11/9/156 (Item 7 from file: 73)

10769043 EMBASE No: 2000247623

Intravascular and endobronchial DNA delivery to murine lung tissue using

a novel, nonviral vector

Kukowska‑Latallo J.F.; Raczka E.; Quintana A.; Chen C.; Rymaszewski M.;

Baker J.R. Jr.

Dr. J.R. Baker Jr., Department of Internal Medicine, 9220 MSRB III, Univ.

of Michigan Medical School, Ann Arbor, MI 48109‑0648 United States

Human Gene Therapy ( HUM. GENE THER. ) ( United States ) 01 JUL 2000 ,

11/10 (1385‑1395)

CODEN: HGTHE ISSN: 1043‑0342

Document Type: Journal ; Article

Language: ENGLISH Summary Language: ENGLISH

Number Of References: 59

Gene transfer to the lung can be achieved via either the airway or the

pulmonary vasculature. We evaluated gene transfer and expression by

intravascular and endobronchial routes, using DNA complexed with G9 PAMAM

dendrimer or naked plasmid DNA. Intravascular tail vein delivery of

dendrimer‑complexed pCF1CAT plasmid resulted in high levels of transgene

expression in the lung at 12 and 24 hr, followed by a second peak of

expression 3 to 5 days after administration. After intravenous

administration of the complexes, CAT expression was never observed in

organs other than the lung. There were only minimal levels of CAT protein

expressed in the lung after intravenous administration of naked plasmid

DNA. Repeated intravascular doses of the dendrimer‑complexed plasmid,

administered four times at 4‑day intervals, maintained expression at

15‑25% of peak concentrations achieved after the initial dose.

Endobronchial delivery of naked pCF1CAT plasmid produced significant

amounts of CAT protein in the lung. Comparison of intratracheal and

intranasal routes resulted in similar expression levels of CAT in the

lung and trachea. However, in juxtaposition to vascular delivery,

intranasal delivery of dendrimer‑complexed plasmid DNA gave lower levels

of CAT expression than that observed with naked plasmid DNA. In situ

localization of CAT enzymatic activity suggested that vascular

administration seemed to achieve expression in the lung parenchyma,

mainly within the alveoli, while endobronchial administration primarily

targeted bronchial epithelium. Our results show that intravenously

administered G9 dendrimer is an effective vector for pulmonary gene

transfer and that transgene expression can be prolonged by repeated

administration of dendrimer‑complexed DNA.

DRUG DESCRIPTORS:

dendrimer; transmembrane conductance regulator; alpha 1 antitrypsin;

liposome

MEDICAL DESCRIPTORS:

* virus vector; *lung parenchyma; *gene transfer; *lung

disease‑‑therapy‑‑th

gene expression; protein expression; enzyme activity; enzyme linked

immunosorbent assay; histochemistry; transgene; plasmid; gene therapy;

nonhuman; mouse; animal experiment; animal cell; article

CAS Registry Number: 9041‑92‑3 (alpha 1 antitrypsin)

Drug Descriptors:

022 Human Genetics

reserved.

11/9/5 (Item 5 from file: 155)

10533454 20360072

Enhanced gene expression in mouse lung after PEI‑DNA aerosol delivery.

Gautam A; Densmore CL; Xu B; Waldrep JC

Department of Molecular Physiology and Biophysics, Baylor College of

Medicine, Houston, Texas 77030, USA.

Mol Ther ( UNITED STATES ) Jul 2000 , 2 (1) p63‑70 , ISSN

1525‑0016 Journal Code: DRT

Languages: ENGLISH

Document type: JOURNAL ARTICLE

Journal Announcement: 0011

Subfile: INDEX MEDICUS

Aerosol gene delivery to the pulmonary system has vast potential for

many diseases, including cystic fibrosis and lung cancer. We recently

reported that polyethyleneimine (PEI), a cationic polymer, holds promise

as a gene delivery vector for transfection in lung by aerosol. To further

optimize the gene expression in the lung by aerosol, we utilized 5% CO(2)

in air for the nebulization of PEI‑DNA complexes. Five percent

CO(2)‑in‑air gave a threefold higher gene expression compared to normal

air using the chloramphenicol acetyl transferase (CAT) reporter gene

delivered by Aerotech II nebulizer. The delivery of DNA by PEI was dose

dependent with the highest expression obtained when 2 mg of DNA in 10 ml

was nebulized at a PEI nitrogen:DNA phosphate (N:P) ratio of 10:1. The

optimal N:P ratio for lung transfection was found to be between 10:1 and

20:1 using the CAT and luciferase reporter genes. The time‑course studies

showed the highest expression at 24 h after aerosol delivery and 40‑50%

of peak level was detectable even after a week. Tissue distribution

indicates the expression to be specific to the lung with no detectable

expression in any other tissue examined. Histological and biochemical

analysis of lungs revealed no evidence of acute inflammation.

