2021 Gene and Cell Therapy Calendar

Vigene Biosciences, an award-winning world leader in plasmid and viral vector development and manufacturing, wishes you a healthy and prosperous 2021.

This 2021 calendar contains figures and tables that have been carefully selected by Vigene from gene and cell therapy articles, all reprinted with permission from Springer Nature.

We hope that it will be a useful reference for researchers in the gene and cell therapy field. Vigene Bioscience has been ranked by Inc. Magazine as one of the fastest growing companies in the USA for the past three years. Vigene offers integrated plasmid and viral vector production and analytical services from its 71,000 sq ft state-of-the-art facility which includes 10 Good Manufacturing Practice (GMP) clean-room suites. Vigene’s mission is to make gene therapy affordable. On the basic research side, Vigene is developing, manufacturing, and distributing adeno-associated virus (AAV), lentivirus, retrovirus, adenovirus, and plasmid-based reagents including Howard Hughes Medical Institute (HHMI)/Janelia Research Campus AAV Biosensors. On the cGMP clinical production side, Vigene combines proven production technologies with rigorous, regulatory compliant cGMP production processes to meet the needs and expectations of clinical and commercial clients.

Vigene offers FDA and EMA compliant cGMP production for AAV, lentivirus, adenovirus, retrovirus, and plasmids to global pharmaceutical and biotech companies, government agencies, and non-profit organizations.

Viral Vector Genome Size Infec on Expression Pote a ons

Retrovirus 7-11 kb (ssRNA) Dividing cells Stable onal mutagenesis poten al

Len virus 9 kb (ssRNA) Dividing & non-dividing cells Stable onal mutagenesis

poten al

Adenovirus 36 kb (dsDNA) Dividing & non-dividing cells Transient Immune response

Adeno-associated Virus, AAV

4.7 kb (ssDNA) Dividing & non-dividing cells long las ng Immune response

Herpes Simplex Virus, HSV 150 kb (dsDNA) Dividing & non-

dividing cells Transient No gene expression during latent infec on

Vaccinia virus 190 kb (dsDNA) Dividing cells Transient Pote al cytopathic eff ects

Vigene integrated vector and plasmid service — Research, preclinical and clinical

Vigene is a world leader in plasmid and viral vector development and manufacturing. Vigene has expertise in development, manufacturing and analytics for plasmid vectors and many viral vectors, including AAV, lentivirus, adenovirus, retrovirus, HSV, and vaccinia virus.

Please see the Jan 2021 promotion here: vigenebio.com/2021/Jan

www.vigenebio.com

2021JANUARY TUESDAY WEDNESDAY THURSDAY FRIDAY SATURDAY SUNDAYMONDAY

64

1311

2018

2725

75

1412

2119

2826

2

9

16

23

30

1

8

15

22

29

3

10

17

24

31

Vigene integrated vector and plasmid service

Figure reprinted with permission from Springer Nature

AAV reference standards

Th e gene and cell therapy fi eld needs good AAV reference standards. Pursuant to the Jan 2020 FDA Gene Th erapy Chemistry Manufacturing and Controls (CMC) guidance, Vigene’s AAV reference standards are as follows:• AAV1, AAV2, AAV5, AAV6, AAV8, and AAV9 serotypes• Full and empty capsids• AAV titer standards

Please view the details and promotions at: vigenebio.com/2021/Feb

Transmission electron microscopy of rAAV. It is clear that full particles (open arrow) stain diff erently from empty particles (darkly stained center; small dark arrow). Viewing numerous fields similar to this will allow determination of the full-to-empty particle ratio. (Figure 8, Nature Protocols 1, 1412–1428 (2006))

www.vigenebio.com

2021FEBRUARY TUESDAY WEDNESDAY THURSDAY FRIDAY SATURDAY SUNDAYMONDAY

1 3

108

1715

2422

42

119

1816

2523

6

13

20

27

5

12

19

26

7

14

21

28

AAV reference standards

ITRITR

Therapeutic transgene Poly(A)

AAV vector packaging(4.5-kb capacitywithout engineering)

