Targeted Gene Transfer Research Articles

Recent advances in various emerging vescicular systems: An overview

Liposomes have been widely investigated since 1970 as drug carriers for improving the delivery of therapeutic agents to specific sites in the body. As a result, numerous improvements have been made to make this technology potential the treatment of certain diseases in the clinics. This review mainly focused on various aspects related to the vesicular system, including method of preparation, stabilization, drawbacks, and applications. Various types of vesicular systems such as liposomes, niosomes, transfersomes, pharmacosomes, and nanoparticle have been discussed briefly along with some other emerging vescicular systems (photosomes, archaesomes, genosomes, cryptosomes, discomes) focusing on cell specific gene transfer, photodynamic therapy and ligand mediated drug targeting. Present applications of the liposomes are in the immunology, dermatology, vaccine adjuvant, eye disorders, brain targeting, infective disease and in tumour therapy. The new developments in this field are of specific binding properties of a drug-carrying liposome to a target cell such as a tumor cell and specific molecules in the body (antibodies, proteins, peptides etc), stealth liposomes which are especially used as carriers for hydrophilic (water soluble) anticancer drugs like doxorubicin, mitoxantrone and bisphosphonate-liposome mediated depletion of macrophages. This review would help researchers working in the area of liposomal drug delivery.

The combination of a synthetic promoter and a CMV promoter improves foreign gene expression efficiency in myocytes

Skeletal muscle is becoming an attractive target tissue for gene therapy. Nevertheless, the low level of gene therapeutic expression in this tissue is the major limitation to it becoming an ideal target for gene transfer. The promoter is important element for gene transcription; however, the gene expression efficiencies and specificities of viral promoters and skeletal muscle-specific promotors are in themselves limiting factors. In this study, we established a dual-promoters system in skeletal muscle using a cytomegalovirus (CMV) promoter and a skeletal muscle-specific synthetic promoter. Mouse myoblast cell line C2C12 cells were transfected with the system. We demonstrated that the dual-promoters system could significantly improve exogenous gene expression rate in vitro when compared with a single CMV promoter system and a skeletal muscle-specific synthetic promoter system in C2C12 cell line, by 69.48% and 41.93%, respectively. Next, we evaluated the system efficiency in vivo, the results showed that the dual-promoters system increased gene expression in mice 1.23-fold and 1.60-fold, respectively compared with expression controlled by the two single promoter vectors. Finally, we tested the dual-promoters system in growth hormone-releasing hormone (GHRH) gene therapy, and revealed that when these two promoters co-drove the GHRH gene expression in vivo animal growth was enhanced significantly. All these results indicate that use of the dual-promoter vector was more efficient for gene expression in skeletal muscle tissue than use of the single promoter vectors. These finding could, hopefully, lead to the development of a high efficiency expression system in myocytes and form an ideal approach for gene therapy.

Targeted gene delivery to the mouse brain by MRI-guided focused ultrasound-induced blood–brain barrier disruption

This study aimed to investigate the feasibility of targeted gene transfer into central nervous system (CNS) by MRI-guided focused ultrasound-induced blood–brain barrier (BBB) disruption. Before each sonication, T2-weighted images were obtained to select the target region. Followed by injecting DNA-loaded microbubbles into the tail vein, sonication was performed. The state of local BBB, distribution of plasmid DNA through the opened BBB, the ultrastructural changes of neurons and BDNF expression were detected. The results showed that MRI-guided focused ultrasound (FUS) could accomplish noninvasive, transient, and local BBB disruption, at 1h after sonication, plasmid DNA across the opened BBB had been internalized into the neurons presenting heterogeneous distribution and numerous transparent vesicles were observed in the cytoplasm of the neurons at the sonicated region, suggesting vesicle-mediated endocytosis. At 48h after sonication, the expressions of exogenous gene pBDNF-EGFP were observed in the cytoplasm of some neurons, and BDNF expressions were markedly enhanced by the combination of ultrasound and pBDNF-EGFP-loaded microbubbles about 20-fold than that of the control group (P<0.01). The method by using MRI-guided FUS to induce the local BBB disruption could accomplish effective targeted exogenous gene transfer in CNS. This technique may provide a new option for the treatment of various CNS diseases.

