The ZyCoV-D vaccine was developed by Cadila Healthcare Limited, Ahmedabad, India.
The spike gene region was selected from submitted Wuhan Hu-1 isolate (Genebank Accession No. MN908947.3). For generation of the SARS-CoV-2 DNA vaccine construct pVAX-1 plasmid vector was used.
Chemically synthesized Spike regions and signal peptide gene were inserted into pVAX-1 plasmid DNA vaccine vector. Following the receipt of the plasmid DNA constructs, transformations of the construct were carried out in DH5-alpha™ chemically competent cells.
The DH5-alpha E coli carrying the plasmid DNA was further propagated for large scale production in manufacturing suite approved by National Regulatory Authority under current Good Manufacturing Practice conditions. Each 0.5 mL of ZyCoV-D vaccine contains–5 mg of DNA plasmid with spike protein gene region insert from SARS-CoV-2 Virus suspended in phosphate buffer saline.
On 20 August, India’s drug regulator authorized the vaccine for people aged 12 and older. The efficacy figure of 67% came from trials involving more than 28,000 participants, which saw 21 symptomatic cases of COVID-19 in the vaccinated group and 60 among people who received a placebo.
ZyCoV-D contains circular strands of DNA known as plasmids, which encode the spike protein of SARS-CoV-2, together with a promoter sequence for turning the gene on. Once the plasmids enter the nuclei of cells, they are converted into mRNA, which travels to the main body of the cell, the cytoplasm, and is translated into the spike protein itself.
The body’s immune system then mounts a response against the protein, and produces tailored immune cells that can clear future infections. Plasmids typically degrade within weeks to months, but the immunity remains.
Both DNA and mRNA vaccines have been under development since the 1990s, says Weiner. The challenge for DNA vaccines is that they need to make it all the way to the cell nucleus, unlike mRNA vaccines, which just need to get to the cytoplasm, says Jameel. So, for a long time, DNA vaccines struggled to induce potent immune responses in clinical trials, which is why they had been approved for use as vaccines only in animals, such as horses, until now.
Injection-free vaccine
To solve this problem, ZyCoV-D is deposited under the skin, as opposed to deep in muscle tissue. The area under the skin is rich in immune cells that gobble up foreign objects, such as vaccine particles, and process them. “This helps capture the DNA far more efficiently than in the muscle,” Jameel says. Unusually, the vaccine is delivered using a needle-free device pressed against the skin, which creates a fine, high-pressure stream of fluid that punctures the surface and is less painful than an injection.
But despite being more potent than previous DNA vaccines, ZyCoV-D requires a minimum of three doses to achieve its initial efficacy. This is likely to add to the logistical challenge of administering the vaccine during the current pandemic, says Jameel.
Although ZyCoV-D’s efficacy seems to be lower than the 90% or higher achieved by some mRNA vaccines, the figures are not comparable, says Jameel. The ZyCoV-D trials in India earlier this year were conducted while the Delta variant of SARS-CoV-2 was the dominant variant in circulation, whereas earlier mRNA vaccine trials were conducted when less transmissible variants were circulating. “The efficacy is essentially against the Delta variant, so that is pretty good,” he says.
Some researchers have criticized a lack of transparency in the approval process, because no late-stage trial results have yet been published. Zydus Cadila says the trial is still under way and it will submit the full analysis for publication shortly. The company says the first doses will start to be administered in India in September and it plans to produce up to 50 million doses by early next year.
DNA VACCINES IN CLINICAL TRIALS
Many DNA vaccines against COVID-19 are currently undergoing clinical trials around the world.
| Vaccine | Developer | Location | Route | Stage of trial |
| ZyCoV-D | Zydus Cadila | India | Skin | Approved for emergency use |
| INO-4800 | Inovio and partners | United States | Skin | Phase II/III |
| AG0302-COVID19 | AnGes, Osaka University, Takara Bio | Japan | Muscle | Phase II/III |
| GX-19N | Genexine | South Korea | Muscle | Phase I/II |
| GLS-5310 | GeneOne Life Science | South Korea | Skin | Phase I/II |
| COVID-eVax | Takis, Rottapharm Biotech | Italy | Muscle | Phase I/II |
| AG0301-COVID19 | AnGes, OSaka University, Takara Bio | Japan | Muscle | Phase I/II |
| Covigenix VAX-001 | Entos Pharmaceuticals | Canada | Muscle | Phase I |
| CORVax12 | OncoSec, Providence Cancer Institute | United States | Skin | Phase I |
| bacTRL-Spike | Symvivo | Canada | Oral | Phase I |
| COVIGEN | BioNet, Technovalia, University of Sydney | Thailand, Australia | Skin or muscle | Phase I |
Source: World Health Organization. COVID-19 Vaccine Tracker and Landscape (WHO, 2021).