Promoter

Positiveeffect onimmuneresponse

Positive effect ontransduction

Engineeringapproach

• Decreases innate immune response to AAV

Use of dual vectors that take advantage of:• AAV genome concatemerization• Homologous recombination• A hybrid dual-vector strategy• Intein-mediated protein trans-splicing technologyCross-packaging of an AAV genome

• Synthetic poly(A)• Reversed poly(A)

• Increases size of transgene that can be packaged

• Decreases innate immune response to AAV

• Omits need for second- strand synthesis of ssDNA

• Mutation of ITRs to generate self- complementary AAV intermediates

• Enhances polyadenylation• Minimizes kilobases of DNA taken up by poly(A)• Avoids rolling circle transcription and Pol II jumping (between concatamers)

• Decreases innate immune response to AAV• Reduces CTL response

• Increases transcription and translation of the transgene

• Codon optimization via codon usage bias• Interference of antigen presentation

• Decreases CTL response to AAV

• Increases tissue-specific transcription of transgene• Minimizes kilobases of DNA taken up by promoter

• Screen for and use small, strong, tissue-specific promoters and enhancer elements

RecombinantAAV virion

5′ 3′ OH

Figure reprinted with permission from Springer Nature

Engineering the AAV cassette. The adeno-associated virus (AAV) cassette can be engineered to enhance AAV transduction and also to enable AAV to escape immune responses. Mutation of one inverted terminal repeat (ITR) on the AAV vector, which prevents the nicking of Rep protein, can generate self-complementary AAV vector to enhance vector transduction. Mutation of ITRs may also decrease the innate response to AAV. Use of small tissue-specifi c promoters increases tissue-specifi c transgene expression and the packaging capacity of the AAV genome and minimizes the cytotoxic T lymphocyte (CTL) immune response to AAV. Optimization of transgene codons increases the transcription and translation of the AAV transgene and decreases the immune response to AAV. Using synthetic poly(A) can increase the nuclear export, translation and stability of mRNA (by enhancing polyadenylation), and using reversed poly(A) can avoid the transcription of ITR; both approaches enhance AAV transduction, and reversed poly(A) decreases the innate response to AAV. Finally, the use of dual AAV vectors or the cross-packaging of the AAV genome enables eff ective and functional expression of large transgenes. Pol II, DNA polymerase II; ssDNA, single-stranded DNA. (Figure 2, Nature Reviews Genetics 21, 255–272 (2020))

Plasmid production – Research, preclinical, clinical

• Research grade & GMP-ReadyTM, GMP production• Ready-to-use, GMP-Ready pHelper & other viral vector packaging plasmids• Endotoxin free & supercoil plasmid homogeneity • Animal component free production process

Please view the details at vigenebio.com/2021/Mar

www.vigenebio.com

2021MARCH TUESDAY WEDNESDAY THURSDAY FRIDAY SATURDAY SUNDAYMONDAY

1 3

108

1715

2422

3129

42

119

1816

2523

30

6

13

20

27

5

12

19

26

7

14

21

28

Plasmid production – preclinical and GMP

AAV production – Research, preclinical, clinical

• Research grade, preclinical, clinical, and commercial GMP AAV production• Proprietary cGMP released MCB of HEK293, 293T adherent and suspension cell lines• Established and proven high productivity AAV production process• >15,000 research batches, >50 preclinical and clinical batches released

Please view the promo at vigenebio.com/2021/Apr

TABLE 1: AAV PRODUCTION OPTIONSPros and cons of various recombinant adeno-associated virus (rAAV) manufacturing strategies. rAAV cannot replicate without a helper virus, and early manufacturing efforts entailed coinfection of host cells with adenovirus or herpesvirus. Newer strategies replace these with plasmids containing key helper virus genes, or combine all necessary genetic elements into an insect cell-specific baculovirus vector. Production is most efficient in free-floating suspension cells, but substrate-attached adherent cell lines can also achieve reasonable viral output.