Ultrasound- and Liposome Microbubble-Mediated Targeted Gene Transfer to Cardiomyocytes In Vivo Accompanied by Polyethylenimine

Gene transfer to cardiomyocytes in vivo has received much research attention in the last decade but remains a substantial hurdle. Gene transfer using ultrasound-targeted microbubble destruction is a promising tool for gene therapy. Little data have shown the feasibility and optimization of this method for primary myocardial disease. In this study, we sought to determine the feasibility and efficiency of in vivo gene transfer to the myocardium mediated by ultrasound-targeted microbubble destruction accompanied by polyethylenimine. Three plasmids (luciferase reporter, red fluorescent protein reporter, and enhanced green fluorescent protein reporter) were used in this study. The ultrasound parameters were also optimized. A solution containing phosphate-buffered saline, a plasmid, plasmid complex, or polyethylenimine/plasmid, and liposome microbubbles was injected via a tail vein with (study) or without (control) transthoracic ultrasound irradiation. The efficiency of reporter gene transfer was determined by detection of luciferase activity or microscopy, and histologic investigations of the tissue specimens were performed. Ultrasound-targeted microbubble destruction significantly increased luciferase activity in vivo compared to plasmids and microbubbles alone (P < .001). More importantly, the increase in transgene expression was significantly related to ultrasound-targeted microbubble destruction in the presence of polyethylenimine (P < .001). In addition, fluorescein expression was present in all sections that received ultrasound-targeted microbubble destruction. The fluorescent reporter genes and luciferase plasmid all had similar results. Regardless of ultrasound exposure, expression in other organs was close to a background level except for the liver and lung. Hematoxylin-eosin staining showed no notable myocardial injury or death in control and treated mice. An atraumatic targeted gene delivery technique based on ultrasound-targeted microbubble destruction and polyethylenimine has been developed to transfect cardiomyocytes in vivo. If a suitable target gene is added, the novel technique could be highly effective in many kinds of heart disease.

Antibody-mediated targeted gene transfer of helper virus-free HSV-1 vectors to rat neocortical neurons that contain either NMDA receptor 2B or 2A subunits

Because of the numerous types of neurons in the brain, and particularly the forebrain, neuron type-specific expression will benefit many potential applications of direct gene transfer. The two most promising approaches for achieving neuron type-specific expression are targeted gene transfer to a specific type of neuron and using a neuron type-specific promoter. We previously developed antibody-mediated targeted gene transfer with Herpes Simplex Virus (HSV-1) vectors by modifying glycoprotein C (gC) to replace the heparin binding domain, which mediates the initial binding of HSV-1 particles to many cell types, with the Staphylococcus A protein ZZ domain, which binds immunoglobulin (Ig) G. We showed that a chimeric gC--ZZ protein is incorporated into vector particles and binds IgG. As a proof-of-principle for antibody-mediated targeted gene transfer, we isolated complexes of these vector particles and an anti-NMDA NR1 subunit antibody, and demonstrated targeted gene transfer to neocortical cells that contain NR1 subunits. However, because most forebrain neurons contain NR1, we obtained only a modest increase in the specificity of gene transfer, and this targeting specificity is of limited utility for physiological experiments. Here, we report efficient antibody-mediated targeted gene transfer to NMDA NR2B- or NR2A-containing cells in rat postrhinal cortex, and a neuron-specific promoter further restricted recombinant expression to neurons. Of note, because NR2A-containing neurons are relatively rare, these results show that antibody-mediated targeted gene transfer with HSV-1 vectors containing neuron type-specific promoters can restrict recombinant expression to specific types of forebrain neurons of physiological significance.

Gene Transfer in Respiratory Epithelium Using Synthetic Vectors and Folic Acid Receptor

Objective: Synthetic vectors may be an interesting alternative way compared with viral vectors. They are potentially less toxic and immunogenic. However, synthetic vectors use an aspecific electrostatic recognition. Cellular targeting is one of the major conditions for development of efficient synthetic vectors for gene therapy. Method: A neutral colipid derived from archaebacteria with acid folic ligand was developed and associated with cationic lipid vector. Its efficiency was tested on human respiratory epithelium cell culture. Results: Safe respiratory epithelium offer few targets for gene transfer. Indeed, using lectin for specific gene transfer is difficult because of numerous different oligosaccharides patterns. We studied the interest of folic acid receptor as a target for gene transfer. These receptors are sparsely represented on this epithelium but have a great affinity. We demonstrated a specific gene transfer using a competitive inhibition by free folic acid in media. For a healthy human respiratory cell line, this new vector is more efficient and less toxic than lipofectamin. Conclusion: Despite a small presence on human respiratory epithelium, folic acid receptor is a pertinent target for nonviral gene transfer. This vector is to be tested on animals. Such a targeting will increase gene transfer because of low electrostatic disturbance. Neutral vector may decrease toxicity.

