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Rabies vaccines: where do we stand, where are we heading? Expert Review of Vaccines Downloaded from informahealthcare.com by Korea University on 01/06/15 For personal use only.
Expert Rev. Vaccines Early online, 1–13 (2014)
Manpreet Kaur1,2, Rajni Garg1, Samer Singh*3 and Rakesh Bhatnagar*1 1 BSL3 Laboratory, School of Biotechnology, Jawaharlal Nehru University, New Delhi – 110067, Delhi, India 2 Vaccine and Infectious Disease Research Center, Translational Health Science and Technology Institute, An autonomous institute of Department of Biotechnology, Govt. of India, Gurgaon – 122016, Haryana, India 3 Department of Microbial Biotechnology, Panjab University, Chandigarh – 160014, India *Authors for correspondence: Tel.: +91 112 670 4079 Fax: +91 112 674 2040 [email protected]; [email protected]
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Rabies being the most lethal zoonotic, vaccine-preventable viral disease with worldwide distribution of reservoir wild animals presents unique challenges for its diagnosis, management and control. Although vaccines available are highly effective, which had played the key role in controlling rabies in North America, western Europe and in a number of Asian and Latin American countries, the requirement of multiple doses along with boosters, associated cost to reduce the incidence in wild animals and prophylactic human vaccination has remained a major impediment towards achieving the same goals in poorer parts of the world such as sub-Saharan Africa and southeast Asia. Current efforts to contain rabies worldwide are directed towards the development of more safe, cheaper and efficacious vaccines along with anti-rabies antibodies for post-exposure prophylaxis. The work presented here provides an overview of the advances made towards controlling the human rabies, particularly in last 10 years, and future perspective. KEYWORDS: encephalitis • glycoprotein • in vivo challenge • protection • rabies virus-neutralizing antibody • survival • therapeutics • vaccines
Rabies (Latin meaning ‘rage or madness’) is one of the oldest zoonosis known to mankind. It is primarily a disease of terrestrial and airborne mammals, including dogs, wolves, foxes, coyotes, jackals, cats, bobcats, lions, mongooses, skunks, badgers, bats, monkeys and humans [1]. It’s an acute, almost invariably fatal progressive encephalitis caused by viruses belonging to genus Lyssavirus of Rhabdoviridae family, within the order Mononegavirales [2]. The great majority (~99%) of human infections is reported to be caused by rabies virus but more than a dozen other members of Lyssavirus genus exist that are supposed to have potential to cause clinical rabies [3,4]. Lyssavirus genus has been classified on the basis of genetic distances and serological cross-reactivity into three phylogroups. The rabies virus belongs to phylogroup I. Most studies on rabies including the development of vaccine against rabies are based on rabies virus, so vaccination and protective strategies developed against rabies virus are not supposed to be protective against rabies caused by viruses belonging to phylogroups II and III of the Lyssavirus genus. Unless specified otherwise any
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reference to ‘rabies’ in the text hereafter refers to rabies caused by rabies virus. Rabies is characterized by neuronal dysfunction and lesions, which are more prominent in the brain stem, as compared with other areas of central nervous system (CNS) [5]. The people who fall victim to death by rabies include mostly people of low socioeconomic background because they cannot afford post-exposure prophylaxis (PEP). Human rabies has been eradicated in some developed countries [6], but it continues to take its toll in developing countries. In Asia and Africa, rabies causes approximately 55,000 deaths annually. Pre-exposure prophylaxis (PrEP) of rabies vaccine is recommended for people who are travelling to high-risk area, or who are occupationally involved with wild animal reservoirs like cave hunters or scientists working on rabies-infected animals. PEP is used for people who are bitten by rabid animals. The cell culture vaccines and embryonated egg-based vaccines (CCEEV) namely, human diploid cell vaccine (HDCV), purified chick embryo cell vaccine (PCECV), purified Vero cell vaccine (PVCV) and purified duck embryo vaccine (PDEV) are recommended by WHO for human
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Kaur, Garg, Singh & Bhatnagar
use [3]. Despite their potency, these vaccines show some side effects such as headache, dizziness and systemic reactions like fever and gastrointestinal symptoms, occurring at low frequencies [7]. Multiple doses of CCEEVs are required for PrEP and PEP along with rabies immunoglobulin (RIG), in case of transdermal bite, scratches, licks on broken skin by a rabid animal, making the rabies control expensive [8]. Focused efforts need to be made in the direction of development of a safe, cost-effective vaccine that could provide consistent protection, desirably in a single dose. This review aims to summarize the current scenario and the future strategies for rabies vaccine development.
Prophylaxis PrEP
PrEP is recommended for people at continued, frequent or increased risk of exposure to the rabies virus either as a result of their residence or occupation. Travelers with extensive outdoor exposure and children living in endemic areas are at particular risk. PrEP may be performed with any of the modern cell-derived vaccines. Booster doses increase and prolong the antibody response. Thus, they should be offered to persons at continuing risk every 1–3 years.
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PEP
Rabies virus
The rabies virion is classic bullet-shaped structure with one end rounded and other planar. It comprises an external viral envelope, the structural unit and a helical ribonucleocapsid core – the internal functional unit. It has a negative sense 12 kb non-segmented ssRNA genome that codes for five proteins (RNA polymerase [L], nucleoprotein [N], phosphoprotein [P], matrix protein [MP], glycoprotein-G [G]) and four intergenic regions. Three proteins, RNA polymerase (L), nucleoprotein (N) and phosphoprotein (P), are tightly associated with the RNA genome to form ribonucleoprotein core [9]. The MP envelopes the ribonucleoprotein, keeps it condensed and targets it to the plasma membrane and incorporates G-protein into the newly forming viral particles [10]. G-protein forms spikes on the viral surface and is the major inducer of apoptosis. Antibody response against G-protein is the primary determinant of protective immunity as it is the primary target of the host humoral [11] and cell-mediated immune responses [12,13]. Additionally, G-protein because of being the only exposed protein on the viral surface is usually targeted for developing vaccines. The clinical spectrum of rabies manifestation is divided into the prodromal phase, acute neurological or excitation phase and coma or terminal phase. The incubation period of rabies is dependent upon the virus involved, the distance of the site of bite from CNS and the size and innervations of the bite site and may vary from 5 days to many years (usually 2–3 months; rarely >1 year). The patient’s age and genetic background also have been speculated to have bearing on incubation period of rabies similar to other neurotropic viruses [14]. Once the symptoms have set in, rabies generally proceeds to the terminal phase after 10 days leading to cardio-respiratory arrest and death. Most of the human rabies cases develop as a result of bite from an infected animal, usually dog. Human-to-human transmission is extremely rare, but has been reported in certain organ transplant cases [5]. Successful implementation of rabies control programs in canines, primarily dogs, has almost eliminated the incidence of human rabies in North America [15] and greatly reduced it in Latin America and the Caribbean [16]. Asia has the highest number of rabies cases with India on the top, contributing to one-third of the total global rabies deaths. doi: 10.1586/14760584.2015.973403
Rabies is almost always fatal, once the clinical symptoms have set in. Recovery and survival is extremely rare. It is thus essential that individuals be given immediate PEP after any case of exposure. This involves instant wound cleaning and immunizations. The immunization initially included five doses of CCEEV along with human RIG (HRIG), in the deltoid area of adults or in the anterolateral thigh of young children [3]. In 2008, Advisory Committee on Immunization Practices recommended to reduce dosage to four, keeping HRIG dose intact because fifth dose was not required according to experimental data and epidemiological surveys [17]. Accordingly, three doses of CCEEV (1/0.5 ml) need to be administered on days 0, 7 and 21 or 28. WHO has still not incorporated these modifications in its recommendations [18]. However, based on extensive research in developing countries; WHO has now recommended reduced dose regimens, thereby enhancing compliance by the patients as well as cost–effectiveness of the program. One alternate intramuscular regimen recommends vaccine (1/0.5 ml) at two sites on day 0, one site on day 7 and day 21 each. Two modified intradermal (i.d.) regimens are also proposed: these are eight-site i.d. regimen containing 0.1 ml injections at eight sites on day 0, four sites on day 7 and one site each on day 28 and day 90, and two-site i.d. regimen containing 0.1 ml immunizations at two sites each on day 0, two sites each on day 3, two sites each on day 7 and two sites each on day 28. HRIG immunization on day 0 still finds a mandatory position in the regimen. Another modification has been proposed in the eight-site i.d. regimen, making it four site, which reduces the hospital visits to three and the amount of vaccine to half [19]. In case of patients previously immunized with standard vaccination regimen, a single-visit four-site i.d. injections of 0.1 ml equally distributed over left and right deltoids or thighs may be an equally viable option [20]. Despite vast usage of HRIG, their safety and availability remains a cause of concern. WHO has proposed recombinant monoclonal antibodies (mAbs) as suitable alternatives for the PEP. Recently, a recombinant human IgG1 anti-rabies mAb (SII RmAb) has been developed in India. In the dose-escalation Phase I study, SII RmAb was found to be safe and the elicited neutralizing serum antibody titers comparable to HRIG when administered in conjunction with rabies vaccine in a simulated PEP regimen [21]. Bakker et al. showed that mAbCL184, generated by combination of mAbs CR57 and CR4098, is safe and Expert Rev. Vaccines
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Rabies vaccines
does not interfere with the development of subsequent vaccineinduced rabies-virus neutralizing antibodies [22]. mAbCL184 Phase II trials were completed in 2008, in children and adolescents in the Philippines [23] and in adults in the USA [24], followed by a third trial in adults in India that was completed in late 2012 [25,26]. These results demonstrate the feasibility and potential of mAbs for PEP. In another study, rabies virusinfected suckling mouse brain was used to generate 11 hybridoma cells secreting mAbs against rabies virus. These mAbs exhibited strong reactivity with rabies-infected Vero cells. Out of these, mAb 4B7 recognized the recombinant nucleoprotein (N) of rabies virus, while seven other reacted specifically with phosphoprotein (P) of rabies virus. Hence, these mAbs may prove useful in studying interactions between rabies virus and its host, and for diagnostic purpose. More importantly, three of the mAbs (1B11, 1C8 and 8H12) showed a neutralizing effect toward rabies virus [27]. Advances in antibody engineering have led to their improved protective efficacy and potent clinical applications. Turki et al. reported the generation of a trivalent single-chain Fv (scFv50AD1-Fd), which showed a high neutralization activity up to 75-fold higher than that in monovalent form at an equivalent concentration [28]. This was more than three fold than that of WHO standard serum recommendations [29]. The multivalency of single-chain Fv not only improved the avidity, but also the biological activity, emphasizing its potential in rabies PEP. Plant-based expression systems provide a safe and economically viable platform for generation of therapeutic and diagnostic mAbs. In a study, mAbP SO57 and its endoplasmic reticulum targeted variant were produced in transgenic tobacco plants (Nicotiana tabacum). Both were found to be effective in neutralizing rabies virus challenge virus standard (CVS)-11 [30]. In another study, the chimeric (mouse-human) version of mAb 62-71-3 was expressed in Nicotiana benthamiana. The mAb exhibited rabies as well as pseudotyped virus neutralization [31]. These studies establish the usage of plant-based expression system for the development of effective and inexpensive antibody products. Vaccines
The development of rabies vaccines has a long and distinguished history. However, the urgent need of safe and efficacious vaccines has stimulated their further refinement. Conventional vaccines
The first rabies vaccine was developed by Louis Pasteur in 1885 by using spinal cord of rabbits infected with rabies. His post-exposure administration of the desiccated spinal cords, which were supposed to contain partially inactivated infectious agent, to a boy named Joseph Meister who was bitten by a rabid dog, protected him from developing rabies. It heralded the era of rabies vaccine development. Based on same principle, David Semple developed and introduced relatively safer nerve tissue vaccine that consisted of phenol-inactivated sheep or goat informahealthcare.com
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brain tissue [32]. For a very long time, these nerve tissue vaccines had remained the pillars of rabies immunization efforts. Though very effective, they were associated with serious side effects like demyelinating allergic encephalitis and required cumbersome and painful 14–21 shots for generating effective immunity [33]. Later, their use was discontinued and they were replaced with modified and safer vaccines. One such newgeneration vaccine was purified duck embryo vaccine (PDEV), which used embryonated duck eggs for propagation of fixed virus strains, which were inactivated by b-propiolactone [34]. HDCV was introduced in 1978, in which Fixed Rabies virus Pitman-Moore L503 strain was used that was adapted for growth in normal human fibroblast cell line. This vaccine proved to be efficacious as well as safer in comparison to previous vaccines. It imparted complete protection to human subjects against silver haired bat virus strain and was as effective as the other available PCECV, which was obtained by growing fixed Flury LEP-50 virus strain in chick fibroblasts [35]. Though owing to ease of production, PCECV is a cheaper alternative; it also produces urticaria and pruritus in few of its recipients. Currently, efficacious and safe vaccines for human use are HDCV, PCECV and purified Vero cell rabies vaccine (PVRV), which uses inactivated Wistar strain of rabies virus grown on Vero cell cultures [36]. Another less frequently used vaccine is primary hamster kidney cell vaccine, which uses the fixed Beijing strain cultured in primary hamster kidney cells [7]. Development of improved vaccines Recombinant live vaccines
The efficacy of PrEP and PEP therapy is often compromised in the poorer areas of the world, where rabies is highly endemic due to lack of vaccination compliance both in human and animal. The poor vaccine compliance partially results from lack of awareness, poor infrastructure, unavailability of vaccines and most importantly the associated high cost. Though oral rabies vaccination of dogs have been known for several years, owing to the risk associated with the dissemination of replicating antigens in young and/or immunodepressed individuals; extensive vaccination program has not been undertaken. Additionally, this does not yield in levels of protective antibody sufficient to break the dog-to-dog transmission cycles or maintain immunity. Other concerns include the preconditioned requisite of suitable bait well accepted by the target population. The artificial or manufactured baits need to be imported, which is an expensive proposition for countries with limited financial resources. Many times, these are not readily accepted by dogs due to unfamiliar taste, odor and texture of baits. This usually affects subsequent vaccination attempts [37]. Locally baits could be probable solution to both cost–effectiveness and general acceptance. The overall safety remains a cause of immense concern. Esh et al. demonstrated that the Evelyn Rokitniki Abelseth strain, generally oral fox vaccination causes fatal rabies in cats [38]. Oral vaccine candidates are usually attenuated derivatives of Street Alabama doi: 10.1586/14760584.2015.973403
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Dufferin (SAD) strain. They are generally achieved by a single amino acid exchange, which may not lead to a stable nonpathogenic phenotype, as elegantly shown by Faber et al. They found that rabies virus attenuation through an amino acid substitution for Arg333 can revert back to pathogenicity upon serial passage of the virus in mouse brain through the introduction of mutations at other positions of the glycoprotein gene [39]. Multiple changes in different components may be the key to development of a stable and safe attenuated live vaccine that may have wider acceptability. Additionally, there is a need to develop more immunogenic and preferably single-dose-vaccine as a remedy to failure of the follow-up vaccination in these endemic areas. However, it is unlikely that a single dose of inactivated vaccines will suffice the need of active immunization, as suggested by Strady et al., who showed that a minimum of three inoculations are required to reduce the percentage of non- or poor-responders (100-fold in many countries as the most of rabies-related deaths occur in communities away from regular hospitals [87,88] and many a times they are misdiagnosed for other neurological diseases [89]. Though neural tissue-based vaccines have been discontinued and are not recommended anymore by WHO due to associated side effects and safety issues, they are still in use for PEP in poorer countries such as Algeria and Ethiopia due to cost and poor availability of current generation rabies vaccines. Outside Africa, China and India had remained the major contributors to global rabies burden till last decade, accounting for 25–40% of the total global mortality from rabies [86].The efforts by Chinese and Indian governments in last decade to implement dog population control measures, vaccination of dogs, making available modern vaccine and improving the accessibility to PEP measures have improved the situation leading to significant decrease in incidences and stabilization [3,86]. To further improve the situation in these countries more efforts are required to educate the population in remote areas, particularly who do not have doi: 10.1586/14760584.2015.973403
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Table 1. Different rabies vaccines. Vaccine name
Composition
Limitations/problems
Ref.