Tags: Animal; Female; Support, Non‑U.S. Gov't

Descriptors: *Aerosols; *Gene Therapy‑‑Methods‑‑MT; *Genetic

Vectors‑‑Administration and Dosage‑‑AD; *Lung‑‑Metabolism‑‑ME;

*Polyethyleneimine ; Carbon Dioxide‑‑Metabolism‑‑ME; Chloramphenicol

O‑Acetyltransferase ‑‑Metabolism‑‑ME; Dose‑Response Relationship, Drug;

DNA‑‑Administration and Dosage‑‑AD; Enzyme‑Linked Immunosorbent Assay;

Gene Expression; Luciferase ‑‑Metabolism‑‑ME; Lung‑‑Pathology‑‑PA; Mice;

Mice, Inbred BALB C; Peroxidase‑‑Metabolism‑‑ME;

Plasmids‑‑Metabolism‑‑ME; Time Factors; Tissue Distribution; Transgenes

CAS Registry No.: 0 (Aerosols); 0 (Genetic Vectors); 0 (Plasmids);

124‑38‑9 (Carbon Dioxide); 9002‑98‑6 (Polyethyleneimine); 9007‑49‑2 (DNA)

Enzyme No.: EC 1.11.1.7 (Peroxidase); EC 1.13.12.‑ (Luciferase); EC

2.3.1.28 (Chloramphenicol O‑Acetyltransferase)

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11/9/20 (Item 20 from file: 155)

10407316 20262156

Gene transfer to adult human lung tissue ex vivo.

McBride S; Rannie D; Harrison DJ

CRC Laboratories, Pathology Department, University of Edinburgh Medical

School, Teviot Place, Edinburgh EH8 9AG, Scotland, UK.

Gene therapy ( ENGLAND ) Apr 2000 , 7 (8) p675‑8 , ISSN 0969‑7128

Journal Code: CCE

Languages: ENGLISH

Document type: JOURNAL ARTICLE

Journal Announcement: 0008

Subfile: INDEX MEDICUS

The potential of gene therapy for treatment of lung disease remains

unrealised. Early model systems often resulted in promising efficiency of

gene transfer, only to prove irreproducible in the clinic. While problems

such as induction of host immune responses and duration of expression

also need to be addressed, it is now widely believed that alternative,

relevant models which more accurately reflect gene transfer efficiencies

in human lungs are urgently required. We report here on a human lung

slice culture system to assess gene transfer to adult lung epithelium. A

lacZ‑expressing adenovirus (AdCA35lacZ) was used as a reporter vector. A

solution of AdCA35lacZ was instilled via bronchioles into resected lung

tissue, a route analogous to clinical administration. Following a 1 h

incubation, the tissue was inflated with a 0.4% agarose solution,

instilled via the same bronchioles. Once solidified, 500 &mgr;m slices of

the tissue were prepared and cultured for 4 days. beta‑Galactosidase

staining revealed lacZ transgene expression in bronchiolar and alveolar

cells of the lung slices throughout the 4 days in culture. This system,

which can also be used to study other viral and liposome vectors, could

prove to be a useful alternative model for assessing gene delivery to

adult human lung epithelium.

Tags: Animal; Human; Support, Non‑U.S. Gov't

Descriptors: *Adenoviridae‑‑Genetics‑‑GE; *Gene Therapy‑‑Methods‑‑MT;

*Gene Transfer; *Genetic Vectors‑‑Administration and Dosage‑‑AD; *Lung ;

Adult; Epithelium; Gene Expression; Lac Operon; Mice; Tissue Culture

CAS Registry No.: 0 (Genetic Vectors)

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rights reserved.

