

3 M HR Bayraktar E G KH Abd-Ellah MF Amero P Chavez-Reyes A and Rodriguez-Aguayo C Exosomes From Garbage Bins to Promising Therapeutic Targets Int J Mol Sci 201718

4 Gallet R Dawkins J Valle J Simsolo E de Couto G Middleton R Tseliou E Luthringer D Kreke M Smith RR Marban L Ghaleh B and Marban E Exosomes secreted by cardiosphere-derived cells reduce scarring attenuate adverse remodelling and improve function in acute and chronic porcine myocardial infarction Eur Heart J 201738201-211

5 Vandergriff AC de Andrade JB Tang J Hensley MT Piedrahita JA Caranasos TG and Cheng K Intravenous Cardiac Stem Cell-Derived Exosomes Ameliorate Cardiac Dysfunction in Doxorubicin Induced Dilated Cardiomyopathy Stem Cells Int 20152015960926

6 Shiue SJ Rau RH Shiue HS Hung YW Li ZX Yang KD and Cheng JK Mesenchymal stem cell exosomes as a cell-free therapy for nerve injury-induced pain in rats Pain 2019160210-223

Upcoming Scientific Meetings

PACT Coordinating Center representatives were planning on presenting at the ASNTR Cell therapy training workshop (23 April 2020) and the Wake Forest Regenerative Medicine Essentials Course (08-11 June 2020) and having materials available at a booth in the 2020 ASGCT Annual Meeting (12-15 May 2020) However due to COVID-19 concerns these meetings have been cancelled

PACT is still planning to have a booth at the 2020 AABB Annual Meeting in Baltimore MD in October 2020 We hope to see you there

Stay healthy

Upcoming Web Seminars

PACT has two upcoming web seminars in development Announcements will be posted on the PACT website and via email to our distribution list Sign up for email updates by emailing pactupdatesemmescom Stay tuned

Page 5 of 6

Page 6 of 6

PACT Program AnnouncementPACT is still accepting applications however the current NHLBI PACT program is funded through June 30 2021 Therefore the PACT program will only accept applica-tions for services that can be completed by this date If you are interested in applying for PACT services please review the materials available on the PACT website wwwpactgroupnet under the Apply to PACT tab

Apply soon

PACT can provide general facility SOPs upon request to assist you in your own cell processing facility SOPs

PLEASE NOTE that these SOPs are for INFORMATIONAL PURPOSES ONLY and therefore require validation by your own facility To see a full list of SOPs available for request go to the PACT website and look under the Resource Center tab

SOP Categories available for requestbull Cleaning Proceduresbull Deviation Managementbull Environmental Monitoringbull Personnel Trainingbull Quality Assurance Quality Controlbull Quality Managementbull Development and Managementbull Validation Process

Below are just a few of the various products and services that PACT offersProgenitor CellsC-kit+ cells Endothelial cells HSCs MSCs iPSCsLymphocytesVirus-specific T-cells Tumor-infiltrating lymphocytes (TILs)EBV-transformed B cell linesGenetically Modified CellsCAR-T-cells Cytotoxic T lymphocytes fibroblastsServicesCell culture isolation expansion and cryopreservation cell depletion cell enrichment cell manufacturing for animal model studies

PACT Cell Processing Facilities

University of Minnesota Interdisciplinary Stem Cell InstituteMolecular and Cellular Cellular Manufacturing ProgramTherapeutics Facility University of Miami

PI David McKenna MD Miller School of MedicineContract Number PI Joshua Hare MD

HHSN268201600014I Contract NumberHHSN268201600012I

Center for Cell and Gene Therapy City of Hope Center for Baylor College of Medicine Biomedicine and GeneticsPI Adrian Gee MI Biol PhD PI Joseph Gold PhD

Contract Number Contract NumberHHSN268201600015I HHSN268201600011I

Moffitt Cancer Center PACT Coordinating CenterPI Linda Kelley PhD The Emmes Company LLC

Contract Number Contract NumberHHSN268201600013I HHSN268201600020C

This project has been funded in whole or in part with Federal funds from the National Heart Lung and Blood Institute National Institutes of Health Department of Health and Human Services