Semple vaccine
Phenol or b-propiolactone inactivated homogenate of rabies virus-infected goat or sheep brain tissue
– Pain, swelling, erythema – Fever, headache, insomnia – Can cause demyelinating allergic encephalitis, due to myelin basic protein – Less immunogenic, therefore, required 14–21 shots – Discontinued by WHO since 1984
[7,93]
Fuenzalida rabies vaccine
Suckling mouse brain tissue
– Pain, swelling, erythema – Fever, headache, insomnia – Decreased myelin basic protein, therefore, decreased allergic reactions – Post-vaccination neuroparalytic syndromes resembling Guillain–Barre´ syndrome
Human diploid cell vaccine
Pitman-Moore L503 or Flury strain of rabies virus grown on MRC-5 human diploid cell culture
– Fever, headache, dizziness – Immune complex-like reaction – Neurological disease
[35]
Purified chick embryo cell vaccine
b-Propiolactone inactivated fixed Flury LEP-25 rabies virus strain grown in primary cultures of chick fibroblasts
– Urticaria, pruritus – Similar but less frequent local, systemic reactions and neurological reactions than HDCV
[35]
Purified Vero cell rabies vaccine
b-Propiolactone inactivated Wistar strain of rabies virus grown in Vero cells
Local, systemic and neurological reactions occurring at a frequency similar to purified chick embryo cell vaccine
[36]
Primary hamster kidney cell vaccine
Formalin inactivated Beijing strain, adsorbed to aluminum hydroxide
– Redness, itching and pain at injection site – Fever, headache, dizziness – Pain in joints and lumbar pain in some
[94]
Purified duck embryo vaccine
b-Propiolactone inactivated fixed virus strain grown in duck embryo cells
– Pain at the injection site – Allergic reactions – Neurological disease, at a rate lower than HDCV
[34]
[7]
HDCV: Human diploid cell vaccine.
immediate access to appropriate care and there is lack of awareness. In South America, particularly remote/hard-to-access areas of Brazil, Colombia, Ecuador and Peru in the Amazon region may be another hotspot of rabies due to presence of human rabies transmitting vampire bats, lack of awareness and at the same time unavailability of proper care [90]. Combined together, to effectively control the disease and eventually eliminate rabies, we would need to focus on reaching remote regions of the world, educate the population, dispelling myths besides making the PrEP and PEP readily available to masses and put in place comprehensive plans to control animal rabies. It could be made possible only by joint cooperative and collaborative efforts involving local populace, volunteers, public healthcare professionals, scientists, local governments, charity institutions, WHO and all the other stakeholders. In this direction, some excellent endeavors have been already put in place in ASEAN countries and Latin American countries that intend to achieve these goals [86]. An attractive strategy to decrease the overall global burden of human rabies, particularly in poorer countries/regions would be to incorporate rabies vaccination into existing child immunization programs [3], as it could reduce the economic, logistical or programmatic obstacles substantially. Any positive step taken by respective doi: 10.1586/14760584.2015.973403
regional cooperative organizations or individual nations with endemic rabies may go a long way in controlling rabies. Some additional points that may not be directly linked to rabies vaccine development but pertinent to our overall goal of rabies control/management are being described briefly. There is dearth of studies replicating overall pathogenesis, disease progression and intensive care requirement of human subjects in animal models due to unavailability of suitable animal models. Urgent efforts need to be made to find suitable animal models then carry out such studies as they may help us better formulate therapeutic intervention strategies to improve the outcome of rabies-patient care. There had been relative lack of interest in developing antiviral therapy for rabies virus. As of now, no commercial antiviral therapy is available for rabies virus. Better understanding of the biology of rabies virus such as interactions among different viral proteins, working of viral replication complex may help us develop small molecule inhibitors for rabies therapy. Research on siRNA that can inhibit rabies virus replication has lot of potential in terms of delaying the onset of rabies symptoms and allowing currently used PEP to take effect [81,91]. Such studies need to be more vigorously pursued as they may be the game changer in saving the life of rabies virus-infected individuals. Expert Rev. Vaccines
Rabies vaccines
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Table 2. Improved rabies vaccines/monoclonal antibodies under clinical development. Vaccine/antibody
Composition
Status
R&D/manufacturer
Ref.
Purified Vero rabies vaccine (PVRV-NG)
Serum-free purified Vero rabies vaccine
Phase II
Sanofi Pasteur
[95]
Chromatographically purified rabies vaccine
Chromatographically purified PVRV, devoid of albumin
Phase II
Pasteur Me´rieux Connaught, France
[96]
Purified Vero cell rabies vaccine
Lyophilized vaccine containing inactivated purified (Pitman Moore, PM3218 as virus strain) produced using Vero ATCC CCL 81 cells
Phase III
Serum Institute of India Ltd
[97]
Anti-rabies monoclonal antibody
Anti-rabies monoclonal antibody
Phase III
Serum Institute of India & Mass Biologics of the University of Massachusetts Medical School
[98]
Rabies immune globulin (human)
Intramuscularly administered rabies immune globulin (human)
Phase I
Grifols Therapeutics Inc
[99]
Rabies vaccines
Purified Vero rabies vaccine and rabies human diploid cell vaccine
Phase II
Sanofi Pasteur
[100]
Rabies vaccine
Purified inactivated Vero rabies vaccine
Phase III
Sanofi Pasteur
[101]
Rabies vaccine-rabies immunoglobulin
Two intradermal rabies vaccine regimens administered with and without human rabies immunoglobulin
Phase III
Novartis
[102]
CL184
Human monoclonal antibody cocktail, containing a 1:1 equipotent mixture of two human monoclonal antibodies (CR57 and CR4098)
Phase II
Sanofi Pasteur
[103]
HRIG-rabies vaccine
HRIG with co-administration of active rabies vaccine
Phase II/III
Kamada Ltd
[104]
HRIG: Human polyclonal anti-rabies immune globulin.
Five-year view
In the last decade, the steady improvement in the availability of safer CCV rabies vaccines and RIGs has greatly enhanced the PEP and PrEP outcomes. It has helped tremendously in controlling spread of rabies and associated mortality at global scale. However, the lack of awareness, cost and availability of vaccines and RIGs and their efficacy, requirement of multiple boosters to mount protective response had been the main impediments toward achieving the desired level of rabies control in the poorer nations. Currently, some of the notable endeavors that are underway and have clear potential to further improve the accessibility and applicability of PEP and PrEP include efforts to cut the cost of available vaccines and RIGs through reduced dosage and altered administration schedule, reduce the PEP and PrEP administration time, make single dose vaccines (TABLES 1 & 2) [3]. Various studies and follow-ups from Thailand, India and other southeast Asian countries have clearly demonstrated the possibility of dose reduction but comparable protection offered by CCV vaccines when given intradermally both in PEP and PrEP scenarios. These reduced dose and less dose regimens would need to be implemented in more rabies-endemic areas. To control the rabies in wild reservoir population, we will have to direct more efforts toward broader application of available oral vaccines. Currently, RIGs are informahealthcare.com
prohibitively expensive. Development of newer biological such as mAbs, which are already at different stages of clinical trials [3,22,92], should become available in near future, effectively reducing the cost and making the passive immunization as a part of PEP accessible to people who can least afford it. In addition to advances envisaged to become available soon that will help improve the PEP and PrEP to control rabies, the development and availability of inexpensive, accurate and easyto-use diagnostics such as the direct rapid immunohistochemical test [80] in poor settings, could tremendously improve the rabies surveillance allowing real-time adjustment and modification in any comprehensive rabies control program. Combined together, all these developments will greatly improve our chances of controlling/eliminating rabies in next 5–10 years. Financial & competing interests disclosure
This work was supported by Department of Biotechnology (DBT), Department of Science and Technology, Council of Scientific and Industrial Research, Indian Council of Medical Research, India and NATP-World Bank. S Singh is a Ramalingaswami Fellow, supported by DBT, India. 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. doi: 10.1586/14760584.2015.973403
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Key issues • Rabies, a zoonosis, is a fatal progressive encephalitis caused by the infection of the members of the Lyssavirus genus. All mammals are susceptible to rabies but animals from the orders of Carnivora and Chiroptera are primary reservoirs, that is, dogs, foxes, jackals, coyotes, skunks, raccoons, mongoose and bats. • Annually about 60,000 people die of rabies. Most of the human rabies cases result from rabid dog bites (>98%), especially in Asia and Africa or bat bites, especially in Americas, Europe and Australia. Very rarely saliva, body fluids and organ transplants originating from infected individuals or bites of other reservoir animals have been reported as a cause of rabies. • Rabies is a 100% preventable disease if proper prophylactic vaccination is provided as a part of the pre-exposure prophylaxis (PrEP) or along with anti-rabies serum (rabies immunoglobins [RIGs]) as a part of the post-exposure prophylaxis (PEP). No one immunized with PrEP and PEP vaccines has died of rabies following coming in contact with a rabid animal. Expert Review of Vaccines Downloaded from informahealthcare.com by Korea University on 01/06/15 For personal use only.
