UNASSIGNED: Monkeypox (Mpox) has become a concern worldwide after spreading into nonendemic countries. The World Health Organization (WHO) has declared this a public health emergency of international concern and recommended to get vaccinated first who are at the highest risk. Risk perception and subjective norms can influence the decision of vaccine uptake. Therefore, we intended to perform a cross-sectional study on the male population in our country to assess their risk perception and subjective norms on Mpox.
UNASSIGNED: We measured participants\' risk perception and subjective norms using Google form. Demographic profile of participants was obtained using a structured questionnaire. We performed a χ 2 test to compare the levels of risk perception and subjective norm perception and multiple logistic regression analysis to determine the association between the study parameters and the sociodemographic profile of the participants.
UNASSIGNED: Among the participants, 93 (23.72%), 288 (73.47%), and 11 (2.81%) had high, medium, and low-risk perceptions, respectively. For subjective norms, we observed 288 (58.16%) participants had a medium, 117 (29.85%) had high, and 47 (11.99%) had low levels of subjective norms, respectively. Most participants possessed medium risk perception (73.47%) and subjective norms (58.16%). Moreover, we observed that moderate risk perception was prevalent in people with body mass index (BMI) level between 18.5 and 25 (73.3%), married (63.5%), low economic background (94.1%), living with a family (77.1%), smokers (68.4%), heterosexuals (99%), people with no/little impact of coronavirus disease 2019 (Covid-19) on life (91%). Proportions of people with moderate subjective norms BMI level of 18.5-25 (73.2%), married (60.5%), low economic status (93.9%), rural (58.8%), living with family (77.2%), nonsmokers (71.1%), and people with no/little impact of Covid-19 in their lives (91.2%).
UNASSIGNED: The majority of participants perceived medium risk perception and subjective norms related to Mpox. Furthermore, we observed a significant association between the study parameters and the sociodemographic characteristics of our study participants. We recommend that further longitudinal studies to yield more accurate results.

This article presents the results of isolation of camel smallpox virus (Poxviridae: Orthopoxvirus: Camelpox virus, CMLPV) and study of its reproductive properties on sensitive biological systems.
The epizootic strain M-96 of the virus as well as its attenuated variants KM-40 and KM-70 obtained by sequential passivation were used in the study. Isolation of the pathogen from suspension of biopsy specimens was performed on cell culture and in embryonated chicken eggs (ECEs). All experiments were performed with the number of replications ensuring obtaining reliable results.
The CMLPV was isolated from the crusts and pox papules of the skin taken from sick camels (Camelus bactrianus) during an outbreak in various districts of the Mangistau region at the end of 2019. The signs of pathogen reproduction on chorio-allantoic membrane (CAM) were observed from 3 passages. The obtained virus caused formation of pathological changes on the CAM in the form of elevated dot or solid white formations separated from the surrounding tissue, with hemorrhagic foci in the center. The reproductive properties of the isolate on sensitive biological systems were determined in comparison with the epizootic CMLPV strain M-96, isolated earlier in the territory of Kazakhstan during the outbreak 23-24 years ago, as well as its attenuated variants. The isolated virus was given the conventional name M-2020.
When studied in two sensitive cultivation systems (cell culture and ECEs), strain M-96 and its attenuated variants KM-40, KM-70, which were used in the experiments as a control, demonstrated high infectious activity with titer 4.75-6.75 lg TCID50/cm3, while for the examined isolate M-2020 of CMLPV had the significantly lower values (3.00-4.75 lg TCID50/cm3, p > 0,05).