Page 3 of 6

Process Development of Large-Scale Mesenchymal Stem Cell Derived Exosome Manufacturing (continued)

To begin cell expansion for exosome production cryopreserved cells were loaded directly onto the Quantum Bioreactor for the P2 expansion to occur over the next seven days For clinical production 30x106 cells were loaded onto the Quantum Bioreactor(s) directly after harvest This critical change eliminated the need for any required testing to establish a cryopre-served master cell bank (MCB) At the completion of expansion medium used for expansion was washed out and replaced by the serum free medium Exosome collection began after this medium replacement Quantum Bioreactorrsquos automated fea-ture allowed fresh medium to flow into the bioreactor core at previously validated inlet rate where the spent medium would be collected as waste During the exosome collection stage the spent conditioned medium that usually would be designated as ldquowasterdquo would therefore be collected as the exosome-rich conditioned medium used for final product collection At an optimized rate approximately 200-500 mL of the exosome-rich conditioned medium would be collected daily This feature allowed for the repetitive collection of exosome-rich conditioned medium over a determined period of time with a simple step of replacing the medium collection bag For our process developmental work exosome-rich conditioned medium was collected up to 96 hours post medium replacement Each day the collected exosome-rich conditioned medium underwent a sequential step of centrifugation and ultracentrifuge precipitation resulting in final purified exosome product The optimal duration for exosome collection is yet to be determined for bone marrow MSC derived exosome as it is closely associated with its temporal exosome potency profile Exosome collected throughout the entire duration will be pooled together as one final exosome product This critical step is to ensure each batch of product has the maximum yield to increase the cost-effec-tiveness associated with required testing Nanosight NS300 nanoparticle tracking analysis was used to provide quantitative and qualitative data on the exosomes collected daily Our developmental exosomes were about 107 nm (Figure 1A) and the daily yield ranged from 1x1012 to over 4x1012 (Figure 1B) particles The presence of exosomal associated markers CD63 CD81 and CD9 was confirmed using flow cytometry (Figure 1C) Importantly the total particle yield collected during each daily time point showed the large-scale capabilities of this system (Figure 1B)

In anticipation of FDArsquos batch release requirements of allogenic more than minimally manipulated exosome products it is critical to harvest a portion of the cells at P3 and final exosome products to satisfy the release requirements These require-ments include testing for fungalbacterialyeast contamination (sterility and endotoxin testing) mycoplasma potency purity (test for residuals and in-vitro adventitious virus testing) and identity (exosome phenotype particle sizeconcentration protein concentration) of the exosome product Future exosome production utilizing the Quantum Bioreactors for clinical application will allow a single yield of large quantity of exosome deeming its economic value as transitional into future com-mercial product A single high yield of exosome product will not only provide enough product to perform the required testing but also reduce the cost to perform repetitive validation testing satisfying the FDA requirements Furthermore production of small batches will require a large percentage of the lot being used only for safety testing resulting in dramatically increased costs and loss of product

In conclusion we have developed a manufacturing process utilizing the Quantum Bioreactor to produce large-scale MSC-de-rived exosomes for use in both animal studies and clinical trials (Figure 2) This methodology has considerable scale up potential compared to standard cell culture flask cultures that require manual tasks Furthermore this system allowed us to move forward with the completion of IND enabling studies and collect relevant data that can satisfy FDA requirements

Figure 1 (A B amp C) Process Validation Data of Large-Scale Exosome Production (A) Nanosight nanoparticle tracking analysis of particle size and concentration distribution in a representative sample (B) The total particle harvest from daily conditioned medium collections and precipitations (C) Flow cytometry analysis of exosome associated markers from a representative sample

Page 3 of 6

Page 4 of 6

Process Development of Large-Scale Mesenchymal Stem Cell Derived Exosome Manufacturing (continued)

Figure 2 Schematic Representation of Manual Cell Culture Flask vs Quantum Bioreactor Methodologies

Acknowledgment We would like to thank Dr Peiman Hematti and Dr John Kink from University of Wisconsin for their support We would also like to thank PACT for proving the funding for this project