• The complications associated with overwhelming replication of Lyssavirus in CNS are supposed to cause rabies-related death. The nerve tissue vaccine that was widely used up to about a decade ago in many parts of the world though highly effective had serious side effects such as demyelinating allergic encephalitis. Now, it has been replaced with more effective and safer cell culture vaccines in most parts of the world. • Despite the availability of effective rabies vaccines, the reach of vaccines to the patients in the poorer regions of the world had been limited by a combination of factors such as cost, availability, access to preventive medical care, awareness/ignorance and misconceptions about rabies. • To effectively control the disease, various awareness programs and vaccination programs including that for the reservoir animals are desired. A number of such programs are in place and many more are being formulated for the endemic areas by local governments, philanthropic institutions and other stakeholders such as WHO. • The animal vaccination program could greatly benefit from the availability of oral rabies vaccines. Some oral vaccines have been successfully used/tested in wild animals. Concerted efforts are required to make them available globally, particularly to poorer countries where rabies is endemic. • Development of a safe, single-dose vaccine and the cost reduction of the existing vaccine can significantly reduce the economic burden of rabies vaccination programs worldwide besides improving the PrEP or PEP compliance and eventually success of rabies control programs. • Vaccines under development such as subunit vaccines, attenuated live virus vaccines have the potential to eliminate the need of boosters and reduce the dosage required for PEP or PrEP to one, while development of plant-based edible-vaccines could significantly improve the reach and reduce the associated cost of vaccine delivery and administration. • Future rabies intervention efforts should include the development of strategies/small molecules to slow down/stop the replication/spread of virus allowing the PEP to take effect. Development of anti-viral drugs or interfering RNA that may delay the viral replication/spread should be made priority. Although it has tremendous potential to bring down mortality from rabies, the efforts in this direction have been scanty. • Future vaccine development efforts need to include targeting of the members of the phylogroup II and III of Lyssavirus genus to make the vaccine ready for any such eventuality that may see zoonosis of these Lyssavirus members or in a rare case emergence of some recombinant viruses. Available rabies vaccines and those under various stages of development primarily target viruses from phylogroup I of Lyssavirus genus and are presumed to be ineffective against the infection of the members of phylogroup II and III.
References
4.
Fooks AR, Banyard AC, Horton DL, et al. Current status of rabies and prospects for elimination. Lancet 2014. [Epub ahead of print]
9.
Wunner WH, Larson JK, Dietzschold B, Smith CL. The molecular biology of rabies viruses. Rev Infect Dis 1988;10(Suppl 4): S771-84
5.
Consales CA, Bolzan VL. Rabies review: immunopathology, clinical aspects and treatment. J. Venom. Anim. Toxins Incl. Trop. Dis 2007;13(1):5-38
10.
6.
Menezes R. Rabies in India. CMAJ 2008; 178(5):564-6
Mebatsion T, Weiland F, Conzelmann KK. Matrix protein of rabies virus is responsible for the assembly and budding of bullet-shaped particles and interacts with the transmembrane spike glycoprotein G. J Virol 1999;73(1):242-50
7.
WHO. Information sheet. Observed rate of vaccine reactions. Rabies vaccine. World Health Organization; Geneva, Switzerland: 2012
11.
•
Describes side effects of existing vaccines.
Cox JH, Dietzschold B, Schneider LG. Rabies virus glycoprotein. II. Biological and serological characterization. Infect Immun 1977;16(3):754-9
8.
McGettigan JP. Experimental rabies vaccines for humans. Expert Rev Vaccines 2010; 9(10):1177-86
12.
Macfarlan RI, Dietzschold B, Koprowski H. Stimulation of cytotoxic T-lymphocyte responses by rabies virus glycoprotein and
Papers of special note have been highlighted as: • of interest •• of considerable interest 1.
Fishbein DB, Robinson LE. Rabies. N Engl J Med 1993;329(22):1632-8
2.
Francki RIB, Fauquet CM, Knudson DL, Brown F. Classification and nomenclature of viruses: fifth report of international committee on taxonomy of viruses. Arch Virol. 1991. 2(Suppl):1-450
3.
••
WHO Expert Consultation on Rabies. Second Report. WHO Technical Report Series 982. World Health Organization; Geneva, Switzerland: 2013 Includes most current data on rabies prevention and control.
doi: 10.1586/14760584.2015.973403
Expert Rev. Vaccines
Rabies vaccines
Expert Review of Vaccines Downloaded from informahealthcare.com by Korea University on 01/06/15 For personal use only.
identification of an immunodominant domain. Mol Immunol 1986;23(7):733-41
35.
Dietzschold B, Hooper DC. Human diploid cell culture rabies vaccine (HDCV) and purified chick embryo cell culture rabies vaccine (PCECV) both confer protective immunity against infection with the silver-haired bat rabies virus strain (SHBRV). Vaccine 1998;16(17):1656-9
36.
Jaiiaroensup W, Lang J, Thipkong P, et al. Safety and efficacy of purified Vero cell rabies vaccine given intramuscularly and intradermally. (Results of a prospective randomized trial). Vaccine 1998;16(16): 1559-62
37.
Estrada R, Vos A, Rc de L. Acceptability of local made baits for oral vaccination of dogs against rabies in the Philippines. BMC Infect Dis 2001;1:19
38.
Esh JB, Cunningham JG, Wiktor TJ. Vaccine-induced rabies in four cats. J Am Vet Med Assoc 1982;180(11):1336-9
39.
Faber M, Faber M-L, Papaneri A, et al. A single amino acid change in rabies virus glycoprotein increases virus spread and enhances virus pathogenicity. J Virol 2005; 79(22):14141-8
40.
Strady C, Andreoletti L, Baumard S, et al. Immunogenicity and booster efficacy of pre-exposure rabies vaccination. Trans R Soc Trop Med Hyg 2009;103(11):1159-64
41.
Prevec L, Campbell JB, Christie BS, et al. A recombinant human adenovirus vaccine against rabies. J Infect Dis 1990;161(1): 27-30
42.
Yarosh OK, Wandeler AI, Graham FL, et al. Human adenovirus type 5 vectors expressing rabies glycoprotein. Vaccine 1996;14(13):1257-64
43.
Chakrabarti P. “Living versus dead”: the Pasteurian paradigm and imperial vaccine research. Bull Hist Med 2010;84(3): 387-423
Shen CF, Lanthier S, Jacob D, et al. Process optimization and scale-up for production of rabies vaccine live adenovirus vector (AdRG1.3). Vaccine 2012;30(2):300-6
44.
Biswas S, Reddy GS, Srinivasan VA, Rangarajan PN. Preexposure efficacy of a novel combination DNA and inactivated rabies virus vaccine. Hum Gene Ther 2001; 12(15):1917-22
Cenna J, Hunter M, Tan GS, et al. Replication-deficient rabies virus-based vaccines are safe and immunogenic in mice and nonhuman primates. J Infect Dis 2009; 200(8):1251-60
45.
Dorfmeier CL, Lytle AG, Dunkel AL, et al. Protective vaccine-induced CD4(+) T cell-independent B cell responses against rabies infection. J Virol 2012;86(21): 11533-40
46.
Dorfmeier CL, Shen S, Tzvetkov EP, McGettigan JP. Reinvestigating the role of IgM in rabies virus postexposure vaccination. J Virol 2013;87(16):9217-22
47.
Chen Z, Zhou M, Gao X, et al. A novel rabies vaccine based on a recombinant parainfluenza virus 5 expressing rabies virus glycoprotein. J Virol 2013;87(6):2986-93
Healthy Adult Subjects. Available from: http://clinicaltrials.gov/show/NCT00656097
13.
Celis E, Karr RW, Dietzschold B, et al. Genetic restriction and fine specificity of human T cell clones reactive with rabies virus. J Immunol 1988;141(8):2721-8
14.
Hoff G, M S. Rabies and Animal Bite investigation. Missouri Health Department, Kansas City, Kansas. 2005
26.
15.
Blanton JD, Palmer D, Rupprecht CE. Rabies surveillance in the United States during 2009. J Am Vet Med Assoc 2010; 237(6):646-57
Nath A, Tyler KL. Novel approaches and challenges to treatment of central nervous system viral infections. Ann Neurol 2013; 74(3):412-22
27.
16.
Epidemiology and burden of disease. Available from: http://www.who.int/rabies/ epidemiology/en/
17.
Rupprecht CE. Use of a Reduced (4-Dose) Vaccine Schedule for Postexposure Prophylaxis to Prevent Human Rabies Recommendations of the Advisory Committee on Immunization Practices. MMWR 2010;59(RR02):1-9
Zhang J, Ruan X, Zan J, et al. Efficient generation of monoclonal antibodies against major structural proteins of rabies virus with suckling mouse brain antigen. Monoclon. Antib. Immunodiagn. Immunother 2014; 33(2):94-100
•
Depicts efforts to reduce vaccine dosage.
18.
Current WHO guide for rabies pre and post-exposure treatment in humans. Available from: www.who.int/rabies/en/ WHO_guide_rabies_pre_post_ exp_treat_humans.pdf
19.
Warrell MJ, Riddell A, Yu L-M, et al. A simplified 4-site economical intradermal post-exposure rabies vaccine regimen: a randomised controlled comparison with standard methods. PLoS Negl Trop Dis 2008;2(4):e224
20.
WHO. Available from: www.who.int/wer (Weekly Epidemiological record, no. 32, 6 august 2010)
21.
Gogtay N, Thatte U, Kshirsagar N, et al. Safety and pharmacokinetics of a human monoclonal antibody to rabies virus: a randomized, dose-escalation phase 1 study in adults. Vaccine 2012;30(50):7315-20
22.
23.
24.
Bakker ABH, Python C, Kissling CJ, et al. First administration to humans of a monoclonal antibody cocktail against rabies virus: safety, tolerability, and neutralizing activity. Vaccine 2008;26(47):5922-7 Randomized Phase II Trial on Safety and Neutralizing Activity of CL184 and Rabies Vaccine Versus Human Rabies Immune Globulin (HRIG) and Rabies Vaccine in Children and Adolescents. Available from: http://clinicaltrials.gov/show/NCT00708084 A Randomized Phase II Trial to Compare the Safety and Neutralizing Activity of CL184 in Combination With Rabies Vaccine vs. HRIG or Placebo in Combination With Rabies Vaccine in
informahealthcare.com
25.
28.
29.
Rabies Virus Neutralizing Activity and Safety of CL184, a Monoclonal Antibody Cocktail, in Simulated Rabies Post-Exposure Prophylaxis in Healthy Adults. Available from: http://clinicaltrials.gov/show/ NCT01228383
Yang L, Cen J, Xue Q, et al. Identification of rabies virus mimotopes screened from a phage display peptide library with purified dog anti-rabies virus serum IgG. Virus Res 2013;174(1-2):47-51 Turki I, Hammami A, Kharmachi H, Mousli M. Engineering of a recombinant trivalent single-chain variable fragment antibody directed against rabies virus glycoprotein G with improved neutralizing potency. Mol Immunol 2014;57(2):66-73
30.
Lee J-H, Park D-Y, Lee K-J, et al. Intracellular reprogramming of expression, glycosylation, and function of a plant-derived antiviral therapeutic monoclonal antibody. PLoS One 2013;8(8):e68772
31.
Both L, van Dolleweerd C, Wright E, et al. Production, characterization, and antigen specificity of recombinant 62-71-3, a candidate monoclonal antibody for rabies prophylaxis in humans. FASEB J 2013; 27(5):2055-65
32.
33.
34.
•
Review
Mahendra BJ, Madhusudana SN, Sampath G, et al. Immunogenicity, safety and tolerance of a purified duck embryo vaccine (PDEV, VAXIRAB) for rabies post-exposure prophylaxis: results of a multicentric study in India. Hum Vaccin 2010;6(9):721-4 Depicts post-exposure prophylactic potential of purified duck embryo vaccine in Indian population.
doi: 10.1586/14760584.2015.973403
Review 48.
49.
Expert Review of Vaccines Downloaded from informahealthcare.com by Korea University on 01/06/15 For personal use only.
50.
51.
52.
