IFNα Subtypes in HIV Infection and Immunity.

Type I interferons (IFN), immediately triggered following most viral infections, play a pivotal role in direct antiviral immunity and act as a bridge between innate and adaptive immune responses. However, numerous viruses have evolved evasion strategies against IFN responses, prompting the exploration of therapeutic alternatives for viral infections. Within the type I IFN family, 12 IFNα subtypes exist, all binding to the same receptor but displaying significant variations in their biological activities. Currently, clinical treatments for chronic virus infections predominantly rely on a single IFNα subtype (IFNα2a/b). However, the efficacy of this therapeutic treatment is relatively limited, particularly in the context of Human Immunodeficiency Virus (HIV) infection. Recent investigations have delved into alternative IFNα subtypes, identifying certain subtypes as highly potent, and their antiviral and immunomodulatory properties have been extensively characterized. This review consolidates recent findings on the roles of individual IFNα subtypes during HIV and Simian Immunodeficiency Virus (SIV) infections. It encompasses their induction in the context of HIV/SIV infection, their antiretroviral activity, and the diverse regulation of the immune response against HIV by distinct IFNα subtypes. These insights may pave the way for innovative strategies in HIV cure or functional cure studies.

Macrophage migration inhibitory factor receptor CD74 expression is associated with expansion and differentiation of effector T cells in COVID-19 patients.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) caused millions of COVID-19 cases and deaths worldwide. Severity of pulmonary pathologies and poor prognosis were reported to be associated with the activation non-virus-specific bystander T cells. In addition, high concentrations of the macrophage migration inhibitory factor (MIF) were found in serum of COVID-19 patients. We hypothesized that these two pathogenic factors might be related and analyzed the expression of receptors for MIF on T cells in COVID-19. T cells from PBMCs of hospitalized patients with mild and severe COVID-19 were characterized. A significantly higher proportion of CD4+ and CD8+ T cells from COVID-19 patients expressed CD74 on the cell surface compared to healthy controls. To induce intracellular signaling upon MIF binding, CD74 forms complexes with CD44, CXCR2, or CXCR4. The vast majority of CD74+ T cells expressed CD44, whereas expression of CXCR2 and CXCR4 was low in controls but increased upon SARS-CoV-2 infection. Hence, T cells in COVID-19 patients express receptors that render them responsive to MIF. A detailed analysis of CD74+ T cell populations revealed that most of them had a central memory phenotype early in infection, while cells with an effector and effector memory phenotype arose later during infection. Furthermore, CD74+ T cells produced more cytotoxic molecules and proliferation markers. Our data provide new insights into the MIF receptor and co-receptor repertoire of bystander T cells in COVID-19 and uncovers a novel and potentially druggable aspect of the immunological footprint of SARS-CoV-2.

Targeted mutations in IFNα2 improve its antiviral activity against various viruses.

During viral infections, type I interferons (IFN) are induced and play a key role in counteracting initial viral spread. Twelve different human IFNα subtypes exist that bind the same receptor; however, they elicit unique host responses and display distinct potencies of antiviral activities. Our previous studies on human immunodeficiency virus (HIV) and hepatitis B virus (HBV) demonstrated that the clinically used IFNα2 is not the most effective one among the IFNα subtypes. By sequence modeling, we identified a region in helix B with mainly conserved residues at the outside facing IFNAR1, but variable residues at the inside facing the core of IFNα, potentially representing a putative tunable anchor to tune pleiotropic IFN responses. Using site-directed mutagenesis, various mutations were introduced into the IFNα2b backbone targeting sites which are important for binding to IFNAR1 and IFNAR2, the putative tunable anchor, or outside these three regions. Selected mutations were based on sequence differences to high antiviral subtypes IFNα6 and IFNα14. Treatment assays against HBV and HIV identified several critical residues for the antiviral activity of IFNα mainly in the IFNAR1 binding region. Combined mutations of the IFNα2 IFNAR1/2 binding sites or the IFNAR1 binding region plus the putative tunable anchor by those of IFNα14 further augmented activation of different downstream signaling cascades providing a molecular correlate for the enhanced antiviral activity. We describe here important functional residues within IFNα subtype molecules, which enabled us to design novel and innovative drugs that may have the potential to be used in clinical trials against a variety of different viral infections.IMPORTANCEThe potency of interferon (IFN)α to restrict viruses was already discovered in 1957. However, until today, only IFNα2 out of the 12 distinct human IFNα subtypes has been therapeutically used against chronic viral infections. There is convincing evidence that other IFNα subtypes are far more efficient than IFNα2 against many viruses. In order to identify critical antiviral residues within the IFNα subtype sequence, we designed hybrid molecules based on the IFNα2 backbone with individual sequence motifs from the more potent subtypes IFNα6 and IFNα14. In different antiviral assays with HIV or HBV, residues binding to IFNAR1 as well as combinations of residues in the IFNAR1 binding region, the putative tunable anchor, and residues outside these regions were identified to be crucial for the antiviral activity of IFNα. Thus, we designed artificial IFNα molecules, based on the clinically approved IFNα2 backbone, but with highly improved antiviral activity against several viruses.