11/9/21 (Item 21 from file: 155)

10400993 20220616

Cationic lipid structure and formulation considerations for optimal

gene transfection of the lung.

Marshall J; Nietupski JB; Lee ER; Siegel CS; Rafter PW; Rudginsky SA;

Chang CD; Eastman SJ; Harris DJ; Scheule RK; Cheng SH

Genzyme Corporation, Framingham, MA 01701‑9322, USA.

john.marshall@genzyme.com

Journal of drug targeting ( SWITZERLAND ) 2000 , 7 (6) p453‑69 ,

ISSN 1061‑186X Journal Code: B3S

Languages: ENGLISH

Document type: JOURNAL ARTICLE

Journal Announcement: 0008

Subfile: INDEX MEDICUS

Enhanced gene transduction to the lung using cationic lipids could be

attained through optimization of the structure of the lipids and the

formulation of the cationic lipid:plasmid DNA (pDNA) complexes. We have

expanded on our earlier observation of the importance of the structural

orientation of the cationic lipid headgroup. Through the synthesis of a

number of matched pairs of cationic lipids differing only in the

configuration of their headgroup, we confirmed that those harboring a

T‑shape headgroup are more active than their linear counterparts, at

least when tested in the lungs of BALB/c mice. Additionally, we

demonstrated that not only are the structural considerations of these

cationic lipids important, but also their protonation state, the free

base being invariably more active than its salt counterpart. The salt

forms of cationic lipids bound pDNA with greater avidity, which may have

affected their subsequent intracellular dissolution and transit of the

pDNA to the nucleus. Inclusion of a number of frequently used solutes in

the vehicle severely inhibited the gene transfection activity of the

cationic lipids. The selection of neutral co‑lipids was also an important

factor for overall transfection activity of the formulation, with

significant gains in transfection activity realized when

diphytanoylphosphatidylethanolamine or

dilinoleoylphosphatidylethanolamine were used in lieu of

dioleoylphosphatidylethanolamine. Finally, we showed that a

transacylation reaction could occur between the cationic lipid and

neutral co‑lipid which reduced the transfection activity of the

complexes. It is the hope that as our understanding of the many factors

that influence the activity of these cationic lipid:pDNA complexes

improves, formulations with much greater potency can be realized for use

in the treatment of pulmonary diseases.

Tags: Animal; Female

Descriptors: *Gene Therapy; *Lipids‑‑Administration and Dosage‑‑AD;

*Lung‑‑Metabolism ‑‑ME; *Transfection ; Drug Stability;

Excipients‑‑Pharmacology‑‑PD; Lipids‑‑Chemistry‑‑CH; Mice; Mice, Inbred

BALB C

CAS Registry No.: 0 (Excipients); 0 (Lipids)

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rights reserved.

11/9/41 (Item 41 from file: 155)

09954356 99316336

Repeated administration of adenoviral vectors in lungs of human CD4

transgenic mice treated with a nondepleting CD4 antibody.

Chirmule N; Truneh A; Haecker SE; Tazelaar J; Gao Gp; Raper SE; Hughes

JV; Wilson JM

Institute for Human Gene Therapy, Department of Medicine, University of

Pennsylvania, Wistar Institute, Philadelphia, PA 19104, USA.

Journal of immunology ( UNITED STATES ) Jul 1 1999 , 163 (1)

p448‑55 , ISSN 0022‑1767 Journal Code: IFB

Contract/Grant No.: P30DK47757‑06, DK, NIDDK; P50DK49136‑05, DK,

NIDDK

Languages: ENGLISH

Document type: JOURNAL ARTICLE

Journal Announcement: 9909

Subfile: AIM; INDEX MEDICUS

The central role of CD4+ T cells in regulation of adenovirus

vector‑mediated immune responses has been documented previously in murine

models. We analyzed the effects of a nondepleting mAb to human CD4 (CD4

mAb; Clenoliximab) on immune functions following intratracheal

administration of adenoviral vectors in murine CD4‑deficient mice

(muCD4KO) expressing a human CD4 transgene (HuCD4 mice). Treatment of

HuCD4 mice with Clenoliximab inhibited both cell‑mediated and humoral

immune responses to adenoviral Ags. Chronic treatment of HuCD4 mice with

Clenoliximab permitted successful readministration of adenoviral vectors

at least four times. The ability to readminister these vectors is

associated with marked suppression of neutralizing Ab responses to viral

capsid proteins. Clenoliximab also inhibited CTL and prolonged expression

of the transgene. T or B cell responses to adenovirus did not emerge

after the effects of a short course of Clenoliximab diminished. These

data illustrate the potential utility of a nondepleting CD4 Ab in

facilitating gene therapy using adenoviral vectors.

