Engineering Humoral Immunity as Prophylaxis or Therapy

Abstract

Purpose of the review

In this review, we will discuss the field of engineered humoral immunity with an emphasis on recent work using viral vectors to produce antibodies in vivo. As an alternative to passive transfer of monoclonal antibody protein, a transgene encoding an antibody is delivered to cells via vector transduction, resulting in expression and secretion by the host cell. This review will summarize the evidence in support of this strategy as an alternative to traditional vaccines against infection and as novel therapeutics for a variety of diseases.

Recent findings

Historically, humoral immunity has been engineered through vaccination and passive transfer of monoclonal antibodies. However, recent work suggests that vectors can be used to deliver transgenes encoding broadly neutralizing antibodies to nonhematopoietic tissues and can mediate long-term expression that is capable of preventing or treating infectious diseases. The production of engineered monoclonal antibodies allows for precise targeting and elimination of aberrant self-proteins that are characteristic of certain neurodegenerative disease. This approach has also been successfully used to combat cancer and addiction in several animal models. Despite the wide array of expression platforms that have been described, adeno-associated virus vectors have emerged as the frontrunner for rapid clinical translation.

Summary

Recent advances in vector-mediated antibody expression have demonstrated the potential for such interventions to prevent infection and treat disease. As such, it offers an alternative to immunogen-based vaccine design and a novel therapeutic intervention by enabling precise manipulation of humoral immunity. Success translating these approaches to patients may enable the development of effective prevention against previously intractable pathogens that evade immunity such as HIV, influenza, malaria or HCV and may also enable new treatment options for neurodegenerative diseases such as Alzheimer’s disease.

Introduction

The humoral immune system is one of the first obstacles encountered by invading pathogens, thus playing a crucial role in preventing infection and maintaining human health. Immunological memory, particularly in the form of pre-existing antibodies, has been shown to form the basis of protection for nearly all vaccines in use today [1]. This ability to generate memory against previously encountered pathogens enabled the first practitioners in Asia to engineer immunity against smallpox in the fifteenth century through a process termed variolation [1]. By intentionally exposing patients to the relatively mild variola minor, they were able to induce protective immunity that prevented life-threatening smallpox infection by variola major [1]. Similarly, modern-day vaccines engineer immunity by exposing a patient to inactivated or attenuated whole pathogens, or recombinant components of pathogens that are known to elicit protective immunity. As a result, effective vaccines have been developed against many of the diseases for which natural infection results in immunity against re-infection. This practice has resulted in the global eradication of smallpox [2] and decreased incidence of diphtheria, measles, mumps, pertussis, poliomyelitis, rubella and tetanus [3]. However a number of diseases for which prior exposure is ineffective at preventing subsequent disease have proven more difficult targets. These include intractable pathogens that evade immunity, such as human immunodeficiency virus (HIV), malaria, hepatitis C virus (HCV), and influenza A virus (IAV) as well as complex diseases of self, such as cancer and neurodegenerative disorders.

Engineered Humoral Immunity Through Passive Transfer

Ideally, vaccination elicits a protective cellular and humoral response, however the protection raised by most of the currently licensed vaccines is largely antibody-dependent [1,4]. Importantly, neutralizing antibodies (nAb) alone have been shown to prevent the spread of many diseases within populations [4,5]. This antibody-based protection underlies passive transfer, whereby the administration of sera or purified antibodies into naïve patients transiently confers immunity. Technological advances including the development of hybridoma technology [6] and the “humanization” of antibodies has spawned a new class of antibody-based drugs which have demonstrated remarkable success in the clinic for a wide range of diseases.

While mAbs have several advantages over conventional small-molecule drugs, there are considerable disadvantages as well. Currently licensed mAbs have a typical half-life ranging from two days to one month in vivo [7], necessitating frequent dosing. However, specific mutations in the antibody Fc region have been described that prolong half-life up to five-fold by increasing the affinity of the antibody for the neonatal Fc receptor (FcRn) [8]. Despite the potential for less frequent dosing, high concentrations of most antibodies are necessary to achieve clinical efficacy, resulting in high materials costs [5,9]. Additionally, antibody use is further complicated by the requirement of cold-chain storage and trained medical personnel for administration [7], making such therapy difficult to implement in low-resource areas. For mAb therapy to become a widely administered intervention on a global scale, improved delivery approaches will be required.

