The impact of the COVID‒19 pandemic on organ donation and transplantation in Hungary in 2020 / A COVID-19-járvány hatása a szervadományozásra és -átültetésre Magyarországon 2020-ban.

Összefoglaló. Bevezetés: A SARS-CoV-2-világjárvány terjedése drasztikus változásokat okozott a mindennapi betegellátásban, amelyek érintették a szervadományozás és -átültetés területét is, így csökkent az élo és az elhunyt donorokból történo donációk és transzplantációk száma világszerte. Az esetszám csökkenése mellett a transzplantált és egyben immunszupprimált betegek védelme érdekében további biztonsági intézkedéseket kellett bevezetni. Módszer: A vizsgálat célja a COVID-19-járvány hazai donációs és transzplantációs aktivitásra gyakorolt hatásának kimutatása volt 2020-ban, a megelozo évvel történo összehasonlításban. A magyar eredményeket összehasonlítottuk elsosorban az Eurotransplant, illetve az Európai Unió tagállamainak adataival is. Eredmények: A lakosságszámra súlyozott, regisztrált COVID-19-fertozöttség és -halálozás tekintetében nem igazoltunk 2020-ban kiemelkedo eltérést itthon az Eurotransplant-tagállamokhoz képest. A hazai szervdonációs potenciál nem csökkent a vizsgált idoszakban, ugyanakkor 38,33%-kal csökkent az agyhalott szervdonorok száma Magyarországon, míg az Eurotransplantban átlagosan 8,64%-kal és 23 adatközlo európai országban 17,55%-kal. Az elhunytból történt szervátültetések száma 29,27%-kal csökkent, különösen a szív- és a májátültetések esetén. A külföldrol kapott szervek száma 21,13%-kal és aránya 12,34%-kal emelkedett. Az élo donoros veseátültetések száma nem változott. 2020-ban 25%-kal kevesebb új beteget regisztráltak, mint 2019-ben, és a várólista-mortalitás 28%-kal növekedett az elozo évhez képest, kifejezetten a veseátültetésre várók között. Következtetés: A hazai szervátültetési program biztonságos: donoreredetu SARS-CoV-2-átvitel nem történt hazánkban. A szervdonációs potenciál és a COVID-19-járvány mellett a szervdonációs és -transzplantációs aktivitás jelentosen csökkent Magyarországon 2020. márciustól az év végéig. A legtöbb európai országban átmeneti és kisebb mértéku szervdonációs csökkenést regisztráltak. A szervátültetések száma nem csökkent olyan mértékben, mint a donorszám, mert az Eurotransplantból több donorszerv érkezett hazánkba, mint amennyit külföldre küldtünk. Orv Hetil. 2021; 162(23): 890-896. INTRODUCTION: The spread of the SARS-CoV-2 pandemic has resulted in drastic changes in day-to-day patient care, which has also affected the field of organ donation and transplantation, thus reducing the number of donations and transplants from living and deceased donors worldwide. In addition to the reduction in the number of cases, additional safety measures had to be introduced to protect transplanted and implicatively immunosuppressed patients. METHOD: The aim of the study was to demonstrate the impact of the COVID-19 epidemic on domestic donation and transplantation activity in 2020, compared to the previous year. We also compared the Hungarian results with the data of the Eurotransplant and the European Union member states. RESULTS: In terms of population-weighted, registered COVID-19 infection and mortality, we did not find a significant difference in Hungary in 2020 compared to the Eurotransplant member states. The national organ donation potential did not diminish in the period under review, however, the number of brain-dead organ donors decreased by 38.33% in Hungary, while in the Eurotransplant it did by 8.64% on average and in 23 reporting European countries by 17.55%. The number of organ transplants from the deceased decreased by 29.27%, especially regarding heart and liver transplants. Both the number and the proportion of organs received from abroad increased by 21.13% and 12.34%, respectively. The number of living donor kidney transplants did not change. In 2020, 25% fewer new patients were registered than in 2019 and the mortality on waiting list increased by 28% compared to the previous year, especially among those waiting for a kidney transplant. CONCLUSION: The national organ transplantation program is safe: donor-derived SARS-CoV-2 transmission did not occur in Hungary. In addition to the organ donation potential and the COVID-19 pandemic, organ donation and transplantation activity decreased significantly in Hungary from March 2020 until the end of the year. Transient and smaller reductions in organ donation rates have been reported in most European countries. The number of organ transplants did not decrease as much as the number of donors, because more donor organs arrived in Hungary from the Eurotransplant than we sent abroad. Orv Hetil. 2021; 162(23): 890-896.