Gene Transfer for Inherited Metabolic Disorders of the Liver: Immunological Challenges

Hepatocytes are a key target for gene transfer directed at correction of inborn errors of metabolism. The theoretical potential of hepatocyte-directed gene transfer contrasts with the hurdles for clinical translation of this technology. Innate immune responses following gene transfer are initiated by recognition of pathogen-associated molecular patterns by pattern recognition receptors like Toll-like receptors. Adaptive immune responses may constitute the most significant hurdle for efficient gene transfer. Besides the challenge imposed by adaptive immune responses against the vector and the potential problem of pre-existing immunity, immune responses against the transgene product may also constitute an obstacle. The liver is a tolerogenic organ. Naive T cells encounter liver antigens initially in the liver, rather than in lymphoid tissue. Lymph nodes and the spleen are anatomical compartments that provide a particular microarchitecture and microenvironment for the induction of immunity. In contrast, antigen presentation in the liver takes place in a completely different microarchitecture and microenvironment. This is a key aspect of the hepatic adaptive immune tolerance induction. Consistent with the tolerogenic nature of the liver microenvironment, the risk of antibody formation against the transgene product may be limited in the setting of hepatocyte-directed gene transfer and specifically by restricting transgene expression to hepatocytes by use of hepatocyte-specific expression cassettes. However, it is unclear to which extent animal experimental data following gene transfer predict immune responses in humans. Extrapolations from animals to humans are required but should be performed with sufficient insight into the dramatic species differences of the immune system.

Ultrasound and Liposome Microbubble-Mediated Targeted Gene Transfer to Cardiomyocytes In Vivo Accompanied by Polyethylenimine

Ultrasound and Liposome Microbubble-Mediated Targeted Gene Transfer to Cardiomyocytes In Vivo Accompanied by Polyethylenimine

Targeted Delivery in Breast Cancer Cells via Iodine: Nuclear Localization Sequence Conjugate

The nonviral vector with iodine-nuclear localization sequence (namely, NLS-I) targeting breast cancer cells was fabricated. Ternary complexes were formed via charge interactions among NLS-I peptides, PEI 1800, and DNA, and we investigated their cellular internalization, nuclear accumulation as well as transfection efficiency. All the experiments were assessed by employing MCF-7 cells that express sodium/iodide symporter and HeLa cells that lack the expression of the symporter. In MCF-7 cells, cell internalization and nuclear accumulation of NLS-I was markedly increased compared to that in NLS. In addition, compared to that of the PEI1800/DNA complex, PEI1800/DNA/NLS-I complexes exhibited much enhanced luciferase reporter gene expression by up to 130-fold. By contrast, in HeLa cells, the evident improvements of cellular internalization, nuclear accumulation, and transfection efficiency by NLS-I were not observed. This study demonstrates an alternative method to construct a nonviral delivery system for targeted gene transfer into breast cancer cells.

Oxidase-deficient neutrophils from X-linked chronic granulomatous disease iPS cells: functional correction by zinc finger nuclease–mediated safe harbor targeting

AbstractWe have developed induced pluripotent stem cells (iPSCs) from a patient with X-linked chronic granulomatous disease (X-CGD), a defect of neutrophil microbicidal reactive oxygen species (ROS) generation resulting from gp91phox deficiency. We demonstrated that mature neutrophils differentiated from X-CGD iPSCs lack ROS production, reproducing the pathognomonic CGD cellular phenotype. Targeted gene transfer into iPSCs, with subsequent selection and full characterization to ensure no off-target changes, holds promise for correction of monogenic diseases without the insertional mutagenesis caused by multisite integration of viral or plasmid vectors. Zinc finger nuclease–mediated gene targeting of a single-copy gp91phox therapeutic minigene into one allele of the “safe harbor” AAVS1 locus in X-CGD iPSCs without off-target inserts resulted in sustained expression of gp91phox and substantially restored neutrophil ROS production. Our findings demonstrate how precise gene targeting may be applied to correction of X-CGD using zinc finger nuclease and patient iPSCs.