DNA vectors have been widely used as a priming of poxvirus vaccine in prime/boost regimens. Whether the number of DNA impacts qualitatively or quantitatively the immune response is not fully explored. With the aim to reinforce T-cell responses by optimizing the prime-boost regimen, the multicentric EV03/ANRS VAC20 phase I/II trial, randomized 147 HIV-negative volunteers to either 3xDNA plus 1xNYVAC (weeks 0, 4, 8 plus 24; n = 74) or to 2xDNA plus 2xNYVAC (weeks 0, 4 plus 20, 24; n = 73) groups. T-cell responses (IFN-γ ELISPOT) to at least one peptide pool were higher in the 3xDNA than the 2xDNA groups (91% and 80% of vaccinees) (P = 0.049). In the 3xDNA arm, 26 (37%) recipients developed a broader T-cell response (Env plus at least to one of the Gag, Pol, Nef pools) than in the 2xDNA (15; 22%) arms (primary endpoint; P = 0.047) with a higher magnitude against Env (at week 26) (P<0.001). In both groups, vaccine regimens induced HIV-specific polyfunctional CD4 and CD8 T cells and the production of Th1, Th2 and Th17/IL-21 cytokines. Antibody responses were also elicited in up to 81% of vaccines. A higher percentage of IgG responders was noted in the 2xDNA arm compared to the 3xDNA arm, while the 3xDNA group tended to elicit a higher magnitude of IgG3 response against specific Env antigens. We show here that the modulation of the prime strategy, without modifying the route or the dose of administration, or the combination of vectors, may influence the quality of the responses.

Every poxvirus mRNA transcribed after viral DNA replication has an evolutionarily conserved, non-templated 5\'-poly(A) leader in the 5\'-UTR. To dissect the role of 5\'-poly(A) leader in mRNA translation during poxvirus infection we developed an in vitro transcribed RNA-based luciferase reporter assay. This reporter assay comprises of four core steps: (1) PCR to amplify the DNA template for in vitro transcription; (2) in vitro transcription to generate mRNA using T7 RNA polymerase; (3) Transfection to introduce in vitro transcribed mRNA into cells; (4) Detection of luciferase activity as the indicator of translation. The RNA-based luciferase reporter assay described here circumvents issues of plasmid replication in poxvirus-infected cells and cryptic transcription from the plasmid. This protocol can be used to determine translation regulation by cis-elements in an mRNA including 5\'-UTR and 3\'-UTR in systems other than poxvirus-infected cells. Moreover, different modes of translation initiation like cap-dependent, cap-independent, re-initiation, and internal initiation can be investigated using this method.

Proteins harbor domains or short linear motifs, which facilitate their functions and interactions. Finding functional motifs in protein sequences could predict the putative cellular roles or characteristics of hypothetical proteins. In this study, we present Shetti-Motif, which is an interactive tool to (i) map UniProt and PROSITE flat files, (ii) search for multiple pre-defined consensus patterns or experimentally validated functional motifs in large datasets protein sequences (proteome-wide), (iii) search for motifs containing repeated residues (low-complexity regions, e.g., Leu-, SR-, PEST-rich motifs, etc.). As proof of principle, using this comparative proteomics pipeline, eleven proteomes encoded by member of Poxviridae family were searched against about 100 experimentally validated functional motifs. The closely related viruses and viruses infect the same host cells (e.g. vaccinia and variola viruses) show similar motif-containing proteins profile. The motifs encoded by these viruses are correlated, which explains why poxviruses are able to interact with wide range of host cells. In conclusion, this in silico analysis is useful to establish a dataset(s) or potential proteins for further investigation or compare between species.

We have conducted a phase 1 study of intravenous vvDD, a Western Reserve strain oncolytic vaccinia virus, on 11 patients with standard treatment-refractory advanced colorectal or other solid cancers. The primary endpoints were maximum tolerated dose and associated toxicity while secondary endpoints were pharmacokinetics, pharmacodynamics, immune responses, and antitumor activity. No dose-limiting toxicities and treatment related severe adverse events were observed. The most common adverse events were grades 1/2 flu-like symptoms. Virus genomes were detectable in the blood 15-30 minutes after virus administration in a dose-dependent manner. There was evidence of a prolonged virus replication in tumor tissues in two patients, but no evidence of virus replication in non-tumor tissues, except a healed injury site and an oral thrush. Over 100-fold of anti-viral antibodies were induced in patients\' sera. A strong induction of inflammatory and Th1, but not Th2 cytokines, suggested a potent Th1-mediated immunity against the virus and possibly the cancer. One patient showed a mixed response on PET-CT with resolution of some liver metastases, and another patient with cutaneous melanoma demonstrated clinical regression of some lesions. Given the confirmed safety, further trials evaluating intravenous vvDD in combination with therapeutic transgenes, immune checkpoint blockade or complement inhibitors, are warranted.

