Immunocompromised host section: Adoptive T-cell therapy for dsDNA viruses in allogeneic hematopoietic cell transplant recipients.

PURPOSE OF REVIEW: Double-stranded DNA (dsDNA) viruses remain important causes of morbidity and mortality after allogeneic hematopoietic cell transplantation (HCT). As treatment options are limited, adoptive therapy with virus-specific T cells (VST) is promising in restoring immunity and thereby preventing and treating virus infections. Here we review current evidence and recent advances in the field of VST for dsDNA viruses in allogeneic HCT recipients. RECENT FINDINGS: Four different protocols for VST generation are currently used in clinical trials, and various products including multivirus-specific and off-the-shelf products are under investigation for prophylaxis, preemptive therapy or treatment. Data from nearly 1400 dsDNA-VST applications in allogeneic HCT patients have been published and demonstrated its safety. Although Epstein-Barr virus, cytomegalovirus, and adenovirus-specific T-cell therapy studies have predominated over the past 25 years, additional human herpes viruses were added to multivirus-specific T cells over the last decade and clinical evidence for polyomavirus-specific VST has just recently emerged. Response rates of around 70-80% have been reported, but cautious interpretation is warranted as data are predominantly from phase 1/2 studies and clinical efficacy needs to be confirmed in phase 3 studies. SUMMARY: Investigation on the 'ideal' composition of VST is ongoing. Several products recently entered phase 3 trials and may allow widespread clinical use in the near future.

Interferon-λ Enhances the Differentiation of Naive B Cells into Plasmablasts via the mTORC1 Pathway.

Type III interferon (interferon lambda [IFN-λ]) is known to be a potential immune modulator, but the mechanisms behind its immune-modulatory functions and its impact on plasmablast differentiation in humans remain unknown. Human B cells and their subtypes directly respond to IFN-λ. Using B cell transcriptome profiling, we investigate the immune-modulatory role of IFN-λ in B cells. We find that IFN-λ-induced gene expression in B cells is steady, prolonged, and importantly, cell type specific. Furthermore, IFN-λ enhances the mTORC1 (mammalian/mechanistic target of rapamycin complex 1) pathway in B cells activated by the B cell receptor (BCR/anti-IgM). Engagement of mTORC1 by BCR and IFN-λ induces cell-cycle progress in B cells. Subsequently, IFN-λ boosts the differentiation of naive B cells into plasmablasts upon activation, and the cells gain effector functions such as cytokine release (IL-6 and IL-10) and antibody production. Our study shows how IFN-λ systematically boosts the differentiation of naive B cells into plasmablasts by enhancing the mTORC1 pathway and cell-cycle progression in activated B cells.

T-Cell-Mediated Cross-Protective Immunity.

Human invasive fungal infections are caused by different mold and yeast species. Hence, cross-reactive T cells that recognize conserved epitopes of various fungal pathogens are of special interest for vaccination protocols or adoptive T cell transfer. Here, we describe an ELISpot-based method to test cross-reactivity of T cell lines or clones to different molds and yeasts.

Host response to fungal infections - how immunology and host genetics could help to identify and treat patients at risk.

In spite of the ever-increasing incidence and poor outcome of invasive fungal infections in immune compromised patients, there is currently no reliable method to accurately predict the risk, to monitor the outcome and to treat these infections. Protective immunity against Candida and Aspergillus depends on a highly coordinated interaction between the innate and adaptive immune systems. Genetic and immunological defects in components of these networks result in increased risk of invasive fungal infections among patients undergoing chemotherapy or transplant recipients. We review the most important genetic and immunological factors that influence human susceptibility to Candida and Aspergillus infections and discuss the potential role of basic research to promote precision medicine for infectious diseases. We discuss how immunogenetic studies can help to provide tools for improved identification of high-risk patients and the development of tailored antifungal therapies.

Immune recovery in HIV-infected patients after Candida esophagitis is impaired despite long-term antiretroviral therapy.