345. Zinc Finger Nuclease Mediated Safe Harbor Targeted Gene Transfer in Patient iPSCs Functionally Corrects X-Linked Chronic Granulomatous Disease

345. Zinc Finger Nuclease Mediated Safe Harbor Targeted Gene Transfer in Patient iPSCs Functionally Corrects X-Linked Chronic Granulomatous Disease

Site-specific gene transfer into the rat spinal cord by photomechanical waves

Nonviral, site-specific gene delivery to deep tissue is required for gene therapy of a spinal cord injury. However, an efficient method satisfying these requirements has not been established. This study demonstrates efficient and targeted gene transfer into the spinal cord by using photomechanical waves (PMWs), which were generated by irradiating a black laser absorbing rubber with 532-nm nanosecond Nd:YAG laser pulses. After a solution of plasmid DNA coding for enhanced green fluorescent protein (EGFP) or luciferase was intraparenchymally injected into the spinal cord, PMWs were applied to the target site. In the PMW application group, we observed significant EGFP gene expression in the white matter and remarkably high luciferase activity only in the spinal cord segment exposed to the PMWs. We also assessed hind limb movements 24 h after the application of PMWs based on the Basso-Beattie-Bresnahan (BBB) score to evaluate the noninvasiveness of this method. Locomotor evaluation showed no significant decrease in BBB score under optimum laser irradiation conditions. These findings demonstrated that exogenous genes can be efficiently and site-selectively delivered into the spinal cord by applying PMWs without significant locomotive damage.

Targeted gene transfer into rat facial muscles by nanosecond pulsed laser-induced stress waves

We investigate the feasibility of using nanosecond pulsed laser-induced stress waves (LISWs) for gene transfer into rat facial muscles. LISWs are generated by irradiating a black natural rubber disk placed on the target tissue with nanosecond pulsed laser light from the second harmonics (532 nm) of a Q-switched Nd:YAG laser, which is widely used in head and neck surgery and proven to be safe. After injection of plasmid deoxyribose nucleic acid (DNA) coding for Lac Z into rat facial muscles, pulsed laser is used to irradiate the laser target on the skin surface without incision or exposure of muscles. Lac Z expression is detected by X-gal staining of excised rat facial skin and muscles. Strong Lac Z expression is observed seven days after gene transfer, and sustained for up to 14 days. Gene transfer is achieved in facial muscles several millimeters deep from the surface. Gene expression is localized to the tissue exposed to LISWs. No tissue damage from LISWs is observed. LISW is a promising nonviral target gene transfer method because of its high spatial controllability, easy applicability, and minimal invasiveness. Gene transfer using LISW to produce therapeutic proteins such as growth factors could be used to treat nerve injury and paralysis.

Cyclic RGD-poly(ethylene glycol)-polyethyleneimine is more suitable for glioblastoma targeting gene transfer in vivo

Arginine-glycine-aspartic acid (RGD) is a widely chosen ligand to improve the specific gene targeting transfection efficiency of polyethyleneimine (PEI) in vivo. However, the optimal RGD conjugating mode, RGD-poly(ethylene glycol)-PEI (RGD-PEG-PEI) or RGD-PEI-methoxyl poly(ethylene glycol) (RGD-PEI-mPEG) still remains controversial. In this study, RGD-PEG-PEI and RGD-PEI-mPEG were synthesized and compared with respects to their glioblastoma cell-binding capability and tumor-targeting ability of their complexes with plasmid DNA. These results demonstrated that RGD-PEG-PEI/plasmid enhanced green fluorescent protein (pEGFP)-N2 complexes had higher binding affinities with U87 cells than RGD-PEI-mPEG/pEGFP-N2 complexes. The gene transfection was also performed on U87 cells in vitro and in vivo. In vitro, both of the RGD-modified PEI derivatives enhanced the gene transfection efficiency to some extent. However, all of the complexes (with or without RGD modification) had high transfection efficiency. The biodistribution of RGD-PEG-PEI/pEGFP-N2 complexes in mice bearing subcutaneous glioblastomas were significantly greater than that of RGD-PEI-mPEG/pEGFP-N2 complexes, suggesting a more efficient gene transfection in vivo. In the RGD-PEG-PEI, the use of a PEG spacer was particularly important. These results indicated that RGD-PEG-PEI was more suitable for targeted gene transfer in vivo.

