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PubMed1993

Protection of cats against feline leukemia virus by vaccination with a canarypox virus recombinant, ALVAC-FL.

Two ALVAC (canarypox virus)-based recombinant viruses expressing the feline leukemia virus (FeLV) subgroup A env and gag genes were assessed for their protective efficacy in cats. Both recombinant viruses contained the entire gag gene. ALVAC-FL also expressed the entire envelope glycoprotein, while ALVAC-FL(dl IS) expressed an env-specific gene product deleted of the putative immunosuppressive region. Although only 50% of the cats vaccinated with ALVAC-FL(dl IS) were protected against persistent viremia after oronasal exposure to a homologous FeLV isolate, all cats administered ALVAC-FL resisted the challenge exposure. Significantly, protection was afforded in the absence of detectable FeLV-neutralizing antibodies. These results represent the first effective vaccination of cats against FeLV with a poxvirus-based recombinant vector and have implications that are relevant not only to FeLV vaccine development but also to developing vaccines against other retroviruses, including human immunodeficiency virus.

Animals↗
PubMed2003

DNA immunization with hepatitis C virus (HCV) polycistronic genes or immunization by HCV DNA priming-recombinant canarypox virus boosting induces immune responses and protection from recombinant HCV-vaccinia virus infection in HLA-A2.1-transgenic mice.

We studied immune responses to hepatitis C virus (HCV) genes delivered as DNA encoding the entire HCV protein coding genome in two polycistronic plasmids encoding HCV capsid-E1-E2-NS2-NS3 and HCV NS3-NS4-NS5 in HLA-A2.1-transgenic mice. Immune responses to HCV DNA prime and recombinant canarypox virus boost were also studied with the above constructs. At 8 weeks after a canarypox virus boost, the DNA prime/canarypox virus boosting regimen induced potent cellular immune responses to HCV structural and nonstructural proteins on target cells expressing the HLA-A2.1 allele. High frequencies of gamma interferon-secreting cells, as detected by enzyme-linked immunospot assay, were obtained in response to several endogenously expressed HCV proteins. We also observed cytotoxic-T-lymphocyte reactivity in response to endogenously expressed HCV proteins in fresh spleen cells without in vitro expansion. Upon challenge with a recombinant vaccinia virus expressing HCV proteins at 2 months postimmunization, the HCV DNA prime/canarypox virus-immunized mice showed a complete reduction in vaccinia virus titers compared to HCV DNA prime/boost- and mock-immunized controls. Immune responses were still detectable 4 months after canarypox virus boost in immunized mice. Interestingly, at 10 months postimmunization (8 months after canarypox virus boost), the protection in HCV DNA prime/boost-immunized mice against recombinant HCV-vaccinia virus challenge was higher than that observed in HCV DNA prime/canarypox virus boost-immunized mice.

Animals↗
PubMed2001

Mature dendritic cells infected with canarypox virus elicit strong anti-human immunodeficiency virus CD8+ and CD4+ T-cell responses from chronically infected individuals.

Recombinant canarypox virus vectors containing human immunodeficiency virus type 1 (HIV-1) sequences are promising vaccine candidates, as they replicate poorly in human cells. However, when delivered intramuscularly the vaccines have induced inconsistent and in some cases transient antigen-specific cytotoxic T-cell (CTL) responses in seronegative volunteers. An attractive way to enhance these responses would be to target canarypox virus to professional antigen-presenting cells such as dendritic cells (DCs). We studied (i) the interaction between canarypox virus and DCs and (ii) the T-cell responses induced by DCs infected with canarypox virus vectors containing HIV-1 genes. Mature and not immature DCs resisted the cytopathic effects of canarypox virus and elicited strong effector CD8+ T-cell responses from chronically infected HIV+ individuals, e.g., cytolysis, and secretion of gamma interferon (IFN-gamma) and beta-chemokines. Furthermore, canarypox virus-infected DCs were >30-fold more efficient than monocytes and induced responses that were comparable to those induced by vaccinia virus vectors or peptides. Addition of exogenous cytokines was not necessary to elicit CD8+ effector cells, although the presence of CD4+ T cells was required for their expansion and maintenance. Most strikingly, canarypox virus-infected DCs were directly able to stimulate HIV-specific, IFN-gamma-secreting CD4 helper responses from bulk as well as purified CD4+ T cells. Therefore, these results suggest that targeting canarypox virus vectors to mature DCs could potentially elicit both anti-HIV CD8+ and CD4+ helper responses in vivo.