National level adjustments to the challenges of the SARS-CoV2 pandemic on blood banking operations.

BACKGROUND: SARS-CoV2 causing coronavirus disease (COVID-19) is responsible for an unprecedented worldwide pandemic severely affecting all activities of societies including blood banking. We aimed to systematically collect key indicators in a nationally centralized blood banking system and to perform comparisons between 2020 and 2019. METHODS: Count data for January-December 2020 and 2019 were extracted from the integrated informatics system of Hungarian National Blood Transfusion Service and analyzed by simple graphics, tabulations, and statistics. RESULTS: Whole blood donation activity showed a highly significant decline due to a sharp decrease in field donations by an average fall of 24% (range:17%-28%) during March-May 2020 compared to identical period of 2019. A second, more moderate decline accompanied the second wave in late fall. The simultaneous increase in institutional donations did not counterbalance this decline. Donor exclusion rates fell significantly by an average of 1,1% (range:0.9%-1.6%) in the three spring lockdown-affected months. First-time and repeat donors showed decreased turn-out in larger proportions compared to highly repeat donors. Interestingly, among repeat and highly repeat donors, females showed less-pronounced declines compared to males while this was not observed among first-time donors. In June-September, a remarkable swing-back was observed among highly repeat female donors. Product utilization fell most notably for RBC (mean:26.2%) but also for PLT (mean:19.8%) and FFP (mean:24.3%) and showed a full recovery in June-September followed by a second decline. CONCLUSION: Trends and reaction patterns of blood banking reported by our study may be useful in future planning and adjustments of blood banking activities.

The First 5 Years of the Newest Eurotransplant Member State: Hungarian Results of International Organ Exchange From 2014 to 2018.

INTRODUCTION: Hungary joined Eurotransplant International (ET) to improve the chance of transplantation for Hungarian patients and patient outcomes, including access and graft and patient survival. After 5 years of full membership, the evaluation of numbers and quality indicators is possible. METHOD: A comparison was made between 5 years prior to a preliminary cooperation agreement (2007-2011) and 5 years after full ET membership (2014-2018). During the 2 study periods, we analyzed numbers and circumstances of deceased organ donors, multiorgan donors, donated organs, and transplantations in Hungary and development of waiting lists along with international organ exchanges. RESULT: The number of actual organ donors increased by 22.09% (729 vs 890), an additional 823 organ removals represents an increase of 42.71% (1927 vs 2750). There were 46.51% more transplants managed in the selected periods (1561 vs 2287). The number of new patients on the waiting list increased (2305 vs 3247; 40.87%). The mean kidney mismatch number decreased from 3.21 to 2.96. CONCLUSION: Joining ET has been an effective and efficient in terms of increasing access to organs and the lives of patients on the Hungarian waiting list posttransplant. It is also a benefit for patients with special needs because the number of organ transplants is greater than the increased number of donors.

[Personnel and material conditions of the Hungarian intensive care units dealing with organ donation]. / Magyarországi intenzív osztályok szervdonációval kapcsolatos személyi és tárgyi feltételei.