Page 5 of 6

Process Development of Large-Scale Mesenchymal Stem Cell Derived Exosome Manufacturing (continued)

References

1 Kishore R and Khan M More Than Tiny Sacks Stem Cell Exosomes as Cell-Free Modality for Cardiac Repair Circ Res 2016118330-43

2 Mathivanan S Ji H and Simpson RJ Exosomes extracellular organelles important in intercellular communication J Proteomics 2010731907-20

3 M HR Bayraktar E G KH Abd-Ellah MF Amero P Chavez-Reyes A and Rodriguez-Aguayo C Exosomes From Garbage Bins to Promising Therapeutic Targets Int J Mol Sci 201718

4 Gallet R Dawkins J Valle J Simsolo E de Couto G Middleton R Tseliou E Luthringer D Kreke M Smith RR Marban L Ghaleh B and Marban E Exosomes secreted by cardiosphere-derived cells reduce scarring attenuate adverse remodelling and improve function in acute and chronic porcine myocardial infarction Eur Heart J 201738201-211

5 Vandergriff AC de Andrade JB Tang J Hensley MT Piedrahita JA Caranasos TG and Cheng K Intravenous Cardiac Stem Cell-Derived Exosomes Ameliorate Cardiac Dysfunction in Doxorubicin Induced Dilated Cardiomyopathy Stem Cells Int 20152015960926

6 Shiue SJ Rau RH Shiue HS Hung YW Li ZX Yang KD and Cheng JK Mesenchymal stem cell exosomes as a cell-free therapy for nerve injury-induced pain in rats Pain 2019160210-223

Upcoming Scientific Meetings

PACT Coordinating Center representatives were planning on presenting at the ASNTR Cell therapy training workshop (23 April 2020) and the Wake Forest Regenerative Medicine Essentials Course (08-11 June 2020) and having materials available at a booth in the 2020 ASGCT Annual Meeting (12-15 May 2020) However due to COVID-19 concerns these meetings have been cancelled

PACT is still planning to have a booth at the 2020 AABB Annual Meeting in Baltimore MD in October 2020 We hope to see you there

Stay healthy

Upcoming Web Seminars

PACT has two upcoming web seminars in development Announcements will be posted on the PACT website and via email to our distribution list Sign up for email updates by emailing pactupdatesemmescom Stay tuned

Page 5 of 6

Page 6 of 6

PACT Program AnnouncementPACT is still accepting applications however the current NHLBI PACT program is funded through June 30 2021 Therefore the PACT program will only accept applica-tions for services that can be completed by this date If you are interested in applying for PACT services please review the materials available on the PACT website wwwpactgroupnet under the Apply to PACT tab

Apply soon

PACT can provide general facility SOPs upon request to assist you in your own cell processing facility SOPs

PLEASE NOTE that these SOPs are for INFORMATIONAL PURPOSES ONLY and therefore require validation by your own facility To see a full list of SOPs available for request go to the PACT website and look under the Resource Center tab

SOP Categories available for requestbull Cleaning Proceduresbull Deviation Managementbull Environmental Monitoringbull Personnel Trainingbull Quality Assurance Quality Controlbull Quality Managementbull Development and Managementbull Validation Process

Below are just a few of the various products and services that PACT offersProgenitor CellsC-kit+ cells Endothelial cells HSCs MSCs iPSCsLymphocytesVirus-specific T-cells Tumor-infiltrating lymphocytes (TILs)EBV-transformed B cell linesGenetically Modified CellsCAR-T-cells Cytotoxic T lymphocytes fibroblastsServicesCell culture isolation expansion and cryopreservation cell depletion cell enrichment cell manufacturing for animal model studies

PACT Cell Processing Facilities

University of Minnesota Interdisciplinary Stem Cell InstituteMolecular and Cellular Cellular Manufacturing ProgramTherapeutics Facility University of Miami

PI David McKenna MD Miller School of MedicineContract Number PI Joshua Hare MD

HHSN268201600014I Contract NumberHHSN268201600012I

Center for Cell and Gene Therapy City of Hope Center for Baylor College of Medicine Biomedicine and GeneticsPI Adrian Gee MI Biol PhD PI Joseph Gold PhD

