Changes to the classification of Viral Vectors · Changes to the classification of Viral Vectors...

Changes to the classification Changes to the classification of Viral Vectorsof Viral Vectors

Review of the Review of the Gene Technology Regulations 2001Gene Technology Regulations 2001

Talk OutlineTalk Outline●●

Overview of changes involving viral vectorsOverview of changes involving viral vectors●●

DefinitionsDefinitions●●

Replication defective (RD) vectorsReplication defective (RD) vectors●●

Not able to transduce human cellsNot able to transduce human cells●●

Able to transduce human cellsAble to transduce human cells●●

NonNon--

retroviralretroviral●●

Retroviral Retroviral ●●

examplesexamples

●●

Tools & ResourcesTools & Resources

Disclaimer: Automatic DNIRsDisclaimer: Automatic DNIRsHost:Host:●●

Risk Group 4 organism 3.1(p)Risk Group 4 organism 3.1(p)

Introduced nucleic acid:Introduced nucleic acid:●●

Toxin or uncharacterised nucleic acid Toxin or uncharacterised nucleic acid from a toxin producing organism from a toxin producing organism 3.1(a3.1(a--c)c)

●●

Novel replication competent virus Novel replication competent virus 3.1(i)3.1(i)

Exempt

PC1 NLRD PC2 NLRD

DNIR

Most RDRetroviral vectors

Some RDNon-

retroviral vectors

Exempt vector

oncogenes

Summary of changes to in vitro dealings

Summary of changes to in vivo dealings

PC2 NLRD DNIR

Some RDretroviruses

Talk terminologyTalk terminology‘‘safesafe’’

‘‘low risklow risk’’

or or ‘‘none of the abovenone of the above’’

genegene●●

Not a toxinNot a toxin●●

Does not restore replication competence or Does not restore replication competence or create a novel viruscreate a novel virus

●●

Not a pathogenic determinant for the vectorNot a pathogenic determinant for the vector●●

Not immunomodulatory in humansNot immunomodulatory in humans●●

Does not confer an oncogenic modificationDoes not confer an oncogenic modification

Oncogenic modificationOncogenic modificationoncogenic modificationoncogenic modification --

means a genetic means a genetic

modification capable of contributing to modification capable of contributing to tumour formation, including modifications tumour formation, including modifications that cause at least 1 of the following:that cause at least 1 of the following:

(a) defects in DNA proofreading and repair(a) defects in DNA proofreading and repair(b) defects in chromosome maintenance(b) defects in chromosome maintenance(c) defects in cell cycle checkpoint mechanisms(c) defects in cell cycle checkpoint mechanisms(d) uncontrolled cell proliferation(d) uncontrolled cell proliferation(e) resistance to apoptosis(e) resistance to apoptosis(f) cellular immortalisation.(f) cellular immortalisation.

Talk terminologyTalk terminology

●●

‘‘lentilenti’’, , ‘‘lentislentis’’

or or ‘‘lentiviral vectorlentiviral vector’’

––

replication defective retroviral vector replication defective retroviral vector system derived from system derived from Human Human immunodeficiency virusimmunodeficiency virus (HIV)(HIV)

●●

‘‘retroretro’’

––

a retrovirus which is not a a retrovirus which is not a

lentiviral vectorlentiviral vector

Immunomodulatory in humanImmunomodulatory in human●●

Includes:Includes:●●

Human cytokines, Human cytokines, chemokineschemokines

and their and their receptorsreceptors

●●

Molecules involved in immune cell differentiation Molecules involved in immune cell differentiation maturation and activation maturation and activation

●●

Animal homologues unless known not to affect Animal homologues unless known not to affect human cellshuman cells

●●

Does not include antigensDoes not include antigens●●

Note: if not known whether the genes are Note: if not known whether the genes are immunomodulatory in humans then should immunomodulatory in humans then should assume they are and assess accordinglyassume they are and assess accordingly