Immunotherapy-induced cytotoxic T follicular helper cells reduce numbers of retrovirus-infected reservoir cells in B cell follicles.

Antiretroviral therapy (ART) transformed HIV from a life-threatening disease to a chronic condition. However, eliminating the virus remains an elusive therapy goal. For several decades, Friend virus (FV) infection serves as a murine model to study retrovirus immunity. Similar to HIV, FV persists at low levels in lymph nodes B cell follicles avoiding elimination by immune cells. Such immune-privileged reservoirs exclude cytotoxic T cells from entry. However, CXCR5+ T cells are permitted to traffic through germinal centers. This marker is predominantly expressed by CD4+ follicular helper T cells (Tfh). Therefore, we explored immunotherapy to induce cytotoxic Tfh, which are rarely found under physiological conditions. The TNF receptor family member CD137 was first identified as a promising target for cancer immunotherapy. We demonstrated that FV-infected mice treatment with αCD137 antibody resulted in an induction of the cytotoxic program in Tfh. The therapy significantly increased numbers of cytotoxic Tfh within B cell follicles and contributed to viral load reduction. Moreover, αCD137 antibody combined with ART delayed virus rebound upon treatment termination without disturbing the lymph node architecture or antibody responses. Thus, αCD137 antibody therapy might be a novel strategy to target the retroviral reservoir and an interesting approach for HIV cure research.

SRSF1 acts as an IFN-I-regulated cellular dependency factor decisively affecting HIV-1 post-integration steps.

Efficient HIV-1 replication depends on balanced levels of host cell components including cellular splicing factors as the family of serine/arginine-rich splicing factors (SRSF, 1-10). Type I interferons (IFN-I) play a crucial role in the innate immunity against HIV-1 by inducing the expression of IFN-stimulated genes (ISGs) including potent host restriction factors. The less well known IFN-repressed genes (IRepGs) might additionally affect viral replication by downregulating host dependency factors that are essential for the viral life cycle; however, so far, the knowledge about IRepGs involved in HIV-1 infection is very limited. In this work, we could demonstrate that HIV-1 infection and the associated ISG induction correlated with low SRSF1 levels in intestinal lamina propria mononuclear cells (LPMCs) and peripheral blood mononuclear cells (PBMCs) during acute and chronic HIV-1 infection. In HIV-1-susceptible cell lines as well as primary monocyte-derived macrophages (MDMs), expression levels of SRSF1 were transiently repressed upon treatment with specific IFNα subtypes in vitro. Mechanically, 4sU labeling of newly transcribed mRNAs revealed that IFN-mediated SRSF1 repression is regulated on early RNA level. SRSF1 knockdown led to an increase in total viral RNA levels, but the relative proportion of the HIV-1 viral infectivity factor (Vif) coding transcripts, which is essential to counteract APOBEC3G-mediated host restriction, was significantly reduced. In the presence of high APOBEC3G levels, however, increased LTR activity upon SRSF1 knockdown facilitated the overall replication, despite decreased vif mRNA levels. In contrast, SRSF1 overexpression significantly impaired HIV-1 post-integration steps including LTR transcription, alternative splice site usage, and virus particle production. Since balanced SRSF1 levels are crucial for efficient viral replication, our data highlight the so far undescribed role of SRSF1 acting as an IFN-modulated cellular dependency factor decisively regulating HIV-1 post-integration steps.

IFNα subtype-specific susceptibility of HBV in the course of chronic infection.

Chronic hepatitis B virus (HBV) infection continues to be a major health problem worldwide and remains hard to be cured. Therapy with interferon (IFN) α is an important method for the clinical treatment of chronic hepatitis B. IFNα exhibits direct antiviral effects as well as immunomodulatory activities, which can induce sustained antiviral responses in part of the treated chronic hepatitis B patients. Numerous IFNα subtypes with high sequence identity between 76-96% exist which are characterized by diverse, non-redundant biological activities. Our previous studies have demonstrated that the clinically approved IFNα2 is not the most effective subtype for the anti-HBV treatment among all IFNα subtypes. So far very little is known about the IFNα subtype expression pattern during early HBV infection and the IFNα subtype-specific susceptibility during persistent HBV infection as well as its related cellular mechanism. Here we determined the Ifna subtype mRNA expression during acute and chronic HBV infection by using the well-established hydrodynamic injection (HDI) mouse model and we revealed a transient but strong expression of a panel of Ifna subtypes in the spleen of HBV persistent replication mice compared to HDI controls. Immunotherapy with distinct IFNα subtypes controlled chronic HBV infection. IFNα subtype-mediated antiviral response and immune activation were comprehensively analyzed in an AAV-HBV persistent infection murine model and murine IFNα2 was identified as the most effective subtype in suppression of HBV replication. Further analysis of the immune response revealed a strong immunomodulatory activity of murine IFNα2 on splenic and intrahepatic NK and T cell activation during persistent HBV infection. Taken together, our data provide IFNα subtype-specific differences in the antiviral and immunomodulatory effector responses and a strong expression of all IFNα subtypes in the spleen during persistent HBV infection in mice. This knowledge will support the development of novel immunotherapeutic strategies for chronic hepatitis B infection.