AIDS Vaccines↗
PubMed2006

Recombinant canarypox virus vaccine co-expressing genes encoding the VP2 and VP5 outer capsid proteins of bluetongue virus induces high level protection in sheep.

We describe the development and preliminary characterization of a recombinant canarypox virus vectored vaccine for protective immunization of ruminants against bluetongue virus (BTV) infection. Sheep (n=6) immunized with recombinant canarypox virus vector (BTV-CP) co-expressing synthetic genes encoding the two outer capsid proteins (VP2 and VP5) of BTV serotype 17 (BTV-17) developed high titers (40-160) of virus-specific neutralizing antibodies and were resistant to challenge with a field strain of BTV-17. In contrast, sheep (n=5) immunized with a commercial recombinant canarypox virus vector expressing the E and preM genes of West Nile virus were seronegative to BTV and developed pyrexia, lymphopenia, and extended, high-titered viremias following challenge exposure to the field strain of BTV-17. These data confirm that the BTV-CP vaccine may be useful for the protective immunization of ruminants against bluetongue, and it may avoid the problems inherent to live-attenuated (LA) BTV vaccines.

Animals↗
PubMed1997

Protection of dogs against canine distemper by vaccination with a canarypox virus recombinant expressing canine distemper virus fusion and hemagglutinin glycoproteins.

OBJECTIVES: To evaluate the safety and efficacy of a live canarypox virus recombinant-canine distemper virus (CDV) combination vaccine against virulent CDV challenge exposure, and to document lack of interference among the other modified-live virus (MLV) components. ANIMALS: 33 specific-pathogen-free (SPF) Beagle pups (7 to 10 weeks old). PROCEDURE: A canarypox virus recombinant-CDV combination vaccine was tested for safety and efficacy along with MLV components (canine adenovirus type 2, canine coronavirus, canine parainfluenza virus, and canine parvovirus) in 26 SPF Beagle pups. The combination vaccine was rehydrated with either Leptospira canicola-L icterohaemorrhagiae combination bacterin (vaccine 1) or sterile diluent (vaccine 2). An additional group of 7 seronegative SPF pups received the control MLV components devoid of the combination vaccine (vaccine 3). Two vaccinations were administered 21 days apart, either IM or SC. The dose of the combination vaccine used to inoculate these pups was 40 times lower than the recommended commercial dose. At 21 days after the booster vaccination, all pups were challenge exposed with a virulent CDV strain, then were observed for 21 days to record morbidity and mortality. RESULTS: Adverse local or generalized reactions were not induced by vaccinations. All vaccinates seroconverted to CDV. Serum antibody titers to MLV components were not different, with or without inclusion of the combination vaccine. After challenge exposure, morbidity and mortality in vaccinates were 0% (0/26); in control dogs, values were 100% morbidity and 86% mortality (6/7). Brain impression smear slides made from all dogs that did not survive challenge exposure were CDV positive by use of a direct fluorescein isothiocyanate method. CONCLUSIONS: The canarypox virus-CDV combination vaccine, administered SC or IM, is a safe product that elicits CDV seroconversion, does not interfere with other vaccine components, and protects vaccinated pups against virulent CDV challenge exposure.

Animals↗
PubMed1998

Hepatitis C virus-specific CTL responses in PBMC from chimpanzees with chronic hepatitis C: determination of CTL and CTL precursor frequencies using a recombinant canarypox virus (ALVAC).