Kaur, Garg, Singh & Bhatnagar
Faber M, Li J, Kean RB, et al. Effective preexposure and postexposure prophylaxis of rabies with a highly attenuated recombinant rabies virus. Proc Natl Acad Sci USA 2009; 106(27):11300-5 Schutsky K, Curtis D, Bongiorno EK, et al. Intramuscular inoculation of mice with the live-attenuated recombinant rabies virus TriGAS results in a transient infection of the draining lymph nodes and a robust, long-lasting protective immune response against rabies. J Virol 2013;87(3):1834-41 Koraka P, Bosch B-J, Cox M, et al. A recombinant rabies vaccine expressing the trimeric form of the glycoprotein confers enhanced immunogenicity and protection in outbred mice. Vaccine 2014;32(36):4644-50 Wang X, Feng N, Ge J, et al. Recombinant canine distemper virus serves as bivalent live vaccine against rabies and canine distemper. Vaccine 2012;30(34):5067-72 Wu Q, Yu F, Xu J, et al. Rabies-virusglycoprotein-pseudotyped recombinant baculovirus vaccine confers complete protection against lethal rabies virus challenge in a mouse model. Vet Microbiol 2014;171(1-2):93-101
53.
MERCK MANUALS. Available from: www.merckmanuals.com/vet/pharmacology/ vaccines_and_immunotherapy/ active_immunization.html
54.
Saxena M, Van TTH, Baird FJ, et al. Pre-existing immunity against vaccine vectors-friend or foe? Microbiology 2013; 159(Pt 1):1-11
55.
56.
Daniell H. Production of biopharmaceuticals and vaccines in plants via the chloroplast genome. Biotechnol J 2006;1(10):1071-9 McGarvey PB, Hammond J, Dienelt MM, et al. Expression of the rabies virus glycoprotein in transgenic tomatoes. Biotechnology 1995;13(13):1484-7
57.
Yusibov V, Hooper DC, Spitsin S V, et al. Expression in plants and immunogenicity of plant virus-based experimental rabies vaccine. Vaccine 2002;20(25-26):3155-64
58.
Ashraf S, Singh PK, Yadav DK, et al. High level expression of surface glycoprotein of rabies virus in tobacco leaves and its immunoprotective activity in mice. J. Biotechnol 2005 119(1):1-14
59.
Loza-Rubio E, Rojas E, Go´mez L, et al. Development of an edible rabies vaccine in maize using the Vnukovo strain. Dev. Biol 2008;131:477-82
60.
Desme´zie`res E, Jacob Y, Saron MF, et al. Lyssavirus glycoproteins expressing immunologically potent foreign B cell and
doi: 10.1586/14760584.2015.973403
cytotoxic T lymphocyte epitopes as prototypes for multivalent vaccines. J Gen Virol 1999;80(Pt 9):2343-51 61.
Da Cruz FW, McBride AJ, Conceic¸a˜o FR, et al. Expression of the B-cell and T-cell epitopes of the rabies virus nucleoprotein in Mycobacterium bovis BCG and induction of an humoral response in mice. Vaccine 2001;20(5-6):731-6
62.
Mansfield KL, Johnson N, Fooks AR. Identification of a conserved linear epitope at the N terminus of the rabies virus glycoprotein. J Gen Virol 2004;85(Pt 11): 3279-83
63.
Xiang ZQ, Spitalnik S, Tran M, et al. Vaccination with a plasmid vector carrying the rabies virus glycoprotein gene induces protective immunity against rabies virus. Virology 1994;199(1):132-40
64.
Perrin P, Jacob Y, Aguilar-Se´tien A, et al. Immunization of dogs with a DNA vaccine induces protection against rabies virus. Vaccine 1999;18(5-6):479-86
65.
Lodmell DL, Ray NB, Parnell MJ, et al. DNA immunization protects nonhuman primates against rabies virus. Nat Med 1998;4(8):949-52
66.
Margalith M, Vilalta A. Sustained protective rabies neutralizing antibody titers after administration of cationic lipid-formulated pDNA vaccine. Genet Vaccines Ther 2006;4:2
67.
Lodmell DL, Ray NB, Ulrich JT, Ewalt LC. DNA vaccination of mice against rabies virus: effects of the route of vaccination and the adjuvant monophosphoryl lipid A (MPL). Vaccine 2000;18(11-12):1059-66
68.
Pinto AR, Reyes-Sandoval A, Ertl HCJ. Chemokines and TRANCE as genetic adjuvants for a DNA vaccine to rabies virus. Cell Immunol 2003;224(2):106-13
69.
Lodmell DL, Ray NB, Ewalt LC. Gene gun particle-mediated vaccination with plasmid DNA confers protective immunity against rabies virus infection. Vaccine 1998;16(2-3): 115-18
70.
Lodmell DL, Ewalt LC. Rabies vaccination: comparison of neutralizing antibody responses after priming and boosting with different combinations of DNA, inactivated virus, or recombinant vaccinia virus vaccines. Vaccine 2000;18(22):2394-8
71.
Lodmell DL, Parnell MJ, Bailey JR, et al. Rabies DNA vaccination of non-human primates: post-exposure studies using gene gun methodology that accelerates induction of neutralizing antibody and enhances neutralizing antibody titers. Vaccine 2002; 20(17-18):2221-8
72.
Osorio JE, Tomlinson CC, Frank RS, et al. Immunization of dogs and cats with a DNA vaccine against rabies virus. Vaccine 1999;17(9-10):1109-16
73.
Kaur M, Rai A, Bhatnagar R. Rabies DNA vaccine: no impact of MHC class I and class II targeting sequences on immune response and protection against lethal challenge. Vaccine 2009;27(15):2128-37
••
Indicates that the directing target sequence is not the exclusive deciding factor for type and extent of immune response elicited and emphasizes on the antigen dependence of immune enhancement strategies.
74.
Kaur M, Saxena A, Rai A, Bhatnagar R. Rabies DNA vaccine encoding lysosome-targeted glycoprotein supplemented with Emulsigen-D confers complete protection in preexposure and postexposure studies in BALB/c mice. FASEB J 2010;24(1):173-83
••
Demonstrates that the optimized DNA vaccine formulation provides an avenue for preventing as well as controlling rabies.
75.
Tao L, Ge J, Wang X, et al. Generation of a recombinant rabies Flury LEP virus carrying an additional G gene creates an improved seed virus for inactivated vaccine production. Virol J 2011;8:454
76.
Liu X, Yang Y, Sun Z, et al. A recombinant rabies virus encoding two copies of the glycoprotein gene confers protection in dogs against a virulent challenge. PLoS One 2014;9(2):e87105
77.
Wenqiang J, Yin X, Lan X, et al. Development of a reverse genetics system for the aG strain of rabies virus in China. Arch Virol 2014;159(5):1033-8
78.
Norton JE, Lytle AG, Shen S, et al. ICAM-1-based rabies virus vaccine shows increased infection and activation of primary murine B cells in vitro and enhanced antibody titers in-vivo. PLoS One 2014; 9(1):e87098
79.
Fenje P, Pinteric L. Potentiation of tissue culture rabies vaccine by adjuvants. Am J Public Health Nations Health 1966;56(12): 2106-13
80.
Ullas PT, Madhusudana SN, Desai A, et al. Enhancement of immunogenicity and efficacy of a plasmid DNA rabies vaccine by nanoformulation with a fourth-generation amine-terminated poly(ether imine) dendrimer. Int J Nanomedicine 2014;9: 627-34
Expert Rev. Vaccines
Rabies vaccines
81.
Expert Review of Vaccines Downloaded from informahealthcare.com by Korea University on 01/06/15 For personal use only.
82.
Gupta PK, Sonwane AA, Singh NK, et al. Intracerebral delivery of small interfering RNAs (siRNAs) using adenoviral vector protects mice against lethal peripheral rabies challenge. Virus Res 2012;163(1):11-18
90.
Wiktor TJ, Macfarlan RI, Reagan KJ, et al. Protection from rabies by a vaccinia virus recombinant containing the rabies virus glycoprotein gene. Proc Natl Acad Sci USA 1984;81(22):7194-8
91.
Israsena N, Mahavihakanont A, Hemachudha T. Rabies virus infection and microRNAs. Adv Virus Res 2011;79:329-44
92.
83.
Cadoz M, Strady A, Meignier B, et al. Immunisation with canarypox virus expressing rabies glycoprotein. Lancet 1992; 339(8807):1429-32
84.
Cliquet F, Gurbuxani JP, Pradhan HK, et al. The safety and efficacy of the oral rabies vaccine SAG2 in Indian stray dogs. Vaccine 2007;25(17):3409-18
85.
86.
Blancou J, Artois M, Brochier B, et al. Safety and efficacy of an antirabies vaccine consisting of recombinant vaccinia-rabies virus administered orally to the fox, dog and cat. Ann Rech Vet 1989;20(2):195-204 World Health Organization. Sustaining the drive to overcome the global impact of neglected tropical diseases. Second WHO report on neglected tropical diseases. WHO, Geneva, Switzerland; 2013
87.
Cleaveland S, Fe`vre EM, Kaare M, Coleman PG. Estimating human rabies mortality in the United Republic of Tanzania from dog bite injuries. Bull World Health Organ 2002;80(4):304-10
88.
Hampson K, Dobson A, Kaare M, et al. Rabies exposures, post-exposure prophylaxis and deaths in a region of endemic canine rabies. PLoS Negl Trop. Dis 2008;2(11): e339
89.
Mallewa M, Fooks AR, Banda D, et al. Rabies encephalitis in malaria-endemic area, Malawi, Africa. Emerg Infect Dis 2007; 13(1):136-9
informahealthcare.com
Schneider MC, Romijn PC, Uieda W, et al. Rabies transmitted by vampire bats to humans: an emerging zoonotic disease in Latin America? Rev Panam Salud Publica 2009;25(3):260-9
Review
org/pub_releases/2012-08/uomm-ctf080712. php 99.
An Open-label, Single-Arm Study to Evaluate the Safety and Rabies Virus Antibody Titers of Intramuscularly Administered Rabies Immune Globulin (Human). Available from: http:// clinicaltrials.gov/show/NCT02139657
Mu¨ller T, Dietzschold B, Ertl H, et al. Development of a mouse monoclonal antibody cocktail for post-exposure rabies prophylaxis in humans. PLoS Negl Trop Dis 2009;3(11):e542
100.
Study of Purified Vero Rabies Vaccine and Rabies Human Diploid Cell Vaccine in a Simulated Rabies Post-exposure Regimen. Available from: http://clinicaltrials.gov/ct2/ show/NCT01877395?term=rabies&rank=1
93.
WHO. Rabies. Available from: http://www. who.int/biologicals/vaccines/rabies/en/
101.
94.
Lin F, Zeng F, Lu L, et al. The primary hamster kidney cell rabies vaccine: adaptation of viral strain, production of vaccine, and pre- and postexposure treatment. J Infect Dis 1983;147(3):467-73
Study of the Purified Vero Rabies Vaccine Serum Free in Comparison With the Reference Purified Vero Rabies Vaccine. Available from: http://clinicaltrials.gov/ct2/ show/NCT01339312?term=rabies&rank=4
102.
Safety and Immunogenicity of Two Intradermal Rabies Vaccine Regimens Administered With and Without Human Rabies Immunoglobulin in Subjects ‡ 1 Years of Age. Available from: http://clinicaltrials.gov/ct2/show/ NCT02177032?term=rabies&rank=8
103.
A Randomized Phase II Trial to Compare the Safety and Neutralizing Activity of CL184 in Combination With Rabies Vaccine vs. HRIG or Placebo in Combination With Rabies Vaccine in Healthy Adult Subjects. Available from: http://clinicaltrials.gov/ct2/show/ NCT00656097?term=rabies&rank=13
104.
Phase II/III Study of the Safety and Effectiveness of HRIG With Co-administration of Active Rabies Vaccine in Healthy Subjects (KAMRAB-003). Available from: http://clinicaltrials.gov/ct2/ show/NCT02040090?term=rabies&rank=20
95.
Pichon S, Guinet-Morlot F, Minutello M, et al. A serum-free, purified vero cell rabies vaccine is safe and as immunogenic as the reference vaccine Verorab for pre-exposure use in healthy adults: results from a randomized controlled phase-II trial. Vaccine 2013;31(18):2295-301
96.
Lang J, Cetre JC, Picot N, et al. Immunogenicity and safety in adults of a new chromatographically purified Vero-cell rabies vaccine (CPRV): a randomized, double-blind trial with purified Vero-cell rabies vaccine (PVRV). Biologicals 1998; 26(4):299-308
97.
Kulkarni PS, Sapru A, D’costa PM, et al. Safety and immunogenicity of a new purified vero cell rabies vaccine (PVRV) administered by intramuscular and intradermal routes in healthy volunteers. Vaccine 2013;31(24):2719-22
98.
Clinical trial for rabies monoclonal antibody. Available from: www.eurekalert.
doi: 10.1586/14760584.2015.973403
Rabies vaccines: where do we stand, where are we heading? Expert Review of Vaccines Downloaded from informahealthcare.com by Korea University on 01/06/15 For personal use only.
