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E Paoletti

Publications and source records attributed to E Paoletti.

At least 37 records · Page 2Linked to original sources
PubMed1997

Replication-defective canarypox (ALVAC) vectors effectively activate anti-human immunodeficiency virus-1 cytotoxic T lymphocytes present in infected patients: implications for antigen-specific immunotherapy.

In the attempt to develop immunotherapeutic strategies for acquired immunodeficiency syndrome capable of activating effector cells in an antigen-specific manner while maintaining the broadest possible T-cell repertoire, we evaluated two canarypox (ALVAC)-based vectors for their capacity to induce ex vivo activation/expansion of human immunodeficiency virus (HIV)-specific CD8+ cytotoxic lymphocyte precursors (CTLp) obtained from HIV-1-infected donors. These two vectors, vCP205 encoding HIV-1 gp120 + TM (28 amino acid transmembrane anchor sequence) in addition to Gag/protease and vCP300 encoding gp120 + Gag/protease as well as Nef and Pol CTL determinants, are pancytotropic but replication incompetent in mammalian cells. Bulk peripheral blood mononuclear cells (PBMCs) or enriched CD8+ T cells were stimulated for 10 days with autologous ALVAC-infected PBMCs in the presence of different cytokine combinations (interleukin-2 [IL-2], IL-4, IL-7, and IL-12). Activation by ALVAC constructs was highly antigen-specific, because vCP205 elicited only Env and Gag CTL, whereas vCP300 elicited broader reactivities against Env, Gag, Pol, and Nef determinants. The ALVAC activation of CTLp was IL-2 dependent and enhanced by the addition of IL-7, whereas IL-4 and IL-12 failed to augment cytotoxic reactivities elicited by these constructs. The expansion of enriched CD8+ T cells after activation with vCP300 was higher in patients with CD4 counts greater than 400 cells/microL. Two rounds of in vitro stimulation (IVS) with vCP300 resulted in nearly an eightfold expansion of CD8+ lymphocytes over a 25-day period. After the second IVS, an average 3.2-fold increase among the different antigen-specific CTL frequencies was achieved. These studies clearly show that HIV-recombinant ALVAC vectors represent powerful polyvalent antigenic stimuli for activation and expansion of the CD8 lymphocyte response that occurs as a result of HIV infection.

Adult↗
PubMed1997

Recognition of dengue virus NS1-NS2a proteins by human CD4+ cytotoxic T lymphocyte clones.

We examined nine dengue virus-specific human CD4+ CD8- cytotoxic T lymphocyte (CTL) clones for protein recognition, using recombinant vaccinia viruses which contain genes coding for dengue virus proteins. These clones were established from peripheral blood mononuclear cells of a donor previously immunized with a live-attenuated experimental dengue-1 vaccine. Of nine CD4+ T cell clones, seven were dengue-1-specific and two were dengue-1-dengue-3 cross-reactive. Four dengue-1-specific clones and one dengue-1-dengue-3 cross-reactive clone recognized epitopes within the NS1 or NS2a proteins. Analysis of human leukocyte antigen (HLA) restriction revealed that three dengue-1-specific clones are HLA-DR1-restricted and one dengue-1-dengue-3 cross-reactive clone is HLA-DPw3-restricted. These results indicate that NS1 and NS2a proteins as well as C, E, and NS3 proteins reported earlier contain one or more epitopes recognized by dengue virus-specific human CD4+ T lymphocytes.

Antigens, Viral↗
PubMed1997

Challenge of chimpanzees immunized with a recombinant canarypox-HIV-1 virus.