●●

Currently exempt host vector Currently exempt host vector system in tissue culturesystem in tissue culture●●

now includes now includes gene(sgene(s) capable of ) capable of inducing an oncogenic inducing an oncogenic modificationmodification

●●

All All in vivoin vivo dealings PC2 NLRDdealings PC2 NLRD

Replication defective viral vectorsReplication defective viral vectorsNot able to transduce human cells

●●

Currently Currently in vitroin vitro dealings with dealings with ‘‘safe genessafe genes’’ PC1 NLRDPC1 NLRD

●●

In vitroIn vitro dealings with vectors derived fromdealings with vectors derived from Human adenovirusHuman adenovirus ((HAdHAd) and ) and Adenovirus Adenovirus

associated virusassociated virus (AAV) still PC1, (AAV) still PC1, all others all others PC2 NLRDPC2 NLRD●●

Long history of safe useLong history of safe use●●

Internationally accepted as Risk Group 1Internationally accepted as Risk Group 1

Replication defective nonReplication defective non--retrovirusesretrovirusesAble to transduce human cells

Replication defective nonReplication defective non--retrovirusesretroviruses

Donor nucleic acid (transgene)

DealingsIn vitro In vivo

2007 2011 2007 2011Immunomodulatory PC1 NLRD

PC2 NLRD DNIROncogenic modification PC2 NLRD

None of the above PC1 NLRD

PC1 NLRD HAd, AAV

PC2 NLRDPC2 NLRD

others

Able to transduce human cells

Lentiviral vectorsLentiviral vectors●●

Common tool used by researchers to Common tool used by researchers to introduce a gene into the genome of a introduce a gene into the genome of a target cell or organismtarget cell or organism

●●

Stable gene incorporation and Stable gene incorporation and expressionexpression

●●

Use of different envelope proteins Use of different envelope proteins allow transduction of a wide range of allow transduction of a wide range of cell typescell types

●●

Derived from HIV Derived from HIV

Gene of interest5’ UTR 3’ UTRRRE

GagPol

RevEnv

Packaging cells

Target cell

GM Lentiviral particles

Replication defective Replication defective retroviruses able retroviruses able to transduce human cells as NLRDsto transduce human cells as NLRDs

●●

Must have Must have bothboth

of the following safety of the following safety features:features:●●

All viral genes removed from vectorAll viral genes removed from vector●●

Packaging proteins supplied Packaging proteins supplied in transin trans from from multiple unlinked loci with minimum sequence multiple unlinked loci with minimum sequence overlapoverlap

●●

Must have Must have at least at least oneone

of the following:of the following:●●

self inactivating deletion in 3self inactivating deletion in 3’’UTR (SIN)UTR (SIN)●●

essential packaging proteins only essential packaging proteins only ––

gag, gag, polpol, (rev , (rev for lentis) and an envelope of some kindfor lentis) and an envelope of some kind

Retroviral vectorsRetroviral vectors

Red = vector

Green = essential genes provided on independent loci

Blue = accessory genes

Replication defective Replication defective retroviruses able retroviruses able to transduce human cells to transduce human cells --

big picturebig picture

●●

2007 Regs PC2 NLRD if:2007 Regs PC2 NLRD if:●●

All structural genes removed from vector; ANDAll structural genes removed from vector; AND●●

Self inactivating Self inactivating ANDAND

Only gag, Only gag, polpol, rev & , rev & envenv●●

Any gene Any gene in vitroin vitro, safe genes , safe genes in vivoin vivo

●●

2011 Regs PC2 NLRD if:2011 Regs PC2 NLRD if:●●

All structural genes removed from vector; ANDAll structural genes removed from vector; AND●●

Self inactivating Self inactivating OROR

Only gag, Only gag, polpol, rev & , rev & envenv●●

Any gene Any gene in vitroin vitro, safe genes , safe genes in vivoin vivo

Disclaimer: Automatic DNIRsDisclaimer: Automatic DNIRsHost:Host:●●

Risk Group 4 organism 3.1(p)Risk Group 4 organism 3.1(p)