Contract Number Contract NumberHHSN268201600015I HHSN268201600011I

Moffitt Cancer Center PACT Coordinating CenterPI Linda Kelley PhD The Emmes Company LLC

Contract Number Contract NumberHHSN268201600013I HHSN268201600020C

This project has been funded in whole or in part with Federal funds from the National Heart Lung and Blood Institute National Institutes of Health Department of Health and Human Services

Page 4 of 6

Process Development of Large-Scale Mesenchymal Stem Cell Derived Exosome Manufacturing (continued)

Figure 2 Schematic Representation of Manual Cell Culture Flask vs Quantum Bioreactor Methodologies

Acknowledgment We would like to thank Dr Peiman Hematti and Dr John Kink from University of Wisconsin for their support We would also like to thank PACT for proving the funding for this project

Page 5 of 6

Process Development of Large-Scale Mesenchymal Stem Cell Derived Exosome Manufacturing (continued)

References

1 Kishore R and Khan M More Than Tiny Sacks Stem Cell Exosomes as Cell-Free Modality for Cardiac Repair Circ Res 2016118330-43

2 Mathivanan S Ji H and Simpson RJ Exosomes extracellular organelles important in intercellular communication J Proteomics 2010731907-20

3 M HR Bayraktar E G KH Abd-Ellah MF Amero P Chavez-Reyes A and Rodriguez-Aguayo C Exosomes From Garbage Bins to Promising Therapeutic Targets Int J Mol Sci 201718

4 Gallet R Dawkins J Valle J Simsolo E de Couto G Middleton R Tseliou E Luthringer D Kreke M Smith RR Marban L Ghaleh B and Marban E Exosomes secreted by cardiosphere-derived cells reduce scarring attenuate adverse remodelling and improve function in acute and chronic porcine myocardial infarction Eur Heart J 201738201-211

5 Vandergriff AC de Andrade JB Tang J Hensley MT Piedrahita JA Caranasos TG and Cheng K Intravenous Cardiac Stem Cell-Derived Exosomes Ameliorate Cardiac Dysfunction in Doxorubicin Induced Dilated Cardiomyopathy Stem Cells Int 20152015960926

6 Shiue SJ Rau RH Shiue HS Hung YW Li ZX Yang KD and Cheng JK Mesenchymal stem cell exosomes as a cell-free therapy for nerve injury-induced pain in rats Pain 2019160210-223

Upcoming Scientific Meetings

PACT Coordinating Center representatives were planning on presenting at the ASNTR Cell therapy training workshop (23 April 2020) and the Wake Forest Regenerative Medicine Essentials Course (08-11 June 2020) and having materials available at a booth in the 2020 ASGCT Annual Meeting (12-15 May 2020) However due to COVID-19 concerns these meetings have been cancelled

PACT is still planning to have a booth at the 2020 AABB Annual Meeting in Baltimore MD in October 2020 We hope to see you there

Stay healthy

Upcoming Web Seminars

PACT has two upcoming web seminars in development Announcements will be posted on the PACT website and via email to our distribution list Sign up for email updates by emailing pactupdatesemmescom Stay tuned

Page 5 of 6

Page 6 of 6

PACT Program AnnouncementPACT is still accepting applications however the current NHLBI PACT program is funded through June 30 2021 Therefore the PACT program will only accept applica-tions for services that can be completed by this date If you are interested in applying for PACT services please review the materials available on the PACT website wwwpactgroupnet under the Apply to PACT tab

Apply soon

PACT can provide general facility SOPs upon request to assist you in your own cell processing facility SOPs

PLEASE NOTE that these SOPs are for INFORMATIONAL PURPOSES ONLY and therefore require validation by your own facility To see a full list of SOPs available for request go to the PACT website and look under the Resource Center tab

SOP Categories available for requestbull Cleaning Proceduresbull Deviation Managementbull Environmental Monitoringbull Personnel Trainingbull Quality Assurance Quality Controlbull Quality Managementbull Development and Managementbull Validation Process

