Perinatal gene delivery to the CNS

483ISSN 2041-599010.4155/TDE.11.16 © 2011 Future Science Ltd Therapeutic Delivery (2011) 2(4), 483–491

Review

Special FocuS: Gene DeliveRy

A powerful research toolSome of the earliest work on perinatal gene delivery to the CNS dates back to the 1980s. At this time, the application was primarily as a research tool for investigating neurological development. Using retroviral vectors to deliver reporter genes to cells of the fetal mouse brain, a series of studies produced data providing a fun-damental insight into neural cell lineages and migration patterns [1–3].The ability of retroviral vectors to integrate a marker gene into the host cell made it an invaluable tool for these early studies and the elucidation of neural progenitor cells and their progeny [4].

Over the years, the application of a range of other viral vectors has proven useful. Shen et al. demonstrated efficient gene delivery to the fetal mouse brain following in utero administration of adenoviral vectors into the cerebral ventri-cles [5,6]. Similar studies using the neurotropic herpes simplex virus (HSV), combined with the TC-1 ‘sleeping beauty’ transposable ele-ment technology that allow for stable integra-tion into the cell genome, have also produced impressive results. In utero intraventricular administration of these hybrid HSV vectors led to specific transduction of neural progeni-tor cells and their derivatives [7]. The inclusion of integrating properties to the HSV vector was an important step forward for this vector to be used for long-term expression. Lentiviral vec-tors, a subclass of retroviruses, are particularly efficient at integrating their genetic payload into host cell genomes and have the additional advantage of being able to transduce nondivid-ing cells. Endo et al. demonstrated that a lenti-viral vector, administered to the amniotic fluid of an E8 fetal mouse, resulted in transduction

of ocular stem cells in the retina, lens and cor-nea [8]. In a subsequent study, the same group injected a lentiviral vector carrying the GFP gene into the murine amniotic cavity at E8 [9]. They demonstrated transduction of the entire nervous system for over 80% of the mouse lifes-pan with all major neural cell types express-ing GFP. Furthermore, gene delivery was also seen in the neural stem cells residing within the subventricular zone and the subgranular zone. This approach represents a powerful and relatively simple means of investigating the function of specific genes in development of the CNS.

Although viral vectors have been seen as the most efficient means of delivering genes to cells of the CNS, nonviral methods have recently emerged that show promise. The use of an electric current to form transient pores through which plasmid DNA carrying a gene can enter cells has been of particular value in the fetal CNS. Kawauchi et al. administered plasmid DNA carrying a reporter gene to the 4th ventricle of E12.5 mouse embryos followed by electroporation to investigate the cellular basis of nuclei and nucleogenesis. Tracking of cells using the reporter gene expression revealed that precerebellar neurons, derived from the lower rhombic lip, go through multiple migra-tion steps before forming nuclei [10]. Soma et al. used electroporation-mediated gene delivery to label cells at E10, E11 and E12 and investi-gate the development of the amygdala, an area important for control of emotions [11]. Tracking of cell migration and differentiation demon-strated that the central nucleus was derived from the neuroepithelium in the ganglionic eminence. However, the basolateral complex

Perinatal gene delivery to the CNS

The relative inaccessibility of the brain compared with other major organs, the highly regulated transfer of molecules across the blood–brain barrier and the limited capacity of neurons to regenerate, make efficient gene delivery to the CNS both challenging and imperative. Perinatal gene delivery to the CNS represents a powerful tool for the investigation of genes in development and disease. However, it may also hold immense therapeutic value for neonatal lethal neurodegenerative diseases for which no treatment is available. This article will focus on the use of perinatal gene delivery as a research tool and the potential it has to develop into a realistic therapy that can be translated to the clinic.