OBJECTIVE: To determine whether 1 of 2 vaccines based on dendritic cells (DCs) and poxvectors encoding CEA (carcinoembryonic antigen) and MUC1 (PANVAC) would lengthen survival in patients with resected metastases of colorectal cancer (CRC).
BACKGROUND: Recurrences after complete resections of metastatic CRC remain frequent. Immune responses to CRC are associated with fewer recurrences, suggesting a role for cancer vaccines as adjuvant therapy. Both DCs and poxvectors are potent stimulators of immune responses against cancer antigens.
METHODS: Patients, disease-free after CRC metastasectomy and perioperative chemotherapy (n = 74), were randomized to injections of autologous DCs modified with PANVAC (DC/PANVAC) or PANVAC with per injection GM-CSF (granulocyte-macrophage colony-stimulating factor). Endpoints were recurrence-free survival overall survival, and rate of CEA-specific immune responses. Clinical outcome was compared with that of an unvaccinated, contemporary group of patients who had undergone CRC metastasectomy, received similar perioperative therapy, and would have otherwise been eligible for the study.
RESULTS: Recurrence-free survival at 2 years was similar (47% and 55% for DC/PANVAC and PANVAC/GM-CSF, respectively) (χ P = 0.48). At a median follow-up of 35.7 months, there were 2 of 37 deaths in the DC/PANVAC arm and 5 of 37 deaths in the PANVAC/GM-CSF arm. The rate and magnitude of T-cell responses against CEA was statistically similar between study arms. As a group, vaccinated patients had superior survival compared with the contemporary unvaccinated group.
CONCLUSIONS: Both DC and poxvector vaccines have similar activity. Survival was longer for vaccinated patients than for a contemporary unvaccinated group, suggesting that a randomized trial of poxvector vaccinations compared with standard follow-up after metastasectomy is warranted. (NCT00103142).

BACKGROUND: Childhood immunology has been suggested to play a role in development of inflammatory bowel disease (IBD) based on the studies of childhood vaccinations, infections, and treatment with antibiotics. Bacille Calmette-Guérin (BCG) and smallpox vaccinations were gradually phased-out in Denmark for children born between 1965 and 1976, hence allowing the study of subsequent risk of Crohn\'s disease and ulcerative colitis in a unique prospective design.
METHODS: The Copenhagen School Health Records Register contains detailed documentation of vaccination. Among the background cohort of individuals born between 1965 and 1976 (N = 47,622), cases with Crohn\'s disease (n = 218) and ulcerative colitis (n = 256) were identified through linkage to the Danish National Patient Registry. The vaccination status of the cases was compared with that of a subcohort (n = 5741) of the background cohort and analyzed in a case-cohort design.
RESULTS: No difference in risk of IBD was observed between individuals vaccinated and unvaccinated with BCG (hazard ratio = 0.95; 95% confidence interval, 0.75-1.19) or smallpox vaccine (hazard ratio = 1.01; 95% confidence interval, 0.77-1.32). This was also the case for Crohn\'s disease and ulcerative colitis separately. However, BCG given before 4 months of age may decrease the risk of IBD (hazard ratio = 0.43; 95% confidence interval, 0.20-0.93).
CONCLUSIONS: This prospective long-term case-cohort study shows that BCG and smallpox vaccination do not cause IBD later in life. These findings are important for the etiological understanding of IBD and of clinical importance because BCG is still one of the most commonly used childhood vaccinations, smallpox vaccine has been reintroduced in the U.S. military, and both vaccines may be used as vectors in new vaccines.