OBJECTIVE: Candida esophagitis belongs to the most common AIDS-defining diseases; however, a comprehensive immune pathogenic concept is lacking. DESIGN: We investigated the immune status of 37 HIV-1-infected patients from the Swiss HIV cohort study at diagnosis of Candida esophagitis, 1 year before, 1 year later and after 2 years of suppressed HIV RNA. We compared these patients with three groups: 37 HIV-1-infected patients without Candida esophagitis but similar CD4 cell counts as the patients at diagnosis (advanced HIV group), 15 HIV-1-infected patients with CD4 cell counts higher than 500 cells/µl, CD4 cell nadirs higher than 350 cells/µl and suppressed HIV RNA under combination antiretroviral therapy (cART) (early cART group) and 20 healthy individuals. METHODS: We investigated phenotype, cytokine production and proliferative capacity of different immune cells by flow cytometry and enzyme-linked immunosorbent spot. RESULTS: We found that patients with Candida esophagitis had nearly abolished CD4 cell proliferation in response to Candida albicans, significantly increased percentages of dysfunctional CD4 cells, significantly decreased cytotoxic natural killer cell counts and peripheral innate lymphoid cell counts and significantly reduced IFN-γ and IL-17 production compared with the early cART group and healthy individuals. Most of these defects remained for more than 2 years despite viral suppression. The advanced HIV group without opportunistic infection showed partly improved immune recovery. CONCLUSION: Our data indicate that Candida esophagitis in HIV-1-infected patients is caused by an accumulation of multiple, partly Candida-specific immunological defects. Long-term immune recovery is impaired, illustrating that specific immunological gaps persist despite cART. These data also support the rationale for early cART initiation to prevent irreversible immune defects.

T cells specific for different latent and lytic viral proteins efficiently control Epstein-Barr virus-transformed B cells.

BACKGROUND AIMS: Epstein-Barr virus (EBV)-associated post-transplant lymphoproliferative disorders (PTLD) belong to the most dreaded complications of immunosuppression. The efficacy of EBV-specific T-cell transfer for PTLD has been previously shown, yet the optimal choice of EBV-derived antigens inducing polyclonal CD4(+) and CD8(+) T cells that cover a wide range of human leukocyte antigen types and efficiently control PTLD remains unclear. METHODS: A pool of 125 T-cell epitopes from seven latent and nine lytic EBV-derived proteins (EBVmix) and peptide pools of EBNA1, EBNA3c, LMP2a and BZLF1 were used to determine T-cell frequencies and to isolate T cells through the use of the interferon (IFN)-γ cytokine capture system. We further evaluated the phenotype and functionality of the generated T-cell lines in vitro. RESULTS: EBVmix induced significantly higher T-cell frequencies and allowed selecting more CD4(+)IFN-γ(+) and CD8(+)IFN-γ(+) cells than single peptide pools. T cells of all specificities expanded similarly in vitro, recognized cognate antigen, and, to a lower extent, EBV-infected cells, exerted moderate cytotoxicity and showed reduced alloreactivity. However, EBVmix-specific cells most efficiently controlled EBV-infected lymphoblastoid cell lines (LCLs). This control was mainly mediated by EBV-specific CD8(+) cells with an oligoclonal epitope signature covering both latent and lytic viral proteins. Notably, EBV-specific CD4(+) cells unable to control LCLs produced significantly less perforin and granzyme B, probably because of limited LCL epitope presentation. CONCLUSIONS: EBVmix induces a broader T-cell response, probably because of its coverage of latent and lytic EBV-derived proteins that may be important to control EBV-transformed B cells and might offer an improvement of T-cell therapies.

Combined PI3K/Akt and Hsp90 targeting synergistically suppresses essential functions of alloreactive T cells and increases Tregs.