Ahad A Rahim†1, Suzanne MK Buckley1, Jerry KY Chan2, Donald M Peebles1 & Simon N Waddington1

1Institute for Women’s Health, University College London, 86–96 Chenies Mews, London, WC1E 6HX, UK 2Experimental Fetal Medicine Group, Department of Obstetrics and Gynaecology, National University of Singapore, NUHS Tower Block, Level 2, 1E Kent Ridge Road, 119228 Singapore †Author for correspondence:Tel.: +44 770 939 9780 E-mail: [email protected]

For reprint orders, please contact [email protected]

Review | Rahim, Buckley, Chan, Peebles & Waddington

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originates from neuroepithelium from the ven-tral and lateral pallia. This data provides an important means of understanding the patho-genesis of emotional disorders. However, while electroporation is certainly a promising tool from a basic research point of view, the clinical translation of the methodology is likely to be limited in its present form. Table 1 summa-rizes the key features associated to the vectors described here.

Although the focus of this article is gene delivery, one cannot ignore the transfer of other genetic material to the perinatal CNS that are proving to be powerful research tools. The important role of miRNA in development and disease has recently been recognized [12–14]. miRNA are small endogenously expressed noncoding RNA that regulate mRNA expres-sion. They achieve this by base pairing with the mRNA strand, which mediates degrada-tion or translational inhibition. A number of miRNAs have been identified that are present only in the CNS and delivery of these miRNAs to the fetal brain is proving vital in elucidat-ing their roles in development. miRNA 9 is one example that is only found in neurogenic areas of the brain during development and in adulthood [15–18]. However, whether it has a role in neural stem cell self-renewal and dif-ferentiation was unknown. To investigate this, Zhao et al. administered miRNA 9 duplexes to neural stem cells in the ventricular zone of E13.5 mouse brains via electroporation [19]. This led to a marked decrease in cell prolif-eration but an increase in cell migration from the ventricular zone to the cortical plate. They were able to show that miRNA 9 targets TLX

nuclear receptor expression and negatively regulates neural stem cell proliferation and accelerates differentiation.

shRNA are similar to miRNA and can be used to silence or reduce gene expression [20]. Unlike miRNA, they are not found endogenously in cells and are a short sequence of RNA that forms a hairpin loop and inhibits gene expression by RNA interference. They can be designed and targeted against a specific gene sequence and have been shown to be very efficient. shRNA against DISC1 gene expression were delivered to the brains of E15 mice using both electropora-tion and lentiviral vectors [21]. This resulted in cell migration defects in the cerebral cortex and confirmed that DISC1 has a developmental role. While shRNA technology is certainly useful, it does have a disadvantage to miRNA. shRNA are transcribed by RNA polymerase III and the mammalian cell can, in some instances, mount an interferon response to protect it from what it perceives as a viral infection. miRNA is tran-scribed by RNA polymerase II and so avoids this potential complication.

Advantages & therapeutic proof of principlePerinatal gene therapy has the potential to treat a number of genetic diseases. Indeed, proof of concept has been demonstrated through the correction of several genetic disease models in mice. For diseases specifically affecting the CNS, Karolewski and Wolfe demonstrated that intracranial administration of adeno-associated virus serotype 2/1 (AAV2/1) car-rying the human b-glucuronidase to the fetal brain resulted in increased survival of a mouse

Table 1. The gene delivery vectors described and their relative key features.

Gene delivery vector Key features Ref.

Retroviral Stable expression, moderate packaging capacity, low toxicity, transduces dividing cells, higher risk of insertional mutagenesis

[1–4,45,49,51]

HSV hybrid Stable expression, large packaging capacity, low toxicity, transduces both dividing/nondividing cells, moderate risk of insertional mutagenesis

[7]

Adenoviral Transient expression, large packaging capacity, moderate toxicity, transduces both dividing/nondividing cells, minimal risk of insertional mutagenesis

[5,6, 25,48]

Lentiviral Stable expression, moderate packaging capacity, low toxicity, transduces both dividing/nondividing cells, higher risk of insertional mutagenesis

[8,9,36,46,51]