Distinct Type I Interferon Subtypes Differentially Stimulate T Cell Responses in HIV-1-Infected Individuals.

The expression of type I interferons (IFNs) is one of the immediate host responses during most viral infections. The type I IFN family consists of numerous highly conserved IFNα subtypes, IFNß, and some others. Although these IFNα subtypes were initially believed to act interchangeably, their discrete biological properties are nowadays widely accepted. Subtype-specific antiviral, immunomodulatory, and anti-proliferative activities were reported explained by differences in receptor affinity, downstream signaling events, and individual IFN-stimulated gene expression patterns. Type I IFNs and increased IFN signatures potentially linked to hyperimmune activation of T cells are critically discussed for chronic HIV (human immunodeficiency virus) infection. Here, we aimed to analyze the broad immunological effects of specific type I IFN subtypes (IFNα2, IFNα14, and IFNß) on T and NK cell subsets during HIV-1 infection in vitro and ex vivo. Stimulation with IFNα14 and IFNß significantly increased frequencies of degranulating (CD107a+) gut-derived CD4+ T cells and blood-derived T and NK cells. However, frequencies of IFNγ-expressing T cells were strongly reduced after stimulation with IFNα14 and IFNß. Phosphorylation of downstream molecules was not only IFN subtype-specific; also, significant differences in STAT5 phosphorylation were observed in both healthy peripheral blood mononuclear cells (PBMCs) and PBMCs of HIV-infected individuals, but this effect was less pronounced in healthy gut-derived lamina propria mononuclear cells (LPMCs), assuming cell and tissue specific discrepancies. In conclusion, we observed distinct type I IFN subtype-specific potencies in stimulating T and NK cell responses during HIV-1-infection.

Differential interferon-α subtype induced immune signatures are associated with suppression of SARS-CoV-2 infection.

Type I interferons (IFN-I) exert pleiotropic biological effects during viral infections, balancing virus control versus immune-mediated pathologies, and have been successfully employed for the treatment of viral diseases. Humans express 12 IFN-alpha (α) subtypes, which activate downstream signaling cascades and result in distinct patterns of immune responses and differential antiviral responses. Inborn errors in IFN-I immunity and the presence of anti-IFN autoantibodies account for very severe courses of COVID-19; therefore, early administration of IFN-I may be protective against life-threatening disease. Here we comprehensively analyzed the antiviral activity of all IFNα subtypes against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to identify the underlying immune signatures and explore their therapeutic potential. Prophylaxis of primary human airway epithelial cells (hAEC) with different IFNα subtypes during SARS-CoV-2 infection uncovered distinct functional classes with high, intermediate, and low antiviral IFNs. In particular, IFNα5 showed superior antiviral activity against SARS-CoV-2 infection in vitro and in SARS-CoV-2-infected mice in vivo. Dose dependency studies further displayed additive effects upon coadministration with the broad antiviral drug remdesivir in cell culture. Transcriptomic analysis of IFN-treated hAEC revealed different transcriptional signatures, uncovering distinct, intersecting, and prototypical genes of individual IFNα subtypes. Global proteomic analyses systematically assessed the abundance of specific antiviral key effector molecules which are involved in IFN-I signaling pathways, negative regulation of viral processes, and immune effector processes for the potent antiviral IFNα5. Taken together, our data provide a systemic, multimodular definition of antiviral host responses mediated by defined IFN-I. This knowledge will support the development of novel therapeutic approaches against SARS-CoV-2.

Immunotherapy With Interferon α11, But Not Interferon Beta, Controls Persistent Retroviral Infection.

Type I Interferons (IFNs), including numerous IFNα subtypes and IFNß, are key molecules during innate and adaptive immune responses against viral infections. These cytokines exert various non-redundant biological activities, although binding to the same receptor. Persistent viral infections are often characterized by increased IFN signatures implicating a potential role of type I IFNs in disease pathogenesis. Using the well-established Friend retrovirus (FV) mouse model, we compared the therapeutic efficacy of IFNα11 and IFNß in acute and chronic retroviral infection. We observed a strong antiviral activity of both IFNs during acute FV infection, whereas only IFNα11 and not IFNß could also control persistent FV infection. The therapeutic treatment with IFNα11 induced the expression of antiviral IFN-stimulated genes (ISG) and improved cytotoxic T cell responses. Finally, dysfunctional CD8+ T cells solely regained cytotoxicity after IFNα11 treatment. Our data provide evidence for opposing activities of type I IFNs during chronic retroviral infections. IFNß was shown to be involved in immune dysfunction in chronic infections, whereas IFNα11 had a strong antiviral potential and reactivated exhausted T cells during persistent retroviral infection. In contrast, during acute infection, both type I IFNs were able to efficiently suppress FV replication.