Adenovirus vaccine therapy with CD137L promotes CD8+ DCs-mediated multifunctional CD8+ T cell immunity and elicits potent anti-tumor activity.

Renal carcinoma progresses aggressively in patients with metastatic disease while curative strategies are limited. Here, we constructed a recombinant non-replicating adenovirus (Ad) vaccine encoding an immune activator, CD137L, and a tumor antigen, CAIX, for treating renal carcinoma. In a subcutaneous tumor model, tumor growth was significantly suppressed in the Ad-CD137L/CAIX vaccine group compared with the single vaccine group. The induction and maturity of CD11C+ and CD8+CD11C+ dendritic cell (DC) subsets were promoted in Ad-CD137L/CAIX co-immunized mice. Furthermore, the Ad-CD137L/CAIX vaccine elicited stronger tumor-specific multifunctional CD8+ T cell immune responses as demonstrated by increased proliferation and cytolytic function of CD8+ T cells. Notably, depletion of CD8+ T cells greatly compromised the effective protection provided by Ad-CD137L/CAIX vaccine, suggesting an irreplaceable role of CD8+ T cells for the immunopotency of the vaccine. In both lung metastatic and orthotopic models, Ad-CD137L/CAIX vaccine treatment significantly decreased tumor metastasis and progression and increased the induction of tumor-specific multifunctional CD8+ T cells, in contrast to treatment with the Ad-CAIX vaccine alone. The Ad-CD137L/CAIX vaccine also augmented the tumor-specific multifunctional CD8+ T cell immune response in both orthotopic and metastatic models. These results indicated that Ad-CD137L/CAIX vaccine elicited a potent anti-tumor activity by inducing CD8+DC-mediated multifunctional CD8+ T cell immune responses. The potential strategy of CD137L-based vaccine might be served as a novel treatment for renal carcinoma or other malignant tumors.

Pediatric acute gastroenteritis associated with adenovirus 40/41 in low-income and middle-income countries.

PURPOSE OF REVIEW: To review the roles of enteric adenovirus types 40 and 41 and nonenteric adenoviruses in the global burden of pediatric diarrhea. RECENT FINDINGS: Large studies using highly sensitive, type-specific molecular diagnostics have demonstrated a substantial and previously under-estimated burden of pediatric diarrheal disease because of enteric infections with adenovirus types 40/41. However, the true epidemiology of adenovirus 40/41 remains incompletely understood. Similarly, additional adenovirus types may also be implicated as agents of community-acquired pediatric gastroenteritis but current data are too limited to elucidate their epidemiological role(s), if any. SUMMARY: Efforts at global diarrhea control in low-income and middle-income countries will require combating pediatric gastroenteritis because of enteric adenovirus infections. Future research in these settings using type-specific molecular diagnostics or strain genotyping to fully characterize the epidemiology of adenovirus 40/41 infections, identify non-40/41 adenoviruses significantly associated with gastroenteritis, and develop vaccines effective at preventing adenovirus diarrhea is warranted.

Adenoviral Infections in Singapore: Should New Antiviral Therapies and Vaccines Be Adopted?

BACKGROUND: A number of serious human adenovirus (HAdV) outbreaks have been recently reported: HAdV-B7 (Israel, Singapore, and USA), HAdV-B7d (USA and China), HAdV-D8, -D54, and -C2 (Japan), HAdV-B14p1 (USA, Europe, and China), and HAdV-B55 (China, Singapore, and France). METHODS: To understand the epidemiology of HAdV infections in Singapore, we studied 533 HAdV-positive clinical samples collected from 396 pediatric and 137 adult patients in Singapore from 2012 to 2018. Genome sequencing and phylogenetic analyses were performed to identify HAdV genotypes, clonal clusters, and recombinant or novel HAdVs. RESULTS: The most prevalent genotypes identified were HAdV-B3 (35.6%), HAdV-B7 (15.4%), and HAdV-E4 (15.2%). We detected 4 new HAdV-C strains and detected incursions with HAdV-B7 (odds ratio [OR], 14.6; 95% confidence interval [CI], 4.1-52.0) and HAdV-E4 (OR, 13.6; 95% CI, 3.9-46.7) among pediatric patients over time. In addition, immunocompromised patients (adjusted OR [aOR], 11.4; 95% CI, 3.8-34.8) and patients infected with HAdV-C2 (aOR, 8.5; 95% CI, 1.5-48.0), HAdV-B7 (aOR, 3.7; 95% CI, 1.2-10.9), or HAdV-E4 (aOR, 3.2; 95% CI, 1.1-8.9) were at increased risk for severe disease. CONCLUSIONS: Singapore would benefit from more frequent studies of clinical HAdV genotypes to identify patients at risk for severe disease and help guide the use of new antiviral therapies, such as brincidofovir, and potential administration of HAdV 4 and 7 vaccine.