Tags: Animal; Female; Human; Male; Support, Non‑U.S. Gov't; Support,

U.S. Gov't, P.H.S.

Descriptors: *Adenoviridae‑‑Genetics‑‑GE; *Antibodies,

Monoclonal‑‑Administration and Dosage‑‑AD; *Antigens, CD4‑‑Genetics‑‑GE;

*Antigens, CD4‑‑Immunology‑‑IM; *Genetic Vectors‑‑Administration and

Dosage‑‑AD; *Lung‑‑Immunology‑‑IM ; Adenoviridae‑‑Immunology‑‑IM;

Antibodies, Monoclonal‑‑Therapeutic Use‑‑TU; Cystic

Fibrosis‑‑Genetics‑‑GE; Cystic Fibrosis‑‑Immunology‑‑IM; Cystic

Fibrosis‑‑Therapy‑‑TH; CD4‑Positive T‑Lymphocytes‑‑Immunology‑‑IM;

CD8‑Positive T‑Lymphocytes‑‑Immunology‑‑IM; Gene Therapy‑‑Methods‑‑MT;

Genetic Vectors‑‑Immunology‑‑IM; Injections, Intraperitoneal; Intubation,

Intratracheal; Lung‑‑Metabolism‑‑ME; Lymphocyte Depletion; Lymphocyte

Transformation‑‑Genetics‑‑GE; Mice; Mice, Inbred C57BL; Mice, Inbred DBA;

Mice, Knockout; Mice, Transgenic; Th1 Cells‑‑Metabolism‑‑ME; Th2 Cells

‑‑Metabolism‑‑ME

CAS Registry No.: 0 (Antibodies, Monoclonal); 0 (Antigens, CD4); 0

(Genetic Vectors)

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rights reserved.

11/9/110 (Item 110 from file: 155)

08826974 96432153

Systematic analysis of repeated gene delivery into animal lungs with a

recombinant adenovirus vector.

Dong JY; Wang D; Van Ginkel FW; Pascual DW; Frizzell RA

Department of Laboratory Medicine, University of California, San

Francisco 94143‑0724, USA.

Human gene therapy ( UNITED STATES ) Feb 10 1996 , 7 (3) p319‑31 ,

ISSN 1043‑0342 Journal Code: A12

Contract/Grant No.: DK/HL46177, DK, NIDDK; DK38518, DK, NIDDK;

CA54430, CA, NCI

Languages: ENGLISH

Document type: JOURNAL ARTICLE

Journal Announcement: 9702

Subfile: INDEX MEDICUS

Adenovirus‑based vectors are promising candidates for genetic therapy

of cystic fibrosis (CF). Because adenoviruses naturally infect airway

cells, they grow to very high titers, and the transgenes carried by the

adenoviruses are expressed at high levels. In addition, adenoviruses are

relatively safe because the disease caused by the wild‑type virus is

self‑limiting. One disadvantage of adenovirual vectors is that the

transgene expression would be transient because adenoviruses do not

integrate their DNA into the genome of the host cells. Adenoviral gene

delivery into the lungs is also complicated by the anatomy of the airways

and the defense mechanisms of the recipient. To assess the feasibility of

adenovirus‑mediated gene therapy for CF, a recombinant adenovirus

carrying a lacZ gene was delivered into animal lungs to study the

efficiency and cellular distribution of gene transfer, the duration of

gene expression, the possible histopathology of the lungs after gene

transfer, and the efficacy of repeated administrations of the viral

agent. The results of these studies demonstrate that (i) efficient gene

transfer into animal lungs can be achieved; (ii) a near‑homogenous

delivery of the vectors can be achieved by airway instillation, although

the pattern of transduction varies among individual animals; (iii)

pathological effects are generally mild in CD1 mice; (iv) gene expression

is transient; (v) repetitive gene transfer is achievable, but becomes

progressively less efficient, and (vi) immune responses are induced

against both the viral and transgene products.