The aim of this study was to evaluate HCV-cytotoxic T lymphocyte response from PBMC in bulk CTL assays and in CTL precursor analyses using in vitro stimulation with canarypox virus (ALVAC) expressing HCV-capsid/E1/E2/NS2/NS3 antigens. Canarypox virus is naturally host-range restricted and does not replicate or cause cytopathology on mammalian cells. PBMC were obtained from four chimpanzees with chronic hepatitis C infection and one uninfected chimpanzee. CTL from bulk culture of PBMC and CTL precursor frequencies were found in three of the four chronically infected chimpanzees using ALVAC in vitro stimulation. No CTL response was detected in PBMC from the uninfected chimpanzee. The precursor frequencies of CTL specific for capsid, NS2 and NS3 proteins ranged between 1/2663 and 1/27202. No correlation was observed between percent cytolysis in bulk culture and CTL precursor frequencies. This method may prove useful in assessing the correlation between HCV-CTL response and virological or histological status.

Animals↗
PubMed2001

Cytokine responses to human immunodeficiency virus type 1 (HIV-1) induced by immunization with live recombinant canarypox virus vaccine expressing HIV-1 genes boosted by HIV-1(SF-2) recombinant GP120.

Vaccine-induced T-cell memory for human immunodeficiency virus type 1 (HIV-1) was assessed by measuring HIV-1 antigen-stimulated cytokine secretion in 72 HIV-1-uninfected subjects, of whom 52 received live recombinant canarypox virus vaccine expressing HIV-1 env, gag, and protease gene products (vCP205) with or without HIV-1(SF-2) recombinant gp120 (SF-2 rgp120) subunit vaccine, and 20 the control. The vCP205 vaccine induced secretion of the Th1 cytokine, interferon-gamma, by peripheral blood mononuclear cells (PBMC) after in vitro stimulation with HIV-1 p24 and envelope glycoprotein. Immunization schedules with both vCP205 and SF-2 rgp120 subunit vaccines induced secretion of Th1 and Th2 cytokines by PBMC to HIV-1 envelope glycoprotein. Hence, vCP205 and SF-2 rgp120 subunit vaccines given together and in a prime-boost sequence appeared to induce a broader cytokine response pattern than vCP205 vaccine given alone.

AIDS Vaccines↗
PubMed1992

Immunisation with canarypox virus expressing rabies glycoprotein.

Poxviruses have many useful features as vectors for genes that carry immunising antigens from other viruses, such as ease of production and induction of cellular and humoral immunity, but there is concern about the safety of vaccinia virus. We turned to an avian poxvirus (canarypox); this virus undergoes abortive replication in mammalian cells that enables presentation of early gene products to the immune system. Canarypox virus was used as a vector for the rabies glycoprotein G gene. The safety and efficacy of the recombinant (ALVAC-RG; vCP65) were tested in several animal species, then it was subjected to a phase 1 clinical trial. Twenty-five volunteers were randomly assigned to subcutaneous injections of the recombinant (three groups [A, B, and C] received two doses each of 10(3.5), 10(4.5), and 10(5.5) tissue-culture infectious doses50, respectively) or of human diploid cell culture vaccine (HDC; 6.52 international potency units per dose). 28 days after the second dose, all nine ALVAC-RG group-C subjects and two of three group-B subjects had rabies neutralising antibody concentrations of at least 0.5 IU/ml, the level associated with protection in animals. Although the geometric mean titre of these antibodies at that time was lower in group C than in the ten HDC recipients (4.4 [range 0.9-12.5] vs 11.5 [4.7-25.3] IU/ml), a single booster dose at 6 months induced a recall response in volunteers primed with either vaccine. Side-effects associated with ALVAC-RG were mild and of short duration and occurred at similar frequency to those of HDC vaccine. This study has shown the potential of non-replicating poxviruses as vectors for vaccination in human beings. Trials of canarypox-virus recombinants at higher doses and by other routes of administration are needed.

Adult↗
PubMed2004

The genome of canarypox virus.