Discussion
We report the findings from Phase 1 part of clinical trial on the safety, tolerability and immunogenicity of ZyCoV-D, a SARS-CoV-2 DNA vaccine encoding the spike protein. This first-in-human Phase 1 study of ZyCoV-D DNA vaccine was carried out in an intensive observational unit with frequent monitoring of vital signs and ECGs for at least 24 h post administration of the first dose of vaccine and for at least 4 h post administration of the second and the third dose of vaccine.
Each vaccination was followed by frequent safety follow-up with subjects till 28 days of the last dose of vaccine. ZyCoV-D vaccine was well-tolerated in 48 healthy adults in all four dose groups with no vaccine-related severe or SAEs. The safety profile of ZyCoV-D vaccine supports further development of ZyCoV-D in at-risk populations who are at more serious risk of complications from SARS-CoV-2 infection, including the elderly and subjects with comorbidities.
Our findings also correlate with previous clinical evaluation of other DNA vaccine candidates which were reported to be safe and well-tolerated in healthy subjects [[17],19, 20–[21]].
The majority of solicited AEs reported in this trial were after the first dose of vaccine, while the second and third dose of vaccination were found to be well-tolerated. The ZyCoV-D Phase 1 safety data further suggest that the vaccine could be a safe booster as there was no increase in frequency of side effects after the third dose compared to the first dose, an important aspect for the safety profile of SARS-CoV-2 vaccines.
One attractive feature of DNA vaccines, like ZyCoV-D, is that the immunizations could be boosted without significant limitations such as dosing-incremented toxicities or anti-vector responses and additional boosting with other DNA vaccines have resulted in higher levels of cellular and humoral immune responses without increased toxicity [[22]].
ZyCoV-D also generated balanced humoral and cellular immune responses in participants displaying either or both antibody or T cell responses following three doses of vaccine. Humoral responses were lower in subjects who received 1 mg vaccine irrespective of method of administration.
The exact reason for this is not known but it is likely that when the vaccine is administered at the low dose of 1 mg at single intradermal site, it may lead to inefficient transfection, in the host cells and thus lower the expression of antigen. Our data corroborates well with the Rhesus Macaques challenge study, where the vaccination of 2 mg dose with Pharmajet NFIS elicited significant SARS CoV-2 specific IgG, NAB titers and lower viral loads in animals post challenge (Data on file). Further; a Phase II study in 1000 subjects is currently ongoing which will provide better understanding of immunogenicity of ZyCoV-D vaccine in a larger sample size.
In our study, three doses of 2 mg ZyCoV-D DNA vaccine administered intradermally at two different sites via NFIS device 28 days apart have shown good humoral and cellular immune response at Day 70 onwards. Presently, correlation of protection for vaccine against SARS-CoV-2 is unknown, and the roles of the specific antibodies or T cells in building effective protection are not yet well-defined.
Therefore, we are only able to demonstrate immune response induction following vaccination and not protection to SARS-CoV-2 following DNA vaccination on the basis of the vaccine-elicited immune responses in this study.
A double-blind, placebo controlled Phase III study in 28,216 subjects aged 12 years and above is also currently ongoing which will help evaluate efficacy of the ZyCoV-D 2 mg dose administered via NFIS device in protection against COVID-19 infection. The study is registered with CTRI/2021/01/030416.
Previous studies investigating SARS and Middle East Respiratory Syndrome (MERS) found that there is a temporary rise in specific antibodies which dropped rapidly in subjects after recovery, and the CD4+ and CD8+ T-cell responses played a vital role in memory response and protection against future exposure to virus [[23]].
A similar rapid decline of the specific antibody amounts in subjects with COVID-19 after recovery was also noted [[23]] suggesting that both specific cellular and humoral immunity are potentially important for a successful COVID-19 vaccine. Here, we report immune response till 28 days after the last dose of vaccine.