Expert Rev. Vaccines Early online, 1–13 (2014)
Manpreet Kaur1,2, Rajni Garg1, Samer Singh*3 and Rakesh Bhatnagar*1 1 BSL3 Laboratory, School of Biotechnology, Jawaharlal Nehru University, New Delhi – 110067, Delhi, India 2 Vaccine and Infectious Disease Research Center, Translational Health Science and Technology Institute, An autonomous institute of Department of Biotechnology, Govt. of India, Gurgaon – 122016, Haryana, India 3 Department of Microbial Biotechnology, Panjab University, Chandigarh – 160014, India *Authors for correspondence: Tel.: +91 112 670 4079 Fax: +91 112 674 2040 [email protected]; [email protected]
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Rabies being the most lethal zoonotic, vaccine-preventable viral disease with worldwide distribution of reservoir wild animals presents unique challenges for its diagnosis, management and control. Although vaccines available are highly effective, which had played the key role in controlling rabies in North America, western Europe and in a number of Asian and Latin American countries, the requirement of multiple doses along with boosters, associated cost to reduce the incidence in wild animals and prophylactic human vaccination has remained a major impediment towards achieving the same goals in poorer parts of the world such as sub-Saharan Africa and southeast Asia. Current efforts to contain rabies worldwide are directed towards the development of more safe, cheaper and efficacious vaccines along with anti-rabies antibodies for post-exposure prophylaxis. The work presented here provides an overview of the advances made towards controlling the human rabies, particularly in last 10 years, and future perspective. KEYWORDS: encephalitis • glycoprotein • in vivo challenge • protection • rabies virus-neutralizing antibody • survival • therapeutics • vaccines
Rabies (Latin meaning ‘rage or madness’) is one of the oldest zoonosis known to mankind. It is primarily a disease of terrestrial and airborne mammals, including dogs, wolves, foxes, coyotes, jackals, cats, bobcats, lions, mongooses, skunks, badgers, bats, monkeys and humans [1]. It’s an acute, almost invariably fatal progressive encephalitis caused by viruses belonging to genus Lyssavirus of Rhabdoviridae family, within the order Mononegavirales [2]. The great majority (~99%) of human infections is reported to be caused by rabies virus but more than a dozen other members of Lyssavirus genus exist that are supposed to have potential to cause clinical rabies [3,4]. Lyssavirus genus has been classified on the basis of genetic distances and serological cross-reactivity into three phylogroups. The rabies virus belongs to phylogroup I. Most studies on rabies including the development of vaccine against rabies are based on rabies virus, so vaccination and protective strategies developed against rabies virus are not supposed to be protective against rabies caused by viruses belonging to phylogroups II and III of the Lyssavirus genus. Unless specified otherwise any
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reference to ‘rabies’ in the text hereafter refers to rabies caused by rabies virus. Rabies is characterized by neuronal dysfunction and lesions, which are more prominent in the brain stem, as compared with other areas of central nervous system (CNS) [5]. The people who fall victim to death by rabies include mostly people of low socioeconomic background because they cannot afford post-exposure prophylaxis (PEP). Human rabies has been eradicated in some developed countries [6], but it continues to take its toll in developing countries. In Asia and Africa, rabies causes approximately 55,000 deaths annually. Pre-exposure prophylaxis (PrEP) of rabies vaccine is recommended for people who are travelling to high-risk area, or who are occupationally involved with wild animal reservoirs like cave hunters or scientists working on rabies-infected animals. PEP is used for people who are bitten by rabid animals. The cell culture vaccines and embryonated egg-based vaccines (CCEEV) namely, human diploid cell vaccine (HDCV), purified chick embryo cell vaccine (PCECV), purified Vero cell vaccine (PVCV) and purified duck embryo vaccine (PDEV) are recommended by WHO for human
2014 Informa UK Ltd
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Kaur, Garg, Singh & Bhatnagar
use [3]. Despite their potency, these vaccines show some side effects such as headache, dizziness and systemic reactions like fever and gastrointestinal symptoms, occurring at low frequencies [7]. Multiple doses of CCEEVs are required for PrEP and PEP along with rabies immunoglobulin (RIG), in case of transdermal bite, scratches, licks on broken skin by a rabid animal, making the rabies control expensive [8]. Focused efforts need to be made in the direction of development of a safe, cost-effective vaccine that could provide consistent protection, desirably in a single dose. This review aims to summarize the current scenario and the future strategies for rabies vaccine development.
Prophylaxis PrEP
PrEP is recommended for people at continued, frequent or increased risk of exposure to the rabies virus either as a result of their residence or occupation. Travelers with extensive outdoor exposure and children living in endemic areas are at particular risk. PrEP may be performed with any of the modern cell-derived vaccines. Booster doses increase and prolong the antibody response. Thus, they should be offered to persons at continuing risk every 1–3 years.
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PEP
Rabies virus
The rabies virion is classic bullet-shaped structure with one end rounded and other planar. It comprises an external viral envelope, the structural unit and a helical ribonucleocapsid core – the internal functional unit. It has a negative sense 12 kb non-segmented ssRNA genome that codes for five proteins (RNA polymerase [L], nucleoprotein [N], phosphoprotein [P], matrix protein [MP], glycoprotein-G [G]) and four intergenic regions. Three proteins, RNA polymerase (L), nucleoprotein (N) and phosphoprotein (P), are tightly associated with the RNA genome to form ribonucleoprotein core [9]. The MP envelopes the ribonucleoprotein, keeps it condensed and targets it to the plasma membrane and incorporates G-protein into the newly forming viral particles [10]. G-protein forms spikes on the viral surface and is the major inducer of apoptosis. Antibody response against G-protein is the primary determinant of protective immunity as it is the primary target of the host humoral [11] and cell-mediated immune responses [12,13]. Additionally, G-protein because of being the only exposed protein on the viral surface is usually targeted for developing vaccines. The clinical spectrum of rabies manifestation is divided into the prodromal phase, acute neurological or excitation phase and coma or terminal phase. The incubation period of rabies is dependent upon the virus involved, the distance of the site of bite from CNS and the size and innervations of the bite site and may vary from 5 days to many years (usually 2–3 months; rarely >1 year). The patient’s age and genetic background also have been speculated to have bearing on incubation period of rabies similar to other neurotropic viruses [14]. Once the symptoms have set in, rabies generally proceeds to the terminal phase after 10 days leading to cardio-respiratory arrest and death. Most of the human rabies cases develop as a result of bite from an infected animal, usually dog. Human-to-human transmission is extremely rare, but has been reported in certain organ transplant cases [5]. Successful implementation of rabies control programs in canines, primarily dogs, has almost eliminated the incidence of human rabies in North America [15] and greatly reduced it in Latin America and the Caribbean [16]. Asia has the highest number of rabies cases with India on the top, contributing to one-third of the total global rabies deaths. doi: 10.1586/14760584.2015.973403
Rabies is almost always fatal, once the clinical symptoms have set in. Recovery and survival is extremely rare. It is thus essential that individuals be given immediate PEP after any case of exposure. This involves instant wound cleaning and immunizations. The immunization initially included five doses of CCEEV along with human RIG (HRIG), in the deltoid area of adults or in the anterolateral thigh of young children [3]. In 2008, Advisory Committee on Immunization Practices recommended to reduce dosage to four, keeping HRIG dose intact because fifth dose was not required according to experimental data and epidemiological surveys [17]. Accordingly, three doses of CCEEV (1/0.5 ml) need to be administered on days 0, 7 and 21 or 28. WHO has still not incorporated these modifications in its recommendations [18]. However, based on extensive research in developing countries; WHO has now recommended reduced dose regimens, thereby enhancing compliance by the patients as well as cost–effectiveness of the program. One alternate intramuscular regimen recommends vaccine (1/0.5 ml) at two sites on day 0, one site on day 7 and day 21 each. Two modified intradermal (i.d.) regimens are also proposed: these are eight-site i.d. regimen containing 0.1 ml injections at eight sites on day 0, four sites on day 7 and one site each on day 28 and day 90, and two-site i.d. regimen containing 0.1 ml immunizations at two sites each on day 0, two sites each on day 3, two sites each on day 7 and two sites each on day 28. HRIG immunization on day 0 still finds a mandatory position in the regimen. Another modification has been proposed in the eight-site i.d. regimen, making it four site, which reduces the hospital visits to three and the amount of vaccine to half [19]. In case of patients previously immunized with standard vaccination regimen, a single-visit four-site i.d. injections of 0.1 ml equally distributed over left and right deltoids or thighs may be an equally viable option [20]. Despite vast usage of HRIG, their safety and availability remains a cause of concern. WHO has proposed recombinant monoclonal antibodies (mAbs) as suitable alternatives for the PEP. Recently, a recombinant human IgG1 anti-rabies mAb (SII RmAb) has been developed in India. In the dose-escalation Phase I study, SII RmAb was found to be safe and the elicited neutralizing serum antibody titers comparable to HRIG when administered in conjunction with rabies vaccine in a simulated PEP regimen [21]. Bakker et al. showed that mAbCL184, generated by combination of mAbs CR57 and CR4098, is safe and Expert Rev. Vaccines
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Rabies vaccines
does not interfere with the development of subsequent vaccineinduced rabies-virus neutralizing antibodies [22]. mAbCL184 Phase II trials were completed in 2008, in children and adolescents in the Philippines [23] and in adults in the USA [24], followed by a third trial in adults in India that was completed in late 2012 [25,26]. These results demonstrate the feasibility and potential of mAbs for PEP. In another study, rabies virusinfected suckling mouse brain was used to generate 11 hybridoma cells secreting mAbs against rabies virus. These mAbs exhibited strong reactivity with rabies-infected Vero cells. Out of these, mAb 4B7 recognized the recombinant nucleoprotein (N) of rabies virus, while seven other reacted specifically with phosphoprotein (P) of rabies virus. Hence, these mAbs may prove useful in studying interactions between rabies virus and its host, and for diagnostic purpose. More importantly, three of the mAbs (1B11, 1C8 and 8H12) showed a neutralizing effect toward rabies virus [27]. Advances in antibody engineering have led to their improved protective efficacy and potent clinical applications. Turki et al. reported the generation of a trivalent single-chain Fv (scFv50AD1-Fd), which showed a high neutralization activity up to 75-fold higher than that in monovalent form at an equivalent concentration [28]. This was more than three fold than that of WHO standard serum recommendations [29]. The multivalency of single-chain Fv not only improved the avidity, but also the biological activity, emphasizing its potential in rabies PEP. Plant-based expression systems provide a safe and economically viable platform for generation of therapeutic and diagnostic mAbs. In a study, mAbP SO57 and its endoplasmic reticulum targeted variant were produced in transgenic tobacco plants (Nicotiana tabacum). Both were found to be effective in neutralizing rabies virus challenge virus standard (CVS)-11 [30]. In another study, the chimeric (mouse-human) version of mAb 62-71-3 was expressed in Nicotiana benthamiana. The mAb exhibited rabies as well as pseudotyped virus neutralization [31]. These studies establish the usage of plant-based expression system for the development of effective and inexpensive antibody products. Vaccines
The development of rabies vaccines has a long and distinguished history. However, the urgent need of safe and efficacious vaccines has stimulated their further refinement. Conventional vaccines
The first rabies vaccine was developed by Louis Pasteur in 1885 by using spinal cord of rabbits infected with rabies. His post-exposure administration of the desiccated spinal cords, which were supposed to contain partially inactivated infectious agent, to a boy named Joseph Meister who was bitten by a rabid dog, protected him from developing rabies. It heralded the era of rabies vaccine development. Based on same principle, David Semple developed and introduced relatively safer nerve tissue vaccine that consisted of phenol-inactivated sheep or goat informahealthcare.com
Review
brain tissue [32]. For a very long time, these nerve tissue vaccines had remained the pillars of rabies immunization efforts. Though very effective, they were associated with serious side effects like demyelinating allergic encephalitis and required cumbersome and painful 14–21 shots for generating effective immunity [33]. Later, their use was discontinued and they were replaced with modified and safer vaccines. One such newgeneration vaccine was purified duck embryo vaccine (PDEV), which used embryonated duck eggs for propagation of fixed virus strains, which were inactivated by b-propiolactone [34]. HDCV was introduced in 1978, in which Fixed Rabies virus Pitman-Moore L503 strain was used that was adapted for growth in normal human fibroblast cell line. This vaccine proved to be efficacious as well as safer in comparison to previous vaccines. It imparted complete protection to human subjects against silver haired bat virus strain and was as effective as the other available PCECV, which was obtained by growing fixed Flury LEP-50 virus strain in chick fibroblasts [35]. Though owing to ease of production, PCECV is a cheaper alternative; it also produces urticaria and pruritus in few of its recipients. Currently, efficacious and safe vaccines for human use are HDCV, PCECV and purified Vero cell rabies vaccine (PVRV), which uses inactivated Wistar strain of rabies virus grown on Vero cell cultures [36]. Another less frequently used vaccine is primary hamster kidney cell vaccine, which uses the fixed Beijing strain cultured in primary hamster kidney cells [7]. Development of improved vaccines Recombinant live vaccines
The efficacy of PrEP and PEP therapy is often compromised in the poorer areas of the world, where rabies is highly endemic due to lack of vaccination compliance both in human and animal. The poor vaccine compliance partially results from lack of awareness, poor infrastructure, unavailability of vaccines and most importantly the associated high cost. Though oral rabies vaccination of dogs have been known for several years, owing to the risk associated with the dissemination of replicating antigens in young and/or immunodepressed individuals; extensive vaccination program has not been undertaken. Additionally, this does not yield in levels of protective antibody sufficient to break the dog-to-dog transmission cycles or maintain immunity. Other concerns include the preconditioned requisite of suitable bait well accepted by the target population. The artificial or manufactured baits need to be imported, which is an expensive proposition for countries with limited financial resources. Many times, these are not readily accepted by dogs due to unfamiliar taste, odor and texture of baits. This usually affects subsequent vaccination attempts [37]. Locally baits could be probable solution to both cost–effectiveness and general acceptance. The overall safety remains a cause of immense concern. Esh et al. demonstrated that the Evelyn Rokitniki Abelseth strain, generally oral fox vaccination causes fatal rabies in cats [38]. Oral vaccine candidates are usually attenuated derivatives of Street Alabama doi: 10.1586/14760584.2015.973403
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Dufferin (SAD) strain. They are generally achieved by a single amino acid exchange, which may not lead to a stable nonpathogenic phenotype, as elegantly shown by Faber et al. They found that rabies virus attenuation through an amino acid substitution for Arg333 can revert back to pathogenicity upon serial passage of the virus in mouse brain through the introduction of mutations at other positions of the glycoprotein gene [39]. Multiple changes in different components may be the key to development of a stable and safe attenuated live vaccine that may have wider acceptability. Additionally, there is a need to develop more immunogenic and preferably single-dose-vaccine as a remedy to failure of the follow-up vaccination in these endemic areas. However, it is unlikely that a single dose of inactivated vaccines will suffice the need of active immunization, as suggested by Strady et al., who showed that a minimum of three inoculations are required to reduce the percentage of non- or poor-responders (100-fold in many countries as the most of rabies-related deaths occur in communities away from regular hospitals [87,88] and many a times they are misdiagnosed for other neurological diseases [89]. Though neural tissue-based vaccines have been discontinued and are not recommended anymore by WHO due to associated side effects and safety issues, they are still in use for PEP in poorer countries such as Algeria and Ethiopia due to cost and poor availability of current generation rabies vaccines. Outside Africa, China and India had remained the major contributors to global rabies burden till last decade, accounting for 25–40% of the total global mortality from rabies [86].The efforts by Chinese and Indian governments in last decade to implement dog population control measures, vaccination of dogs, making available modern vaccine and improving the accessibility to PEP measures have improved the situation leading to significant decrease in incidences and stabilization [3,86]. To further improve the situation in these countries more efforts are required to educate the population in remote areas, particularly who do not have doi: 10.1586/14760584.2015.973403
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Table 1. Different rabies vaccines. Vaccine name
Composition
Limitations/problems
Ref.