AAV MANUFACTURING TECHNOLOGY

KEY STRENGTHS

KEY DRAWBACKS

PRODUCTION CELL LINE CHOICES

ADHERENT SUSPENSION

Helper virus

• Highly scalable• Serum-free media• Efficient

production in suspension culture

• Helper virus contamination

• Long lead time for cell line and virus seed generation

• May require serum- containing media

HEK293/293T HeLa

HEK293/293T-s HeLa-s

Helper-free triple transfection

• No helper virus contamination

• Rapidly produce virus in small scale

• Simple procedure

• May require serum- containing media

• Large proportion of empty capsids

• Supply of plasmids for large-scale production can be costly

HEK293/293T HEK293/293T-s

Baculovirus

• Highly scalable Serum-free media

• Efficient production in suspension culture

• Baculovirus virus contamination

• Baculovirus instability

• Long lead time for cell line and virus seed generation

— sf9

www.vigenebio.com

2021APRIL TUESDAY WEDNESDAY THURSDAY FRIDAY SATURDAY SUNDAYMONDAY

75

1412

2119

2826

1

86

1513

2220

2927

3

10

17

24

2

9

16

23

30

4

11

18

25

AAV production – preclinical and GMP

CAR

Target cell

Virus-mediated transduction is auseful tool to generate varioustypes of next-generation stem cells

gf

d

b cOptogenetic actuator-embeddedstem cell derivative

Gene editing tools can be used to knockout expression of HLA genes to reduce theimmunogenicity of allogeneic stem cells

a Prodrug-converting enzymesor oncolytic virus delivery viatumour-homing NSCs/MSCs

Stem cell

HSC

e Cytokine/growth factor-overexpressingstem cells to enhance endogenous repair

Improved delivery of anticancer agents to sites of action using clickchemistry to tether them to stem cells

Azide

DBCO

Anti-PD-L1

Platelet

Gene therapy of stem cells to providelong-term functional compensationfor disease-causing, inherited mutation

CAR-expressing HSCs or PSC-derived NK cells

Virus as a key tool�Click chemistryNuclease or AAV-based gene editing

Oncolytic virusparticles (or prodrug-converting enzymes)

Lentivirus production – Research, preclinical, clinical• Research grade, preclinical, clinical, and commercial GMP lentivirus production• High titer, purifi ed, ready to use in vitro and animal studies — research grade• Chromatography based commercial ready lenti production — GMP • High transduction effi ciency

Please view the promo at vigenebio.com/2021/May

The stem cell toolkit and its application in developing next-generation stem cells. Virus-mediated transduction of stem cells (target cell), particularly with self-inactivating lentiviruses, is a useful tool for the creation/development of next-generation stem cells. Viruses can be used to engineer: prodrug-converting enzymes, oncolytic viruses and other anticancer drugs into neural stem cells (NSCs) and mesenchymal stem cells (MSCs) (part a); optogenetically enhanced stem cell derivatives to provide light-inducible control over the activity of trans-planted stem cells/progenitors (part b); chimeric antigen receptor (CAR)-expressing haematopoietic stem cells (HSCs) and pluripotent stem cell (PSC)-derived natural killer (NK) cells for immune-oncology applications (part c); gene therapy in HSC, skin and muscle progenitors to treat inherited diseases (part d); and cytokine/growth factor delivery in MSCs/neural progenitors to stimulate endogenous tissue repair (part e). Click chemistry is another engineering tool that can be used to tether anticancer agents to stem cells for improved delivery to hard-to-reach cancers, such as leukaemia cells residing deep in the bone marrow (part f). Advances in gene editing technology have made this a versatile tool to precisely edit specifi c loci within the genome, and gene editing technology has become the tool of choice for the creation/development of universally immunocompatible PSC lines and derivatives (part g). Several of these next-generation stem cell-based therapies have already reached clinical testing, whereas others in preclinical development are not far behind. AAV, adeno-associated virus; DBCO, dibenzocyclooctyne; HLA, human leukocyte antigen; PD-L1, programmed cell death protein 1 ligand. (Figure 1, Nature Reviews Drug Discovery 19, 463–479 (2020))