To evaluate the potential protective efficacy of a live recombinant human immunodeficiency virus type 1 (HIV-1) canarypox vaccine candidate, two chimpanzees were immunized five times with ALVAC-HIV-1 vCP250, a recombinant canarypox virus that expresses the HIV-1[IIIB(LAI)] gp120/TM, gag, and protease gene products. One month after the last booster inoculation, the animals were challenged by intravenous injection of cell-associated virus in the form of peripheral blood mononuclear cells from an HIV-1[IIIB(LAI)]-infected chimpanzee. One chimpanzee with a neutralizing antibody titer to HIV-1[IIIB(LAI)] of 128 at the time of challenge was protected, whereas both the second animal, with a neutralizing antibody titer of 32, and a naive control animal became infected. At 5 months after challenge, the protected chimpanzee and a third animal, previously immunized with various HIV-1[MN] antigens, were given a booster inoculation. The two animals were challenged intravenously 5 weeks later with twenty 50% tissue culture infectious doses of cell-free HIV-1[DH12], a heterologous subtype B isolate. Neither chimpanzee had neutralizing antibodies to HIV-1[DH12], and neither one was protected from infection with this isolate. The immune responses elicited by vaccination against HIV-1[IIIB(LAI)] or HIV-1[MN] did not, therefore, protect the animals from challenge with the heterologous cell-free HIV-1[DH12].

AIDS Vaccines↗
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↗
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↗
PubMed1997

Prolonged therapy with ACE inhibitors induces a regression of left ventricular hypertrophy of dialyzed uremic patients independently from hypotensive effects.

Left ventricular hypertrophy (LVH), which frequently occurs in chronic uremia, may be due in part to factors other than arterial hypertension, chronic anemia, and/or other well-known loading conditions inherent to the uremic state. Angiotensin-converting enzyme (ACE) inhibitors may be able to reverse LVH by mechanisms independent of their antihypertensive effects. In this study, 18 subjects free of arterial hypertension or severe anemia were selected from 170 chronically hemodialyzed uremic patients after fulfilling the criterion of a supranormal left ventricular mass (LVM). Ten subjects agreed to undergo treatment with 2.5 to 20 mg lisinopril every other day over a period of 2 years, during which annual determinations of the LVM by echocardiography and of the 24-hour blood pressure with a portable device were carried out. Eight patients unwilling to undergo the treatment served as controls. The average resting left ventricular mass index (LVMi) of the overall group was 178 +/- 30 g/m2 body surface area (+/- SD), and did not differ between the two subgroups. Lisinopril treatment significantly decreased the LVM of eight of 10 treated subjects and actually even completely normalized it in three. The LVM of the untreated group remained unchanged. Systolic and diastolic blood pressures were 138 +/- 5 mm Hg and 78 +/- 6 mm Hg in the treated group and 133 +/- 9 mm Hg and 75 +/- 4 mm Hg in the untreated group, respectively (P = NS), and did not vary over the following 2 years. This study indicates that a mild degree of LVH, which is seemingly independent of arterial blood pressure load, does exist in a tight subset of uremic patients. This study also demonstrates that this type of LVH is apparently nonprogressive. ACE inhibitors given at doses not affecting blood pressure are able to reverse it.

Aged↗
PubMed1997

Canine distemper virus (CDV) infection of ferrets as a model for testing Morbillivirus vaccine strategies: NYVAC- and ALVAC-based CDV recombinants protect against symptomatic infection.

Canine distemper virus (CDV) infection of ferrets causes an acute systemic disease involving multiple organ systems, including the respiratory tract, lymphoid system, and central nervous system (CNS). We have tested candidate CDV vaccines incorporating the fusion (F) and hemagglutinin (HA) proteins in the highly attenuated NYVAC strain of vaccinia virus and in the ALVAC strain of canarypox virus, which does not productively replicate in mammalian hosts. Juvenile ferrets were vaccinated twice with these constructs, or with an attenuated live-virus vaccine, while controls received saline or the NYVAC and ALVAC vectors expressing rabies virus glycoprotein. Control animals did not develop neutralizing antibody and succumbed to distemper after developing fever, weight loss, leukocytopenia, decreased activity, conjunctivitis, an erythematous rash typical of distemper, CNS signs, and viremia in peripheral blood mononuclear cells (as measured by reverse transcription-PCR). All three CDV vaccines elicited neutralizing titers of at least 1:96. All vaccinated ferrets survived, and none developed viremia. Both recombinant vaccines also protected against the development of symptomatic distemper. However, ferrets receiving the live-virus vaccine lost weight, became lymphocytopenic, and developed the erythematous rash typical of CDV. These data show that ferrets are an excellent model for evaluating the ability of CDV vaccines to protect against symptomatic infection. Because the pathogenesis and clinical course of CDV infection of ferrets is quite similar to that of other Morbillivirus infections, including measles, this model will be useful in testing new candidate Morbillivirus vaccines.

