Bi-Reporter Vaccinia Virus for Tracking Viral Infections In Vitro and In Vivo

Despite the eradication of variola virus (VARV), the causative agent of smallpox, poxviruses still represent an important threat to human health due to their possible use as bioterrorism agents and the emergence of zoonotic poxvirus diseases. Recombinant vaccinia viruses (rVV) expressing easily traceable fluorescent or luciferase reporter genes have significantly contributed to the progress of poxvirus research.

Double Recombinant Vaccinia Virus: A Candidate Drug against Human Glioblastoma

Glioblastoma is one of the most aggressive brain tumors. Given the poor prognosis of this disease, novel methods for glioblastoma treatment are needed. Virotherapy is one of the most actively developed approaches for cancer therapy today. VV-GMCSF-Lact is a recombinant vaccinia virus with deletions of the viral thymidine kinase and growth factor genes and insertions of the granulocyte–macrophage colony-stimulating factor and oncotoxic protein lactaptin genes. The virus has high cytotoxic activity against human cancer cells of various histogenesis and antitumor efficacy against breast cancer. In this work, we show VV-GMCSF-Lact to be a promising therapeutic agent for glioblastoma treatment. VV-GMCSF-Lact effectively decreases the viability of glioblastoma cells of both immortalized and patient-derived cultures in vitro, crosses the blood–brain barrier, selectively replicates into orthotopically transplanted human glioblastoma when intravenously injected, and inhibits glioblastoma xenograft and metastasis growth when injected intratumorally.

The Early Development of the Vaccinia–Rabies Recombinant Vaccine Raboral®

The recombinant vaccinia–rabies vaccine, now known as Raboral®, has been widely used in Europe and North America to control/eliminate rabies in the principal wildlife vectors, and thus prevent human transmission. The origins of this vaccine are sometimes forgotten, although the formulation has not changed substantially in almost four decades. This groundbreaking vaccine was assembled by a team at a very young (at that time) genetic engineering company, Transgène, in Strasbourg, France. The joint leaders of the rabies vaccine team reflect, 36 years later, on the trials and tribulations that went hand in hand with the construction of the vaccine.

Preparation of Class Y Immunoglobulins that Neutralize the Marburg Virus

The aim was to study the possibility of inducing Marburg-neutralizing chicken antibodies (MARV) using various immunogens.Materials and methods. Recombinant vaccinia virus expressing the surface glycoprotein (GP) transgene MARV of Musoke strain and pseudovirus particles exhibiting GP of three strains of MARV – Popp, Musoke and DRC2000 based on lentivirus and recombinant strain of vesicular stomatitis virus (VSV) were used as immunogens. Two groups of birds were involved in the study. Chickens were immunized 9 times: first time they were injected with the recombinant vaccinia virus, and then 8 times – with pseudovirus particles (based on lentivirus and a recombinant strain of the vesicular stomatitis virus). The accumulation of specific antibodies was evaluated by enzyme-linked immunosorbent assay (ELISA). We used recombinant VSV exhibiting GP MARV, and natural MARV strain Popp for the analysis of accumulation of neutralizing antibodies.Results and discussion. We have developed an effective immunization schedule for chickens with three recombinant constructs presenting GP MARV, which results in the induction of chicken IgY antibodies against Marburg virus with a titer in ELISA from 1:100 to 1:1 million. The obtained IgY neutralize MARV pseudoviruses (Popp, DRC2000, Musoke) at a dilution of 1/256 to 1/1024 and the natural MARV virus of the Popp strain at a dilution of 1/8. More stable results were demonstrated by immunization using Freund’s incomplete adjuvant.

Development of DNA Vaccine Targeting E6 and E7 Proteins of Human Papillomavirus 16 (HPV16) and HPV18 for Immunotherapy in Combination with Recombinant Vaccinia Boost and PD-1 Antibody