Semple vaccine
Phenol or b-propiolactone inactivated homogenate of rabies virus-infected goat or sheep brain tissue
– Pain, swelling, erythema – Fever, headache, insomnia – Can cause demyelinating allergic encephalitis, due to myelin basic protein – Less immunogenic, therefore, required 14–21 shots – Discontinued by WHO since 1984
[7,93]
Fuenzalida rabies vaccine
Suckling mouse brain tissue
– Pain, swelling, erythema – Fever, headache, insomnia – Decreased myelin basic protein, therefore, decreased allergic reactions – Post-vaccination neuroparalytic syndromes resembling Guillain–Barre´ syndrome
Human diploid cell vaccine
Pitman-Moore L503 or Flury strain of rabies virus grown on MRC-5 human diploid cell culture
– Fever, headache, dizziness – Immune complex-like reaction – Neurological disease
[35]
Purified chick embryo cell vaccine
b-Propiolactone inactivated fixed Flury LEP-25 rabies virus strain grown in primary cultures of chick fibroblasts
– Urticaria, pruritus – Similar but less frequent local, systemic reactions and neurological reactions than HDCV
[35]
Purified Vero cell rabies vaccine
b-Propiolactone inactivated Wistar strain of rabies virus grown in Vero cells
Local, systemic and neurological reactions occurring at a frequency similar to purified chick embryo cell vaccine
[36]
Primary hamster kidney cell vaccine
Formalin inactivated Beijing strain, adsorbed to aluminum hydroxide
– Redness, itching and pain at injection site – Fever, headache, dizziness – Pain in joints and lumbar pain in some
[94]
Purified duck embryo vaccine
b-Propiolactone inactivated fixed virus strain grown in duck embryo cells
– Pain at the injection site – Allergic reactions – Neurological disease, at a rate lower than HDCV
[34]
[7]
HDCV: Human diploid cell vaccine.
immediate access to appropriate care and there is lack of awareness. In South America, particularly remote/hard-to-access areas of Brazil, Colombia, Ecuador and Peru in the Amazon region may be another hotspot of rabies due to presence of human rabies transmitting vampire bats, lack of awareness and at the same time unavailability of proper care [90]. Combined together, to effectively control the disease and eventually eliminate rabies, we would need to focus on reaching remote regions of the world, educate the population, dispelling myths besides making the PrEP and PEP readily available to masses and put in place comprehensive plans to control animal rabies. It could be made possible only by joint cooperative and collaborative efforts involving local populace, volunteers, public healthcare professionals, scientists, local governments, charity institutions, WHO and all the other stakeholders. In this direction, some excellent endeavors have been already put in place in ASEAN countries and Latin American countries that intend to achieve these goals [86]. An attractive strategy to decrease the overall global burden of human rabies, particularly in poorer countries/regions would be to incorporate rabies vaccination into existing child immunization programs [3], as it could reduce the economic, logistical or programmatic obstacles substantially. Any positive step taken by respective doi: 10.1586/14760584.2015.973403
regional cooperative organizations or individual nations with endemic rabies may go a long way in controlling rabies. Some additional points that may not be directly linked to rabies vaccine development but pertinent to our overall goal of rabies control/management are being described briefly. There is dearth of studies replicating overall pathogenesis, disease progression and intensive care requirement of human subjects in animal models due to unavailability of suitable animal models. Urgent efforts need to be made to find suitable animal models then carry out such studies as they may help us better formulate therapeutic intervention strategies to improve the outcome of rabies-patient care. There had been relative lack of interest in developing antiviral therapy for rabies virus. As of now, no commercial antiviral therapy is available for rabies virus. Better understanding of the biology of rabies virus such as interactions among different viral proteins, working of viral replication complex may help us develop small molecule inhibitors for rabies therapy. Research on siRNA that can inhibit rabies virus replication has lot of potential in terms of delaying the onset of rabies symptoms and allowing currently used PEP to take effect [81,91]. Such studies need to be more vigorously pursued as they may be the game changer in saving the life of rabies virus-infected individuals. Expert Rev. Vaccines
Rabies vaccines
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Table 2. Improved rabies vaccines/monoclonal antibodies under clinical development. Vaccine/antibody
Composition
Status
R&D/manufacturer
Ref.
Purified Vero rabies vaccine (PVRV-NG)
Serum-free purified Vero rabies vaccine
Phase II
Sanofi Pasteur
[95]
Chromatographically purified rabies vaccine
Chromatographically purified PVRV, devoid of albumin
Phase II
Pasteur Me´rieux Connaught, France
[96]
Purified Vero cell rabies vaccine
Lyophilized vaccine containing inactivated purified (Pitman Moore, PM3218 as virus strain) produced using Vero ATCC CCL 81 cells
Phase III
Serum Institute of India Ltd
[97]
Anti-rabies monoclonal antibody
Anti-rabies monoclonal antibody
Phase III
Serum Institute of India & Mass Biologics of the University of Massachusetts Medical School
[98]
Rabies immune globulin (human)
Intramuscularly administered rabies immune globulin (human)
Phase I
Grifols Therapeutics Inc
[99]
Rabies vaccines
Purified Vero rabies vaccine and rabies human diploid cell vaccine
Phase II
Sanofi Pasteur
[100]
Rabies vaccine
Purified inactivated Vero rabies vaccine
Phase III
Sanofi Pasteur
[101]
Rabies vaccine-rabies immunoglobulin
Two intradermal rabies vaccine regimens administered with and without human rabies immunoglobulin
Phase III
Novartis
[102]
CL184
Human monoclonal antibody cocktail, containing a 1:1 equipotent mixture of two human monoclonal antibodies (CR57 and CR4098)
Phase II
Sanofi Pasteur
[103]
HRIG-rabies vaccine
HRIG with co-administration of active rabies vaccine
Phase II/III
Kamada Ltd
[104]
HRIG: Human polyclonal anti-rabies immune globulin.
Five-year view
In the last decade, the steady improvement in the availability of safer CCV rabies vaccines and RIGs has greatly enhanced the PEP and PrEP outcomes. It has helped tremendously in controlling spread of rabies and associated mortality at global scale. However, the lack of awareness, cost and availability of vaccines and RIGs and their efficacy, requirement of multiple boosters to mount protective response had been the main impediments toward achieving the desired level of rabies control in the poorer nations. Currently, some of the notable endeavors that are underway and have clear potential to further improve the accessibility and applicability of PEP and PrEP include efforts to cut the cost of available vaccines and RIGs through reduced dosage and altered administration schedule, reduce the PEP and PrEP administration time, make single dose vaccines (TABLES 1 & 2) [3]. Various studies and follow-ups from Thailand, India and other southeast Asian countries have clearly demonstrated the possibility of dose reduction but comparable protection offered by CCV vaccines when given intradermally both in PEP and PrEP scenarios. These reduced dose and less dose regimens would need to be implemented in more rabies-endemic areas. To control the rabies in wild reservoir population, we will have to direct more efforts toward broader application of available oral vaccines. Currently, RIGs are informahealthcare.com
prohibitively expensive. Development of newer biological such as mAbs, which are already at different stages of clinical trials [3,22,92], should become available in near future, effectively reducing the cost and making the passive immunization as a part of PEP accessible to people who can least afford it. In addition to advances envisaged to become available soon that will help improve the PEP and PrEP to control rabies, the development and availability of inexpensive, accurate and easyto-use diagnostics such as the direct rapid immunohistochemical test [80] in poor settings, could tremendously improve the rabies surveillance allowing real-time adjustment and modification in any comprehensive rabies control program. Combined together, all these developments will greatly improve our chances of controlling/eliminating rabies in next 5–10 years. Financial & competing interests disclosure
This work was supported by Department of Biotechnology (DBT), Department of Science and Technology, Council of Scientific and Industrial Research, Indian Council of Medical Research, India and NATP-World Bank. S Singh is a Ramalingaswami Fellow, supported by DBT, India. 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. doi: 10.1586/14760584.2015.973403
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Kaur, Garg, Singh & Bhatnagar
Key issues • Rabies, a zoonosis, is a fatal progressive encephalitis caused by the infection of the members of the Lyssavirus genus. All mammals are susceptible to rabies but animals from the orders of Carnivora and Chiroptera are primary reservoirs, that is, dogs, foxes, jackals, coyotes, skunks, raccoons, mongoose and bats. • Annually about 60,000 people die of rabies. Most of the human rabies cases result from rabid dog bites (>98%), especially in Asia and Africa or bat bites, especially in Americas, Europe and Australia. Very rarely saliva, body fluids and organ transplants originating from infected individuals or bites of other reservoir animals have been reported as a cause of rabies. • Rabies is a 100% preventable disease if proper prophylactic vaccination is provided as a part of the pre-exposure prophylaxis (PrEP) or along with anti-rabies serum (rabies immunoglobins [RIGs]) as a part of the post-exposure prophylaxis (PEP). No one immunized with PrEP and PEP vaccines has died of rabies following coming in contact with a rabid animal. Expert Review of Vaccines Downloaded from informahealthcare.com by Korea University on 01/06/15 For personal use only.