Animals↗
PubMed1996

Applications of pox virus vectors to vaccination: an update.

Recombinant pox viruses have been generated for vaccination against heterologous pathogens. Amongst these, the following are notable examples. (i) The engineering of the Copenhagen strain of vaccinia virus to express the rabies virus glycoprotein. When applied in baits, this recombinant has been shown to vaccinate the red fox in Europe and raccoons in the United States, stemming the spread of rabies virus infection in the wild. (ii) A fowlpox-based recombinant expressing the Newcastle disease virus fusion and hemagglutinin glycoproteins has been shown to protect commercial broiler chickens for their lifetime when the vaccine was administered at 1 day of age, even in the presence of maternal immunity against either the Newcastle disease virus or the pox vector. (iii) Recombinants of canarypox virus, which is restricted for replication to avian species, have provided protection against rabies virus challenge in cats and dogs, against canine distemper virus, feline leukemia virus, and equine influenza virus disease. In humans, canarypox virus-based recombinants expressing antigens from rabies virus, Japanese encephalitis virus, and HIV have been shown to be safe and immunogenic. (iv) A highly attenuated vaccinia derivative, NYVAC, has been engineered to express antigens from both animal and human pathogens. Safety and immunogenicity of NYVAC-based recombinants expressing the rabies virus glycoprotein, a polyprotein from Japanese encephalitis virus, and seven antigens from Plasmodium falciparum have been demonstrated to be safe and immunogenic in early human vaccine studies.

AIDS Vaccines↗
PubMed1996

Multiple immunizations with attenuated poxvirus HIV type 2 recombinants and subunit boosts required for protection of rhesus macaques.

Vaccine protocols involving multiple immunizations with molecularly attenuated vaccinia virus (NYVAC) or naturally attenuated canarypox virus (ALVAC) HIV-2 recombinants and subunit boosts have conferred longlasting protection against HIV-2 infection of macaques. Similar complex protocols using HIV-1 NYVAC and ALVAC recombinants and subunit boosts have provided cross-protection against HIV-2 challenge. Here a simplified three-immunization regimen over 24 weeks was tested in 18 juvenile rhesus macaques. Twelve macaques were immunized twice with NYVAC or ALVAC recombinants carrying HIV-2 env, gag, and pol genes. Subsequently, macaques in groups of three received either an additional recombinant immunization or an HIV-2 gp160 boost. Six control macaques received three immunizations of NYVAC or ALVAC vector alone and additionally alum at the third immunization. Macaques primed with ALVAC recombinant exhibited sporadic T cell proliferative activity, and all but one failed to develop neutralizing antibodies. In contrast, macaques primed with NYVAC recombinants had no T cell proliferative activity but exhibited neutralizing antibody titers (highest in the three recombinant group) that declined by the time of challenge. None of the macaques exhibited significant cytotoxic T lymphocyte activity. Following challenge at 32 weeks with HIV-2SBL6669 all macaques became infected. Thus, the three-immunization regimen is not sufficient to confer protective immunity in the HIV-2 rhesus macaque model. However, delayed infection in macaques immunized with the NYVAC-HIV-2 recombinant may have been associated with the development of memory B cells capable of providing a neutralizing antibody response on challenge.

AIDS Vaccines↗
PubMed1996

p53 as a target for cancer vaccines: recombinant canarypox virus vectors expressing p53 protect mice against lethal tumor cell challenge.

The p53 protein is an attractive target for immunotherapy, because mutations in the p53 gene are the most common genetic alterations found in human tumors. These mutations result in high levels of p53 protein in the tumor cell, whereas the expression level of wild-type p53 in nonmalignant tissue is usually much lower. Several canarypox virus recombinants expressing human or murine p53 in wild-type or mutant form were constructed. Immunization with these viruses protected BALB/c mice from a challenge with an isogenic and highly tumorigenic mouse fibroblast tumor cell line expressing high levels of mutant p53. The tumor protection was equally effective regardless of whether wild-type or mutant p53 was used for the immunization, indicating that the immunologic response was not dependent on any particular p53 mutation and that immunization with this live virus vaccine works effectively against mutant p53 protein expressed in a tumor cell. In tumors escaping immunologic rejection, the expression of the p53 protein was commonly down-regulated.