ZyCoV-D vaccine also induced cellular response as measured by IFN-γ ELISPOT which was maintained till Day 84 in subjects who received vaccination 1 mg or 2 mg via NFIS device. This clearly indicates that vaccination with ZyCoV-D induces cellular response with fold rise. However, the sample size per arm is too small to reach a definitive conclusion on the levels of IFN-γ in different arms and the results should be interpreted in the context of variability of the immunological responses among individuals enrolled in the trial.
Phase II data with a higher sample size will help to understand cellular response obtained with ZyCoV-D vaccine.
This first-in-human study of ZyCoV-D DNA vaccine has some limitations. First, this open-label, non-randomized Phase 1 trial report is based on a modest sample size (48) in all vaccine arms and, therefore lacks a comparator group.
Larger sample-sized randomized placebo controlled blinded trials may be needed to show the true immunogenicity difference between the dose groups. Second, this report only involves healthy Indian male subjects aged between 18 and 55 years. This is due to societal limitation, COVID 19 related lockdown and completion of recruitment with male subjects at study center.
The results of this study are not generalizable to other ethnic groups and female subjects. In this regard, female subjects were part of Phase 2 and 3 studies. SARS-CoV-2 infection has more severe and fatal outcomes in older individuals. In this regard, the Phase 3 trial will evaluate individuals of higher age group.
Third, only data within the first 84 days of vaccination is being reported, and this report does not include data about the durability of the vaccine-induced immunity. In previous clinical trials with similar DNA vaccines, durable immune responses up to 1 year following vaccination were reported [[14],[22]].
Fourth, the study showed good humoral and cellular immune response at Day 70 onwards after administration of the third dose, while most other approved vaccines showed immune response after administration of the second dose.
In this study, two different vaccination strategies were used. One is injection and needle, and the second is needle-free injection. i.e. NFIS device. This technology has evolved significantly over the last 50 years and is now accepted in many routine immunization settings as a safe and effective vaccine delivery method. Disposable syringe jet injectors are now being used for the delivery of vaccines to eradicate polio, measles, mumps, rubella and influenza, and are showing promising results in vaccine clinical trials for the Zika virus and human papillomavirus.
Vaccine administration using NFIS device offers some distinct advantages compared to the conventional method of vaccination using needle and syringe, like improved compliance and better coverage; no needle trash and needle stick injuries; higher immunogenic response; calibrated for specific volume with minimal vaccine wastage; auto disabling and eliminating possibility of re-use; efficient vaccine delivery; and the workflow is 25% faster than a conventional needle-syringe and is less painful [[24]].
Tebas et al. reported better immune response after administration of two doses with intradermal DNA vaccine followed by electroporation (EP) technique [[17]]. However, Pharmajet Tropis device has been used in DNA vaccine clinical trials and has been reported as a better administration technique in terms of ease of administration, reliability, and precision. Use of Tropis is also reported to be cost-effective and have better local tolerance compared with EP [[25]].
Our data suggests that ZyCoV-D demonstrates a good safety profile and that vaccination induces both cellular and humoral responses, supporting its further development to prevent infection and death related to COVID-19 in the global population. The safety and immunogenic profile are important parameters for vaccination for high-risk populations, such as the elderly and those living with co-morbid conditions.
Over the past decade, the vaccine industry and clinical research centers have been asked to provide urgent responses to epidemics of emerging infectious diseases, such as H1N1 influenza, Ebola virus, Zika, MERS, and now SARS-CoV-2 [[26]]. The risk of COVID-19 caused by SARS-CoV-2 is ongoing, making the need for effective vaccines even more urgent [[27]].
Previous findings suggested that those vaccines expressing full-length spike glycoprotein can induce good immune responses and protective efficacy. The full-length spike was chosen in most of the viral vectored, mRNA, or DNA COVID-19 vaccines in development [[23]].
There have been recent reports of emergence of new SARS-CoV-2 viral strains like B.1.1.7 in UK, B.1.351 in South Africa, P.1 in Brazil [[28]]. The emergence of new strains of virus has raised the doubts about efficacy of vaccines which were already approved for emergency use authorization.
Currently ongoing clinical trials with ZyCoV-D vaccine will provide important insights into efficacy and safety of DNA vaccine platform. DNA vaccines are based on plug and play platform, which allows rapid development of new constructs in case mutant strains develop, and possibility of generating a new vaccine candidate in very short time, thus providing protection against mutated viral strains.
reference link :https://www.thelancet.com/journals/eclinm/article/PIIS2589-5370(21)00300-X/fulltext