Acute graft-versus-host disease is still a major cause of transplant-related mortality after allogeneic stem cell transplantation. It requires immunosuppressive treatments that broadly abrogate T cell responses, including beneficial ones directed against tumor cells or infective pathogens. Inhibition of the heat shock protein of 90 kDa has been demonstrated to eliminate tumor cells, as well as alloreactive T cells while preserving antiviral T cell immunity. Here, we show that the suppressive effects of heat shock protein of 90 kDa inhibition on alloreactive T cells were synergistically enhanced by concomitant inhibition of the PI3K/Akt signaling pathway, which is also strongly activated upon allogeneic stimulation. Molecular analyses revealed that this antiproliferative effect was mainly mediated by induction of cell-cycle arrest and apoptosis. In addition, we observed an increased proportion of activated regulatory T cells, which critically contribute to acute graft-versus-host disease control, upon combined heat shock protein of 90 kDa/Akt isoforms 1 and 2 or heat shock protein of 90 kDa/PI3K/p110δ isoform inhibition. Moreover, antiviral T cell immunity was functionally preserved after combined heat shock protein of 90 kDa/Akt isoforms 1 and 2 inhibition. Taken together, our data suggest that the combined heat shock protein of 90 kDa/PI3K/Akt inhibition approach represents a reasonable dual strategy to suppress residual tumor growth and efficiently deplete alloreactive T cells and thus, provide a rationale to prevent and treat acute graft-versus-host disease selectively without impairing pathogen-specific T cell immunity.

Immune Reconstitution After Allogeneic Hematopoietic Stem Cell Transplantation and Association With Occurrence and Outcome of Invasive Aspergillosis.

BACKGROUND: Invasive aspergillosis (IA) remains a leading cause of morbidity and mortality in patients receiving allogeneic hematopoietic stem cell transplantation (HSCT). To date, no reliable immunological biomarkers for management and outcome of IA exist. Here, we investigated reconstitution of antifungal immunity in patients during the first 12 months after HSCT and correlated it with IA. METHODS: Fifty-one patients were included, 9 with probable/proven IA. We determined quantitative and qualitative reconstitution of polymorphonuclear (PMN), CD4, CD8, and natural killer (NK) cells against Aspergillus fumigatus over 5 time points and compared the values to healthy donors. RESULTS: Absolute CD4 and CD8 cell counts, antigen-specific T-cell responses, and killing capacity of PMN against A. fumigatus were significantly decreased in all patients over 12 months. In patients with probable/proven IA, reactive oxygen species (ROS) production tended to be lower compared to patients without IA, and absolute NK-cell counts remained below 200 cells/µL. Patients with well-controlled IA showed significantly higher ROS production and NK-cell counts compared to patients with poor outcome. CONCLUSIONS: This study highlights the importance of functional PMN, T-cell, and NK-cell immunity for the outcome of IA. Larger multicenter studies should address the potential use of NK-cell counts for the management of antifungal therapy.

Multispecific Aspergillus T cells selected by CD137 or CD154 induce protective immune responses against the most relevant mold infections.

BACKGROUND: Aspergillus and Mucorales species cause severe infections in patients after hematopoietic stem cell transplantation (HSCT). Induction of antifungal CD4(+) T-helper type 1 (Th1) immunity is an appealing strategy to combat these infections. Immunotherapeutic approaches are so far limited because of a lack of antigens inducing protective T cells, their elaborate production, and the need of targeting a broad spectrum of pathogenic fungi. METHODS: We examined the response to different Aspergillus fumigatus proteins in healthy individuals and patients after HSCT and compared rapid selection protocols for fungus-specific T cells based on CD137 or CD154 expression. RESULTS: The A. fumigatus proteins Crf1, Gel1, and Pmp20 induced strong Th1 responses in healthy individuals. T cells specific for these antigens expanded in patients with active invasive aspergillosis, indicating their contribution to infection control. Th1 cells specific for the 3 proteins can be selected with similar specificity within 24 hours, based on CD137 or CD154 expression. These cells recognize naturally processed A. fumigatus and the multispecific T-cell lines, directed against all 3 proteins, especially those selected by CD154, additionally cross-react to different Aspergillus and Mucorales species. CONCLUSIONS: These findings may form the basis for adoptive T-cell transfer for prophylaxis or treatment in patients with these devastating infections.