Vectored Antibody Gene Delivery for Infectious Diseases

One alternative to passive transfer utilizes a vector for the delivery of transgenes encoding previously characterized antibodies. These transgenes direct the production of mAbs in non-hematopoetic cells, which secrete mAbs into the circulation or the local environment. A wide variety of vectors, each with distinct expression profiles, have been considered for this approach (Table 1).

Table 1.

Summary of Monoclonal Antibody Expression Vectors

	Vector	Ab Type	Species of Fc	Isotype	Promoter	Furin/2A Cleavage	Animal Model	Vector Dose	Route	Ab Clone	Peak Serum Concentration	References
HIV	plasmid	Fab	mouse	IgG1	CMV	N/A	BALB/c mice	25 μg	IM	VRC01	3.3 μg/mL	Muthumani et al 2013
	Lentivirus	Full-length	human	IgG1	human μ heavy chain	Y	NSG humanized mice	MOI 1000	ex vivo B cell transduction	b12	>1 μg/mL	Luo et al 2009
	Lentivirus	Full-length	human	IgG1	hPGK	Y	NSG humanized mice	100 ng p24	ex vivo B cell transduction	2G12	40 ng/mL	Joseph et al 2010
	rAAV2	Full-length	human	IgG1	CMV, EF1α	N	Rag-1 mice	5×1011 GC	IM	b12	8 μg/mL	Lewis et al 2002
	scAAV2	Immunoadhesin	rhesus	IgG2	CMV	N/A	Rhesus macaques	2×1013 GC	IM	4L6 5L7	400-500 μg/mL 200-300 μg/mL	Johnson et al 2009
										b12 2G12	100 μg/mL >250 μg/mL
	rAAV8	Full-length	human	IgG1	CASI	Y	NSG humanized mice	1×1011 GC	IM	2F5 VRC01 4E10	20 μg/mL >250 μg/mL 20 μg/mL	Balazs et al 2012
										b12 VRC01 VRC07 3BNC117 12A12	115 μg/mL 100 μg/mL 130 μg/mL 24 μg/mL 23 μg/mL
	rAAV8	Full-length	human	IgG1	CASI	Y	NSG and BLT humanized mice	1×1011 GC	IM	VRC-PG04	30 μg/mL	Balazs et al 2014
										VRC07G54W	74 μg/mL
										NIH45-46G54W	40 μg/mL
										PGT121	256 μg/mL
										PGT128	50 μg/mL
										PG9	390 μg/mL
	rAAV8	Full-length	human	IgG1	human thyroglobulin	Y	NSG humanized mice	2.5×1011 GC 2.0×1011 GC 2.5×1013 particles	IV IV	10-1074 3BNC117 eCD4-Ig 3BNC117	300 μg/mL 20 μg/mL 75-180 μg/mL	Horwitz et al 2013
	scAAV1	Full-length	rhesus	IgG2	CMV	N/A	Rhesus macaques	1×1013 particles	M	PGT121 10-1074 NIH45-46	-- -- --	Gardner et al 2015
Influenza	Ad5	Single-domain	--	--	CMV	N	BALB/c	1×108 PFU 1×107 PFU 1×106 PFU	Intranasal	aHA-7	--	Tutykhina et al 2013
	rAAV8	Full-length	human	IgG1	CASI	Y	BALB/c and NSG mice, ferrets	1×1011 GC (mice) 5×1012 GC/kg (ferret)	IM	F10 CR6261	200 μg/mL (mice) 10-100 μg/mL	Balazs et al 2013