Novel Cytotoxic Vectors Based on Adeno-Associated Virus

Vectors based on adeno-associated virus (AAV) are promising tools for gene therapy. The production of strongly toxic vectors, for example for cancer-directed gene transfer, is often unfeasible due to uncontrolled expression of toxic genes in vector-producing cells. Using an approach based on transcriptional repression, we have created novel AAV vectors carrying the genes coding for diphtheria toxin A (DTA) and the pro-apoptotic PUMA protein. The DTA vector had a significant toxic effect on a panel of tumor cell lines, and abrogation of protein synthesis could be shown. The PUMA vector had a toxic effect on HeLa and RPMI 8226 cells, and sensitized transduced cells to doxorubicin. To permit targeted gene transfer, we incorporated the DTA gene into a genetically modified AAV-2 capsid previously developed by our group that mediates enhanced transduction of murine breast cancer cells in vitro. This vector had a stronger cytotoxic effect on breast cancer cells than DTA vectors with wildtype AAV capsid or vectors with a random capsid modification. The vector production and application system presented here allows for easy exchange of promotors, transgenes and capsid specificity for certain target cells. It will therefore be of great possible value in a broad range of applications in cytotoxic gene therapy and significantly broadens the spectrum of available tools for AAV-based gene therapy.

Antibody‐directed lentiviral gene transduction in early immature hematopoietic progenitor cells

The specific and efficient transduction of retroviral particles remains problematic for in vivo and ex vivo gene therapy studies, where the targeting cell population is a heterogeneous bulk population. Pseudotyping lentiviral particles with Sindbis virus envelope (Env) proteins modified with an immunoglobulin Fc-binding domain presents a method of selecting cells within a mixed population through antibody (Ab)-mediated targeting. Conditions were tested for targeted lentiviral gene delivery to hematopoietic progenitor cells via Ab-conjugated envelopes independent of CD34. Conditions to optimize the efficiency of gene delivery were established using the ABCG2 multidrug resistance protein, associated with stem cell phenotypes, as the cell surface target. By varying the proportion of ABCG2 expressing cells in a population, ABCG2-targeted gene delivery was detectable by flow cytometry when ABCG2(+) cells comprised greater than 5% of the population. Conditions that increased the efficiency of gene transfer, including cholesterol independent Env proteins and pH, increased nonspecific gene delivery. The feasibility of this cell-Ab-virus sandwich system in targeting transduction in a mixed population was tested in cells derived from human cord blood (CB). Conjugation of viral particles with anti-CD133 and anti-ABCG2 hematopoietic stem cell-associated Ab resulted in targeted gene transfer into early immature hematopoietic progenitor cells. Enhancement was found when the hematopoietic progenitor cells were enriched from CB cells via the depletion of lineage(+) committed cells. Gene transfer to lineage(-) early immature hematopoietic progenitors from human umbilical CB was obtained using CD133, ABCG2 or HLA-1 antibodies conjugated to lentiviruses pseudotyped with modified Sindbis viral Env proteins.

Specific gene transfer to neurons, endothelial cells and hematopoietic progenitors with lentiviral vectors

We present a flexible and highly specific targeting method for lentiviral vectors based on single-chain antibodies recognizing cell-surface antigens. We generated lentiviral vectors specific for human CD105(+) endothelial cells, human CD133(+) hematopoietic progenitors and mouse GluA-expressing neurons. Lentiviral vectors specific for CD105 or for CD20 transduced their target cells as efficiently as VSV-G pseudotyped vectors but discriminated between endothelial cells and lymphocytes in mixed cultures. CD133-targeted vectors transduced CD133(+) cultured hematopoietic progenitor cells more efficiently than VSV-G pseudotyped vectors, resulting in stable long-term transduction. Lentiviral vectors targeted to the glutamate receptor subunits GluA2 and GluA4 exhibited more than 94% specificity for neurons in cerebellar cultures and when injected into the adult mouse brain. We observed neuron-specific gene modification upon transfer of the Cre recombinase gene into the hippocampus of reporter mice. This approach allowed targeted gene transfer to many cell types of interest with an unprecedented degree of specificity.

