Dataset Information

Live-Attenuated Respiratory Syncytial Virus Vaccine With Deletion of RNA Synthesis Regulatory Protein M2-2 and Cold Passage Mutations Is Overattenuated.

ABSTRACT:

Background

The live respiratory syncytial virus (RSV) candidate vaccine LIDcpΔM2-2 is attenuated through deletion of M2-2 and 5 cold-passage mutations.

Methods

RSV-seronegative children aged 6-24 months received a single intranasal dose of 10⁵ plaque-forming units (PFU) of LIDcpΔM2-2 or placebo. RSV serum antibodies, vaccine infectivity, and reactogenicity were assessed.

Results

Four of 11 (36%) vaccinees shed vaccine virus with median peak titers of 1.6 log₁₀ PFU/mL by quantitative culture and 4.5 log₁₀ copies/mL by polymerase chain reaction; 45% had ≥4-fold rise in serum-neutralizing antibodies. Respiratory symptoms or fever were common in vaccinees (64%) and placebo recipients (6/6, 100%).

Conclusions

RSV LIDcpΔM2-2 is overattenuated. Clinical Trial Numbers. NCT02890381, NCT02948127.

SUBMITTER: Cunningham CK

PROVIDER: S-EPMC6559275 | biostudies-literature | 2019 Jun

REPOSITORIES: biostudies-literature

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Publications

Live-Attenuated Respiratory Syncytial Virus Vaccine With Deletion of RNA Synthesis Regulatory Protein M2-2 and Cold Passage Mutations Is Overattenuated.

Cunningham Coleen K CK Karron Ruth R Muresan Petronella P McFarland Elizabeth J EJ Perlowski Charlotte C Libous Jennifer J Thumar Bhagvanji B Gnanashanmugam Devasena D Moye Jack J Schappell Elizabeth E Barr Emily E Rexroad Vivian V Aziz Mariam M Deville Jaime J Rutstein Richard R Yang Lijuan L Luongo Cindy C Collins Peter P Buchholz Ursula U

Open forum infectious diseases 20190506 6

<h4>Background</h4>The live respiratory syncytial virus (RSV) candidate vaccine LIDcpΔM2-2 is attenuated through deletion of M2-2 and 5 cold-passage mutations.<h4>Methods</h4>RSV-seronegative children aged 6-24 months received a single intranasal dose of 10<sup>5</sup> plaque-forming units (PFU) of LIDcpΔM2-2 or placebo. RSV serum antibodies, vaccine infectivity, and reactogenicity were assessed.<h4>Results</h4>Four of 11 (36%) vaccinees shed vaccine virus with median peak titers of 1.6 log<sub> ...[more]

PMID: 31211158

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Project description:Human respiratory syncytial virus (RSV) is the most important viral cause of serious pediatric respiratory illness worldwide. Currently, the most promising live-attenuated vaccine candidate is a temperature-sensitive (ts) cDNA-derived virus named rA2cp248/404/1030ΔSH, in reference to its set of attenuating mutations. In a previous clinical study, more than one-third of postvaccination nasal wash isolates exhibited partial loss of the ts phenotype. Most of this instability appeared to be due to reversion at a missense point mutation called 1030. This 1030 mutation is a single-nucleotide tyrosine-to-asparagine substitution at position 1321 (Y1321N) of the polymerase L protein that contributes to the ts and attenuation phenotypes of the vaccine candidate. The goals of the present study were to identify a reversion-resistant codon at position 1321 conferring a comparable level of attenuation and to use this to develop a genetically stable version of the vaccine virus. We modified wild-type (wt) RSV to insert each of the 20 possible amino acids at position 1321; 19 viruses were recoverable. We also investigated small deletions at or near this position, but these viruses were not recoverable. Phenotypic analysis identified alternative attenuating amino acids for position 1321. Several of these amino acids were predicted, based on the genetic code, to be refractory to deattenuation. Classical genetics, using temperature stress tests in vitro combined with nucleotide sequencing, confirmed this stability but identified a second site with a compensatory mutation at position 1313. It was possible to stabilize the 1313 site as well, providing a stable 1030 mutation. Further stress tests identified additional incidental mutations, but these did not reverse the ts/attenuation phenotype. An improved version of the vaccine candidate virus was constructed and validated in vitro by temperature stress tests and in vivo by evaluation of attenuation in seronegative chimpanzees. In addition to developing an improved version of this promising live-attenuated RSV vaccine candidate, this study demonstrated the propensity of an RNA virus to escape from attenuation but also showed that, through systematic analysis, genetics can be used to cut off the routes of escape.

Project description:We recently developed a system for the generation of infectious bovine respiratory syncytial virus (BRSV) from cDNA. Here, we report the recovery of fully viable chimeric recombinant BRSVs (rBRSVs) that carry human respiratory syncytial virus (HRSV) glycoproteins in place of their BRSV counterparts, thus combining the replication machinery of BRSV with the major antigenic determinants of HRSV. A cDNA encoding the BRSV antigenome was modified so that the complete G and F genes, including the gene start and gene end signals, were replaced by their HRSV A2 counterparts. Alternatively, the BRSV F gene alone was replaced by that of HRSV Long. Each antigenomic cDNA directed the successful recovery of recombinant virus, yielding rBRSV/A2 and rBRSV/LongF, respectively. The HRSV G and F proteins or the HRSV F in combination with BRSV G were expressed efficiently in cells infected with the appropriate chimeric virus and were efficiently incorporated into recombinant virions. Whereas BRSV and HRSV grew more efficiently in bovine and human cells, respectively, the chimeric rBRSV/A2 exhibited intermediate growth characteristics in a human cell line and grew better than either parent in a bovine line. The cytopathology induced by the chimera more closely resembled that of BRSV. BRSV was confirmed to be highly restricted for replication in the respiratory tract of chimpanzees, a host that is highly permissive for HRSV. Interestingly, the rBRSV/A2 chimeric virus was somewhat more competent than BRSV for replication in chimpanzees but remained highly restricted compared to HRSV. This showed that the substitution of the G and F glycoproteins alone was not sufficient to induce efficient replication in chimpanzees. Thus, the F and G proteins contribute to the host range restriction of BRSV but are not the major determinants of this phenotype. Although rBRSV/A2 expresses the major neutralization and protective antigens of HRSV, chimpanzees infected with this chimeric virus were not significantly protected against subsequent challenge with wild-type HRSV. This suggests that the growth restriction of rBRSV/A2 was too great to provide adequate antigen expression and that the capacity of this chimeric vaccine candidate for replication in primates will need to be increased by the importation of additional HRSV genes.

Dataset Information

Live-Attenuated Respiratory Syncytial Virus Vaccine With Deletion of RNA Synthesis Regulatory Protein M2-2 and Cold Passage Mutations Is Overattenuated.

Background

Methods

Results

Conclusions

Publications

Live-Attenuated Respiratory Syncytial Virus Vaccine With Deletion of RNA Synthesis Regulatory Protein M2-2 and Cold Passage Mutations Is Overattenuated.

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