INTRODUCTION: At the end of 2016, the number of patients on the domestic transplant waiting list was twice as much as the number of the organ transplantations accomplished that year. The institutional prerequisites for functional organ donation programs are the sufficient number of personnel and the adequate material conditions to be provided in relation to the needs. AIM: The goal of the current study was to evaluate the professional environment in Hungary. METHOD: The Organ Coordination Office at the Hungarian National Blood Transfusion Service compiled a questionnaire survey on the personnel and material conditions of the intensive care units (ICUs) in Hungary in regards to organ donations. The survey applied an online questionnaire including 43 questions. In addition to the number of beds and employees, we investigated the tools needed for the legal and the medical diagnosis of brain death as well as the accessibility of examinations on the donor information form. The data collection spanned from 12 December 2016 to 30 June 2017. RESULTS: 59 intensive care units completed the questionnaire; the investigation involved 640 hospital beds, 816 physicians and 1252 nurses. In the daytime shift, 0.25 doctors and 0.41 nurses work on a patient bed at an average, while in the night shift, the figures are 0.11 and 0.33, respectively. 51.7% of the doctors are registered to access the National Non-Donor Registry, and brain death diagnosis committee is available in 83% of the hospitals. Among the medical imaging methods (cranial, abdominal-thoracic), CT scan in 71-73%, abdominal ultrasound in 75%, transthoracic echocardiograpy (TTE) in 37%, transoesophageal echocardiography (TEE) in 4%, bronchoscopy in 49%, coronarography in 19% are non-stop available, with instant interpretation in 75% of the cases. Transcranial Doppler (TCD) in 30%, four-vessel angiography in 45% and SPECT in 14% of the cases are available. More than 90% of the laboratory examinations on the donor information form are available 24 hours a day. CONCLUSION: The number of doctors and nurses did not change compared to our 2008 survey (0.18 doctors, 0.37 nurses/ICU beds in 2008), but the care of potential donors needs more resources and time. The standby availability of personnel and material conditions is a prerequisite for organ donation programs in order to save lives. Orv Hetil. 2018; 159(33): 1360-1367.

Vaccine induced antibodies to the first variable loop of human immunodeficiency virus type 1 gp120, mediate antibody-dependent virus inhibition in macaques.

The role of antibodies directed against the hyper variable envelope region V1 of human immunodeficiency virus type 1 (HIV-1), has not been thoroughly studied. We show that a vaccine able to elicit strain-specific non-neutralizing antibodies to this region of gp120 is associated with control of highly pathogenic chimeric SHIV(89.6P) replication in rhesus macaques. The vaccinated animal that had the highest titers of antibodies to the amino terminus portion of V1, prior to challenge, had secondary antibody responses that mediated cell killing by antibody-dependent cellular cytotoxicity (ADCC), as early as 2 weeks after infection and inhibited viral replication by antibody-dependent cell-mediated virus inhibition (ADCVI), by 4 weeks after infection. There was a significant inverse correlation between virus level and binding antibody titers to the envelope protein, (R=-0.83, p=0.015), and ADCVI (R=-0.84 p=0.044). Genotyping of plasma virus demonstrated in vivo selection of three SHIV(89.6P) variants with changes in potential N-linked glycosylation sites in V1. We found a significant inverse correlation between virus levels and titers of antibodies that mediated ADCVI against all the identified V1 virus variants. A significant inverse correlation was also found between neutralizing antibody titers to SHIV(89.6) and virus levels (R=-0.72 p=0.0050). However, passive inoculation of purified immunoglobulin from animal M316, the macaque that best controlled virus, to a naïve macaque, resulted in a low serum neutralizing antibodies and low ADCVI activity that failed to protect from SHIV(89.6P) challenge. Collectively, while our data suggest that anti-envelope antibodies with neutralizing and non-neutralizing Fc(R-dependent activities may be important in the control of SHIV replication, they also demonstrate that low levels of these antibodies alone are not sufficient to protect from infection.

Regulatory T-cell markers, indoleamine 2,3-dioxygenase, and virus levels in spleen and gut during progressive simian immunodeficiency virus infection.