Introduced DNA:Introduced DNA:●●

Toxin or uncharacterised nucleic acid Toxin or uncharacterised nucleic acid from a toxin producing organism from a toxin producing organism 3.1(a3.1(a--c)c)

●●

Novel replication competent virus Novel replication competent virus 3.1(i)3.1(i)

Replication defective retrovirusesReplication defective retroviruses

NoOncogenic /

immunomodulatory PC2 NLRDDNIR

None of the above PC2 NLRD

Yes

Oncogenic / immunomodulatory

DNIRPC2

NLRDDNIR

None of the above DNIR PC2 NLRD

Able to transduce human cells and contains a self inactivating deletion

Packaging plasmids contain

‘extra’ genes


Dealings

In vitro In vivo

2007 2011 2007 2011


No


Lenti DNIRPC2

NLRD DNIRretro PC2 NLRD

None of the aboveLenti DNIR

PC2 NLRD

Lenti DNIRPC2 NLRDretro PC2

NLRDretro PC2

NLRDYes Any DNIR

Packaging plasmids contain

‘extra’ genes


Dealings

In vitro In vivo

2007 2011 2007 2011

Able to transduce human cells but does not

contain a self inactivating deletion

Assessing dealings with Assessing dealings with viral vectorsviral vectors

Factors to consider when assessing Factors to consider when assessing dealings with viral vectorsdealings with viral vectors

∙∙ Replication competent or defectiveReplication competent or defective

∙∙ Able or unable to transduce human cellsAble or unable to transduce human cells

∙∙ Retroviral or nonRetroviral or non--retroviralretroviral

∙∙ If retroviral, presence of safety featuresIf retroviral, presence of safety features

∙∙ Characteristics (source & function) of the Characteristics (source & function) of the introduced nucleic acidintroduced nucleic acid

∙∙ Dealings in tissue culture or animalsDealings in tissue culture or animals

Example 1Example 1ApplicationApplication●●

RD RD lentilenti

systemsystem●●


33’’UTR contains self inactivating deletionUTR contains self inactivating deletion●●

3 packaging plasmids:3 packaging plasmids:●●

Gag/Gag/polpol,,●●

Rev, other HIV genesRev, other HIV genes●●

VSVVSV--G envelope proteinG envelope protein●●

Genes of interest cell growth and metastasisGenes of interest cell growth and metastasis●●

Tissue culture and in miceTissue culture and in mice


Yes Yes

NoOncogenic /

immunomodulatory PC2 NLRD DNIR

None of the above PC2 NLRD PC2 NLRD

Yes

Oncogenic / immunomodulatory DNIR PC2

NLRD DNIR

None of the above DNIR PC2 NLRD DNIR PC2

NLRD

Can enter

human cells

SIN ‘extra’ genes


Dealings

In vitro In vivo

2007 2011 2007 2011

Example 1Example 1AssessmentAssessment●●

Tissue culture dealings were DNIR, will Tissue culture dealings were DNIR, will become PC2 NLRDbecome PC2 NLRD

●●

Animal dealings still DNIRAnimal dealings still DNIR

●●

Partial downgrade for existing licencePartial downgrade for existing licence

●●

Or NLRD and licence for new projectOr NLRD and licence for new project


RD RD lentilenti

systemsystem●●


no self inactivating deletionno self inactivating deletion●●


gagpolgagpol,,●●

RevRev●●

HCV envelope proteinHCV envelope protein●●

Genes of interest mouse cytokines, and their Genes of interest mouse cytokines, and their receptors receptors --

known not to be active in humans known not to be active in humans

●●

Tissue culture & miceTissue culture & mice


Yes No No


Lenti DNIRPC2



PC2 NLRD

Lenti DNIRPC2

NLRDretro PC2 NLRD

retro PC2 NLRD

Yes No Yes Any DNIR

Able to enter

human cells



Dealings

In vitro In vivo

2007 2011 2007 2011


Tissue culture dealings DNIR to PC2 Tissue culture dealings DNIR to PC2 NLRDNLRD

●●

Animal dealings DNIR to PC2 NLRDAnimal dealings DNIR to PC2 NLRD

●●

Complete downgradeComplete downgrade

●●

Note: if not known whether the genes Note: if not known whether the genes are immunomodulatory in humans are immunomodulatory in humans then should assume they are and then should assume they are and assess accordinglyassess accordingly