Non-integrating lentiviral Transient expression, moderate packaging capacity, low toxicity, transduces both dividing/nondividing cells, minimal risk of insertional mutagenesis

[27,47]

AAV Transient expression, limited packaging capacity, low toxicity, transduces both dividing/nondividing cells, minimal risk of insertional mutagenesis

[22–24,26,28,31,37–41]

Nonviral (electroporation) Transient expression, unlimited packaging capacity, moderate toxicity, transfects both dividing/nondividing cells, minimal risk of insertional mutagenesis

[10,11]

AAV: Adeno-associated virus; HSV: Herpes simplex virus.

Key Terms

Perinatal: The period before or after birth. Here, it refers to the late gestation and early neonatal phase.

Gene therapy: The concept of introducing therapeutic genetic material to cells to treat a genetic defect.

Perinatal gene delivery to the CNS | Review

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model of mucopolysaccharidosis (MPS)-VII [22]. Dejneka et al. used the same viral vector to deliver retinal pigment epithelia 65 (RPE65) to the retina following in utero administration, result-ing in restored eyesight in the RPE65-/- mouse model [23]. Passini et al. also demonstrated that direct administration of an AAV8 vector carry-ing the survival motor neuron gene to the neo-natal CNS significantly improved the median lifespan of a mouse model of spinal muscular atrophy (SMA) [24].

These perinatal approaches are attractive due to the unique advantages they confer over adult delivery; for example, overcoming the host immune response to both vector and expressed foreign protein. The relative immaturity of the immune system in both the developing fetus and early neonates provides the opportunity for the development of immune tolerance to an expressed protein. The body perceives the for-eign protein as being ‘self ’ and so an immune response is either attenuated or not triggered. This may not be the case in adults where a fully functional immune system would clear the protein from the body. Waddington et al. demonstrated that in utero injection of an adenovirus expressing human factor IX (FIX) into FIX-/- mice resulted in immune tolerance to the protein as adults [25]. Repeated challeng-ing by injection of FIX protein into the adult mice resulted in persistence within the circula-tion and an absence of anti-FIX antibodies in five of nine mice. By contrast, mice that were not administered in utero cleared injected FIX as adults and mounted an immune response measurable as anti-FIX antibodies. In a similar study, Sabatino et al. were able to demonstrate that following in utero and neonatal administra-tion of AAV1 expressing human FIX in knock-out mice, readministration as adults resulted in persistent gene expression with no measur-able humoral or cellular immune response [26]. Given that the vast majority of gene delivery vectors are viral in origin (e.g., AAV, lentivirus, adenovirus and HSV), pre-existing antibodies in adults would result in elimination of the vector. The fetus and neonate are less likely to have been exposed to these viruses and so the likelihood of antibody-mediated clearance of vector is low.

Perinatal gene delivery is also advantageous in terms of realistic scale-up. Owing to the smaller size of fetal and neonatal brains, the vector:cell ratio is much greater than in adults resulting in increased transduction efficiency.

The spread of vector through the brain follow-ing intracranial administration is also thought to be greater during the perinatal stage. Rahim et al. recently demonstrated that intracranial administration of a non-integrating lentiviral vector to the fetal mouse brain resulted in sig-nificantly enhanced dissemination through the parenchyma when compared with adult administration [27]. Similarly, Passini and Wolfe showed in an earlier study that injection of AAV2 into the ventricle of neonatal brains led to structure-specific delivery of a reporter gene [28]. If the expressed therapeutic protein is soluble and secreted into the extracellular space, uptake by neighboring cells via the appropriate receptor could allow for ‘cross-correction’ and increase the therapy’s range. Widespread gene delivery is required for those diseases that have a global effect throughout the brain and is vital to the success of the treatment. Of course, in some cases this is not required and a more local-ized area of gene delivery is sufficient, such as in the case of subretinal injection to treat the retinopathies [23].