High levels of viral replication occur in gut-associated lymphoid tissue (GALT) and other lymphoid tissues (LT) since the early phase of human/simian immunodeficiency virus (HIV/SIV) infection. Regulatory T cells (T(reg)), a subset of immunosuppressive T cells expressing CTLA-4 and the FoxP3 transcription factor, accumulate in LT during HIV/SIV infection. Here we show that FoxP3 and CTLA-4 mRNA are increased in leukocytes from the spleens, lymph nodes (LN), and mucosal sites of chronically SIV-infected macaques with high viremia (SIV(HI)) compared to animals with low viremia (SIV(LO)). FoxP3 and CTLA-4 correlated with SIV RNA levels in tissues; SIV virus levels in the spleen, inguinal LN, mesenteric LN, colon, and jejunum directly correlated with the plasma virus level. Importantly, CTLA-4 and FoxP3 mRNA were predominantly increased in the CD25(-) subpopulation of leukocytes from SIV(HI), further challenging the classical definition of T(reg) as CD4(+) CD25(+) T cells. Similar to CTLA-4 and FoxP3, expression of indoleamine 2,3-dioxygenase (IDO), an immunosuppressive enzyme induced by T(reg) in antigen-presenting cells, was increased in the spleens, mesenteric LN, colons, and jejuna from SIV(HI) compared to SIV(LO) and directly correlated to SIV RNA in the same tissues. Accordingly, plasma kynurenine/tryptophan, a marker for IDO enzymatic activity, was significantly higher in SIV(HI) compared to SIV(LO) and correlated with plasma viral levels. Increased T(reg) and IDO in LT of SIV-infected macaques may be the consequence of increased tissue inflammation and/or may favor virus replication during the chronic phase of SIV infection.

Interleukin-15 but not interleukin-7 abrogates vaccine-induced decrease in virus level in simian immunodeficiency virus mac251-infected macaques.

The loss of CD4(+) T cells and the impairment of CD8(+) T cell function in HIV infection suggest that pharmacological treatment with IL-7 and IL-15, cytokines that increase the homeostatic proliferation of T cells and improve effector function, may be beneficial. However, these cytokines could also have a detrimental effect in HIV-1-infected individuals, because both cytokines increase HIV replication in vitro. We assessed the impact of IL-7 and IL-15 treatment on viral replication and the immunogenicity of live poxvirus vaccines in SIV(mac251)-infected macaques (Macaca mulatta). Neither cytokine augmented the frequency of vaccine-expanded CD4(+) or CD8(+) memory T cells, clonal recruitment to the SIV-specific CD8(+) T cell pool, or CD8(+) T cell function. Vaccination alone transiently decreased the viral set point following antiretroviral therapy suspension. IL-15 induced massive proliferation of CD4(+) effector T cells and abrogated the ability of vaccination to decrease set point viremia. In contrast, IL-7 neither augmented nor decreased the vaccine effect and was associated with a decrease in TGF-beta expression. These results underscore the importance of testing immunomodulatory approaches in vivo to assess potential risks and benefits for HIV-1-infected individuals.

CTLA-4 blockade decreases TGF-beta, IDO, and viral RNA expression in tissues of SIVmac251-infected macaques.

Regulatory T (T(reg)) cells are a subset of CD25(+)CD4(+) T cells that constitutively express high levels of cytotoxic T lymphocyte antigen-4 (CTLA-4) and suppress T-cell activation and effector functions. T(reg) cells are increased in tissues of individuals infected with HIV-1 and macaques infected with simian immunodeficiency virus (SIV(mac251)). In HIV-1 infection, T(reg) cells could exert contrasting effects: they may limit viral replication by decreasing immune activation, or they may increase viral replication by suppressing virusspecific immune response. Thus, the outcome of blocking T(reg) function in HIV/SIV should be empirically tested. Here, we demonstrate that CD25(+) T cells inhibit virus-specific T-cell responses in cultured T cells from blood and lymph nodes of SIV-infected macaques. We investigated the impact of CTLA-4 blockade using the anti-CTLA-4 human antibody MDX-010 in SIV-infected macaques treated with antiretroviral therapy (ART). CTLA-4 blockade decreased expression of the tryptophan-depleting enzyme IDO and the level of the suppressive cytokine transforming growth factor-beta (TGF-beta) in tissues. CTLA-4 blockade was associated with decreased viral RNA levels in lymph nodes and an increase in the effector function of both SIV-specific CD4(+) and CD8(+) T cells. Therefore, blunting T(reg) function in macaques infected with SIV did not have detrimental virologic effects and may provide a valuable approach to complement ART and therapeutic vaccination in the treatment of HIV-1 infection.