Below are just a few of the various products and services that PACT offersProgenitor CellsC-kit+ cells Endothelial cells HSCs MSCs iPSCsLymphocytesVirus-specific T-cells Tumor-infiltrating lymphocytes (TILs)EBV-transformed B cell linesGenetically Modified CellsCAR-T-cells Cytotoxic T lymphocytes fibroblastsServicesCell culture isolation expansion and cryopreservation cell depletion cell enrichment cell manufacturing for animal model studies

PACT Cell Processing Facilities

University of Minnesota Interdisciplinary Stem Cell InstituteMolecular and Cellular Cellular Manufacturing ProgramTherapeutics Facility University of Miami

PI David McKenna MD Miller School of MedicineContract Number PI Joshua Hare MD

HHSN268201600014I Contract NumberHHSN268201600012I

Center for Cell and Gene Therapy City of Hope Center for Baylor College of Medicine Biomedicine and GeneticsPI Adrian Gee MI Biol PhD PI Joseph Gold PhD

Contract Number Contract NumberHHSN268201600015I HHSN268201600011I

Moffitt Cancer Center PACT Coordinating CenterPI Linda Kelley PhD The Emmes Company LLC

Contract Number Contract NumberHHSN268201600013I HHSN268201600020C

This project has been funded in whole or in part with Federal funds from the National Heart Lung and Blood Institute National Institutes of Health Department of Health and Human Services

Page 5 of 6

Process Development of Large-Scale Mesenchymal Stem Cell Derived Exosome Manufacturing (continued)

References

1 Kishore R and Khan M More Than Tiny Sacks Stem Cell Exosomes as Cell-Free Modality for Cardiac Repair Circ Res 2016118330-43

2 Mathivanan S Ji H and Simpson RJ Exosomes extracellular organelles important in intercellular communication J Proteomics 2010731907-20

3 M HR Bayraktar E G KH Abd-Ellah MF Amero P Chavez-Reyes A and Rodriguez-Aguayo C Exosomes From Garbage Bins to Promising Therapeutic Targets Int J Mol Sci 201718

4 Gallet R Dawkins J Valle J Simsolo E de Couto G Middleton R Tseliou E Luthringer D Kreke M Smith RR Marban L Ghaleh B and Marban E Exosomes secreted by cardiosphere-derived cells reduce scarring attenuate adverse remodelling and improve function in acute and chronic porcine myocardial infarction Eur Heart J 201738201-211

5 Vandergriff AC de Andrade JB Tang J Hensley MT Piedrahita JA Caranasos TG and Cheng K Intravenous Cardiac Stem Cell-Derived Exosomes Ameliorate Cardiac Dysfunction in Doxorubicin Induced Dilated Cardiomyopathy Stem Cells Int 20152015960926

6 Shiue SJ Rau RH Shiue HS Hung YW Li ZX Yang KD and Cheng JK Mesenchymal stem cell exosomes as a cell-free therapy for nerve injury-induced pain in rats Pain 2019160210-223

Upcoming Scientific Meetings

PACT Coordinating Center representatives were planning on presenting at the ASNTR Cell therapy training workshop (23 April 2020) and the Wake Forest Regenerative Medicine Essentials Course (08-11 June 2020) and having materials available at a booth in the 2020 ASGCT Annual Meeting (12-15 May 2020) However due to COVID-19 concerns these meetings have been cancelled

PACT is still planning to have a booth at the 2020 AABB Annual Meeting in Baltimore MD in October 2020 We hope to see you there

Stay healthy

Upcoming Web Seminars

PACT has two upcoming web seminars in development Announcements will be posted on the PACT website and via email to our distribution list Sign up for email updates by emailing pactupdatesemmescom Stay tuned

Page 5 of 6

Page 6 of 6

PACT Program AnnouncementPACT is still accepting applications however the current NHLBI PACT program is funded through June 30 2021 Therefore the PACT program will only accept applica-tions for services that can be completed by this date If you are interested in applying for PACT services please review the materials available on the PACT website wwwpactgroupnet under the Apply to PACT tab

Apply soon

PACT can provide general facility SOPs upon request to assist you in your own cell processing facility SOPs