Here we present the genomic sequence, with analysis, of a canarypox virus (CNPV). The 365-kbp CNPV genome contains 328 potential genes in a central region and in 6.5-kbp inverted terminal repeats. Comparison with the previously characterized fowlpox virus (FWPV) genome revealed avipoxvirus-specific genomic features, including large genomic rearrangements relative to other chordopoxviruses and novel cellular homologues and gene families. CNPV also contains many genomic differences with FWPV, including over 75 kbp of additional sequence, 39 genes lacking FWPV homologues, and an average of 47% amino acid divergence between homologues. Differences occur primarily in terminal and, notably, localized internal genomic regions and suggest significant genomic diversity among avipoxviruses. Divergent regions contain gene families, which overall comprise over 49% of the CNPV genome and include genes encoding 51 proteins containing ankyrin repeats, 26 N1R/p28-like proteins, and potential immunomodulatory proteins, including those similar to transforming growth factor beta and beta-nerve growth factor. CNPV genes lacking homologues in FWPV encode proteins similar to ubiquitin, interleukin-10-like proteins, tumor necrosis factor receptor, PIR1 RNA phosphatase, thioredoxin binding protein, MyD116 domain proteins, circovirus Rep proteins, and the nucleotide metabolism proteins thymidylate kinase and ribonucleotide reductase small subunit. These data reveal genomic differences likely affecting differences in avipoxvirus virulence and host range, and they will likely be useful for the design of improved vaccine vectors.

Animals↗
PubMed2004

Comparison of systemic and mucosal delivery of 2 canarypox virus vaccines expressing either HIV-1 genes or the gene for rabies virus G protein.

BACKGROUND: Since the primary routes of human immunodeficiency type 1 (HIV-1) infection are across mucosal barriers, a randomized trial of canarypox virus-based vectors was conducted in 84 individuals, with delivery of vaccine by mucosal routes, and was accompanied by a detailed analysis of humoral, cellular, and mucosal immune responses. METHODS: Over the course of 6 months, HIV-1-specific (vCP 205) and rabies (vCP 65) canarypox virus vectors were delivered systemically and/or mucosally into the nose, mouth, vagina, or rectum in a 4-dose schedule, followed by 2 doses of HIV-1 MN recombinant glycoprotein (rgp) 120 or subunit rabies vaccine administered by the intramuscular route. RESULTS: Administration of vaccine and collection of samples were well tolerated. Serum IgG HIV-1-specific antibodies to rgp120 were rarely seen after either systemic or mucosal delivery of canarypox virus vaccine. In contrast, serum IgG rabies and canarypox antibodies were detected in all individuals after systemic, but rarely after mucosal, delivery of vaccine. Suggestions of mucosal recognition of HIV-1 antigen included a cytotoxic T lymphocyte response in 4 of 8 individuals after administration of vaccine by the intrarectal route and a limited immunoglobulin A response at the same site. CONCLUSIONS: Each of the routes of vaccine administration was feasible in the context of a phase 1 study with motivated individuals. However, with the doses and routes of administration used, canarypox virus was not an effective mucosal immunogen.

AIDS Vaccines↗
PubMed1992

Nonreplicating viral vectors as potential vaccines: recombinant canarypox virus expressing measles virus fusion (F) and hemagglutinin (HA) glycoproteins.

The development of canarypox virus (CPV) recombinants expressing the hemagglutinin (HA) and fusion (F) glycoproteins of measles virus (MV) is described. Inoculation of the CPV-MV recombinants into avian or nonavian tissue culture substrates led to the expression of authentic MVF and MVHA as determined by radioimmunoprecipitation and surface immunofluorescence. In contrast to avian-derived tissue culture, no productive replication of the CPV recombinant was evident in tissue culture cells derived from nonavian origin. On inoculation of dogs, a species restricted for avipoxvirus replication, the recombinants elicited a protective immune response against a lethal canine distemper virus (CDV) challenge. The level of MV neutralizing antibodies and the level of protection induced against CDV challenge achieved by the host-restricted CPV vector were equivalent to that obtained by vaccinia virus vectors expressing the same MV antigens.

Animals↗
PubMed1997

Effect of canarypox virus (ALVAC)-mediated cytokine expression on murine prostate tumor growth.