	rAAV9	Immunoadhesin	human	IgG1	CAG	N/A	BALB/c mice, ferret and rhesus macaques	1×1011 GC (mice) 1×1012 GC (ferret) 1×1013 GC (rhesus)	Intranasal	F16	0.5 μg/mL (nose), 2.0 μg/mL(lung)	Limberis et al 2013
Malaria	rAAV8	Full-length	human	IgG1	CASI	Y	C57BL/6NCr	1×1011 GC	IM	2A10 2C11	>1000 μg/mL 100-500 μg/mL	Deal et al 2014
HCV	rAAV9	Full-length	human	IgG1	CASI	Y	Rosa26-Fluc, FNRG humanized mice	1×1011 GC	IM	AR3A AR3B AR4A	1000-3000 μg/mL	de Jong et al 2014
RSV	Ad5 rAAVrh.10	Full-length	mouse	IgG1	CMV	N Y	BALB/c mice	5×1010 PFU 1×1011 GC	IV Intrapleural	murine palivizumab precursor	--	Skaricic et al 2008
Anthrax	Ad5 rAAVrh.10	Ful-length	mouse	IgG1	CMV	N Y	C57BL/6 mice	1×1011 PFU 1×1011 GC	IV Intrapleural	14B7-1H	--	De et al 2008
HD	rAAV2	scFv intrabody	--	--	CBA	N/A	C57BL/6, BACHD, R6/2, N171-82Q and YAC128 mice	1×1010 GC	Intrastriatal	VL12.3 Happ1	--	Southwell et al 2009
	rAAV1	scFv intrabody	--	--	--	N/A	B6.HDR6/1 mice	2×1010 GC	Intrastriatal	C4	--	Snyder-Keller 2010
AD	rAAV1 rAAV1	scFv	--	--	CMV	N/A	3xTg-AD mice	1×109 GC	Hippocampal	Aβ-scFv	--	Ryan et al 2010
	rAAV2 rAAV5	scFv	--	--	CAG	N/A	C57BL/6 and TgAβPPswe/PS1 dE9 mice	3×1010 GC	intracranial	scFv59	--	Kou et al 2011
	rAAV2	scFv	--	--	CMV	N/A	APPswe/PS1dE9 mice	5×1010 GC	IM, Intraventricular	scFv	--	Wang et al 2010, 2009
	rAAV1	Full-length	--	--	CMV	Y	Tg2576 mice	3×1010 GC	IM	IIA2	300 μg/mL	Shimada et al 2013
ALS	scAAV1	scFv	--	--	CMV	N/A	SOD1G93A, GAP- 43-luc/gfp/ SOD1G93A mice	3×109 GC	Intrathecal	D3H5	--	Patel et al 2014
Prion	rAAV9	scFv	-- --	-- --	--	N/A	CD1 mice	1.4×1012 GC	Intracranial	D18 D18	--	Moda et al 2012
	rAAV2	scFv	-- -- --	-- -- --	CMV	N/A	mice	9×109 GC	Intracranial	3:3 6:4 6:6	--	Wuertzer et al 2008
Cancer	rAAV8	Full-length	rat	IgG1	CAG	Y	NCr nu/nu mice	2×1011 GC	IV	DC101	>8000 μg/mL	Fang et al 2005
	rAAV8	Full-length	rat	IgG1	ZgIL-2P	Y	C57BL/6 mice	2.5×1011 GC	IV	DC101	>1000 μg/mL	Fang et al 2007
	rAAVrh.10	Full-length	mouse	IgG1	CMV	Y	C57BL/6 mice	1×1011 GC	IV	4D5	30-40 μg/mL	Wang et al 2010
	rAAV8	scFv	humanized	IgG1	CMV	N/A	SCID-BEIGE	2×1011 GC	IV	h1567	96 μg/mL	Han et al 2012
Addiction	rAAVrh.10	Full-length	--	--	CAG	Y	BALB/c	1×1011 GC	IV	GNC92H2	--	Rosenberg et al 2012
	rAAVrh.10	Full-length	--	--	CAG	Y	C57BL/6 mice	1×1011 GC	IV	NIC9D9	1300 μg/mL	Hicks et al 2012