Amino Acid Sequence↗
PubMed1996

MAGE-1-specific precursor cytotoxic T-lymphocytes present among tumor-infiltrating lymphocytes from a patient with breast cancer: characterization and antigen-specific activation.

A potential target for development of tumor-specific immunotherapeutic strategies is the MAGE-1 gene. We have utilized a recently developed recombinant canarypox (ALVAC) virus vector containing the MAGE-1 gene (vCP235) to activate CTLs from a breast cancer patient bearing a MAGE-1+ tumor. Tumor-infiltrating lymphocytes (TILs) obtained from the tumor of a patient were stimulated in vitro with irradiated autologous peripheral blood mononuclear cells acutely infected with the vCP235 construct. These TILs preferentially expanded approximately 6-fold over a 16-day culture period and specifically recognized an allogeneic transformed B-cell line acutely infected with a vaccinia-MAGE-1 recombinant targeting vector (vP1188) in the context of HLA-A2 and/or B7. TCR V beta analysis of in vitro expanded T cells by a quantitative multiprobe RNase protection assay revealed preferential expansion of TCR V beta 6.3 and V beta 6.4. In addition, homologous T-cell receptor beta CDR3 joining sequences were found in the in vitro stimulated cultures. These results suggest that tumor antigen-specific, MHC-restricted CTLs may be derived from precursor CTLs present in TILs obtained from patients with MAGE-1+ tumors by in vitro stimulation with recombinant avipox MAGE-1 virus-infected autologous cells. Collectively, these findings provide a rationale for tumor-associated antigen-based immunization as a means of activating precursor CTLs residing in patients with tumors expressing defined tumor-associated antigens such as MAGE-1.

Amino Acid Sequence↗
PubMed1996

Comparison of numerous delivery systems for the induction of cytotoxic T lymphocytes by immunization.

A variety of vaccine delivery systems including peptides with various adjuvants, recombinant particles, live recombinant viruses and bacteria and plasmid DNA were tested for their ability to induce CD8+ cytotoxic T lymphocytes (CTL) against a well-defined epitope (amino acids 252-260) from the circumsporozoite (CS) protein of Plasmodium berghei. We compared routes of immunization that would be applicable for the administration of a malaria vaccine in humans. The majority of these vaccines did not induce high CTL responses in the spleens of immunized mice. However, both a yeast-derived Ty virus-like particle expressing the optimal nine-amino acid epitope SYIPSAEKI from the CS protein (CSP-VLP) and a lipid-tailed peptide of this same sequence induced high levels of the major histocompatibility complex (MHC) class I-restricted CTL with one and three subcutaneous immunizations, respectively. Moreover, these CTL were able to recognize naturally processed antigen expressed by a recombinant vaccinia virus. The levels of CTL induced by CSP-VLP could be augmented by co-immunization with certain cytokines. Target cells pulsed with CSP-VLP were recognized and lysed, showing that the particles were effectively processed and presented through MHC class I presentation pathway. The levels of CTL induced using CSP-VLP and lipopeptides are comparable to those observed after immunization with multiple doses of irradiated sporozoites.

Amino Acid Sequence↗
PubMed1996

Host-range restriction of vaccinia virus E3L-specific deletion mutants.

The vaccinia virus (VV) E3L gene product functions as a dsRNA binding protein that is involved in conferring an interferon-resistant phenotype upon the virus. Studies with a vaccinia virus (VV) E3L- deletion mutant (vP1080) have also demonstrated that the E3L gene product is critical for productive replication on certain cell substrates. While E3L was found to be nonessential for replication in chick embryo fibroblasts (CEFs), virus specifically deleted of E3L was found to be replication deficient in Vero, HeLa, and murine L929 cells. Further, the temporal block in replication appears to differ in these cell systems, as evidenced by the observed timing of protein synthesis inhibition. In Vero cells infected with the VV E3L- mutant, there was no detectable protein synthesis after 2 hr post-infection, whereas in L929 cells normal protein patterns were observed even at late times post-infection. Expression of a heterologous dsRNA binding protein, the reovirus sigma 3 protein, by the E3L- mutant virus restored near wild-type growth characteristics, suggesting the critical nature for regulating dsRNA levels in VV-infected cells.