Tags: Animal; Female; Male; Support, Non‑U.S. Gov't; Support, U.S.

Gov't, P.H.S.

Descriptors: *beta‑Galactosidase‑‑Genetics‑‑GE;

*Adenoviridae‑‑Genetics‑‑GE; *Defective Viruses‑‑Genetics‑‑GE; *DNA,

Recombinant‑‑Administration and Dosage‑‑AD; *Gene Therapy‑‑Methods‑‑MT;

*Genetic Vectors‑‑Genetics‑‑GE; *Lung; *Recombinant Fusion

Proteins‑‑Biosynthesis‑‑BI ; beta‑Galactosidase‑‑Biosynthesis‑‑BI;

Adenoviridae‑‑Pathogenicity‑‑PY; Administration, Intranasal; Defective

Viruses‑‑Pathogenicity‑‑PY; DNA, Recombinant‑‑Genetics‑‑GE;

Epithelium‑‑Metabolism‑‑ME; Feasibility Studies; Gene Expression

Regulation, Viral; Genes, Reporter; Genetic Vectors ‑‑Toxicity‑‑TO;

Instillation, Drug; Lung‑‑Injuries‑‑IN; Lung‑‑Metabolism ‑‑ME;

Lung‑‑Pathology‑‑PA; Mice; Pneumonia, Viral‑‑Etiology‑‑ET; Time Factors;

Tissue Distribution; Trachea

CAS Registry No.: 0 (DNA, Recombinant); 0 (Genetic Vectors); 0

(Recombinant Fusion Proteins)

Enzyme No.: EC 3.2.1.23 (beta‑Galactosidase)

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11/9/46 (Item 46 from file: 155)

09916442 99261922

Effect of immune response on gene transfer to the lung via systemic

administration of cationic lipidic vectors.

Li S; Wu SP; Whitmore M; Loeffert EJ; Wang L; Watkins SC; Pitt BR;

Huang L

Department of Pharmacology, University of Pittsburgh School of

Medicine, Pittsburgh, Pennsylvania 15261, USA.

American journal of physiology ( UNITED STATES ) May 1999 , 276 (5

Pt 1) pL796‑804 , ISSN 0002‑9513 Journal Code: 3U8

Contract/Grant No.: CA‑59327, CA, NCI; CA‑64654, CA, NCI; CA‑71731,

CA, NCI; +

Languages: ENGLISH

Document type: JOURNAL ARTICLE

Journal Announcement: 9908

Subfile: INDEX MEDICUS

Cationic lipid‑mediated intravenous gene delivery shows promise in

treating pulmonary diseases including lung tumor metastases, pulmonary

hypertension, and acute respiratory distress syndrome. Nevertheless,

clinical applications of cationic lipidic vectors via intravenous

administration are limited by their transient gene expression. In

addition, repeated dosing is not effective at frequent intervals. In an

effort to elucidate the mechanism of gene inactivation, we report in this

study that cationic lipid‑protamine‑DNA (LPD) complexes, but not each

component alone, can induce a high level of cytokine production,

including interferon‑gamma and tumor necrosis factor‑alpha. Furthermore,

we demonstrate that LPD administration triggers apoptosis in the lung, a

phenomenon that may be mediated in part by the two cytokines. Treatment

of mice with antibodies against the two cytokines prolongs the duration

of gene expression and also improves lung transfection on a second

administration of LPD. Although the mechanism underlying LPD‑induced

cytokine production is unclear, methylation of the DNA significantly

decreased the level of both interferon‑gamma and tumor necrosis

factor‑alpha, suggesting that unmethylated CpG sequences in plasmid DNA

play an important role. These data suggest that decreasing the

CpG‑mediated immune response while not affecting gene expression may be a

useful therapeutic strategy to improve cationic lipid‑mediated

intravenous gene delivery to the lung.

Tags: Animal; Female; Support, Non‑U.S. Gov't; Support, U.S. Gov't,

P.H.S.