Abbreviations: rAAV, recombinant aden-associated virus; scAAV, self-complementary adeno-associated virus; Ad5, adenovirus serotype 5;, fragment-antigen binding; scFv, single-chain variable fragment; CMV, cytomegalovirus; hPGK, human phosphoglycerate kinase promoter; EF1α, elongation factor 1α; N/A, not applicable; MOI, multiplicity of infection; GC, genome copy; PFU, plaque forming unit; IM, intramuscular; IV, intravenous

Naked plasmid DNA offers simplicity of use, lack of immunogenicity and ease of large-scale production. Improvements in electroporation techniques has allowed for enhanced transfection of specific tissues in vivo. Electroporation of various monoclonal antibody transgenes into muscle has yielded production of mAb light and heavy chains (ie. Fab fragments), peaking at 50-200ng/mL in mice and 30-50ng/mL in sheep [10]. Optimization of the expression plasmid and electroporation conditions yielded 2-3μg/mL of the VRC01 HIV broadly neutralizing antibody (bNAb) in the plasma of mice 12 days post administration [11], demonstrating that electroporation of plasmid DNA encoding antibody can be used to rapidly produce mAb in vivo. However, the modest concentrations and transient nature of expression obtained with existing protocols limit the clinical potential of this technique.

Viruses have been exploited as vectors for many years owing to their highly evolved mechanisms for efficient delivery of genetic material to host cells. Lentiviral vectors, consisting of an extensively modified HIV-1-derived genome pseudotyped with vesicular stomatitis virus (VSV) G protein, represent an attractive means of integrating transgenes into the host genome, enabling long term gene expression in a wide variety of both dividing and nondividing cells. Such vectors have been used to transduce primary hematopoietic stem cells (HSC) with a transgene encoding the HIV bNAb b12, allowing for their differentiation into plasmablasts that secreted b12 in vitro [12]. Using a similar approach, B cells were engineered to secrete HIV bNAb 2G12 in a humanized mouse model, achieving concentrations of approximately 40ng/mL in plasma, which was sufficient to inhibit HIV infection in vivo [13]. However, all of these studies utilized ex vivo transduction, making widespread implementation of this approach challenging.

While lentivirus is well suited for long-term expression of mAb, adenoviral vectors have been shown to exhibit transient, but rapid gene expression ideal for responding to infectious disease outbreaks. Adenovirus serotype 5 (Ad5) encoding Palivizumab, a respiratory syncytial virus (RSV) mAb, produced detectable antibody expression as early as four days post-transduction and resulted in a 5.4 fold decrease in RSV titers in the lung four days post-challenge as compared to controls [14]. Ad5 has also been used to express a single-domain antibody specific for H5N1 influenza A virus (IAV) hemagglutinin (HA), which protected mice when administered 14 days prior to, or even 48 hours after, infection [15]. In another study, a mAb targeting the protective antigen of Bacillus anthracis was delivered by Ad5 that protected mice from toxin challenge between 1 day and 8 weeks post-administration, but which was no longer protective at 6 months [16].

Adeno-associated virus (AAV) has never been associated with any disease in humans and recombinant vectors derived from AAV (rAAV) result in stable gene expression in the absence of integration through formation of extrachromosomal concatamers of the delivered transgene sequences [17]. The serotype used to package the vector strongly influences its ability to transduce different tissues [17,18] and has been shown to play a significant role in the immunogenicity of the vector in various animal models [19,20]. AAV1 delivering lipoprotein lipase (LPL) was recently approved in Europe as the first ever gene therapy product for humans [21] and recent clinical trials testing AAV8 for the delivery of Factor IX for hemophilia has met with considerable success [22]. Given their clinical efficacy and favorable expression profile, AAV vectors have been extensively characterized as a platform for the delivery of mAbs in vivo. However, the packaging capacity of AAV is limited to 5 kilobases, presenting a significant obstacle to the efficient expression of both heavy and light chain. As a result, some groups have turned to smaller bivalent single chain antibodies (scFv) or immunoadhesins, chimeric antibody-like molecules combining the functional domain with the immunoglobulin constant domain [23-26]. However, others have employed heterologous viral sequences such as foot and mouth disease virus-derived 2A self-processing sequence (F2A) to express full-length antibodies from a single open reading frame [27], yielding greater than 1000 μg/mL of sustained mAb serum levels in vivo [27,28].