Decreased number of CD4+ and CD8+ T cells that express the interleukin-7 receptor in blood and tissues of SIV-infected macaques.

Acute HIV/SIV (human/simian immunodeficiency virus) infection results in severe CD4(+) T cell depletion in lymphoid compartments. During the chronic phase of infection, CD4(+) T cell numbers rebound in blood but remain low in the gut-associated lymphoid tissue (GALT), even when viral replication is suppressed by antiretroviral therapy (ART). Thus, strategies to repopulate lymphoid compartments may ameliorate the clinical outcome of HIV/SIV infection. Interleukin (IL)-7 is a key cytokine for the maintenance of homeostatic proliferation of T cells. In HIV/SIV infection, IL-7 expression is increased, likely to compensate for T cell loss, suggesting that supraphysiological administration of IL-7 could provide additional benefit. However, the ability of T cells to respond to IL-7 is dependent on the level of expression of the IL-7 receptor (IL-7R) in T cells in various body compartments. In here, we investigated the proportion of IL-7R(+) T cells in blood, spleen, gut, and genitourinary tract of healthy and SIV-infected macaques with various degrees of CD4(+) T cell depletion. We found that the percentage of T cells expressing IL-7R was significantly lower in both CD4(+) and CD8(+) T cell subsets in SIV-infected macaques than in healthy animals and this decrease directly correlated with the CD4(+) T cell number. Importantly, the proportion of CD4(+) and CD8(+) T cells expressing IL-7R in blood paralleled that found in tissues. IL-7R(+) T cells within the SIV-specific CD8(+) T cells varied and were lowest in most tissues of viremic macaques, likely reflecting continuous antigen stimulation of effector cells.

Systemic immunization with an ALVAC-HIV-1/protein boost vaccine strategy protects rhesus macaques from CD4+ T-cell loss and reduces both systemic and mucosal simian-human immunodeficiency virus SHIVKU2 RNA levels.

Transmission of human immunodeficiency virus type 1 (HIV-1) occurs primarily via the mucosal route, suggesting that HIV-1 vaccines may need to elicit mucosal immune responses. Here, we investigated the immunogenicity and relative efficacy of systemic immunization with two human ALVAC-HIV-1 recombinant vaccines expressing Gag, Pol, and gp120 (vCP250) or Gag, Pol, and gp160 (vCP1420) in a prime-boost protocol with their homologous vaccine native Env proteins. The relative efficacy was measured against a high-dose mucosal exposure to the pathogenic neutralization-resistant variant SHIV(KU2) (simian-human immunodeficiency virus). Systemic immunization with both vaccine regimens decreased viral load levels not only in blood but unexpectedly also in mucosal sites and protected macaques from peripheral CD4+ T-cell loss. This protective effect was stronger when the gp120 antigen was included in the vaccine. Inclusion of recombinant Tat protein in the boosting phase along with the Env protein did not contribute further to the preservation of CD4+ T cells. Thus, systemic immunization with ALVAC-HIV-1 vaccine candidates elicits anti-HIV-1 immune responses able to contain virus replication also at mucosal sites in macaques.

Improved vaccine protection from simian AIDS by the addition of nonstructural simian immunodeficiency virus genes.