Figure reprinted with permission from Springer Nature

2021

www.vigenebio.com

MAY TUESDAY WEDNESDAY THURSDAY FRIDAY SATURDAY SUNDAYMONDAY

53

1210

1917

2624

31

64

1311

2018

2725

1

8

15

22

29

7

14

21

28

2

9

16

23

30

Lentivirus research production

Oncolyticvirus

ROS

ROS

• ER stress• Genotoxic stress

Cancer cell

Viraloncolysis

CD8+ T cell

CD4+ T cell

NK cell

Antigen presenting cell

Cytokinereceptors

Cytokinereceptors

CD28

Cytotoxicity

Cytotoxicity

Activation

TCR

TCR

IL-2R

IL-2

MHC MHCTLR

MHC

CD40

CD40L

• Type I IFNs• DAMPs/PAMPs

DAMPs/PAMPs

• Type I IFNs• Cytokines

• Viral proteins• Viral genome

• Viral antigens• TAAs/neoantigens

• CD80/CD86• Chemokine receptors

PAMPs• Viral capsids• Viral DNA• Viral dsRNA/ssRNA• Viral proteins

DAMPs• HSPs• HMGB1• Calreticulin• ATP• Uric acid

Cytokines• Type I interferons• TNFα• IFNγ• IL-12

Infection

Release Release

Antigenuptake

Release/secrete

Adenovirus production - Research, preclinical, clinical• Research grade, preclinical, clinical, and commercial GMP adenovirus production• High titer, purifi ed, ready to use in vitro and animal studies – research grade• Chromatography based commercial ready adenovirus production – GMP • Oncolytic virus and vaccine production process ready

Please view the promo at vigenebio.com/2021/June

The induction of local and systemic anti-tumour immunity by oncolytic viruses. The therapeutic effi cacy of oncolytic viruses is determined by a combination of direct cancer cell lysis and indirect activation of anti-tumour immune responses. Upon infection with an oncolytic virus, cancer cells initiate an antiviral response that consists of endoplasmic reticulum (ER) and genotoxic stress. This response leads to the upregulation of reactive oxygen species (ROS) and the initiation of antiviral cytokine production. ROS and cytokines, specifi cally type I interferons (IFNs), are released from the infected cancer cell and stimulate immune cells (antigen presenting cells, CD8+ T cells, and natural killer (NK) cells). Subsequently, the oncolytic virus causes oncolysis, which releases viral progeny, pathogen-associated molecular patterns (PAMPs), danger-associated molecular pattern signals (DAMPs), and tumour associated antigens (TAAs) including neo-antigens. The release of viral progeny propagates the infection with the oncolytic virus. The PAMPs (consisting of viral particles) and DAMPs (comprising host cell proteins) stimulate the immune system by triggering activating receptors such as Toll-like receptors (TLRs). In the context of the resulting immune-stimulatory environment, TAAs and neo-antigens are released and taken up by antigen presenting cells. Collectively, these events result in the generation of immune responses against virally infected cancer cells, as well as de novo immune responses against TAAs/neo-antigens displayed on un-infected cancer cells. CD40L, CD40 ligand; dsRNA, double-stranded RNA; HMGB1, high mobility group box 1; HSP, heat shock protein; IL-2, interleukin-2; IL-2R, IL-2 receptor; MHC, major histocompatibility complex; ssRNA, single-stranded RNA; TCR, T cell receptor; TNFα, tumour necrosis factor-α. (Figure 2, Nature Reviews Drug Discovery 14, 642–662 (2015))

Figure reprinted with permission from Springer Nature

www.vigenebio.com

2021JUNE TUESDAY WEDNESDAY THURSDAY FRIDAY SATURDAY SUNDAYMONDAY

2

97

1614

2321

3028

31

108

1715

2422

29

5

12

19

26

4

11

18

25

6

13

20

27

Adenovirus production 

Table 1 | Properties and clinical use of AAV serotypes

AAV serotype

Origin of isolation

Primary receptor

Co-receptor Tissue tropism Condition (ClinicalTrials.gov identifier)

Approved drug

AAV1 Monkey Sialic acid AAVR Muscle, CNS, heart Muscle diseases (NCT01519349) None

Heart failure (NCT01643330)

AAT deficiency (NCT01054339, NCT00430768)

AAV2 Human Heparin Integrin, FGFR , HGFR , LamR , AAVR

Liver, CNS, muscle Eye diseases (NCT00643747) Luxturna for Leber congenital amaurosis

Haemophilia (NCT00515710)

CNS diseases (NCT00400634)

AAT deficiency (NCT00377416)