Descriptors: *Cytokines‑‑Immunology‑‑IM; *DNA‑‑Immunology‑‑IM; *Gene

Transfer; *Genetic Vectors; *Lipids; *Lung‑‑Metabolism‑‑ME ;

Antibodies‑‑Pharmacology‑‑PD; Apoptosis; Cations; DNA‑‑Administration and

Dosage‑‑AD; DNA Methylation; Gene Expression; Immunization, Passive;

Injections, Intravenous; Interferon Type II‑‑Immunology‑‑IM; Lung

Diseases ‑‑Therapy‑‑TH; Mice; Plasmids‑‑Genetics‑‑GE; Tumor Necrosis

Factor ‑‑Immunology‑‑IM

CAS Registry No.: 0 (Antibodies); 0 (Cations); 0 (Cytokines); 0 (Genetic

Vectors); 0 (Lipids); 0 (Plasmids); 0 (Tumor Necrosis Factor); 82115‑62‑6

(Interferon Type II); 9007‑49‑2 (DNA)

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rights reserved.

11/9/153 (Item 4 from file: 73)

10905960 EMBASE No: 2000393474

Biodistribution and transgene expression with nonviral cationic

vector/DNA complexes in the lungs

Bragonzi A.; Dina G.; Villa A.; Calori G.; Biffi A.; Bordignon C.; Assael

B.M.; Conese M.

M. Conese, Inst. for Experimen. Cystic Fibrosis, San Raffaele Scientific

Institute, Via Olgettina 58, Milano 20132 Italy

Gene Therapy ( GENE THER. ) ( United Kingdom ) 2000 , 7/20 (1753‑1760)

CODEN: GETHE ISSN: 0969‑7128

Document Type: Journal ; Article

Language: ENGLISH Summary Language: ENGLISH

Number Of References: 36

Biodistribution of nonviral cationic vector/DNA complexes was studied

after systemic or intratracheal administration to the lungs and

correlated with transgene expression. Intravenous injection in C57BI/6

mice gave maximal and significant luciferase expression in the lungs with

the cationic polymer PEI 22K/DNA complexes at the highest ratios of

positive/negative charges versus DNA alone. While DOTAP/DNA complexes

with high charge ratio determined lower but still significant luciferase

activity versus uncomplexed DNA, GL‑67A and PEI 25K mediated negligible

luciferase expression. Labelled PEI 22K and DOTAP complexes were evenly

distributed in the alveolar region, where GFP expression was revealed,

while PEI 25K and GL‑67A complexes were not detected, suggesting a

different interaction of these complexes with the plasma membrane of

endothelial cells. Following an intratracheal injection, the highest and

significant levels of transfection were obtained with slightly positive

PEI complexes as compared with DNA alone, whereas cationic lipid‑based

vectors. DOTAP and GL‑67A, gave not significant luciferase activity. Both

types of polyplexes gave similar levels of lung luciferase expression by

targeting different airway cell populations. PEI 25K complexes determined

high levels of GFP in the bronchial cells, confirming confocal data on

fluorescent complexes internalization. PEI 22K complexes gave mainly high

GFP signal in the distal tract of the bronchial tree, where tagged

complexes were recovered. Fluorescent lipid complexes were found in

aggregates in the lumen of bronchi totally (DOTAP) or partially (GL‑67A)

co‑localizing with surfactant protein A. Results indicated that cationic

polymers could overcome the surfactant barrier which inhibited airway

cell transfection mediated by cationic lipids.

Brand Name/Manufacturer Name: Exgen 500/Euromedex/France

Manufacturer Names: Euromedex/France; Sigma Aldrich/United States;

Hoffmann La Roche/Germany; Genzyme/United States

DRUG DESCRIPTORS:

* DNA‑‑intrathecal drug administration‑‑tl; *DNA‑‑intravenous drug

administration‑‑iv; *polyethyleneimine‑‑intrathecal drug

administration‑‑tl ; *polyethyleneimine‑‑intravenous drug

administration‑‑iv

luciferase‑‑endogenous compound‑‑ec; cation‑‑intrathecal drug

administration‑‑tl; cation‑‑intravenous drug administration‑‑iv;

liposome; macrogol; green fluorescent protein; surfactant protein

A‑‑endogenous compound‑‑ec

MEDICAL DESCRIPTORS:

* lung; *transgene

complex formation; gene expression; mouse strain; protein expression;

enzyme activity; tissue distribution; lung alveolus; cell membrane;

endothelium cell; genetic transfection; expression vector; gene

targeting; cell population; bronchus; gene transfer; human; nonhuman;

mouse; animal experiment; animal model; controlled study; human cell;

animal tissue; article; priority journal

Drug Terms (Uncontrolled): Exgen 500

CAS Registry Number: 9007‑49‑2 (DNA); 74913‑72‑7 (polyethyleneimine);