PLEASE NOTE that these SOPs are for INFORMATIONAL PURPOSES ONLY and therefore require validation by your own facility To see a full list of SOPs available for request go to the PACT website and look under the Resource Center tab

SOP Categories available for requestbull Cleaning Proceduresbull Deviation Managementbull Environmental Monitoringbull Personnel Trainingbull Quality Assurance Quality Controlbull Quality Managementbull Development and Managementbull Validation Process

Below are just a few of the various products and services that PACT offersProgenitor CellsC-kit+ cells Endothelial cells HSCs MSCs iPSCsLymphocytesVirus-specific T-cells Tumor-infiltrating lymphocytes (TILs)EBV-transformed B cell linesGenetically Modified CellsCAR-T-cells Cytotoxic T lymphocytes fibroblastsServicesCell culture isolation expansion and cryopreservation cell depletion cell enrichment cell manufacturing for animal model studies

PACT Cell Processing Facilities

University of Minnesota Interdisciplinary Stem Cell InstituteMolecular and Cellular Cellular Manufacturing ProgramTherapeutics Facility University of Miami

PI David McKenna MD Miller School of MedicineContract Number PI Joshua Hare MD

HHSN268201600014I Contract NumberHHSN268201600012I

Center for Cell and Gene Therapy City of Hope Center for Baylor College of Medicine Biomedicine and GeneticsPI Adrian Gee MI Biol PhD PI Joseph Gold PhD

Contract Number Contract NumberHHSN268201600015I HHSN268201600011I

Moffitt Cancer Center PACT Coordinating CenterPI Linda Kelley PhD The Emmes Company LLC

Contract Number Contract NumberHHSN268201600013I HHSN268201600020C

This project has been funded in whole or in part with Federal funds from the National Heart Lung and Blood Institute National Institutes of Health Department of Health and Human Services

Page 6 of 6

PACT Program AnnouncementPACT is still accepting applications however the current NHLBI PACT program is funded through June 30 2021 Therefore the PACT program will only accept applica-tions for services that can be completed by this date If you are interested in applying for PACT services please review the materials available on the PACT website wwwpactgroupnet under the Apply to PACT tab

Apply soon

PACT can provide general facility SOPs upon request to assist you in your own cell processing facility SOPs

PLEASE NOTE that these SOPs are for INFORMATIONAL PURPOSES ONLY and therefore require validation by your own facility To see a full list of SOPs available for request go to the PACT website and look under the Resource Center tab

SOP Categories available for requestbull Cleaning Proceduresbull Deviation Managementbull Environmental Monitoringbull Personnel Trainingbull Quality Assurance Quality Controlbull Quality Managementbull Development and Managementbull Validation Process

Below are just a few of the various products and services that PACT offersProgenitor CellsC-kit+ cells Endothelial cells HSCs MSCs iPSCsLymphocytesVirus-specific T-cells Tumor-infiltrating lymphocytes (TILs)EBV-transformed B cell linesGenetically Modified CellsCAR-T-cells Cytotoxic T lymphocytes fibroblastsServicesCell culture isolation expansion and cryopreservation cell depletion cell enrichment cell manufacturing for animal model studies

PACT Cell Processing Facilities

University of Minnesota Interdisciplinary Stem Cell InstituteMolecular and Cellular Cellular Manufacturing ProgramTherapeutics Facility University of Miami

PI David McKenna MD Miller School of MedicineContract Number PI Joshua Hare MD

HHSN268201600014I Contract NumberHHSN268201600012I

Center for Cell and Gene Therapy City of Hope Center for Baylor College of Medicine Biomedicine and GeneticsPI Adrian Gee MI Biol PhD PI Joseph Gold PhD

Contract Number Contract NumberHHSN268201600015I HHSN268201600011I

Moffitt Cancer Center PACT Coordinating CenterPI Linda Kelley PhD The Emmes Company LLC

Contract Number Contract NumberHHSN268201600013I HHSN268201600020C

This project has been funded in whole or in part with Federal funds from the National Heart Lung and Blood Institute National Institutes of Health Department of Health and Human Services