AAV3 Human Heparin FGFR , HGFR LamR , AAVR

Muscle, stem cells No trials underway None

AAV4 Monkey Sialic acid Unknown Eye, CNS Eye diseases (NCT01496040) None

AAV5 Human Sialic acid PDGFR , AAVR CNS, lung, eye Haemophilia (NCT03520712) None

Eye diseases (NCT02781480)

AIP (NCT02082860)

AAV6 Human Heparin, sialic acid

EGFR , AAVR Muscle, CNS, heart, lung

Haemophilia (NCT03061201) None

CNS diseases (NCT02702115)

AAV7 Monkey Unknown Unknown Muscle, CNS No trials underway None

AAV8 Monkey Unknown LamR , AAVR Liver, muscle, pancreas, CNS

Eye diseases (NCT03066258) None

Haemophilia (NCT00979238)

Muscle diseases (NCT03199469)

AAV9 Human Galactose LamR , AAVR Every tissue CNS diseases (NCT02122952) Zolgensma for spinal muscular atrophy

Muscle diseases (NCT03362502)

AAV10 Monkey Unknown Unknown Muscle No trials underway None

AAV11 Monkey Unknown Unknown Unknown No trials underway None

AAV12 Human Unknown Unknown Nasal No trials underway NoneAAT, α1-antitrypsin; AAV, adeno-associated virus; AAVR , AAV receptor ; AIP, acute intermittent porphyria; CNS, central nervous system; EGFR , epidermal growth factor receptor ; FGFR , fibroblast growth factor receptor ; HGFR , hepatocyte growth factor receptor ; LamR , laminin receptor 1; PDGFR , platelet-derived growth factor receptor.

Properties and clinical use of AAV serotypes

Table reprinted with permission from Springer Nature

AAT, α1-antitrypsin; AAV, adeno-associated virus; AAVR, AAV receptor; AIP, acute intermittent porphyria; CNS, central nervous system; EGFR, epidermal growth factor receptor; FGFR, fi broblast growth factor receptor; HGFR, hepatocyte growth factor receptor; LamR, laminin receptor 1; PDGFR, platelet-derived growth factor receptor. (Table 1, Nature Reviews Genetics 21, 255–272 (2020))

AAV, lentivirus and adenovirus controls

• Reporters available: RFP, GFP, mCherry, Luciferase, Cre, LacZ• Promoters available: ALB, aMHC, c-Fos, CAG, CaMKIIa, CK0.4, CK1.3, CMV, cTnT, EF1a, EFFS,

GFAP, HCRApoE, MBP, MCK, MeCP2, NSE, PDX1, PGK, Rpe65, SST, Syn, TBG, UBC, DIO-GFP, DIO-mCherry, DIO-RFP, DIO-LacZ

Please view the promo at vigenebio.com/2021/July

www.vigenebio.com

2021JULY TUESDAY WEDNESDAY THURSDAY FRIDAY SATURDAY SUNDAYMONDAY

75

1412

2119

2826

1

86

1513

2220

2927

3

10

17

24

31

2

9

16

23

30

4

11

18

25

AAV controls and lentivirus control

Table 1 Example of characterization testing for an HEK293 master cell bank

noitacfiicepSdohteMtseT

MicrobialypocsorcimnortcelenoissimsnarTnoitanimatnoclaiborcimroF

fungi, yeasts, bacteriaBacteriostatic/fungistatic activity of test articleBacterial and fungal contaminantsAgar cultivable and noncultivable mycoplasmas

General virusesInapparent viruses In vivo Not detectedViral contaminants In vitro assay for the presence of

viral contaminantsNot detected

Specific human virusesevitageNRCPVMCevitageNRCPVBEevitageNRCP-TRVAHevitageNRCPVBHevitageNRCP-TRVCHevitageNRCP6-VHHevitageNRCP7-VHHevitageNRCP8-VHHevitageNRCP2/1-VIHevitageNRCP91BsurivovrapnamuHevitageNRCPII/IVLTH

TREP-QsesurivorteR < 5.0 10 7 U ml 1

Specific simian virusesevitageNRCPVFSevitageNRCP-TRVRSevitageNRCPVLTSevitageNRCP04VS