BACKGROUND: Canarypox virus, ALVAC, does not replicate in infected mammalian cells and has potential as a vector for gene therapy in the treatment of cancer. PURPOSE: Recombinant viruses carrying DNA sequences encoding interleukin 2 (ALVAC-IL-2), interferon gamma (ALVAC-IFN gamma), tumor necrosis factor-alpha (ALVAC-TNF-alpha), or the co-stimulatory molecule B7-1 (ALVAC-B7-1) were investigated as agents for the treatment of a newly defined mouse prostate tumor model. METHODS: RM-1 mouse prostate cancer cells, which are syngeneic (i.e., same genetic background) to C57BL/6 mice, were used. The expression of foreign gene products in vitro in infected RM-1 cells was measured by immunoprecipitation, bioassay, or flow cytometry. The effects of foreign gene product expression on RM-1 tumor cell growth in C57BL/6 mice were measured after subcutaneous injection (in the back) of 5 x 10(5) uninfected or infected cells; measurements included determinations of time to a measurable tumor size, tumor size as a function of time, and survival. The induction of protective immunity by uninfected and infected RM-1 cells was tested by injection of lethally irradiated (70 Gy) cells and subsequent challenge with uninfected cells. The generation of cytotoxic T cells was monitored by use of a 51Cr release assay. Severe combined immunodeficient (SCID) mice were used to determine whether T or B lymphocytes were involved in ALVAC vector-mediated antitumor responses. Data were analyzed by use of Pearson's modification of the chi-squared test and Kaplan-Meier survival methods. Reported P values are two-sided. RESULTS: The level of foreign gene product expression in ALVAC-infected RM-1 cells was dependent on the multiplicity of virus infection used; a multiplicity of five viruses per infected cell was chosen for subsequent experiments. RM-1 tumor growth in C57BL/6 mice was not affected by tumor cell expression of IL-2 alone, IFN gamma alone, or B7-1 alone; however, expression of TNF-alpha alone significantly delayed tumor growth at early time points (compared with parental ALVAC-infected tumors, P = .0001 at day 21 and P = .037 at day 28). Tumor cell expression of both TNF-alpha and IL-2 completely inhibited tumor growth in 60%-100% of treated mice. No protection against subsequent tumor challenge was detected in mice previously exposed to RM-1 cells expressing both TNF-alpha and IL-2. Cytotoxic T-lymphocyte activity toward RM-1 cells was not observed in C57BL/6 mice that rejected tumors. Tumor cell expression of TNF-alpha and IL-2 also resulted in tumor growth inhibition in SCID mice. CONCLUSIONS: RM-1 mouse prostate cancer cells are readily infected by ALVAC vectors, and foreign gene products are efficiently expressed. Inhibition of RM-1 tumor growth by tumor cell expression of TNF-alpha and IL-2 appears to involve nonspecific antitumor activity.

Animals↗
PubMed1999

Identification of the canarypox virus thymidine kinase gene and insertion of foreign genes.

We mapped the canarypox virus (CaPV) thymidine kinase (TK) gene within a 5.8-kbp XbaI fragment of the genome by Southern blotting using the fowlpox virus (FPV) TK gene as a probe. Nucleotide sequence analysis of the fragment revealed seven open reading frames (ORFs) showing gene organization similar to that of FPV. The TK gene contained in this region had an ORF of 179 amino acids encoding a polypeptide with a putative molecular mass of 20.0 kDa. An A/T-rich region and a transcription termination signal, TTTTTAT, were found upstream and at the end of the ORF, which is consistent with poxvirus early gene regulation. The consensus sequence of the late promoter TAAAT also overlapped with the initiation codon of the ORF. The amino acid sequence similarity between the TK genes of CaPV and FPV, avipoxviruses, was 64.2%, which was lower than the similarities between vaccinia and variola orthopoxviruses (97.2%) and between Shope fibroma and myxoma leporipoxviruses (82.6%). However, the monophyly of avian clades of CaPV and FPV was supported by phylogenetic analysis. We then inserted the genes encoding lacZ, luciferase (luci), and envelope of human T-lymphotropic virus type 1 (HTLV-1 env) into the TK gene of CaPV to evaluate its suitability as an expression vector. The recombinant viruses obtained were unstable, although the foreign genes were expressed efficiently in the mammalian cells infected with the viruses.