In many cases, the best cell type to transduce, with an integrating or episomally maintained vector, would be a stem or progenitor cell that can differentiate and multiply into a number of neural daughter cells, all of which would carry the delivered gene. The fetal brain would pro-vide the ideal environment to maximize upon this with active neurogenesis and astrogenesis taking place.

All these advantages make perinatal gene delivery very attractive for a range of CNS disorders. However, in utero intervention is uniquely placed to address those diseases that result in neuro degenerative neonatal death. In many of these cases, the disease manifests itself during gestation and, thus, postnatal interven-tion may be ineffective. One of the rationales behind in utero gene delivery is to intervene, therapeutically, before pathology can manifest and prevent early neonatal death. This is argu-ably most significant in the context of the CNS where damaged neurons have a limited ability to regenerate. So disease pathology is both dev-astating and irreversible. An example of this is in type II Gaucher disease, a lysosomal storage disorder caused by mutation in the glucocer-ebrosidase gene. The disease pathology begins during gestation [29] resulting in progressive neurodegeneration and death, usually before 2 years of age [30]. Although enzyme replace-ment therapy is available for treating the

Key Term

Gene delivery vector: A construct that transports genetic material into cells. They can broadly be divided into viral and nonviral vectors.



visceral pathology associated with this disease, it is, ultimately, only palliative as the imperme-able BBB prevents enzymes from accessing the CNS. Therefore, no effective clinical treatment is available for this disease. Fetal gene therapy would be a potential option for addressing this disease.

Of course, not all CNS diseases affect the entire brain, some are more localized and pathology is seen in discrete areas. In such cir-cumstances, a more targeted approach would be desirable and the expanding repertoire of gene delivery vectors is starting to make this possi-ble. In an impressive study by Cearley et al., 17 different novel serotypes of AAV carrying the GFP reporter gene were evaluated for their neu-ral tropisms by administration into the lateral ventricles of neonatal mice. The brains were then evaluated revealing unique transduction patterns of the different serotypes within dis-crete anatomical areas. The areas evaluated and the tropisms of the individual viruses are too numerous to list here, but the reader is referred to Table 2 of the study [31]. An extension of this study to other gene delivery vectors would provide useful information. Furthermore, an evaluation of a wide range of vectors within the fetal brain would be invaluable in terms of selecting the correct vector to address a par-ticular disease affecting a defined region of the brain.

Translation to the clinicAdministration of vector to the brain can be technically challenging. This is due to the protective encapsulation of the brain within the skull and the impermeable nature of the BBB preventing delivery via the bloodstream. Therefore, direct intracranial injection of vec-tor to the brain has been the preferred route of delivery. This has been demonstrated to be safe and effective in neonatal [28,32] and fetal mice [27] resulting in widespread gene delivery. However, would this approach be clinically feasible? At present, this is highly unlikely for in utero administration. The main obstacle lies in the complexity of neuro surgery. This is perhaps best exemplified by the surgical pro-tocols required in clinical trials for late infan-tile neuronal lipofuscinosis [33] and Canavan disease [34]. In these pediatric trials, children required an MRI presurgical scan, a head frame, six bur-holes and injection without stereotaxis into 12 sites of the brain in order to achieve sufficient viral spread. The skill and nuances