Other specific virusesBovine viruses In vitro assay for the presence of

bovine viruses (9CFR)Not detected

Porcine viruses In vitro assay for the presence ofporcine viruses (9CFR)

Not detected

IdentitytnetsisnocsecnatsidnoitargimemyzneosIsisylanaemyzneosInoitac�itnedilleC

with cells of human origin

Abbreviations: CFR, Code of Federal Regulations; CMV, cytomegalovirus; EBV, Epstein–Barr virus; HAV, hepatitis A virus; HBV, hepatitis Bvirus; HCV, hepatitis C virus; HHV, human herpes virus; HIV-1/2, human immunode�ciency virus types 1 and 2; HTLV I/II, human T-celllymphotropic virus types I and II; SFV, simian foamy virus; SRV, simian retroviruses; STLV, simian T-lymphotropic virus; SV40, simian virus 40.US Dept Health Human Services, Food and Drug Administration. 23

No viruses, virus-like particles, mycoplasmas,

No bacterial and fungal activityFour media, direct inoculationFour media, direct inoculation1993 points to consider

NegativeNegative

Quality control analytical services• Plasmid QC

◆ Purity & identity: % supercoil (HPLC), sequencing, residual host DNA/RNA/proteins ◆ Safety: sterility, endotoxin, mycoplasma, etc.

• Viral vector QC ◆ Purity and impurities: residual IDX (HPLC), residual host DNA/RNA/proteins ◆ Strength & safety: vector genome titer (qPCR, ddPCR), total particle titer (ELISA), sterility, mycoplasma, etc.

Please view the promo at vigenebio.com/2021/Aug

Example of characterization testing for an HEK293 master cell bank

Abbreviations: CFR, Code of Federal Regulations; CMV, cytomegalovirus; EBV, Epstein–Barr virus; HAV, hepatitis A virus; HBV, hepatitis B virus; HCV, hepatitis C virus; HHV, human herpes virus; HIV-1/2, human immunodeficiency virus types 1 and 2; HTLV I/II, human T-cell lymphotropic virus types I and II; SFV, simian foamy virus; SRV, simian retroviruses; STLV, simian T-lymphotropic virus; SV40, simian virus 40. US Dept Health Human Services, Food and Drug Administration23. (Table 1, Gene Therapy 15, 840–848 (2008))

Table reprinted with permission from Springer Nature

2021AUGUST

www.vigenebio.com

TUESDAY WEDNESDAY THURSDAY FRIDAY SATURDAY SUNDAYMONDAY

42

119

1816

2523

30

53

1210

1917

2624

31

7

14

21

28

6

13

20

27

1

8

15

22

29

Viral vector and plasmid analytical services

Table 1 Selected BCMA CAR-T trials.

Institution/developer

Vector/co-stimulatory domain BCMA positivity requirement No of patients Median prior lines(range)

EfficacyORR*/PFS(months)

SafetyCRS/ICANS

NCI γ-retrovirus/CD28 >50% 24 9 (3–13) 81%/7.2 94% (38% grade ≥ 3)/NE

UPenn Lentivirus/4–1BB Not required 25 7 (3–13) Cohort 1 44%/2.2Cohort 2 20%/1.9Cohort 3 64%/4.2

88% (32% grade ≥ 3)/32%(12% grade ≥ 3)

Bb2121 Lentivirus/4-1BB ≥50% in dose escalation, NR indose expansion

33 7 (3–14)8 (3–23)

85%/11.8 70% (6% grade ≥ 3)/42%(3.3% grade ≥ 3)

Bb21217 Lentivirus/4-1BB; PI3K inh during in vivoexpansion

>50% 22 7 (4–17) 83%/NR 59% (4.5% grade ≥ 3)/22%(9% grade≥3)

LCAR-B38 M Lentivirus/41-BB Required 57 3 (1–9) 88%/15 90% (grade 3 ≥ 7%)/2%

LCAR-B38M Lentivirus/4-1BB Required 17 4 (3–11) 88%/NR 100% (grade ≥ 3 35%)/NR

Poseida (P-BCMA 101) PiggyBAC/4-1BB Not required 23 6 (3–11) 63%/NR 9.5%/4.8% (grade ≥ 3 4.8%)