An HIV-1 vaccine able to induce broad CD4+ and CD8+ T cell responses may provide long-term control of viral replication. In this study we directly assess the relative benefit of immunization with vaccines expressing three structural Ags (Gag, Pol, and Env), three early regulatory proteins (Rev, Tat, and Nef), or a complex vaccine expressing all six Ags. The simultaneous administration of all six Ags during vaccination resulted in Ag competition manifested by a relative reduction of CD8+ T cell and lymphoproliferative responses to individual Ags. Despite the Ag competition, vaccination with all six Ags resulted in a delay in the onset and a decrease in the extent of acute viremia after mucosal challenge exposure to highly pathogenic SIV(mac251). Reduced levels of acute viremia correlated with lower post-set point viremia and long-term control of infection. In immunized animals, virus-specific CD4+ T cell and lymphoproliferative responses were preserved during acute viremia, and the maintenance of these responses predicted the long-term virological outcome. Taken together, these results suggest that the breadth of the immune response is probably more important than high frequency responses to a limited number of epitopes. These data provide the first clear evidence of the importance of nonstructural HIV Ags as components of an HIV-1 vaccine.

Correlation between viral RNA levels but not immune responses in plasma and tissues of macaques with long-standing SIVmac251 infection.

Plasma virus in human immunodeficiency virus type 1/simian immunodeficiency virus (HIV-1/SIV) infection most likely results from the combination of viruses produced in different tissues. As immunological pressure may be higher in effector sites than secondary lymphoid tissues, we investigated quantitative and qualitative changes in viral RNA in blood and tissues of 10 Mamu-A*01-positive SIV-infected macaques in parallel with the frequency of CD8+ T cells recognizing the dominant Gag181-189 CM9 epitope. The plasma virus level in these macaques directly correlated with the viral RNA levels in lymph nodes, spleen, lungs, colon, and jejunum. In contrast, the frequency of the Gag181-189 CM9 tetramer did not correlate with SIV RNA levels in any compartment. We investigated the presence of viral immune escape in RNA from several tissues. The complete substitution of wild-type genotype with viral immune-escape variant within the Gag181-189 CM9 epitope was associated with low tetramer response in all tissues and blood of two macaques. In one macaque, the replacement of wild type with an immune-escape mutant was asynchronous. While the mutant virus was prevalent in blood and effector tissues (lungs, jejunum, and colon), secondary lymphoid organs such as spleen and lymph nodes still retained 80% and 40%, respectively, of the wild-type virus. These results may imply that there are differences in the immunological pressure exerted by cytotoxic T lymphocytes (CTLs) in tissue compartments of SIVmac251-infected macaques.

Contrasting effects of low-dose IL-2 on vaccine-boosted simian immunodeficiency virus (SIV)-specific CD4+ and CD8+ T cells in macaques chronically infected with SIVmac251.

IL-2, the first cytokine discovered with T cell growth factor activity, is now known to have pleiotropic effects on T cells. For example, it can promote growth, survival, and differentiation of Ag-selected cells, or facilitate Ag-induced cell death of T cells when Ag persists, and in vivo, it is thought to contribute to the regulation of the size of adaptive T cell response. IL-2 is deficient in HIV-1 infection and has been used in the management of HIV-1-infected individuals undergoing antiretroviral therapy. In this study, we investigated how continuous low-dose IL-2 affected the CD4+ and CD8+ T cell response induced by two inoculations of a canarypox recombinant SIV-based vaccine candidate in healthy macaques chronically infected with SIVmac251. These macaques had normal levels of CD4+ T cells at the beginning of antiretroviral therapy treatment. Vaccination in the presence of IL-2 significantly augmented Gag-specific CD8+ T cell responses, but actually reduced Gag-specific CD4+ T cell responses. Although IL-2 at low doses did not change the overall concentration of circulating CD4+ or CD8+ T cells, it expanded the frequency of CD4+CD25+ T cells. Depletion of the CD4+CD25+ T cells in vitro, however, did not result in a reconstitution of Gag-specific CD4+ responses or augmentation of SIV-specific CD8+ T cell responses. Thus, we conclude that the decrease in virus-specific CD4+ T cell response may be due to IL-2-promoted redistribution of cells from the circulation, or due to Ag-induced cell death, rather than suppression by a T regulatory population.

Avipox-based simian immunodeficiency virus (SIV) vaccines elicit a high frequency of SIV-specific CD4+ and CD8+ T-cell responses in vaccinia-experienced SIVmac251-infected macaques.