The earliest forms of AAV mediated gene antibody transfer were implemented as a dual-promoter vector whereby the heavy and light chain genes were transcribed independently. This yielded up to 8μg/mL of biologically active an HIV bNAb for over 6 months in immunodeficient Rag mice [29]. In rhesus macaques, expression of SIV gp120-specific immunoadhesins peaked 3-4 weeks post transduction at 200μg/mL and was sustained at 20μg/mL for at least 4 years [30]. Out of nine monkeys challenged with SIV one month after AAV administration, six were completely protected from challenge. The three immunized macaques that became infected were later found to have developed an immune response against the immunoadhesin one week prior to challenge, suggesting that an anti-immunoadhesin response led to the observed failure of protection [30]. AAV vectors were also employed in a similar approach coined ‘Vectored ImmunoProphylaxis’ (VIP), whereby full-length human IgG bNAbs against HIV were expressed from an optimized transgene that utilized the F2A sequence to allow for the expression of independent heavy and light chains under a muscle-optimized promoter [31]. This transgene was packaged with an AAV8 capsid [19], leading to the production of mAb at serum concentrations greater than 100μg/mL for at least 52 weeks [31]. Using this system, two different humanized mouse models were protected from either IV challenge with a laboratory strain of HIV [31] or repetitive low-dose vaginal challenge with a more clinically relevant transmitted founder strain (REJO.c) [32]. Decreasing doses of AAV vector led to dose-dependent antibody expression, enabling a determination of the minimum protective dose in vivo for a number of antibodies [31,32]. AAV-vectored bNAbs have also been shown to work in conjunction with passive mAb transfer and HAART to maintain suppression of HIV replication in humanized mice [33]. Most recently, a synthetic fusion of CD4-Ig with a small CCR5-mimetic sulfopeptide (eCD4-Ig) was delivered by AAV and protected rhesus macaques from several infectious SHIV challenges suggesting that AAV-vectored synthetic proteins may be able to create effective HIV prophylaxis [34].

In addition to HIV, successful VIP has also been demonstrated against a number of other infectious diseases. Recently, sterilizing immunity was generated against a murine model of Plasmodium falciparum infection, the malaria parasite responsible for the highest mortality in humans [35], demonstrating the first known example where a parasitic disease was prevented by antibodies alone. Likewise, VIP-mediated expression of bNAbs against HCV conferred protection against viral challenge in humanized mice [36] and was able to abrogate ongoing HCV infection both in vitro and in vivo. Additionally, sera taken from mice given VIP expressing different bNAbs targeting IAV hemagglutinin (HA), were able to neutralize all H1, H2 and H5 strains tested, and antibody expression lasted well over a year post-AAV injection [37]. Interestingly, protection against influenza by VIP did not appear to inhibit the elicitation of novel immune responses, suggesting that VIP may be capable of augmenting immunity without abrogating endogenous immune responses [37]. Notably, this method also protected immunodeficient and older mice from disease, representing two particularly vulnerable human patient populations who are inadequately protected by traditional vaccination [37]. While intramuscular (IM) injection of AAV for expression of antibodies has been most common, other routes of administration and target sites have also been used successfully. One study used AAV9 to deliver IAV bNAb F16 by intranasal administration, which resulted in protection at the primary site of challenge [38]. AAV has also been used to generate long-term expression of an RSV antibody [14]. One study tested a combination of AAV and Ad5 to generate rapid protection against anthrax that lasted at least 26 weeks [16].