JCARH125 Lentivirus/4-1BB Not required 44 7 (3–23) 82%/NR 80% (9% grade ≥ 3)/25%(grade ≥ 3 7%)

MCARH171 Retrovirus/4-1BB/tEGFR Required 11 6 (4–14) 64%/NR 60% (20% grade ≥ 3)/NR

Han et al Lentivirus/4-1BB/Alpaca VHH Not required 16 10 (NR) 100%/NR NR (12.5% grade ≥ 3)/NR

FCARH143 Lentivirus/4-1BB/tEGFR ≥5% 7 8 (6–11) 100%/NR 86%/0%

CARTITUDE-1 Lentivirus/4-1BB Not required 25 5 (3–16) 91%/NR 80% (8% grade ≥ 3)/12%(4% grade ≥ 3)

CT053 Lentivirus/4-1BB ≥50% 16 NR 100%/NR 18% (6% grade ≥ 3)/NR

CT103 Lentivirus/4-1BB NR 16 4 (3–5) 100%/NR 100% (37.5% grade ≥ 3)/0%

Cowan et al Lentivirus/4-1BBGSI (JSMD194)/tEGFR

Required 8 10 (4–23) 100%/NR 100%/70%

C-CAR088 Lentivirus/4-1BB NR 3 7 (NR) 100%/NR NR

HRAIN biotechnology γ-retrovirus-4-1BB/tEGFR >5% 17 NR 79%/NR NR

CRS cytokine release syndrome, ICANS immune effector cell associated neurotoxicity syndrome, ORR overall response rate, PFS Progression free survival, BCMA B cell maturation antigen, GSIgamma secretase inhibitor, EGFR epidermal growth factor receptor.

*responses assessed after 30 days.

B.Dhakalet

al.

Selected BCMA CAR-T trials

Retrovirus production – Research, preclinical, clinical

• Research grade, preclinical, clinical, and commercial GMP retrovirus production• High titer, purifi ed, ready to use in vitro and animal studies – research grade• Chromatography based commercial ready retrovirus production – GMP • High transduction effi ciency

Please view the promo at vigenebio.com/2021/Sept

CRS cytokine release syndrome, ICANS immune eff ector cell associated neurotoxicity syndrome, ORR overall response rate, PFS Progression free survival, BCMA B cell maturation antigen, GSI gamma secretase inhibitor, EGFR epidermal growth factor receptor. *responses assessed after 30days. (Table 1, Bone Marrow Transplantation https://dx.doi.org/10.1038/s41409-020-01023-w (2020))

Table reprinted with permission from Springer Nature

www.vigenebio.com

2021SEPTEMBER TUESDAY WEDNESDAY THURSDAY FRIDAY SATURDAY SUNDAYMONDAY

1

86

1513

2220

2927

2

97

1614

2321

3028

4

11

18

25

3

10

17

24

5

12

19

26

Retrovirus production 

AAV & LVV plasmid ITR/LTR repair and production service

Plasmid design and cloning (identical ITR vs hybrid)• Genetic stability study & ITR sequencing analysis with NGS• AAV plasmid & LVV plasmid clonal screening using diff erent E. coli strains

for stable and reproducible ITR/LTR plasmid production

Please view the promo at vigenebio.com/2021/Oct

5ʹ

5ʹ

3ʹ5ʹ3ʹ

5ʹ3ʹ 3ʹ

4.6 kb

4.6 kb

++

––

wtTR

wtTR

2.3 kb

ΔTR

OpenOpen

OpenOpen

4.6 kb

4.6 kb

4.6 kb

wtTRScAAVConventional ssAAV

Graphical representation of the suggested portion of the transgene plasmid to be used as the template of the probe for dot-blot analysis. The black lines represent sequences in the bacterial backbone; the blue and red lines represent sequences of the coding and complementary sequence of the transgene expression cassette, respectively. (Figure 7, Nature Protocols 1, 1412–1428 (2006))