In vitro evaluation of the therapeutic effectiveness of EBV-LMP2 recombinant adenovirus vaccine in nasopharyngeal carcinoma.

Immunotherapeutic strategies based on Epstein-Barr virus (EBV) latent membrane protein 2 (LMP2) antigen-specific cytotoxic T lymphocytes (CTLs) have been proven to boost LMP2-specific CTL responses in patients with nasopharyngeal carcinoma (NPC). Such strategies can produce clinical benefits in some patients with NPC. Currently, the major challenge limiting the use of immunotherapy for NPC is its low clinical response rate. The efficacy of immunotherapy based on EBV-LMP2 specific CTLs depends mainly on their cytotoxic activity, but no studies have been conducted to elucidate this activity. In this study, laser confocal scanning microscopy (LCSM) and real-time cell analysis (RTCA) were used to evaluate the killing function and its underlying mechanism of LMP2-specific CTLs. LCSM showed that LMP2-specific CTLs recognize and kill target cells expressing viral escape protein LMP2, and that the killing rate is related to the number of CTLs adhering to the target cells. LMP2-specific CTL-mediated cytotoxicity is rate limited by the time required for effective contact and recognition between CTLs and target cells. RTCA showed that the protective effect of LMP2-specific CTLs required an appropriate effector-to-target ratio, and that LMP2-specific CTLs could not eradicate residual target cells at a low effector-to-target ratio. Moreover, our results revealed that LMP2-specific CTL responses involve two independent but complementary mechanisms: the perforin/granzyme and Fas/FasL pathways. Therefore, we have elucidated, for the first time, the selective cytotoxicity and mechanism by which LMP2-specific CTLs induced by the rAd-LMP2 vaccine kill target cells and have explored the killing mode and several key parameters of killing mediated by LMP2-specific CTLs. Our study will contribute to the knowledge of vaccines targeting EBV-LMP2 and to the improvement of immunotherapeutic strategies.

Longitudinal quantification of adenovirus neutralizing responses in Zambian mother-infant pairs: Impact of HIV-1 infection and its treatment.

Vaccination offers the most cost-effective approach to limiting the adverse impact of infectious and neoplastic diseases that reduce the quality of life in sub-Saharan Africa (SSA). However, it is unclear what vaccine vectors would be most readily implementable in the setting and at what age they should be applied for maximal efficacy. Adenoviruses (Ad) and Ad-based vectors have been demonstrated to induce effective humoral and cellular immune responses in animal models and in humans. However, because immunity associated with Ad infection is lifelong, there exists a debate as to whether pre-existing immunity might decrease the efficacy of Ad vectored vaccines. In order to begin to rationally develop vaccination strategies for SSA, we have quantified neutralizing antibodies (nAb) against Ad4, Ad5, Ad7, Ad26, Ad28, Ad45 and Ad48 in 67 adult women and their infants. We are the first to define the decay kinetics of transferred maternal nAb in infants as well as the apparent initiation of de novo Ad responses. Our findings demonstrate that in Zambian adults, robust nAb responses exist against each of the Ads tested and are efficiently transferred to newborns. With few exceptions, neither the HIV-1 infection status of the mothers or the antiretroviral therapy (ART) treatment of HIV-1 disease had significant impact on maternal Ad nAb responses or their transfer to infants. However, maternal Ad nAb decays in infants to a nadir at 12 months of age such that any of the seven Ad types could function as vaccine vectors. The definition of this 'window of opportunity' provides important foundational data for rational design and implementation of Ad vectors in this setting.

Adenovirus 4/7 Vaccine's Effect on Disease Rates Is Associated With Disappearance of Adenovirus on Building Surfaces at a Military Recruit Base.

BACKGROUND: Febrile respiratory illness resulting from adenovirus types 4 and 7 (Ad4/7) was endemic at military training camps, but controlled by an Ad4/7 vaccine from the 1970s to 1999, the year it was discontinued. Thereafter, rates returned to prevaccine levels. Rates dropped after reintroduction of an Ad4/7 vaccine in 2011. METHODS: Surfaces of the barracks and medical clinic of a training camp were swabbed in 3 studies in 2004 and 1 study in 2007, and tested with culture and polymerase chain reaction (PCR). Similar swabbing was done in 2013 and 2015 and tested with PCR. FINDINGS: In the studies before 2011 (prevaccine), 12% of samples were Ad4/7 positive by culture and 27% positive by PCR. In the 2 studies after 2011 (postvaccine), no samples were Ad4/7 positive. DISCUSSION/IMPACT/RECOMMENDATIONS: The Ad 4/7 vaccine has resulted in the near elimination of Ad4/7-related disease and the disappearance of Ad4/7 from surfaces in a military basic training camp. Renewed transmission of Ad4/7 in this setting would likely require new importation from military recruits and an immunologically naive cohort, which the current vaccination program prevents.