Vectored Antibody Delivery for Neurodegenerative Diseases

In addition to engineering immunity against infectious disease, the use of AAV as a means of creating desirable antibody specificities in vivo enables targeting of “self” proteins that would be difficult or impossible to target safely through traditional vaccination. As a result, there has been growing interest in utilizing this approach for the treatment and prevention of neurodegenerative diseases, which represent an increasing share of the healthcare burden in developed nations. For many such diseases, aggregation of misfolded proteins has been suggested as the underlying mechanism, making them ideal targets for mAbs that recognize the misfolded variants and prevent the formation of these aggregates. Small scFv antibody fragments lacking the Fc region, including intrabodies (iAb) that target antigens intracellularly, can distinguish highly homologous proteins, different conformations of the same protein and, in the case of some iAb, target proteins to distinct cellular compartments [39-41]. The genes encoding these scFv have been delivered by AAV vectors, representing a powerful new tool for treating or preventing neurodegenerative disease [42,43].

Huntington’s disease (HD) is caused by a mutation in the huntingtin protein (HTT) and is a model for numerous neurodegenerative disorders due to its simple autosomal dominant inheritance. Several anti-HTT iAbs have been generated [44] and two were tested in transgenic HD models using intrastriatal AAV2 delivery [45]. Of the iAbs tested, Happ1 ameliorated neuropathology in cell lines and conferred significant beneficial effects in a variety of motor and cognitive assays, significantly prolonging life span by 10 weeks in a mouse model of disease [45]. Another group delivering iAb C4 via intrastriatal AAV1 delivery demonstrated that early treatment of an HD mouse model, prevented cells from exhibiting nuclear aggregates and delayed aggregate accumulation [46].

Alzheimer’s disease (AD) is a disorder characterized by the diffuse loss of neurons and accumulation of amyloid beta proteins (Aβ) in the brain. Recently, scFvs have been developed that target Aβ, which could reduce Aβ burden and possibly alleviate symptoms [47]. A transgenic mouse model for AD, given an intrahippocampal infusion of AAV1 encoding an Aβ-scFv exhibited lower levels of insoluble Aβ, increased numbers of microglia and demonstrated improved cognitive function [48]. While most studies have administered AAV vectors directly into the brain, this may pose a safety risk as intraventricular delivery of AAV5 led to an increase in hemorrhaging [23]. As an alternate route, IM injection of AAV2 expressing an Aβ-scFv was found to be as effective as intracranial administration in reducing physiologic and behavioral effects of AD without producing detectable inflammatory responses or microhemorrhages in the brain [24,49]. In a separate study, IM injection of AAV1 expressing a full-length Aβ-mAb resulted in sustained anti-Aβ levels above 100μg/mL in serum that were maintained for up to 64 weeks post-injection [50]. These levels were found to be effective in decreasing Aβ levels in the brain both prophylactically and therapeutically [50].

Emerging evidence suggests that misfolding of superoxide dismutase 1 (SOD1) is a common pathogenic event in amyotrophic lateral sclerosis (ALS)[51]. AAV encoding an scFv specific for misfolded SOD1, was injected intrathecally into an ALS mouse model, resulting in reduced neuronal stress, reduced levels of misfolded SOD1 in the spinal cord and attenuation of motor neuron loss (Table 1) [25]. Overall, this led to delayed disease onset and increased life span that directly correlated with antibody titer.

Similarly, this approach has also been investigated to combat prion disease, which is a neurodegenerative disorder caused by a conversion of cellular prion protein (PrP^c) into the misfolded, insoluble, PrP^sc. Mice inoculated peripherally with infectious prions were given an AAV vector expressing scFv targeting PrP^sc. These mice exhibited decreased PrP^sc burden and delayed onset of prion pathogenesis as determined by improvements of clinical signs (Table 1) [26,52].