RD RD lentilenti

systemsystem●●


no self inactivating deletionno self inactivating deletion●●


Gag/Gag/polpol●●

PolPol, other HIV genes, other HIV genes●●

Rev, other HIV genesRev, other HIV genes●●

Inducer for other gene expressionInducer for other gene expression●●

GM and wild type envelope proteins from human GM and wild type envelope proteins from human and animal virusesand animal viruses

●●

Marker genesMarker genes●●

Tissue culture, mice, rats and rabbitsTissue culture, mice, rats and rabbits


No Yes or no

Yes or no

Oncogenic modification

PC1 NLRD exempt DNIR PC2

NLRDPathogenic determinant PC2 NLRD PC2 NLRD

None of the above Exempt PC2 NLRD

VectorDonor

nucleic acid (transgene)

Dealings

Able to enter

human cells

SIN Extra genes

In vitro In vivo

2007 2011 2007 2011

Unable to transduce human cells


Yes No No


Lenti DNIRPC2



PC2 NLRD

Lenti DNIRPC2

NLRDretro PC2 NLRD

retro PC2 NLRD

Yes No Yes Any DNIR

Able to enter

human cells



Dealings

In vitro In vivo

2007 2011 2007 2011


Tissue culture dealings with envelope genes Tissue culture dealings with envelope genes that canthat can’’t transduce human cells, still t transduce human cells, still exemptexempt

●●

Animal dealings with envelope genes that Animal dealings with envelope genes that cancan’’t transduce human cells, still PC2 NLRDt transduce human cells, still PC2 NLRD

●●

All dealings with envelope genes that can All dealings with envelope genes that can transduce human cells, still DNIRtransduce human cells, still DNIR

●●

No changeNo change

SummarySummary●●

Exempt virus + oncogene Exempt virus + oncogene in vitroin vitro = exempt= exempt●●

In vitroIn vitro dealings with replication defective dealings with replication defective nonnon--retroviral vectors able to transduce retroviral vectors able to transduce human cells now PC2 NLRD, except human cells now PC2 NLRD, except HAdHAd

or or

AAVAAV●●

Tissue culture dealings with replication Tissue culture dealings with replication defective lentis systems that are SIN defective lentis systems that are SIN oror

‘‘essential genes onlyessential genes only’’

are now PC2 NLRDsare now PC2 NLRDs●●

Animal dealings with replication defective Animal dealings with replication defective lentis systems that are SIN lentis systems that are SIN oror

‘‘essential essential

genes onlygenes only’’

are now PC2 NLRDs are now PC2 NLRDs unless unless oncogenic or immumodulatory in humansoncogenic or immumodulatory in humans

ResourcesResources

ResourcesResourcesOn website:On website:

●●

Gene Technology Amendment Regulations 2011Gene Technology Amendment Regulations 2011●●

Updated viral vector tableUpdated viral vector table●●

Updated changes to dealings with viral vectors Updated changes to dealings with viral vectors tables (was appendix 1 in discussion paper 3)tables (was appendix 1 in discussion paper 3)

●●

New viral vector classification flow chartNew viral vector classification flow chartContact usContact us

●●

OGTR inbox OGTR inbox [email protected]@health.gov.au●●

Free call 1800 181 030Free call 1800 181 030

mailto:[email protected]

CopyrightCopyright©© Office of the Gene Technology Regulator 2011.Office of the Gene Technology Regulator 2011.

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Changes to the classification of Viral Vectors · Changes to the classification of Viral Vectors...

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