required for this type of invasive surgery are immense and have been reviewed elsewhere in greater detail [35]. In the context of in utero sur-gery, these difficulties would only be magnified even though it is conceivable that fewer injec-tions sites may be required due to enhanced vector spread in the fetal brain. It is, there-fore, vital that an alternative means of vector delivery is found that retains the efficiency of intracranial administration but is less invasive. Mazarakis et al. demonstrated that lentiviral vectors pseudo typed with the rabies-G envelope had the ability to transduce motor neurons of the lumbar spinal cord following intramuscu-lar injection into the gastrocnemius muscle in rats [36]. Although promising and minimally invasive, this approach has yet to be confirmed as being efficient enough for correction of ani-mal models of CNS disease. The ideal route of vector administration would be via intra-venous administration since the circulation of the human fetus is already accessed for blood sampling and transfusion. As previously men-tioned, the impermeable BBB has historically prevented gene delivery vectors from entering the CNS. However, a number of recent studies have provided promising and encouraging data that have led to a revival in interest in intrave-nous administration. McCarty et al. recently demonstrated that pretreatment of a mouse model of MPS IIIB with mannitol allowed AAV2 expressing a-N-acetylglucosaminidase (NaGlu) to cross the BBB and enter the brain. The subsequent expression of NaGlu resulted in extended survival, reduction in brain pathology and improved behavioral performance in these mice [37]. Perhaps the most compelling data has been provided by a highly elegant series of studies using AAV9. Foust et al. were able to show that intravenous injection of AAV9 car-rying the GFP reporter gene to adult and neo-natal mice resulted in extensive transduction of neural cells within the brain and spine [38]. Neonatal administration led to preferential neuronal transduction while adult administra-tion favored astrocytes. Similarly, Duque et al. demonstrated efficient transduction of motor neurons in the spine of adult mice and neonatal and adult cats [39]. In a subsequent study by Foust et al., the same vector carrying the SMN gene was administered neonatally at postnatal day 1 (P1) to a mouse model of SMA, and motor function, lifespan and neuromuscular physiol-ogy were rescued [40]. Interestingly, administra-tion at P5 resulted in only partial correction



and P10 led to little effect highlighting the importance and increased efficacy of perinatal intervention. Similar results were reported in a separate study using the same vector to pro-long the lifespan of SMA mice [41]. The ability of AAV9 to cross the BBB was also confirmed in neonatal macaques following intravenous administration. Although the exact mecha-nism of how AAV9 traverses the BBB is not fully understood, these are milestone studies. They do more than just confirm the feasibility of perinatal gene delivery to correct a disease of the CNS. They have also highlighted AAV9 as a vector that could overcome the procedural and surgical difficulties associated with poten-tial translation of in utero gene delivery to the clinic. One may now consider that a vector that crosses the BBB could be administered into the fetal circulation using the same technology of ultrasound guidance that permits minimally invasive protocols such as fetal blood transfu-sion or blood sampling (cordocentesis). These are routine clinical protocols used by obstetri-cians worldwide if the fetus is diagnosed to be anemic or suspected of an abnormality, respec-tively. Of course, further studies are required to confirm whether AAV9 would cross the fetal BBB but the prospect is certainly exciting and full of promise.

The inclusion of nonhuman primate stud-ies is important in evaluating perinatal gene delivery and its applicability to the clinic. This is because the differences between rodent and human neural development is a complicated issue. They are likely to vary depending on the criteria used for comparison, for example, ana-tomical, immuno logical or physiological devel-opmental differences. If we take one example of anatomical markers, such as the cortical plate and subplate, then diffusion tensor MRI has shown that the thickening patterns of these areas during human gestation occur in the second tri-mester while the equivalent in mice occurs later at E14–E18 [42,43]. Potential differences do not diminish the value of mice for preclinical experi-mentation but do highlight the need for accom-panying nonhuman primate studies to ‘bridge’ the potential developmental gap.

The risksAlthough perinatal gene therapy for disorders affecting the CNS is very attractive, the poten-tial hazards associated with this must be consid-ered. These concerns are not just confined to the CNS but are widely applicable to other organs