Vectored Antibody Delivery for Cancer

As a result of improved understanding the molecular basis of cancer, antibody-based drugs have become the standard of care for many types of tumors. Human epidermal growth factor receptor type 2 (HER2) overexpression is associated with reduced survival in cases of human breast cancer. Clinical trials of trastuzumab, an mAb that targets an extracellular region of HER2, have been successful at steady state serum concentrations of greater than 10μg/mL [53]. The murine precursor to trastuzumab was encoded in an AAV rh.10 vector [54] and administered to C57BL/6 mice, resulting in serum concentrations near 35μg/mL within twelve weeks that were sustained for at least 56 weeks post injection. A single injection of this vector increased the survival of Balb/c nu/nu mice injected subcutaneously with Calu-3 tumor cells over-expressing HER2, demonstrating its efficacy at inhibiting cancer in vivo [54].

Similarly, cutaneous T-cell lymphoma (CTCL), exhibits over-expression of chemokine receptor 4 (CCR4), whose expression is limited amongst non-malignant cells [55]. AAV8 was used to express a humanized anti-CCR4 mAb in a tumor mouse model, resulting in reduced CCR4⁺ tumor growth and increased survival. A single IV injection resulted in 96μg/mL of mAb in serum, which was able to significantly reduce tumor growth and increase life-span as compared to control animals [56]. While these results have been promising, clinical translation of vectored antibody delivery for cancer will require the use of transient vectors or platforms for regulated mAb delivery.

Vectored Antibody Delivery for Addiction

Substance abuse creates a substantial healthcare burden and is the target of numerous pharmacological and behavioral interventions. Antibodies that target the addictive substance and prevent receptor signaling could offer a potential treatment, however repeated administration of mAb protein is impractical. Previous studies showed that AAVrh.10 expressing a high affinity anti-nicotine mAb, resulted in a serum concentration of 1.3 mg/mL that lasted for at least 18 weeks [57]. AAV-NIC9D9 mice had a majority of serum nicotine bound to the Fab within one minute after challenge with nicotine, resulting in brain concentrations of nicotine that were only 15% of what was observed in naïve controls. Importantly, the expression of this antibody also blocked nicotine-mediated alterations in arterial blood pressure, heart rate and locomoter activity, demonstrating its ability to obviate the physiological effects of nicotine [57]. Using a similar approach against cocaine, AAVrh.10 was engineered to express the high affinity anti-cocaine mAb GNC92H2 [58], leading to the expression of anti-cocaine antibodies for at least 24 weeks after IV administration and resulting in a 31-fold reduction in the ratio of brain to blood cocaine levels and reduced hyperactivity in treated mice as compared to controls [58].

Conclusion

Successful translation of vectored antibody gene delivery to patients is poised to redefine the landscape of immunological interventions by enabling precise engineering of the specificity and intensity of a desirable humoral response. It goes well beyond the ability of traditional vaccines to enable production of non-natural antibody architectures capable of discriminating between normal and aberrant forms of self-proteins. By circumventing the natural immune system, vectored antibody delivery has the potential to yield protection regardless of immune-status or age, allowing it to reach currently vulnerable populations of patients who cannot respond to vaccines and offering a possible alterative to existing therapies delivered by passive transfer.

Key Points.

Vectored antibody delivery represents a new strategy for the production of specific monoclonal antibodies in vivo.

Broadly neutralizing antibodies have been shown to prevent HIV, IAV, HCV, RSV, malaria and anthrax infection in various animal models.

Vectored antibody delivery has the potential to be used as a therapeutic for neurodegenerative diseases, cancer and substance abuse.

This approach represents an alternative vaccine strategy for vulnerable populations that cannot respond to vaccines or for diseases that are able to evade immunity.

Abbreviations

rAAV

recombinant aden-associated virus

scAAV

selfcomplementary adeno-associated virus

Ad5

adenovirus serotype 5;, fragmentantigen binding

scFv

single-chain variable fragment

CMV

cytomegalovirus

hPGK

human phosphoglycerate kinase promoter

EF1α

elongation factor 1α

N/A

not applicable

MOI

multiplicity of infection

GC

genome copy

PFU

plaque forming unit

IM

intramuscular

IV

intravenous