The C2-streptavidin delivery system promotes the uptake of biotinylated molecules in macrophages and T-leukemia cells

Macrophages are tightly associated with inflammatory diseases as well as carcinogenesis, and therefore represent promising targets for drug delivery and gene transfer. We have recently established a novel protein delivery system based on the binary C2 toxin of Clostridium botulinum and streptavidin, allowing the uptake of exogenous biotinylated molecules into mammalian cells. Here, we applied this C2-streptavidin delivery system to macrophages and other leukocytes. First, the effect of wild-type C2 toxin on different leukocyte cell lines was tested, indicating no differences in sensitivity. Next, the uptake and stability of the engineered C2-streptavidin was analyzed in macrophages and Jurkat T-cells, showing internalization into the cytosol of both cell types with similar kinetics. The transporter did not exhibit any cytotoxic effect and did not interfere with phagocytosis in primary human macrophages. The C2-streptavidin system promoted specific uptake of biotinylated fluorophores into the cytosol of macrophages as revealed by confocal microscopy. In addition, flow cytometry analysis showed a significantly enhanced uptake of biotinylated fluorescent tracers in Jurkat leukemia cells mediated by the C2-streptavidin transporter. Our results demonstrate that C2-streptavidin is a functional delivery system for transport of biotinylated molecules into macrophages and other leukocytes without compromising cell viability and intrinsic functions such as phagocytosis.

Nonviral regenerative gene therapy of mammalian cutaneous damages

The rate and character of superficial tissue regeneration after wounds, burns, and other traumas depend on cell proliferation within the damaged area. The acceleration of wound healing via the stimulation of cell proliferation and extracellular matrix synthesis is one of the most important tasks of modern medicine. There are gene therapy approaches to wound treatment, such as the transfer of genes that encode mitogenic growth factors to the wound area. The most important step in the development of the gene therapy approaches is the design of gene delivery tools. Despite the high efficiency of viral vectors, the nonviral approaches have some advantages (low toxicity, low immunogenity, safety, and the absence of side effects). Among the nonviral gene delivery tools molecular conjugates are the most popular due to their efficiency, simplicity, and the capacity for targeted gene transfer. In the present work, we have developed two molecular conjugates, NLS-TSF7 and NLS-TSF12, which consist of the modified signal of the nuclear localization of the T-antigen of the SV40 virus (cationic part) and the peptide ligands of the mammalian transferrin receptor (ligand part). Those conjugates bind to plasmid DNA via the formation of polyelectrolytic complexes and are able to deliver plasmid DNA into cells that express transferrin receptors through receptor-mediated endocytosis. The transfer of the expression vector of the luciferase gene in the complex with the molecular conjugate NLS-TSF7 to murine surface tissues led to about the 100-fold increase of luciferase activity in comparison with the transfer of the free expression vector. The treatment of mice with incised wounds with complexes of the expression vector of the synthetic human gene that encodes insulin-like growth factor 1 with molecular conjugate NLS-TSF7 led to the acceleration of wound healing in comparison with mice treated with the free expression vector. The obtained results confirm the high efficiency of the developed approach to regenerative gene therapy for treating the superficial tissue damage of mammals.

Antibody-mediated targeted gene transfer to NMDA NR1-containing neurons in rat neocortex by helper virus-free HSV-1 vector particles containing a chimeric HSV-1 glycoprotein C–Staphylococcus A protein

Because of the heterogeneous cellular composition of the brain, and especially the forebrain, cell type-specific expression will benefit many potential applications of direct gene transfer. The two prevalent approaches for achieving cell type-specific expression are using a cell type-specific promoter or targeting gene transfer to a specific cell type. Targeted gene transfer with Herpes Simplex Virus (HSV-1) vectors modifies glycoprotein C (gC) to replace the heparin binding domain, which binds to many cell types, with a binding activity for a specific cell surface protein. We previously reported targeted gene transfer to nigrostriatal neurons using chimeric gC–glial cell line-derived neurotrophic factor or gC–brain-derived neurotrophic factor protein. Unfortunately, this approach is limited to cells that express the cognate receptor for either neurotrophic factor. Thus, a general strategy for targeting gene transfer to many different types of neurons is desirable. Antibody-mediated targeted gene transfer has been developed for targeting specific virus vectors to specific peripheral cell types; a specific vector particle protein is modified to contain the Staphylococcus A protein ZZ domain, which binds immunoglobulin (Ig) G. Here, we report antibody-mediated targeted gene transfer of HSV-1 vectors to a specific type of forebrain neuron. We constructed a chimeric gC–ZZ protein, and showed this protein is incorporated into vector particles and binds Ig G. Complexes of these vector particles and an antibody to the NMDA receptor NR1 subunit supported targeted gene transfer to NR1-containing neocortical neurons in the rat brain, with long-term (2months) expression.