Generation of a multipathogen-specific T-cell product for adoptive immunotherapy based on activation-dependent expression of CD154.

Viral and fungal infections remain a leading cause of mortality in patients after hematopoietic stem cell transplantation (HSCT). Adoptive transfer of multipathogen-specific T cells is promising in restoring immunity and thereby preventing and treating infections, but approaches are currently limited because of time-consuming and laborious procedures. Therefore, we investigated a new strategy to simultaneously select T cells specific for viral and fungal pathogens based on activation-dependent expression of CD154. Single- and multipathogen-specific T-cell lines with high specificity for adenovirus (AdV), Epstein-Barr virus (EBV), cytomegalovirus (CMV), Candida albicans, and/or Aspergillus fumigatus could be readily generated within 14 days irrespective of the precursor frequency. The T-cell lines responded reproducibly to endogenously processed antigen and specifically proliferated upon antigenic stimulation. Although isolation based on CD154 favors enrichment of CD4(+) T cells, AdV-, EBV- and CMV-specific CD8(+) T cells could be expanded and demonstrated lysis of target cells. Conversely, T cell-mediated alloreactivity was almost abrogated compared with the starting fraction. This selection and/or expansion strategy may form the basis for future adoptive immunotherapy trials in patients at risk for multiple infections and may be translated to other antigens.

Cross-protective TH1 immunity against Aspergillus fumigatus and Candida albicans.

T cell-mediated heterologous immunity to different pathogens is promising for the development of immunotherapeutic strategies. Aspergillus fumigatus and Candida albicans, the 2 most common fungal pathogens causing severe infections in immunocompromised patients, are controlled by CD4+ type 1 helper T (T(H)1) cells in humans and mice, making induction of fungus-specific CD4+ T(H)1 immunity an appealing strategy for antifungal therapy. We identified an immunogenic epitope of the A fumigatus cell wall glucanase Crf1 that can be presented by 3 common major histocompatibility complex class II alleles and that induces memory CD4+ T(H)1 cells with a diverse T-cell receptor repertoire that is cross-reactive to C albicans. In BALB/c mice, the Crf1 protein also elicits cross-protection against lethal infection with C albicans that is mediated by the same epitope as in humans. These data illustrate the existence of T cell-based cross-protection for the 2 distantly related clinically relevant fungal pathogens that may foster the development of immunotherapeutic strategies.

Selective depletion of alloreactive T cells by targeted therapy of heat shock protein 90: a novel strategy for control of graft-versus-host disease.

Graft-versus-host disease (GVHD) is a major cause of morbidity and mortality in patients with hematologic malignancies undergoing allogeneic hematopoietic stem cell transplantation. Current treatment of GVHD relies on immunosuppressive regimens, considerably increasing the incidence of opportunistic infections. As T cells mediate both GVHD as well as protection against viral infections and the malignant disease, strategies to selectively target host-reactive T cells without impairing pathogen- and disease-specific immunity are highly warranted. Activation of T cells is accompanied by increased expression of the chaperone heat shock protein of 90 kDa (Hsp90), which stabilizes several key signaling pathways crucial for T-cell activation. In this study, selective targeting of Hsp90 in activated T lymphocytes with pharmacologic inhibitors already applied successfully in anticancer therapy resulted in induction of apoptosis predominantly in activated cells. Moreover, if T cells were stimulated with allogeneic dendritic cells, alloreactive T cells were selectively eliminated. In contrast, third party reactions including antiviral T-cell immunity were quantitatively and functionally fully preserved. These data suggest that Hsp90 represents a novel target for selective depletion of alloreactive T cells, and provide the rationale for application of Hsp90 inhibitors as potential approach to selectively prevent and treat GVHD in hematopoietic stem cell transplantation recipients without impairing pathogen- and disease-specific T-cell immunity.