Animals↗
PubMed1996

Human safety and immunogenicity of a canarypox-rabies glycoprotein recombinant vaccine: an alternative poxvirus vector system.

Avian poxvirus recombinants undergo abortive replication in nonavian cells, yet can achieve expression of extrinsic gene products. Canarypox-vectored vaccines have been innocuous and immunogenic in several mammalian species. ALVAC-RG, a canarypox recombinant expressing the rabies glycoprotein gene, was inoculated intramuscularly into adult volunteers on days 0, 28, and 180. Sequential cohorts received 10(3.5), 10(4.5), and 10(5.5) 50% tissue culture infective doses (TCID50); additional volunteers received the standard human diploid cell rabies vaccine (HDCV) on the same schedule. Reactogenicity of ALVAC-RG was minimal. The lowest dose of ALVAC-RG induced little antibody to rabies virus by ELISA or rapid fluorescent focus inhibition test (RFFIT), but 10(4.5) and 10(5.5) TCID50 doses elicited significant responses in both assays. All recipients of 10(4.5) and 10(5.5) TCID50 of ALVAC-RG attained RFFIT values above the presumed protective level. Canarypox-specific immune responses did not inhibit boosting of rabies-specific antibodies by the day 180 dose of ALVAC-RG. T cell proliferation in response to inactivated rabies virus in vitro was similar in HDCV and ALVAC-RG recipients after the first and second doses, although HDCV yielded superior results after the third dose. ALVAC-RG was safe in humans, induced functional antibody to rabies glycoprotein, elicited cellular responses to rabies virus, and could be used successfully for booster dosing at a 6 month interval.

Adolescent↗
PubMed1996

Memory cytotoxic T lymphocyte responses in human immunodeficiency virus type 1 (HIV-1)-negative volunteers immunized with a recombinant canarypox expressing gp 160 of HIV-1 and boosted with a recombinant gp160.

A vaccine against human immunodeficiency virus (HIV) should induce virus-specific cytotoxic T lymphocyte (CTL) activity. Immunization of uninfected volunteers with a canarypox virus expressing HIV envelope was carried out in a phase I trial. Two injections of canarypox expressing HIV-1MN gp 160 (months 0 and 1) were followed by two boosts of recombinant envelope protein (months 3 and 6). HIV envelope-specific CTL were detected in peripheral blood mononuclear cells stimulated with autologous HIV-1-infected blast cells. T cell lines were obtained from 18 of 20 donors: CTL were detected at least once following immunization in 7 (39%) of these 18. This activity was mediated by major histocompatibility complex class I-restricted CD3+CD8+ T cells. For two subjects, this activity was still present 2 years after the initial immunization. The CTL responses with this prime-boost regimen are the best observed with any HIV vaccine tested in humans.

AIDS Vaccines↗
PubMed1996

Immunogenicity and protective efficacy of a human immunodeficiency virus type 2 recombinant canarypox (ALVAC) vaccine candidate in cynomolgus monkeys.

The efficacy of a recombinant human immunodeficiency virus (HIV) type 2 canarypox (ALVAC HIV-2) vaccine candidate given alone or in combination with HIV-2 envelope gp125 or HIV-2 V3 synthetic peptides was investigated in 14 cynomolgus monkeys. High antibody titers to HIV-2 gp125 were demonstrated in monkeys given booster immunizations with gp125. Neutralizing antibody titers were low (< or = 20) in all monkeys except 2. Significant lymphocyte proliferative responses to killed HIV-2 virions were observed in monkeys given booster immunizations with gp125. HIV-2-specific cytotoxic T lymphocytes were demonstrated prior to viral challenge in 3 of 12 monkeys. After challenge with homologous cell-free HIV-2 propagated in monkey cells, 4 of 10 monkeys immunized with ALVAC HIV-2 plus HIV-2 gp125 or V3 peptides were protected, as determined by negative virus isolation and polymerase chain reaction for viral DNA. Four monkeys immunized with ALVAC HIV-2 alone were not protected. All 12 control monkeys became infected. There was no correlation between the immunologic parameters studied and protection against infection in the vaccinated monkeys.

AIDS Vaccines↗