The ability of ALVAC- or fowlpox-based simian immunodeficiency virus (SIV) vaccines to boost SIV-specific CD4+ and CD8+ T-cell responses was tested in 10 vaccinia-experienced macaques infected with SIVmac251. The CD8+ T-cell response to the dominant Gag(181-189) CM9 was quantitated in seven Mamu-A*01-positive macaques by tetramer staining, by ex vivo cytotoxic T-lymphocyte (CTL) activity, and by intracellular cytokine staining (ICS) with the specific Gag(181-189) CM9 peptide. The overall CD8+ T-cell response to Gag was assessed using a peptide pool encompassing the entire Gag protein followed by measurement of TNF-alpha production in ICS assay. Similarly, virus-specific CD4+ T-cell responses were measured by ICS for TNF-alpha following stimulation with the Gag-overlapping peptide and by proliferative response following stimulation with purified p27 Gag. The two vaccine modalities effectively boosted both CD4+ and CD8+ SIV-specific T-cell response despite prior exposure to the vaccinia-derivative NYVAC vector, suggesting that sequential boosting with either avipox-based vector vaccine candidate is a realistic approach in immune therapy of human immunodeficiency virus type 1 (HIV-1)-infected individuals.

Modeling a safer smallpox vaccination regimen, for human immunodeficiency virus type 1-infected patients, in immunocompromised macaques.

We have modeled smallpox vaccination with Dryvax (Wyeth) in rhesus macaques that had depletion of CD4(+) T cells induced by infection with simian immunodeficiency virus or simian/human immunodeficiency virus. Smallpox vaccination induced significantly larger skin lesions in immunocompromised macaques than in healthy macaques. Unexpectedly, "progressive vaccinia" was infrequent. Vaccination of immunocompromised macaques with the genetically-engineered, replication-deficient poxvirus NYVAC, before or after retrovirus infection, was safe and lessened the severity of Dryvax-induced skin lesions. Neutralizing antibodies to vaccinia were induced by NYVAC, even in macaques with severe CD4(+) T cell depletion, and their titers inversely correlated with the time to complete resolution of the skin lesions. Together, these results provide the proof of concept, in macaque models that mirror human immunodeficiency virus type 1 infection, that a prime-boost approach with a highly attenuated poxvirus followed by Dryvax increases the safety of smallpox vaccination, and they highlight the importance of neutralizing antibodies in protection against virulent poxvirus.

Prior DNA immunization enhances immune response to dominant and subdominant viral epitopes induced by a fowlpox-based SIVmac vaccine in long-term slow-progressor macaques infected with SIVmac251.

A therapeutic vaccine for individuals infected with HIV-1 and treated with antiretroviral therapy (ART) should be able to replenish virus-specific CD4+ T-cells and broaden the virus-specific CD8+ T-cell response in order to maintain CD8+ T-cell function and minimize viral immune escape after ART cessation. Because a combination of DNA and recombinant poxvirus vaccine modalities induces high levels of virus-specific CD4+ T-cell response and broadens the cytolytic activity in naive macaques, we investigated whether the same results could be obtained in SIVmac251-infected macaques. The macaques studied here were long-term nonprogressors that naturally contained viremia but were nevertheless treated with a combination of antiviral drugs to assess more carefully the effect of vaccination in the context of ART. The combination of a DNA expressing the gag and pol genes (DNA-SIV-gp) of SIVmac239 followed by a recombinant fowlpox expressing the same SIVmac genes (FP-SIV-gp) was significantly more immunogenic than two immunizations of FP-SIV-gp in SIVmac251-infected macaques treated with ART. The DNA/FP combination significantly expanded and broadened Gag-specific T-cell responses measured by tetramer staining, ELISPOT, and intracellular cytokine staining and measurement of ex vivo cytolytic function. Importantly, the combination of these vaccine modalities also induced a sizeable expansion in most macaques of Gag-specific CD8-(CD4+) T-cells able to produce TNF-alpha. Hopefully, this modality of vaccine combination may be useful in the clinical management of HIV-1-infected individuals.