Amino Acid Sequence↗
PubMed1997

A recombinant canarypox virus protects rabbits against a lethal rabbit hemorrhagic disease virus (RHDV) challenge.

An ALVAC (canarypox)-based recombinant virus ALVAC-RHDV (vCP309) expressing a native rabbit hemorrhagic disease virus (RHDV) capsid protein was derived and assessed for its protective efficacy in rabbits. Protection against a lethal RHDV challenge was demonstrated in rabbits inoculated twice with either high (10(7) p.f.u.) or low (10(5) p.f.u.) doses of vCP309. However, animals in the high dose group developed significantly higher antibody response. These results have implications that are relevant to the development of a safe rabbit hemorrhagic disease (RHD) vaccine and further illustrate the utility of the ALVAC vector system to elicit protective immune responses in nonavian species.

Animals↗
PubMed2003

Safety of intravenous administration of a canarypox virus encoding the human wild-type p53 gene in colorectal cancer patients.

Overexpression of p53 occurs in more than 50% of colorectal cancers. Therefore, p53 represents an attractive target antigen for immunotherapy. We assessed the safety of a canarypox virus encoding the human wild-type p53 gene given intravenously to end-stage colorectal cancer patients in a three-step dose escalation study aimed at inducing p53 immune responses. Patients with metastatic disease of p53-overexpressing colorectal cancers were vaccinated three times at 3-week intervals, each time with 10(6.5) CCID(50) (CCID(50)=cell culture infectious dose 50%; group 1, n=5), 10(7.0) CCID(50) (group 2, n=5) or 10(7.5) CCID(50) (group 3, n=6). Vital signs and the occurrence of adverse events were monitored and blood was analyzed for biochemical and hematological parameters as well as signs of auto-immune safety. In all, 16 patients were enrolled and 15 patients completed three vaccinations. No anaphylactic reaction or unwanted auto-immune reactions were observed. A total of 16 serious adverse events (SAEs) occurred: 10 in group 1, three in group 2 and three in group 3. All SAEs were tumor-related complications. There was no difference in the frequency of adverse events between the three groups, except for fever. Fever was the only vaccination-related adverse event consistently observed and was most frequent and outspoken in the group 3 patients. The majority was a grade 1 or 2 fever (93%) and grade 3 fever (7%) was observed in three patients of group 3. Some patients showed humoral and cellular responses against p53, following vaccinations. After having completed his initial treatment cycle, one patient (group 2) received a second treatment cycle of three doses of 10(7.5) CCID(50) and subsequently showed stable disease. All other patients showed progressive disease. We conclude that ALVAC-p53 can be administered intravenously to colorectal cancer patients without serious toxicity or pathological autoimmunity and can induce immune responses against p53.

Adult↗
PubMed2005

Phase I study of the intratumoral administration of recombinant canarypox viruses expressing B7.1 and interleukin 12 in patients with metastatic melanoma.

The objective of this study was to evaluate the safety and activity of the intratumoral administration of the immune costimulatory molecule, B7.1, encoded by a vector derived from the canarypox virus, ALVAC (ALVAC-B7.1), alone and with the intratumoral injection of ALVAC encoding the immune-stimulatory cytokine, interleukin 12 (ALVAC-IL-12). Fourteen patients with metastatic melanoma who had s.c. nodules received intratumoral injections on days 1, 4, 8, and 11. Nine patients were given escalating doses of up to 25 x 10(8) plaque-forming units of ALVAC-B7.1. Five patients were given 25 x 10(8) plaque-forming units of ALVAC-B7.1 combined with ALVAC-IL-12 50% tissue culture infective dose of 2 x 10(6). Toxicity was mild to moderate and consisted of inflammatory reactions at the injection site and fever, chills, myalgia, and fatigue. Higher levels of B7.1 mRNA were observed in ALVAC-B7.1-injected tumors compared with saline-injected control tumors. Higher levels of intratumoral vascular endothelial growth factor and IL-10, cytokines with immune suppressive activities, were also observed in ALVAC-B7.1- and ALVAC-IL-12-injected tumors compared with saline-injected controls. Serum levels of vascular endothelial growth factor increased at day 18 and returned to baseline at day 43. All patients developed antibody to ALVAC. Intratumoral IL-12 and IFN-gamma mRNA decreased. Changes in serum IL-12 and IFN-gamma levels were not observed. Tumor regressions were not observed. The intratumoral injections of ALVAC-B7.1 and ALVAC-IL-12 were well tolerated at these dose levels and at this schedule and resulted in measurable biological response. This response included the production of factors that may suppress the antitumor immunologic activity of these vectors.