and systems. There are obvious risks associated with any in utero surgery such as infection, pre-term labor and fetal loss. In practice, the risks associated with the use of a fine needle under ultrasound guidance are small. Cordocentesis, as previously mentioned, is an example of a pro-cedure with an excellent safety profile if the pre-ferred intravenous route of vector administration is considered [44]. Additional concerns are associ-ated with the choice of gene delivery vector. A criticism that can be made of integrating vec-tors, such as retroviruses, is that they may cause insertional mutagenesis and oncogenicity. This is based on adverse side effects associated with clinical trials for X-linked severe combined immunodeficiency disorder caused by integra-tion into the promoter sequence of a proto-oncogene [45]. It is possible that the increased levels of cellular proliferation, growth factor and gene activity associated with growth and differentiation in the fetus may exacerbate this. Themis et al. demonstrated a high incidence of hepatocarcinoma following fetal gene delivery to the mouse using the equine infectious ane-mia virus carrying the LacZ reporter gene and FIX [46]. This did not occur in fetuses injected with vectors with an HIV backbone and equine infectious anemia virus has not been reported to cause oncogenesis in adult models. Moreover, a mechanism for this has not yet been identified. Nevertheless, this may suggest that the fetus is more sensitive to mutagenesis. To date, there have been no reports of insertional mutagenesis from any of the animal studies involving direct administration of vectors to the fetal or neonatal CNS. The use of non-integrating vectors could minimize the risks of insertional mutagenesis. These include vectors such as AAV, adenovirus, HSV and non-integrating lentivirus. To improve the safety profile of lentiviral vectors, a non-inte-grating version was developed that included a point mutation within the integrase gene [47]. This rendered the virus unable to integrate its genome into that of the host cell. Instead, the viral genome forms a circular piece of DNA that exists as an episome. In utero administra-tion of this vector carrying the GFP gene to the fetal brain resulted in efficient and widespread gene delivery with long-term expression within neurons [27]. A consequence of this is that gene expression in actively dividing cells would be lost owing to dilution of viral copy numbers. Therefore, the use of non-integrating vectors may not be ideal for transducing astrocytes but could be suitable for postmitotic neurons.

Key Term

Insertional mutagenesis: Mutation in an organism’s genome through insertion or addition of one of more DNA bases. It can be caused by viruses that integrate their DNA into a host genome.



Overexpression of a therapeutic gene dur-ing gestation may also cause problems in the develop ment of the target organ. Gonzaga et al. demonstrated that in utero delivery of an adeno-virus expressing FGF10 to the proximal tracheo-bronchial tree of rats during the pseudoglandular phase resulted in the formation of large cysts [48]. These were lined with columnar epithelium com-posed primarily of Clara cells. Overexpression of FGF10 in the distal lung parenchyma during the canalicular phase resulted in small cysts lined with cuboidal epithelial cells composed primar-ily of type II pneumocytes. These lung malfor-mations emphasize the thorough research that is required to test therapeutic transgene overex-pression and its effect on organ development. If the vector is delivered systemically then a com-plete ana lysis of other organs will be needed due to vector spread.

Gene therapy is targeted towards the correc-tion of somatic cells. However, the inadvertent transmission of genetic material to the germ line is a concern from both a safety and ethical point of view. A number of studies have inves-tigated this issue. Porada et al. studied this in sheep using retroviruses that were administered by in utero intraperitoneal injection to the fetus. Using breeding experiments, PCR ana lysis of purif ied sperm and immunohistochemical

ana lysis of reproductive tissues, they were able to show that, although transmission was detect-able in the germ cells of the injected sheep, this was very low level and well below the upper limits placed by the US FDA [49]. Germline transmission has also been investigated in non-human primates that received an HIV-1-based vector delivered in utero via the intrapulmo-nary, intraperitoneal and intracardiac routes to the fetus [50]. Examination of tissues and cells showed no germ line gene transfer following intrapulmonary and intracardiac administra-tion in the injected primates. However, evi-dence of gene delivery was present in female oocytes following intraperitoneal administra-tion. At present, it is unlikely that low-level transmission to the germ line can be completely excluded following systemic delivery; particu-larly in the case of integrating vectors. This may improve with continuing progress in targeting and tissue specificity. However, the potential of gene therapy to alter the genome requires further ethical debate.