• The complications associated with overwhelming replication of Lyssavirus in CNS are supposed to cause rabies-related death. The nerve tissue vaccine that was widely used up to about a decade ago in many parts of the world though highly effective had serious side effects such as demyelinating allergic encephalitis. Now, it has been replaced with more effective and safer cell culture vaccines in most parts of the world. • Despite the availability of effective rabies vaccines, the reach of vaccines to the patients in the poorer regions of the world had been limited by a combination of factors such as cost, availability, access to preventive medical care, awareness/ignorance and misconceptions about rabies. • To effectively control the disease, various awareness programs and vaccination programs including that for the reservoir animals are desired. A number of such programs are in place and many more are being formulated for the endemic areas by local governments, philanthropic institutions and other stakeholders such as WHO. • The animal vaccination program could greatly benefit from the availability of oral rabies vaccines. Some oral vaccines have been successfully used/tested in wild animals. Concerted efforts are required to make them available globally, particularly to poorer countries where rabies is endemic. • Development of a safe, single-dose vaccine and the cost reduction of the existing vaccine can significantly reduce the economic burden of rabies vaccination programs worldwide besides improving the PrEP or PEP compliance and eventually success of rabies control programs. • Vaccines under development such as subunit vaccines, attenuated live virus vaccines have the potential to eliminate the need of boosters and reduce the dosage required for PEP or PrEP to one, while development of plant-based edible-vaccines could significantly improve the reach and reduce the associated cost of vaccine delivery and administration. • Future rabies intervention efforts should include the development of strategies/small molecules to slow down/stop the replication/spread of virus allowing the PEP to take effect. Development of anti-viral drugs or interfering RNA that may delay the viral replication/spread should be made priority. Although it has tremendous potential to bring down mortality from rabies, the efforts in this direction have been scanty. • Future vaccine development efforts need to include targeting of the members of the phylogroup II and III of Lyssavirus genus to make the vaccine ready for any such eventuality that may see zoonosis of these Lyssavirus members or in a rare case emergence of some recombinant viruses. Available rabies vaccines and those under various stages of development primarily target viruses from phylogroup I of Lyssavirus genus and are presumed to be ineffective against the infection of the members of phylogroup II and III.
References
4.
Fooks AR, Banyard AC, Horton DL, et al. Current status of rabies and prospects for elimination. Lancet 2014. [Epub ahead of print]
9.
Wunner WH, Larson JK, Dietzschold B, Smith CL. The molecular biology of rabies viruses. Rev Infect Dis 1988;10(Suppl 4): S771-84
5.
Consales CA, Bolzan VL. Rabies review: immunopathology, clinical aspects and treatment. J. Venom. Anim. Toxins Incl. Trop. Dis 2007;13(1):5-38
10.
6.
Menezes R. Rabies in India. CMAJ 2008; 178(5):564-6
Mebatsion T, Weiland F, Conzelmann KK. Matrix protein of rabies virus is responsible for the assembly and budding of bullet-shaped particles and interacts with the transmembrane spike glycoprotein G. J Virol 1999;73(1):242-50
7.
WHO. Information sheet. Observed rate of vaccine reactions. Rabies vaccine. World Health Organization; Geneva, Switzerland: 2012
11.
•
Describes side effects of existing vaccines.
Cox JH, Dietzschold B, Schneider LG. Rabies virus glycoprotein. II. Biological and serological characterization. Infect Immun 1977;16(3):754-9
8.
McGettigan JP. Experimental rabies vaccines for humans. Expert Rev Vaccines 2010; 9(10):1177-86
12.
Macfarlan RI, Dietzschold B, Koprowski H. Stimulation of cytotoxic T-lymphocyte responses by rabies virus glycoprotein and
Papers of special note have been highlighted as: • of interest •• of considerable interest 1.
Fishbein DB, Robinson LE. Rabies. N Engl J Med 1993;329(22):1632-8
2.
Francki RIB, Fauquet CM, Knudson DL, Brown F. Classification and nomenclature of viruses: fifth report of international committee on taxonomy of viruses. Arch Virol. 1991. 2(Suppl):1-450
3.
••
WHO Expert Consultation on Rabies. Second Report. WHO Technical Report Series 982. World Health Organization; Geneva, Switzerland: 2013 Includes most current data on rabies prevention and control.
doi: 10.1586/14760584.2015.973403
Expert Rev. Vaccines
Rabies vaccines
Expert Review of Vaccines Downloaded from informahealthcare.com by Korea University on 01/06/15 For personal use only.
identification of an immunodominant domain. Mol Immunol 1986;23(7):733-41
35.
Dietzschold B, Hooper DC. Human diploid cell culture rabies vaccine (HDCV) and purified chick embryo cell culture rabies vaccine (PCECV) both confer protective immunity against infection with the silver-haired bat rabies virus strain (SHBRV). Vaccine 1998;16(17):1656-9
36.
Jaiiaroensup W, Lang J, Thipkong P, et al. Safety and efficacy of purified Vero cell rabies vaccine given intramuscularly and intradermally. (Results of a prospective randomized trial). Vaccine 1998;16(16): 1559-62
37.
Estrada R, Vos A, Rc de L. Acceptability of local made baits for oral vaccination of dogs against rabies in the Philippines. BMC Infect Dis 2001;1:19
38.
Esh JB, Cunningham JG, Wiktor TJ. Vaccine-induced rabies in four cats. J Am Vet Med Assoc 1982;180(11):1336-9
39.
Faber M, Faber M-L, Papaneri A, et al. A single amino acid change in rabies virus glycoprotein increases virus spread and enhances virus pathogenicity. J Virol 2005; 79(22):14141-8
40.
Strady C, Andreoletti L, Baumard S, et al. Immunogenicity and booster efficacy of pre-exposure rabies vaccination. Trans R Soc Trop Med Hyg 2009;103(11):1159-64
41.
Prevec L, Campbell JB, Christie BS, et al. A recombinant human adenovirus vaccine against rabies. J Infect Dis 1990;161(1): 27-30
42.
Yarosh OK, Wandeler AI, Graham FL, et al. Human adenovirus type 5 vectors expressing rabies glycoprotein. Vaccine 1996;14(13):1257-64
43.
Chakrabarti P. “Living versus dead”: the Pasteurian paradigm and imperial vaccine research. Bull Hist Med 2010;84(3): 387-423
Shen CF, Lanthier S, Jacob D, et al. Process optimization and scale-up for production of rabies vaccine live adenovirus vector (AdRG1.3). Vaccine 2012;30(2):300-6
44.
Biswas S, Reddy GS, Srinivasan VA, Rangarajan PN. Preexposure efficacy of a novel combination DNA and inactivated rabies virus vaccine. Hum Gene Ther 2001; 12(15):1917-22
Cenna J, Hunter M, Tan GS, et al. Replication-deficient rabies virus-based vaccines are safe and immunogenic in mice and nonhuman primates. J Infect Dis 2009; 200(8):1251-60
45.
Dorfmeier CL, Lytle AG, Dunkel AL, et al. Protective vaccine-induced CD4(+) T cell-independent B cell responses against rabies infection. J Virol 2012;86(21): 11533-40
46.
Dorfmeier CL, Shen S, Tzvetkov EP, McGettigan JP. Reinvestigating the role of IgM in rabies virus postexposure vaccination. J Virol 2013;87(16):9217-22
47.
Chen Z, Zhou M, Gao X, et al. A novel rabies vaccine based on a recombinant parainfluenza virus 5 expressing rabies virus glycoprotein. J Virol 2013;87(6):2986-93
Healthy Adult Subjects. Available from: http://clinicaltrials.gov/show/NCT00656097
13.
Celis E, Karr RW, Dietzschold B, et al. Genetic restriction and fine specificity of human T cell clones reactive with rabies virus. J Immunol 1988;141(8):2721-8
14.
Hoff G, M S. Rabies and Animal Bite investigation. Missouri Health Department, Kansas City, Kansas. 2005
26.
15.
Blanton JD, Palmer D, Rupprecht CE. Rabies surveillance in the United States during 2009. J Am Vet Med Assoc 2010; 237(6):646-57
Nath A, Tyler KL. Novel approaches and challenges to treatment of central nervous system viral infections. Ann Neurol 2013; 74(3):412-22
27.
16.
Epidemiology and burden of disease. Available from: http://www.who.int/rabies/ epidemiology/en/
17.
Rupprecht CE. Use of a Reduced (4-Dose) Vaccine Schedule for Postexposure Prophylaxis to Prevent Human Rabies Recommendations of the Advisory Committee on Immunization Practices. MMWR 2010;59(RR02):1-9
Zhang J, Ruan X, Zan J, et al. Efficient generation of monoclonal antibodies against major structural proteins of rabies virus with suckling mouse brain antigen. Monoclon. Antib. Immunodiagn. Immunother 2014; 33(2):94-100
•
Depicts efforts to reduce vaccine dosage.
18.
Current WHO guide for rabies pre and post-exposure treatment in humans. Available from: www.who.int/rabies/en/ WHO_guide_rabies_pre_post_ exp_treat_humans.pdf
19.
Warrell MJ, Riddell A, Yu L-M, et al. A simplified 4-site economical intradermal post-exposure rabies vaccine regimen: a randomised controlled comparison with standard methods. PLoS Negl Trop Dis 2008;2(4):e224
20.
WHO. Available from: www.who.int/wer (Weekly Epidemiological record, no. 32, 6 august 2010)
21.
Gogtay N, Thatte U, Kshirsagar N, et al. Safety and pharmacokinetics of a human monoclonal antibody to rabies virus: a randomized, dose-escalation phase 1 study in adults. Vaccine 2012;30(50):7315-20
22.
23.
24.
Bakker ABH, Python C, Kissling CJ, et al. First administration to humans of a monoclonal antibody cocktail against rabies virus: safety, tolerability, and neutralizing activity. Vaccine 2008;26(47):5922-7 Randomized Phase II Trial on Safety and Neutralizing Activity of CL184 and Rabies Vaccine Versus Human Rabies Immune Globulin (HRIG) and Rabies Vaccine in Children and Adolescents. Available from: http://clinicaltrials.gov/show/NCT00708084 A Randomized Phase II Trial to Compare the Safety and Neutralizing Activity of CL184 in Combination With Rabies Vaccine vs. HRIG or Placebo in Combination With Rabies Vaccine in
informahealthcare.com
25.
28.
29.
Rabies Virus Neutralizing Activity and Safety of CL184, a Monoclonal Antibody Cocktail, in Simulated Rabies Post-Exposure Prophylaxis in Healthy Adults. Available from: http://clinicaltrials.gov/show/ NCT01228383
Yang L, Cen J, Xue Q, et al. Identification of rabies virus mimotopes screened from a phage display peptide library with purified dog anti-rabies virus serum IgG. Virus Res 2013;174(1-2):47-51 Turki I, Hammami A, Kharmachi H, Mousli M. Engineering of a recombinant trivalent single-chain variable fragment antibody directed against rabies virus glycoprotein G with improved neutralizing potency. Mol Immunol 2014;57(2):66-73
30.
Lee J-H, Park D-Y, Lee K-J, et al. Intracellular reprogramming of expression, glycosylation, and function of a plant-derived antiviral therapeutic monoclonal antibody. PLoS One 2013;8(8):e68772
31.
Both L, van Dolleweerd C, Wright E, et al. Production, characterization, and antigen specificity of recombinant 62-71-3, a candidate monoclonal antibody for rabies prophylaxis in humans. FASEB J 2013; 27(5):2055-65
32.
33.
34.
•
Review
Mahendra BJ, Madhusudana SN, Sampath G, et al. Immunogenicity, safety and tolerance of a purified duck embryo vaccine (PDEV, VAXIRAB) for rabies post-exposure prophylaxis: results of a multicentric study in India. Hum Vaccin 2010;6(9):721-4 Depicts post-exposure prophylactic potential of purified duck embryo vaccine in Indian population.
doi: 10.1586/14760584.2015.973403
Review 48.
49.
Expert Review of Vaccines Downloaded from informahealthcare.com by Korea University on 01/06/15 For personal use only.
50.
51.
52.