ABSTRACT Immunotherapy for cervical cancer should target high-risk human papillomavirus types 16 and 18, which cause 50% and 20% of cervical cancers, respectively. Here, we describe the construction and characterization of the pBI-11 DNA vaccine via the addition of codon-optimized human papillomavirus 18 (HPV18) E7 and HPV16 and 18 E6 genes to the HPV16 E7-targeted DNA vaccine pNGVL4a-SigE7(detox)HSP70 (DNA vaccine pBI-1). Codon optimization of the HPV16/18 E6/E7 genes in pBI-11 improved fusion protein expression compared to that in DNA vaccine pBI-10.1 that utilized the native viral sequences fused 3′ to a signal sequence and 5′ to the HSP70 gene of Mycobacterium tuberculosis. Intramuscular vaccination of mice with pBI-11 DNA better induced HPV antigen-specific CD8+ T cell immune responses than pBI-10.1 DNA. Furthermore, intramuscular vaccination with pBI-11 DNA generated stronger therapeutic responses for C57BL/6 mice bearing HPV16 E6/E7-expressing TC-1 tumors. The HPV16/18 antigen-specific T cell-mediated immune responses generated by pBI-11 DNA vaccination were further enhanced by boosting with tissue-antigen HPV vaccine (TA-HPV). Combination of the pBI-11 DNA and TA-HPV boost vaccination with PD-1 antibody blockade significantly improved the control of TC-1 tumors and extended the survival of the mice. Finally, repeat vaccination with clinical-grade pBI-11 with or without clinical-grade TA-HPV was well tolerated in vaccinated mice. These preclinical studies suggest that the pBI-11 DNA vaccine may be used with TA-HPV in a heterologous prime-boost strategy to enhance HPV 16/18 E6/E7-specific CD8+ T cell responses, either alone or in combination with immune checkpoint blockade, to control HPV16/18-associated tumors. Our data serve as an important foundation for future clinical translation. IMPORTANCE Persistent expression of high-risk human papillomavirus (HPV) E6 and E7 is an obligate driver for several human malignancies, including cervical cancer, wherein HPV16 and HPV18 are the most common types. PD-1 antibody immunotherapy helps a subset of cervical cancer patients, and its efficacy might be improved by combination with active vaccination against E6 and/or E7. For patients with HPV16+ cervical intraepithelial neoplasia grade 2/3 (CIN2/3), the precursor of cervical cancer, intramuscular vaccination with a DNA vaccine targeting HPV16 E7 and then a recombinant vaccinia virus expressing HPV16/18 E6-E7 fusion proteins (TA-HPV) was safe, and half of the patients cleared their lesions in a small study (NCT00788164). Here, we sought to improve upon this therapeutic approach by developing a new DNA vaccine that targets E6 and E7 of HPV16 and HPV18 for administration prior to a TA-HPV booster vaccination and for application against cervical cancer in combination with a PD-1-blocking antibody.

Injection site vaccinology of a recombinant vaccinia-based vector reveals diverse innate immune signatures

Poxvirus systems have been extensively used as vaccine vectors. Herein a RNA-Seq analysis of intramuscular injection sites provided detailed insights into host innate immune responses, as well as expression of vector and recombinant immunogen genes, after vaccination with a new multiplication defective, vaccinia-based vector, Sementis Copenhagen Vector. Chikungunya and Zika virus immunogen mRNA and protein expression was associated with necrosing skeletal muscle cells surrounded by mixed cellular infiltrates. The multiple adjuvant signatures at 12 hours post-vaccination were dominated by TLR3, 4 and 9, STING, MAVS, PKR and the inflammasome. Th1 cytokine signatures were dominated by IFNγ, TNF and IL1β, and chemokine signatures by CCL5 and CXCL12. Multiple signatures associated with dendritic cell stimulation were evident. By day seven, vaccine transcripts were absent, and cell death, neutrophil, macrophage and inflammation annotations had abated. No compelling arthritis signatures were identified. Such injection site vaccinology approaches should inform refinements in poxvirus-based vector design.

Systems vaccinology analysis of a recombinant vaccinia-based vector reveals diverse innate immune signatures at the injection site

Poxvirus systems have been extensively used as vaccine vectors. Herein a systems vaccinology analysis of intramuscular injection sites provides detailed insights into host innate immune responses, as well as expression of vector and recombinant immunogen genes, after vaccination with a new multiplication defective, vaccinia-based vector, Sementis Copenhagen Vector. Chikungunya and Zika virus immunogen mRNA and protein expression was associated with necrosing skeletal muscle cells surrounded by mixed cellular infiltrates. Adjuvant signatures at 12 hours post-vaccination were dominated by TLR3, 4 and 9, STING, MAVS, PKR and the inflammasome. Th1 cytokine signatures were dominated by IFNγ, TNF and IL1β, and chemokine signatures by CCL5 and CXCL12. Multiple signatures associated with dendritic cell stimulation were evident. By day seven, vaccine transcripts were absent, and cell death, neutrophil, macrophage and inflammation annotations had abated. No compelling arthritis signatures were identified. Such innate systems vaccinology approaches should inform refinements in poxvirus-based vector design.