Adult↗
PubMed2001

Canarypox virus expressing wild type p53 for gene therapy in murine tumors mutated in p53.

The antitumor activity of a recombinant canarypox virus expressing wild type murine p53 (ALVAC-p53) was investigated in two murine syngeneic tumors harboring an endogenous p53 mutation (CMS4 and TS/A). Direct intratumor injections of ALVAC-p53 in CMS4 pre-established subcutaneous tumors induced total tumor regression in 66% of mice. Furthermore, 100% of the cured mice was protected against a contralateral subsequent challenge with the parental tumor cells. The intravenous treatment of experimental lung metastasis by ALVAC-p53 also induced significant tumor growth inhibition in both models. The antitumor effect of ALVAC-p53 was only observed in immunocompetent animals and was associated with the generation of a specific antitumor immune response. ALVAC-p53 induced the expression of a functional p53 wild type protein as demonstrated by up-regulation of p21waf1 and induction of apoptosis. A vaccine strategy using intravenous or subcutaneous ALVAC-p53/NYVAC-p53 prime boost protocol failed to induce CTL against p53 wild type used as target tumor antigen, and failed to protect mice against challenge with the mutated tumor cells. The mechanism of the curative and protective effects observed after direct intratumor injections results from the induction of a specific antitumor response directed against other antigens than p53. Our results suggest that the local induction of tumor apoptosis, combined with the adjuvant effect of ALVAC vector, enhances the immunogenicity of the intratumor environment and allows induction of specific antitumor immune response.

Animals↗
PubMed2001

Immunization with type 5 adenovirus recombinant for a tumor antigen in combination with recombinant canarypox virus (ALVAC) cytokine gene delivery induces destruction of established prostate tumors.

Prostate-specific antigen (PSA) is expressed by prostate epithelial cells and has a highly restricted tissue distribution. Prostatic malignancies in 95% of patients continue to express PSA, making this antigen a good candidate for targeted immunotherapy. The goals of our studies are to generate a recombinant PSA adenovirus type 5 (Ad5-PSA) that is safe and effectively activates a PSA-specific T-cell response capable of eliminating prostate cancer cells, and to characterize the immunologic basis for this rejection. Here we show that immunization of mice with Ad5-PSA induced PSA-specific cellular and humoral immunity that was protective against a subcutaneous challenge with RM11 prostate cancer cells expressing PSA (RM11psa), but not mock-transfected RM11 tumor cells (RM11neo). Mice immunized with recombinant adenovirus type 5 encoding beta-galactosidase (Ad5-lacZ) did not generate protective immunity. Antitumor activity was predominantly mediated by CD8(+) T lymphocytes. Although Ad5-PSA immunization prior to RM11psa challenge was protective, Ad5-PSA immunization alone was not able to control the growth of existing RM11psa tumors. In contrast, established RM11psa tumors ranging in size from 500 to 1,000 mm(3) were efficiently eliminated if Ad5-PSA priming was followed 7 days later by intratumoral injection of recombinant canarypox viruses (ALVAC) encoding interleukin-12 (IL-12), IL-2, and tumor necrosis factor-alpha. In this case, antitumor immunity was still dominated by CD8(+) T lymphocytes, but natural killer cells became necessary for a maximal response. These data provide information on the effector cell populations in a protective immune response to prostate cancer and demonstrate the utility of an Ad5-PSA vaccine combined with cytokine gene delivery to eliminate large established tumors that are refractory to other interventional methods.

Adenoviridae↗