Emergence of cytotoxic T lymphocyte escape mutants following antiretroviral treatment suspension in rhesus macaques infected with SIVmac251.

Structured treatment interruption (STI) of antiretroviral drugs has been proposed as an alternative approach for managing patients infected with HIV-1. While STI is thought to spare drug-related side effects and enhance the HIV-1-specific immune response, the long-lasting clinical benefit of this approach remains uncertain, particularly in patients with long-standing HIV-1 infection. Here, we investigated the basis of unabated virological replication following different STI regimens in rhesus macaques that expressed the MHC class I Mamu-A*01 molecule treated during acute and long-standing infection with SIVmac251. An amino acid change at the anchor residue within the immunodominant Mamu-A*01-restricted Gag(181-189) CM9 epitope (T --> A) in one of six macaques with acute SIVmac251 infection and in three of four macaques with long-standing SIVmac251 infection (T --> A; T --> S; S --> C) was found in the majority of plasma virus. These amino acid changes have been shown to severely decrease binding of the corresponding peptides to the Mamu-A*01 molecule and, in the case of the T --> A change, escape from CTL. In one macaque with long-standing SIVmac251 infection, a mutation emerged that conferred resistance to one of the antiretroviral drugs (PMPA) as well. These results provide insights into the mechanism underlying the limited capacity of repeated interruption of antiretroviral therapy as an approach to restrain viral replication. In addition, these data also suggest that interruption of therapy may be less effective in chronic infection because of preexisting immune escape and that immune escape is a risk of interruption of therapy.

Human immunodeficiency virus type-1 Tat/co-activator acetyltransferase interactions inhibit p53Lys-320 acetylation and p53-responsive transcription.

Patients with AIDS are at increased risk for developing various neoplasms, including Hodgkin's and non-Hodgkin's lymphomas, Kaposi's sarcomas, and anal-rectal carcinomas, suggestive that human immunodeficiency virus type-1 infection might promote establishment of AIDS-related cancers. Tat, the viral trans-activator, can be endocytosed by uninfected cells and has been shown to inhibit p53 functions, providing a candidate mechanism through which the human immunodeficiency virus type-1 might contribute to malignant transformation. Because Tat has been shown to interact with histone acetyltransferase domains of p300/cAMP-responsive element-binding protein (CREB)-binding protein and p300/CREB-binding protein-associated factor, we have investigated whether Tat might alter p53 acetylation and tumor suppressor-responsive transcription. Here, we demonstrate that both Tat and p53 co-localize with p300/CREB-binding protein-associated factor and p300 in nuclei of IMR-32 human neuroblastoma cells and in PC-12 pheochromocytoma cells. Further, p53 trans-activation of the 14-3-3varsigma promoter was markedly repressed by Tat-histone acetyltransferase interactions, and p53 acetylation by p300/CREB-binding protein-associated factor on residue Lys(320) was diminished as a result of Tat-histone acetyltransferase binding in vivo and in vitro. Tat also inhibited p53 acetylation by p300 in a dosage-dependent manner in vitro. Finally, HIV-1-infected Molt-4 cells displayed reduced p53 acetylation on lysines 320 and 373 in response to UV irradiation. Our results allude to a mechanism whereby the human immunodeficiency virus type-1 trans-activator might impair tumor suppressor functions in immune/neuronal-derived cells, thus favoring the establishment of neoplasia during AIDS.

Immune intervention strategies for HIV-1 infection of humans in the SIV macaque model.

Studies in the SIVmac macaque model have demonstrated that the extent of virus-specific CD4+ and CD8+ T-cell responses induced by vaccination prior to virus-challenge exposure correlate with viremia containment following establishment of infection. These findings led to the hypothesis that active immunization with vaccines able to induce virus-specific T-cell responses following the establishment of infection could also ameliorate the virological outcome. Here, we will review the relative effect of ART and vaccination during primary SIVmac infection of macaques.