BACKGROUND: Therapeutic cancer vaccines have shown activity in metastatic castration-resistant prostate cancer (mCRPC), and methods are being assessed to enhance their efficacy. Ipilimumab is an antagonistic monoclonal antibody that binds cytotoxic T-lymphocyte-associated protein 4, an immunomodulatory molecule expressed by activated T cells, and to CD80 on antigen-presenting cells. We aimed to assess the safety and tolerability of ipilimumab in combination with a poxviral-based vaccine targeting prostate-specific antigen (PSA) and containing transgenes for T-cell co-stimulatory molecule expression, including CD80.
METHODS: We did a phase 1 dose-escalation trial, with a subsequent expansion phase, to assess the safety and tolerability of escalating doses of ipilimumab in combination with a fixed dose of the PSA-Tricom vaccine. Patients with mCRPC received 2×10(8) plaque-forming units of recombinant vaccinia PSA-Tricom subcutaneously on day 1 of cycle 1, with subsequent monthly boosts of 1×10(9) plaque-forming units, starting on day 15. Intravenous ipilimumab was given monthly starting at day 15, in doses of 1, 3, 5, and 10 mg/kg. Our primary goal was to assess the safety of the combination. This study is registered with ClinicalTrials.gov, number NCT00113984.
RESULTS: We completed enrolment with 30 patients (24 of whom had not been previously treated with chemotherapy) and we did not identify any dose-limiting toxic effects. Grade 1 and 2 vaccination-site reactions were the most common toxic effects: three of 30 patients had grade 1 reactions and 26 had grade 2 reactions. 21 patients had grade 2 or greater immune-related adverse events. Grade 3 or 4 immune-related adverse events included diarrhoea or colitis in four patients and grade 3 rash (two patients), grade 3 raised aminotransferases (two patients), grade 3 endocrine immune-related adverse events (two patients), and grade 4 neutropenia (one patient). Only one of the six patients previously treated with chemotherapy had a PSA decline from baseline. Of the 24 patients who were chemotherapy-naive, 14 (58%) had PSA declines from baseline, of which six were greater than 50%.
CONCLUSIONS: The use of a vaccine targeting PSA that also enhances co-stimulation of the immune system did not seem to exacerbate the immune-related adverse events associated with ipilimumab. Randomised trials are needed to further assess clinical outcomes of the combination of ipilimumab and vaccine in mCRPC.
BACKGROUND: US National Institutes of Health.

OBJECTIVE: PANVAC is a recombinant poxviral vaccine that contains transgenes for MUC-1, CEA, and 3 T-cell costimulatory molecules. This study was conducted to obtain preliminary evidence of clinical response in metastatic breast and ovarian cancer patients.
METHODS: Twenty-six patients were enrolled and given monthly vaccinations. Clinical and immune outcomes were evaluated.
RESULTS: These patients were heavily pretreated, with 21 of 26 patients having 3 or more prior chemotherapy regimens. Side effects were largely limited to mild injection-site reactions. For the 12 breast cancer patients enrolled, median time to progression was 2.5 months (1-37+) and median overall survival was 13.7 months. Four patients had stable disease. One patient had a complete response by RECIST and remained on study for 37 months or more, with a significant drop in serum interleukin (IL)-6 and IL-8 by day 71. Another patient with metastatic disease confined to the mediastinum had a 17% reduction in mediastinal mass and was on study for 10 months. Patients with stable or responding disease had fewer prior therapies and lower tumor marker levels than patients with no evidence of response. For the ovarian cancer patients (n = 14), the median time to progression was 2 months (1-6) and median overall survival was 15.0 months. Updated data are presented here for one patient treated with this vaccine in a previous trial, with a time to progression of 38 months.
CONCLUSIONS: Some patients who had limited tumor burden with minimal prior chemotherapy seemed to benefit from the vaccine. Further studies to confirm these results are warranted.