Adenovirus-Associated Acute Appendicitis: An Under-Recognized Relationship?

OBJECTIVE: Acute appendicitis (AA) is one of the most common causes of a surgical abdomen worldwide, occurring most frequently in those age 10 to 29 years. Adenovirus (ADV) is a rare but reported cause of AA in children and a well-recognized cause of intussusception in infants and young children. Annually, about 36,000 basic military trainees (BMTs) undergo initial training at Joint Base San Antonio Lackland, Texas. Before reintroduction of the ADV 4/7 vaccine in November 2011, one-third of BMTs developed an adenoviral upper respiratory tract infection (URI) during the 8.5 weeks of training. We hypothesized that ADV may be a common cause of AA in the BMT population given their young age and high incidence of adenoviral URIs. The objective of this study was to determine the frequency with which ADV, cytomegalovirus (CMV), Epstein-Barr virus (EBV), and enterovirus were associated with AA in a population of young adults. MATERIALS AND METHODS: This study was a retrospective review of patient charts and existing pathological tissue specimens of all BMTs who underwent appendectomy at the Wilford Hall Medical Center from January 1, 2003, to August 31, 2011. Pathological tissue samples from 112 BMTs were assayed by quantitative polymerase chain reaction (qPCR) and immunohistochemistry (IHC) for viral targets. RESULTS: ADV DNA was detected in 16 of 112 samples (14%) via qPCR: ADV 4 in 13 cases, ADV B14 in 1 case, and nontypable ADV in 2 cases. IHC was positive in only the ADV B14 case (0.9%). All cases were negative for CMV, EBV, and enterovirus. CONCLUSION: By using qPCR, this study demonstrated an association between ADV and AA higher than has been previously reported: ADV was detected in 14% of AA cases in this series versus in only 0.23% of AA cases in previous studies (p < 0.01). There was no evidence of CMV, EBV, or enterovirus association with AA in this study. Comparison of qPCR to IHC shows that histologic analysis may overlook evidence of ADV in appendiceal tissue: qPCR is significantly more sensitive than light microscopy and IHC for detecting ADV in this setting. Because ADV 4 was detected in 81% of those with positive qPCR, the recently licensed live oral ADV vaccine might be useful for primary prevention against AA. Prospective studies evaluating young adults presenting with AA for evidence of infection with ADV are needed to determine if a causal relationship exists.

Safety evaluation of adenovirus type 4 and type 7 vaccine live, oral in military recruits.

Before the widespread adoption of vaccination, adenovirus type 4 and type 7 were long associated with respiratory illnesses among military recruits. When supplies were depleted and vaccination was suspended in 1999 for approximately a decade, respiratory illnesses due to adenovirus infections resurged. In March 2011, a new live, oral adenovirus vaccine was licensed by the US Food and Drug Administration and was first universally administered to military recruits in October 2011, leading to rapid, dramatic elimination of the disease within a few months. As part of licensure, a postmarketing study (Sentinel Surveillance Plan) was performed to detect potential safety signals within 42days after immunization of military recruits. This study retrospectively evaluated possible adverse events related to vaccination using data from the Armed Forces Health Surveillance Branch Defense Medical Surveillance System (DMSS) database. Among 100,000 recruits who received the adenovirus vaccine, no statistically significant greater risk of prespecified medical events was observed within 42days after vaccination when compared with a historical cohort of 100,000 unvaccinated recruits. In an initial statistical analysis of International Classification of Disease, 9th Revision, Clinical Modification codes, a statistically significant higher risk for 19 other (not prespecified) medical events occurring in 5 or more recruits was observed among vaccinated compared with unvaccinated groups. After case record data abstraction for attribution and validation, two events (psoriasis [21 vs 7 cases] and serum reactions [12 vs 4 cases]) occurred more frequently in the vaccinated cohort. A causal relation of these rare events with adenovirus vaccination could not be established given confounding factors in the DMSS, such as coadministration of other vaccines and incomplete or inaccurate medical information, for some recruits. Prospective surveillance assessing these uncommon, but potentially relevant, immune-related symptoms may be beneficial in defining potential causal association with adenovirus vaccination.

The generation and analyses of a novel combination of recombinant adenovirus vaccines targeting three tumor antigens as an immunotherapeutic.