The safety of the mother in the case of in utero gene delivery is of utmost importance. It is conceivable that all of the outlined risks may also apply to the mother if a gene delivery vector is able to cross the placenta. An early study by Tarantal et al. involved the in utero

Executive summary

A powerful research tool

� A range of viral and nonviral vectors have been found to be efficient in delivering genetic material to the developing brain.

� The study of neural precursors and cell lineages can be achieved by delivering reporter genes to the fetal brain.

� The function of genes and regulatory sequences in brain development and disease can be elucidated through delivery of genetic material to the fetal brain.

Advantages & therapeutic proof of principle

� Gene delivery to the perinatal brain offers unique advantages in terms of avoiding an immune response, increased vector:cell ratio and greater chances of progenitor cell transduction.

� Perinatal gene therapy is particularly well suited to neonatal lethal neurodegenerative diseases where intervention is required before irreplaceable neurons are lost.

� Therapeutic proof of principle of perinatal gene therapy has been achieved in an animal model of various CNS diseases.

Translation to the clinic?

� Perinatal gene delivery in the clinic is likely to be more technically difficult than current pediatric gene therapy clinical trials.

� Intravenous delivery of vectors is the ideal route of administration for fetal gene therapy.

� New vectors have the ability to cross the blood–brain barrier following intravenous injection.

� Vectors that cross the blood–brain barrier represent a significant solution for overcoming procedural concerns associated with perinatal gene therapy for CNS diseases.

The risks

� In utero surgery has associated risks such as infection, preterm labor and fetal loss.

� The potential for insertional mutagenesis when using integrating gene delivery vectors is a concern.

� The overexpression of therapeutic genes in the fetus may cause developmental abnormalities in target organs.

� Inadvertent germ-line transduction and transmission.



administration of both retroviruses and lenti-virus to fetal rhesus monkeys [51]. In a follow-up of the mothers (up to 2 years), they were observed to be healthy and all clinical chem-istry panels were normal. However, in a lim-ited number of cases, traces of transgene pro-tein were detectable in some maternal blood samples, suggesting that transplacental passage of virus may have occurred from those fetuses that received relatively high doses of virus. An alternative explanation is that these findings are a consequence of fetal cells or DNA in the maternal blood. Although this study reports very low level and occasional evidence for trans-placental passage of virus, it is feasible that this may vary with the choice of gene delivery vec-tor and requires more study to characterize the associated risk.

Future perspectivePerinatal gene delivery has huge potential both as a research tool and as a therapeutic application. The delivery of genes and other genetic material to the developing CNS will play a vital role in our continuing understanding of CNS development and the molecular basis of disease. The advan-tages and therapeutic potential of this as part of a fetal gene therapy protocol are clear. It is an approach that is uniquely placed to address neo-natal lethal neurodegenerative diseases. Many of these diseases, such as type II Gaucher disease, have no treatment available and are well suited as targets for fetal gene therapy. The evidence supporting the possible translation to the clinic

is becoming more compelling. However, the case for this should be dealt with responsibly, taking into account the associated risks. It is likely that, as the gene delivery vectors improve, the risks will be reduced and the levels of transduction efficiency increased and targeted to the organs of interest. However, technological advances in vector design are not the sole obstacles in mov-ing this concept to the clinic. Outside of the gene therapy community, progress is required in areas such as prenatal diagnosis. This is, of course, vital and our increasing understanding of the human genome and advances in next-generation sequencing will improve prenatal screening programs. Furthermore, advances in medical imaging will help increase the resolution of accuracy with which the administration of vector is carried out.

Financial & competing interests disclosureThis work was supported by a UK Medical Research Council Grant (G1000709), a grant from the Newlife Foundation for Disabled Children (09-10/15), a Wellcome Trust grant (WT089806MA) and a European Research Council Starting Grant (SomaBio). Rahim and Waddington have also received support from the UK Gauchers Association (funds donated by the family of Ellie Carter). The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.

No writing assistance was utilized in the production of this manuscript.

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Perinatal gene delivery to the CNS