Figure reprinted with permission from Springer Nature

www.vigenebio.com

2021OCTOBER TUESDAY WEDNESDAY THURSDAY FRIDAY SATURDAY SUNDAYMONDAY

11

64

13

2018

2725

75

1412

2119

2826

2

9

16

23

30

1

8

15

22

29

3

10

17

24

31

AAV plasmid ITR repair and production service

Gene X

Packaging and payload

Genetic access Infectivity and toxicity Transgene expression

Delivery Tropism

Virus particle

Localinjection

Globalinjection

Infected cell Uninfected cell

All cells Selected cells

Entry(susceptibility) Infectivity

Transgeneexpression(permissivity) Toxicity

AAAm7G

Healthycell

Damagedcell

Duration ofexpression

Tran

sgen

e ex

pres

sion

Decay ortoxicity

Onset

Virusinjection

Figure reprinted with permission from Springer Nature

Key principles for viral-mediated gene transfer in neuroscience. Schematic demonstrating six key principles essential for the neuroscientist: viral packaging limit (how much nucleic acid a virus particle can carry) and payload (the length and type of genomic material that can be successfully packaged into a virus particle), delivery methods (local versus global injections), tropism (specifi city of a virus for a given cell type(s)), access (ability of a virus to enter a cell type and express its gene product(s)), infectivity and toxicity (how effi ciently a virus infects a cell and how harmful it is to the cell), and transgene expression dynamics (time course of onset and persistence of transgene expression). These principles play a key role in determining a neuroscientist’s choice of virus by weighing the advantages and disadvantages of a given virus. AAA, 3´ poly(A) tail for mRNA; Gene X, a transgene being packaged into a virus particle; m7G, 7-methylguanosine (5´ cap for mRNA). (Figure 1, Nature Reviews Neuroscience 21, 669–681(2020))

Gene therapy bioinformatics & resources central

• Technology handbooks• Bioinformatics tools• Webinar tutorials

Please view the promo at vigenebio.com/2021/Nov

www.vigenebio.com

2021NOVEMBER TUESDAY WEDNESDAY THURSDAY FRIDAY SATURDAY SUNDAYMONDAY

29

31

108

1715

2422

42

119

1816

2523

30

6

13

20

27

5

12

19

26

7

14

21

28

Gene therapy tools

Bottlenecks Solutions

2D cell factories Microcarrier culture (1) Subculture —bead-to-bead transfer

(2) Dissociation

Enzymaticdigestion

Centrifugalseparation

Modified surfaces

Solidmicrocarrier

Degradablesurface

Nn

O

Engineered substratesPNIPAM

T < 32 °C T > 32 °C

a b c d

Figure reprinted with permission from Springer Nature

Process optimization for the expansion of cells and for cell collection from microcarriers. a, Production of clinical lots by using adherent MSCs in 2D cell-culture plates. Issues with the scaling of costs and labour effi ciency make 2D culture unlikely to meet an estimated demand of > 1012

viable cells per year, necessary for treating prevalent adult indications. b, Suspension culture systems for MSCs use microcarriers and stirred tank bioreactors and are a scalable and sustainable approach for cell expansion at high density. c, Unit operations identifi ed as major bioprocessing bottlenecks: (1) bead-to-bead transfer for MSC subculturing and expansion; (2) the need for enzymatic digestion and centrifugal separation to isolate the MSCs from the microcarriers. d, Materials-science innovations in microcarrier substrates can improve product purity, identity and potency through degradable and temperature (T)-sensitive materials (such as poly(N-isopropylacrylamide), PNIPAM) that remove the need for additional enzymatic dissociation processes. (Figure 1, Nature Biomedical Engineering 2, 362–376 (2018))

Bioprocess and GMP production

• FDA/EMA compliant with 17-yr GMP production records• Established AAV, LVV, adenovirus, retrovirus, and plasmid GMP production processes• >10 GMP suites, >100,000 sq ft GMP & lab space for adherent and suspension cell

production (50 L to 1,000 L per batch)

Please view the promo at vigenebio.com/2021/Dec

www.vigenebio.com

2021DECEMBER TUESDAY WEDNESDAY THURSDAY FRIDAY SATURDAY SUNDAYMONDAY

1

86

1513

2220

2927

2

97

1614

2321

3028

4

11

18

25

3

10

17

24

31

5

12

19

26

Gene therapy tools