61970‑00‑1, 9014‑00‑0 ( luciferase); 25322‑68‑3 (macrogol)

Drug Descriptors:

015 Chest Diseases, Thoracic Surgery and Tuberculosis

022 Human Genetics

037 Drug Literature Index

reserved

11/9/47 (Item 47 from file: 155)

09910534 99041640

Direct gene transfer to the respiratory tract of mice with pure plasmid

and lipid‑formulated DNA.

McCluskie MJ; Chu Y; Xia JL; Jessee J; Gebyehu G; Davis HL

Loeb Research Institute, Ottawa, Canada.

Antisense & nucleic acid drug development ( UNITED STATES ) Oct 1998

, 8 (5) p401‑14 , ISSN 1087‑2906 Journal Code: CJY

Languages: ENGLISH

Document type: JOURNAL ARTICLE

Journal Announcement: 9908

Subfile: INDEX MEDICUS

Direct gene transfer into the respiratory system could be carried out

for either therapeutic or immunization purposes. Here we demonstrate that

cells in the lung can take up and express plasmid DNA encoding a

luciferase reporter gene whether it is administered in naked form or

formulated with cationic liposomes. Depending on the lipid used, the

transfection efficiency with liposome‑formulated DNA may be higher, the

same as, or less than that with pure plasmid DNA.

Tetramethyltetraalkylspermine analogs with alkyl groups of 16 or 18

carbons and DMRIE/cholesterol formulations proved particularly effective.

Similar results for reporter gene expression in the lung were obtained

whether the DNA (naked or lipid formulated) was administered by indirect,

noninvasive intranasal delivery (inhaled or instilled) or by invasive,

direct intratracheal delivery (injected or via a cannula). Reporter gene

expression peaks around 4 days, then falls off dramatically by 9 days.

The dose‑response is linear, at least up to 100 microg plasmid DNA,

suggesting better transfection efficiencies might be realized if there

was not a volume limitation. For a given dose of DNA, the best results

are obtained when the DNA is mixed with the minimum amount of lipid that

can complex it completely. These results are discussed in the context of

direct gene transfer for either gene therapy or delivery of a mucosal DNA

vaccine.

Tags: Animal; Support, Non‑U.S. Gov't

Descriptors: *DNA‑‑Metabolism‑‑ME; *Gene Transfer;

*Liposomes‑‑Metabolism‑‑ME; *Plasmids ‑‑Genetics‑‑GE; *Respiratory

System‑‑Metabolism‑‑ME ; beta‑Galactosidase‑‑Genetics‑‑GE;

beta‑Galactosidase‑‑Immunology‑‑IM; beta‑Galactosidase‑‑Metabolism‑‑ME;

Administration, Inhalation; Antibody Formation‑‑Immunology‑‑IM;

Catheterization, Peripheral; Dose‑Response Relationship, Immunologic;

DNA‑‑Administration and Dosage‑‑AD; DNA ‑‑Chemistry‑‑CH;

DNA‑‑Genetics‑‑GE; Gene Expression‑‑Genetics‑‑GE; Genes,

Reporter‑‑Genetics‑‑GE; Injections; Interferon Type II‑‑Genetics‑‑GE; Lac

Operon‑‑Genetics‑‑GE; Liposomes‑‑Chemistry‑‑CH;

Liposomes‑‑Immunology‑‑IM; Luciferase‑‑Genetics‑‑GE;

Luciferase‑‑Metabolism‑‑ME; Lung‑‑Immunology‑‑IM ; Lung‑‑Metabolism‑‑ME;

Mice; Mice, Inbred BALB C; Mice, Knockout; Plasmids‑‑Administration and

Dosage‑‑AD; Plasmids‑‑Immunology‑‑IM; Respiratory System‑‑Immunology‑‑IM

CAS Registry No.: 0 (Plasmids); 82115‑62‑6 (Interferon Type II);

9007‑49‑2 (DNA)

Enzyme No.: EC 1.13.12.‑ (Luciferase); EC 3.2.1.23 (beta‑Galactosidase)

MEDLINE(R) (Dialog« File 155): (c) format only 2000 Dialog Corporation. All

rights reserved.