Kaur, Garg, Singh & Bhatnagar
Faber M, Li J, Kean RB, et al. Effective preexposure and postexposure prophylaxis of rabies with a highly attenuated recombinant rabies virus. Proc Natl Acad Sci USA 2009; 106(27):11300-5 Schutsky K, Curtis D, Bongiorno EK, et al. Intramuscular inoculation of mice with the live-attenuated recombinant rabies virus TriGAS results in a transient infection of the draining lymph nodes and a robust, long-lasting protective immune response against rabies. J Virol 2013;87(3):1834-41 Koraka P, Bosch B-J, Cox M, et al. A recombinant rabies vaccine expressing the trimeric form of the glycoprotein confers enhanced immunogenicity and protection in outbred mice. Vaccine 2014;32(36):4644-50 Wang X, Feng N, Ge J, et al. Recombinant canine distemper virus serves as bivalent live vaccine against rabies and canine distemper. Vaccine 2012;30(34):5067-72 Wu Q, Yu F, Xu J, et al. Rabies-virusglycoprotein-pseudotyped recombinant baculovirus vaccine confers complete protection against lethal rabies virus challenge in a mouse model. Vet Microbiol 2014;171(1-2):93-101
53.
MERCK MANUALS. Available from: www.merckmanuals.com/vet/pharmacology/ vaccines_and_immunotherapy/ active_immunization.html
54.
Saxena M, Van TTH, Baird FJ, et al. Pre-existing immunity against vaccine vectors-friend or foe? Microbiology 2013; 159(Pt 1):1-11
55.
56.
Daniell H. Production of biopharmaceuticals and vaccines in plants via the chloroplast genome. Biotechnol J 2006;1(10):1071-9 McGarvey PB, Hammond J, Dienelt MM, et al. Expression of the rabies virus glycoprotein in transgenic tomatoes. Biotechnology 1995;13(13):1484-7
57.
Yusibov V, Hooper DC, Spitsin S V, et al. Expression in plants and immunogenicity of plant virus-based experimental rabies vaccine. Vaccine 2002;20(25-26):3155-64
58.
Ashraf S, Singh PK, Yadav DK, et al. High level expression of surface glycoprotein of rabies virus in tobacco leaves and its immunoprotective activity in mice. J. Biotechnol 2005 119(1):1-14
59.
Loza-Rubio E, Rojas E, Go´mez L, et al. Development of an edible rabies vaccine in maize using the Vnukovo strain. Dev. Biol 2008;131:477-82
60.
Desme´zie`res E, Jacob Y, Saron MF, et al. Lyssavirus glycoproteins expressing immunologically potent foreign B cell and
doi: 10.1586/14760584.2015.973403
cytotoxic T lymphocyte epitopes as prototypes for multivalent vaccines. J Gen Virol 1999;80(Pt 9):2343-51 61.
Da Cruz FW, McBride AJ, Conceic¸a˜o FR, et al. Expression of the B-cell and T-cell epitopes of the rabies virus nucleoprotein in Mycobacterium bovis BCG and induction of an humoral response in mice. Vaccine 2001;20(5-6):731-6
62.
Mansfield KL, Johnson N, Fooks AR. Identification of a conserved linear epitope at the N terminus of the rabies virus glycoprotein. J Gen Virol 2004;85(Pt 11): 3279-83
63.
Xiang ZQ, Spitalnik S, Tran M, et al. Vaccination with a plasmid vector carrying the rabies virus glycoprotein gene induces protective immunity against rabies virus. Virology 1994;199(1):132-40
64.
Perrin P, Jacob Y, Aguilar-Se´tien A, et al. Immunization of dogs with a DNA vaccine induces protection against rabies virus. Vaccine 1999;18(5-6):479-86
65.
Lodmell DL, Ray NB, Parnell MJ, et al. DNA immunization protects nonhuman primates against rabies virus. Nat Med 1998;4(8):949-52
66.
Margalith M, Vilalta A. Sustained protective rabies neutralizing antibody titers after administration of cationic lipid-formulated pDNA vaccine. Genet Vaccines Ther 2006;4:2
67.
Lodmell DL, Ray NB, Ulrich JT, Ewalt LC. DNA vaccination of mice against rabies virus: effects of the route of vaccination and the adjuvant monophosphoryl lipid A (MPL). Vaccine 2000;18(11-12):1059-66
68.
Pinto AR, Reyes-Sandoval A, Ertl HCJ. Chemokines and TRANCE as genetic adjuvants for a DNA vaccine to rabies virus. Cell Immunol 2003;224(2):106-13
69.
Lodmell DL, Ray NB, Ewalt LC. Gene gun particle-mediated vaccination with plasmid DNA confers protective immunity against rabies virus infection. Vaccine 1998;16(2-3): 115-18
70.
Lodmell DL, Ewalt LC. Rabies vaccination: comparison of neutralizing antibody responses after priming and boosting with different combinations of DNA, inactivated virus, or recombinant vaccinia virus vaccines. Vaccine 2000;18(22):2394-8
71.
Lodmell DL, Parnell MJ, Bailey JR, et al. Rabies DNA vaccination of non-human primates: post-exposure studies using gene gun methodology that accelerates induction of neutralizing antibody and enhances neutralizing antibody titers. Vaccine 2002; 20(17-18):2221-8
72.
Osorio JE, Tomlinson CC, Frank RS, et al. Immunization of dogs and cats with a DNA vaccine against rabies virus. Vaccine 1999;17(9-10):1109-16
73.
Kaur M, Rai A, Bhatnagar R. Rabies DNA vaccine: no impact of MHC class I and class II targeting sequences on immune response and protection against lethal challenge. Vaccine 2009;27(15):2128-37
••
Indicates that the directing target sequence is not the exclusive deciding factor for type and extent of immune response elicited and emphasizes on the antigen dependence of immune enhancement strategies.
74.
Kaur M, Saxena A, Rai A, Bhatnagar R. Rabies DNA vaccine encoding lysosome-targeted glycoprotein supplemented with Emulsigen-D confers complete protection in preexposure and postexposure studies in BALB/c mice. FASEB J 2010;24(1):173-83
••
Demonstrates that the optimized DNA vaccine formulation provides an avenue for preventing as well as controlling rabies.
75.
Tao L, Ge J, Wang X, et al. Generation of a recombinant rabies Flury LEP virus carrying an additional G gene creates an improved seed virus for inactivated vaccine production. Virol J 2011;8:454
76.
Liu X, Yang Y, Sun Z, et al. A recombinant rabies virus encoding two copies of the glycoprotein gene confers protection in dogs against a virulent challenge. PLoS One 2014;9(2):e87105
77.
Wenqiang J, Yin X, Lan X, et al. Development of a reverse genetics system for the aG strain of rabies virus in China. Arch Virol 2014;159(5):1033-8
78.
Norton JE, Lytle AG, Shen S, et al. ICAM-1-based rabies virus vaccine shows increased infection and activation of primary murine B cells in vitro and enhanced antibody titers in-vivo. PLoS One 2014; 9(1):e87098
79.
Fenje P, Pinteric L. Potentiation of tissue culture rabies vaccine by adjuvants. Am J Public Health Nations Health 1966;56(12): 2106-13
80.
Ullas PT, Madhusudana SN, Desai A, et al. Enhancement of immunogenicity and efficacy of a plasmid DNA rabies vaccine by nanoformulation with a fourth-generation amine-terminated poly(ether imine) dendrimer. Int J Nanomedicine 2014;9: 627-34
Expert Rev. Vaccines
Rabies vaccines
81.
Expert Review of Vaccines Downloaded from informahealthcare.com by Korea University on 01/06/15 For personal use only.
82.
Gupta PK, Sonwane AA, Singh NK, et al. Intracerebral delivery of small interfering RNAs (siRNAs) using adenoviral vector protects mice against lethal peripheral rabies challenge. Virus Res 2012;163(1):11-18
90.
Wiktor TJ, Macfarlan RI, Reagan KJ, et al. Protection from rabies by a vaccinia virus recombinant containing the rabies virus glycoprotein gene. Proc Natl Acad Sci USA 1984;81(22):7194-8
91.
Israsena N, Mahavihakanont A, Hemachudha T. Rabies virus infection and microRNAs. Adv Virus Res 2011;79:329-44
92.
83.
Cadoz M, Strady A, Meignier B, et al. Immunisation with canarypox virus expressing rabies glycoprotein. Lancet 1992; 339(8807):1429-32
84.
Cliquet F, Gurbuxani JP, Pradhan HK, et al. The safety and efficacy of the oral rabies vaccine SAG2 in Indian stray dogs. Vaccine 2007;25(17):3409-18
85.
86.
Blancou J, Artois M, Brochier B, et al. Safety and efficacy of an antirabies vaccine consisting of recombinant vaccinia-rabies virus administered orally to the fox, dog and cat. Ann Rech Vet 1989;20(2):195-204 World Health Organization. Sustaining the drive to overcome the global impact of neglected tropical diseases. Second WHO report on neglected tropical diseases. WHO, Geneva, Switzerland; 2013
87.
Cleaveland S, Fe`vre EM, Kaare M, Coleman PG. Estimating human rabies mortality in the United Republic of Tanzania from dog bite injuries. Bull World Health Organ 2002;80(4):304-10
88.
Hampson K, Dobson A, Kaare M, et al. Rabies exposures, post-exposure prophylaxis and deaths in a region of endemic canine rabies. PLoS Negl Trop. Dis 2008;2(11): e339
89.
Mallewa M, Fooks AR, Banda D, et al. Rabies encephalitis in malaria-endemic area, Malawi, Africa. Emerg Infect Dis 2007; 13(1):136-9
informahealthcare.com
Schneider MC, Romijn PC, Uieda W, et al. Rabies transmitted by vampire bats to humans: an emerging zoonotic disease in Latin America? Rev Panam Salud Publica 2009;25(3):260-9
Review
org/pub_releases/2012-08/uomm-ctf080712. php 99.
An Open-label, Single-Arm Study to Evaluate the Safety and Rabies Virus Antibody Titers of Intramuscularly Administered Rabies Immune Globulin (Human). Available from: http:// clinicaltrials.gov/show/NCT02139657
Mu¨ller T, Dietzschold B, Ertl H, et al. Development of a mouse monoclonal antibody cocktail for post-exposure rabies prophylaxis in humans. PLoS Negl Trop Dis 2009;3(11):e542
100.
Study of Purified Vero Rabies Vaccine and Rabies Human Diploid Cell Vaccine in a Simulated Rabies Post-exposure Regimen. Available from: http://clinicaltrials.gov/ct2/ show/NCT01877395?term=rabies&rank=1
93.
WHO. Rabies. Available from: http://www. who.int/biologicals/vaccines/rabies/en/
101.
94.
Lin F, Zeng F, Lu L, et al. The primary hamster kidney cell rabies vaccine: adaptation of viral strain, production of vaccine, and pre- and postexposure treatment. J Infect Dis 1983;147(3):467-73
Study of the Purified Vero Rabies Vaccine Serum Free in Comparison With the Reference Purified Vero Rabies Vaccine. Available from: http://clinicaltrials.gov/ct2/ show/NCT01339312?term=rabies&rank=4
102.
Safety and Immunogenicity of Two Intradermal Rabies Vaccine Regimens Administered With and Without Human Rabies Immunoglobulin in Subjects ‡ 1 Years of Age. Available from: http://clinicaltrials.gov/ct2/show/ NCT02177032?term=rabies&rank=8
103.
A Randomized Phase II Trial to Compare the Safety and Neutralizing Activity of CL184 in Combination With Rabies Vaccine vs. HRIG or Placebo in Combination With Rabies Vaccine in Healthy Adult Subjects. Available from: http://clinicaltrials.gov/ct2/show/ NCT00656097?term=rabies&rank=13
104.
Phase II/III Study of the Safety and Effectiveness of HRIG With Co-administration of Active Rabies Vaccine in Healthy Subjects (KAMRAB-003). Available from: http://clinicaltrials.gov/ct2/ show/NCT02040090?term=rabies&rank=20
95.
Pichon S, Guinet-Morlot F, Minutello M, et al. A serum-free, purified vero cell rabies vaccine is safe and as immunogenic as the reference vaccine Verorab for pre-exposure use in healthy adults: results from a randomized controlled phase-II trial. Vaccine 2013;31(18):2295-301
96.
Lang J, Cetre JC, Picot N, et al. Immunogenicity and safety in adults of a new chromatographically purified Vero-cell rabies vaccine (CPRV): a randomized, double-blind trial with purified Vero-cell rabies vaccine (PVRV). Biologicals 1998; 26(4):299-308
97.
Kulkarni PS, Sapru A, D’costa PM, et al. Safety and immunogenicity of a new purified vero cell rabies vaccine (PVRV) administered by intramuscular and intradermal routes in healthy volunteers. Vaccine 2013;31(24):2719-22
98.
Clinical trial for rabies monoclonal antibody. Available from: www.eurekalert.
doi: 10.1586/14760584.2015.973403