Phenotypic heterogeneity of human carcinoma lesions, including heterogeneity in expression of tumor-associated antigens (TAAs), is a well-established phenomenon. Carcinoembryonic antigen (CEA), MUC1, and brachyury are diverse TAAs, each of which is expressed on a wide range of human tumors. We have previously reported on a novel adenovirus serotype 5 (Ad5) vector gene delivery platform (Ad5 [E1-, E2b-]) in which regions of the early 1 (E1), early 2 (E2b), and early 3 (E3) genes have been deleted. The unique deletions in this platform result in a dramatic decrease in late gene expression, leading to a marked reduction in host immune response to the vector. Ad5 [E1-, E2b-]-CEA vaccine (ETBX-011) has been employed in clinical studies as an active vaccine to induce immune responses to CEA in metastatic colorectal cancer patients. We report here the development of novel recombinant Ad5 [E1-, E2b-]-brachyury and-MUC1 vaccine constructs, each capable of activating antigen-specific human T cells in vitro and inducing antigen-specific CD4+ and CD8+ T cells in vaccinated mice. We also describe the use of a combination of the three vaccines (designated Tri-Ad5) of Ad5 [E1-, E2b-]-CEA, Ad5 [E1-, E2b-]-brachyury and Ad5 [E1-, E2b-]-MUC1, and demonstrate that there is minimal to no "antigenic competition" in in vitro studies of human dendritic cells, or in murine vaccination studies. The studies reported herein support the rationale for the application of Tri-Ad5 as a therapeutic modality to induce immune responses to a diverse range of human TAAs for potential clinical studies.

ΔE1 and high-capacity adenoviral vectors expressing full-length codon-optimized merozoite surface protein 1 for vaccination against Plasmodium falciparum.

BACKGROUND: The merozoite surface protein (MSP)-1 of Plasmodium falciparum, the causative agent of malaria tropica, is considered to be a promising vaccine candidate. Although its stable cloning and expression has been difficult in the past, adenoviral vectors expressing the complex protein are described in the present study. METHODS: Codon-optimized msp-1 was used to construct a set of first generation (ΔE1Ad) and high-capacity adenovirus (HC-Ad) vectors, and cellular and humoral immune responses induced by the vectors were characterized in detail in mice. RESULTS: Generation of stable ΔE1Ad and HC-Ad vectors expressing full-length MSP-1 and their production to high vector titers was found to be feasible. Epitope identification and analysis of frequencies of specific CD8 T-cells revealed that MSP-1 expressing HC-Ad vectors induced higher frequencies of interferon-γ + CD8 T-cells than ΔE1 vectors. Irrespective of the vector format, higher titers of MSP-1 specific antibodies were generated by Ad vectors expressing MSP-1 from a chicken ß-actin (CAG) promoter comprising the cytomegalovirus early enhancer element and the chicken ß-actin promoter. CONCLUSIONS: The findings of the present study suggest that Ad vectors expressing full-length codon-optimized MSP-1 are promising candidate vaccines against P. falciparum infections. Use of the HC-Ad vector type for delivery, as well as the CAG promoter to control MSP-1 expression, may further increase the efficacy of this vaccine candidate.

High-capacity adenoviral vectors circumvent the limitations of ΔE1 and ΔE1/ΔE3 adenovirus vectors to induce multispecific transgene product-directed CD8 T-cell responses.

BACKGROUND: The ability to induce cytotoxic T lymphocyte (CTL) responses that are multispecific is considered to comprise an essential feature for an efficacious genetic vaccine against many pathogens including HIV and hepatitis C virus. ΔE1Ad vectors are promising vectored vaccines but have been shown to induce antigen-specific CTLs with only limited multispecificity. In the present study, we investigated the applicability of gene-deleted high-capacity adenovirus (HC-Ad) vectors and focused on the induction of multispecific CTL responses. METHODS: We generated Δ E1 and HC-Ad vectors expressing hepatitis B virus small surface antigen (HBsAg). We comparatively analyzed the CTL profiles against various transgene product- and vector-derived epitopes in several mouse strains and HBsAg- and vector-directed antibody responses. RESULTS: HC-Ad vectors efficiently induced multispecific HBsAg-directed CTLs. By contrast, ΔE1Ad vectors mainly primed CTLs against one immunodominant epitope of HBsAg. This absence of multispecific CTL responses correlated with the induction of CTLs against viral epitopes generated by de novo expression of Ad genes from the ΔE1Ad vector. However, Ad-specific CTLs induced in trans did not impair HC-AdS-induced multispecific CTL responses against HBsAg. Finally, HC-Ad vectors also induced higher HBsAg antibody titers compared to ΔE1Ad vectors. CONCLUSIONS: De novo expression of viral genes from ΔE1Ad vector genomes restricts the multispecificity of transgene product-specific CTLs by immunodominance effects. HC-Ad vectors devoid of Ad genes are favorable for the induction of both multispecific CD8 T-cell responses and high antibody responses. Our results suggest the deletion of Ad genes as an important means for developing potent Ad-based vectored vaccines.