11/9/51 (Item 51 from file: 155)

09842414 99145145

Contribution of plasmid DNA to inflammation in the lung after

administration of cationic lipid:pDNA complexes.

Yew NS; Wang KX; Przybylska M; Bagley RG; Stedman M; Marshall J;

Scheule RK ; Cheng SH

Genzyme Corporation, Framingham, MA 01701‑9322, USA.

Human gene therapy ( UNITED STATES ) Jan 20 1999 , 10 (2) p223‑34 ,

ISSN 1043‑0342 Journal Code: A12

Languages: ENGLISH

Document type: JOURNAL ARTICLE

Journal Announcement: 9906

Subfile: INDEX MEDICUS

Cationic lipid‑mediated gene transfer to the mouse lung induces a

dose‑dependent inflammatory response that is characterized by an influx

of leukocytes and elevated levels of the cytokines interleukin 6 (IL‑6),

tumor necrosis factor alpha (TNF‑alpha), and interferon gamma

(IFN‑gamma). We have examined the contribution of plasmid DNA (pDNA) to

this observed toxicity, specifically the role of unmethylated CpG

dinucleotides, which have been previously shown to be immunostimulatory.

We report here that complexes of cationic lipid GL‑67 and unmethylated

pDNA (pCF1‑CAT) instilled into the lungs of BALB/c mice induced highly

elevated levels of the cytokines TNF‑alpha, IFN‑gamma, IL‑6, and IL‑12 in

the bronchoalveolar lavage fluids (BALF). In contrast, BALF of animals

administered either GL‑67 alone or GL‑67 complexed with SssI‑methylated

pDNA contained low levels of these cytokines. Similar results were

observed using a plasmid (pCF1‑null) that does not express a transgene,

demonstrating that expression of chloramphenicol acetyltransferase (CAT)

was not responsible for the observed inflammation. The response observed

was dose dependent, with animals receiving increasingly higher amounts of

unmethylated pDNA exhibiting progressively higher levels of the

cytokines. Concomitant with this increase in cytokine levels were also

elevated numbers of neutrophils in the BALF, suggesting a possible cause‑

and‑effect relationship between neutrophil influx and generation of

cytokines. Consistent with this proposal is the observation that

reduction of neutrophils in the lung by administration of antibodies

against Mac‑1alpha and LFA‑1 also diminished cytokine levels. This

reduction in cytokine levels in the BALF was accompanied by an increase

in transgene expression. In an attempt to abate the inflammatory

response, sequences in the pDNA encoding the motif RRCGYY, shown to be

most immunostimulatory, were selectively mutagenized. However,

instillation of a plasmid in which 14 of the 17 CpG sites were altered

into BALF/c mice did not reduce the levels of cytokines in the BALF

compared with the unmodified vector. This suggests that other

unmethylated motifs, in addition to RRCGYY, may also contribute to the

inflammatory response. Together, these findings indicate that

unmethylated CpG residues in pDNA are a major contributor to the

induction of specific proinflammatory cytokines associated with

instillation of cationic lipid:pDNA complexes into the lung. Strategies

to abate this response are warranted to improve the efficacy of this

nonviral gene delivery vector system for the treatment of chronic

diseases.

Tags: Animal

Descriptors: *DNA‑‑Administration and Dosage‑‑AD; *Plasmids;

*Pneumonia‑‑Genetics‑‑GE ; Bronchoalveolar Lavage Fluid; Cations; CpG

Islands; DNA‑‑Metabolism‑‑ME; DNA Methylation; Interferon Type

II‑‑Metabolism‑‑ME; Interleukin‑6 ‑‑Metabolism‑‑ME; Lung‑‑Metabolism‑‑ME;

Mice; Mice, Inbred BALB C; Neutrophils‑‑Cytology‑‑CY;

Pneumonia‑‑Metabolism‑‑ME; Tumor Necrosis Factor ‑‑Metabolism‑‑ME

CAS Registry No.: 0 (Cations); 0 (Interleukin‑6); 0 (Plasmids); 0 (Tumor

Necrosis Factor); 82115‑62‑6 (Interferon Type II); 9007‑49‑2 (DNA)

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