Latent human herpesvirus 6 is reactivated in CAR T cells.

Cell therapies have yielded durable clinical benefits for patients with cancer, but the risks associated with the development of therapies from manipulated human cells are understudied. For example, we lack a comprehensive understanding of the mechanisms of toxicities observed in patients receiving T cell therapies, including recent reports of encephalitis caused by reactivation of human herpesvirus 6 (HHV-6)1. Here, through petabase-scale viral genomics mining, we examine the landscape of human latent viral reactivation and demonstrate that HHV-6B can become reactivated in cultures of human CD4+ T cells. Using single-cell sequencing, we identify a rare population of HHV-6 'super-expressors' (about 1 in 300-10,000 cells) that possess high viral transcriptional activity, among research-grade allogeneic chimeric antigen receptor (CAR) T cells. By analysing single-cell sequencing data from patients receiving cell therapy products that are approved by the US Food and Drug Administration2 or are in clinical studies3-5, we identify the presence of HHV-6-super-expressor CAR T cells in patients in vivo. Together, the findings of our study demonstrate the utility of comprehensive genomics analyses in implicating cell therapy products as a potential source contributing to the lytic HHV-6 infection that has been reported in clinical trials1,6-8 and may influence the design and production of autologous and allogeneic cell therapies.

Retrospective study on the association of human herpesvirus reactivation with severe DRESS: A description of blood and skin reactivations.

BACKGROUND: Drug Reaction with Eosinophilia and Systemic Symptoms (DRESS) is a severe adverse event (mortality of 10%). Its pathophysiology involves herpesviruses, particularly HHV-6, but the exact mechanisms are still poorly understood. OBJECTIVE: To describe severe cases of DRESS and especially their association with herpesvirus reactivation. METHODS: This study was a multicentre case series conducted between 2007 and 2021 at five University Hospital Centres in France. The study included patients who had severe DRESS, which was defined as death, transfer to the intensive care unit (ICU), or severe damage to internal organs. We excluded patients without blood PCR sample, without a drug formally attributed or with RegiSCAR score < 6. We collected data on severity, causative drug, associated visceral damage and results of viral blood PCRs. HHV-6 reactivation was studied in skin biopsies by detection of small non-coding transcripts (HHV-6 miR-aU14) and a late viral protein (GP82/105). RESULTS: Fifty-two patients were included (29 female, median age 62, interquartile range (IQR) [37;72]). Eight patients (15%) died, 13 (27%) were admitted to ICU. Most patients (n = 34; 65%) had multisystem involvement: most frequent was liver (n = 46; 88%), then renal failure (n = 24; 46%). Forty patients (77%) had at least one blood viral reactivation among HHV-6, EBV or CMV, of which 21 (53%) had at least two. Median time of blood HHV-6 reactivation was 24 days (IQR [20;35]). HHV-6 reactivation was demonstrated in 15 out of 20 skin biopsies, with a median time of 11 days [9;17]. CONCLUSIONS: We confirmed the high rate of HHV-6 reactivation in severe DRESS and demonstrated cutaneous HHV-6 reactivation using small non-coding transcripts (HHV-6 miR-aU14), which preceded viral PCR positivity in blood. These results suggest that HHV-6 reactivation during DRESS may start in skin. Furthermore, search for miR-aU14 in skin biopsy could become a useful diagnostic tool for early detection of HHV-6 reactivation.

Immunopathogenic Insights on Preferential Human Herpesvirus-6 Reactivation in Drug Rash With Eosinophilia and Systemic Symptoms: A Scoping Review.

INTRODUCTION: Human herpesvirus-6 (HHV-6) is a ubiquitous lymphotropic betaherpesvirus that can reactivate in drug rash with eosinophilia and systemic symptoms (DRESS). Despite recent publications advancing our understanding of HHV-6 in DRESS, the exact role of HHV-6 in disease pathogenesis remains unclear. METHODS: A scoping review with the PubMed query "(HHV 6 AND (drug OR DRESS OR DIHS)) OR (HHV6 AND (drug OR DRESS OR DIHS))" was conducted in accordance with PRISMA guidelines. Articles containing original data on at least one DRESS patient with HHV-6 testing were included. RESULTS: Our search returned a total of 373 publications, of which 89 met eligibility criteria. HHV-6 reactivation occurred in 63% of DRESS patients (n = 748), which was significantly more often than other herpesviruses. HHV-6 reactivation was associated with worse outcomes and greater severity in controlled studies. Case reports have demonstrated sometimes fatal HHV-6-related multi-organ involvement. Temporally, HHV-6 reactivation typically occurs 2 to 4 weeks after DRESS onset and has been linked to markers of immunologic signaling, such as OX40 (CD134), an HHV-6 entry receptor. Efficacy of antiviral or immunoglobulin treatment has only been demonstrated anecdotally, and steroid use may affect HHV-6 reactivation. CONCLUSION: HHV-6 is implicated in DRESS more than in any other dermatologic condition. It is still unclear whether HHV-6 reactivation is cause or consequence of DRESS dysregulation. Similar pathogenic mechanisms precipitated by HHV-6 in other contexts may be relevant in DRESS. Future randomized controlled studies to assess effects of viral suppression on clinical outcomes is needed.

Human herpesvirus 6-specific T-cell immunity in allogeneic hematopoietic stem cell transplant recipients.

Human herpesvirus 6 (HHV-6) can reactivate after allogeneic hematopoietic stem cell transplant (allo-HSCT) and may lead to severe symptoms. HHV-6-specific immune responses after HSCT are largely unexplored. We conducted a prospective observational study on 208 consecutive adult patients who received allo-HSCT to investigate HHV-6 reactivations and specific immune responses. Interferon gamma-producing HHV-6-specific T cells were quantified using enzyme-linked immunospot assay (ELISpot). HHV-6 reactivation occurred in 63% of patients, at a median of 25 days from allo-HSCT. Only 40% of these presented a clinically relevant infection, defined by the presence of classical HHV-6 end-organ diseases (EODs), based on European Conference on Infections in Leukaemia (ECIL) guidelines, and other possible HHV6-related EODs. Using multivariate analysis, we identified risk factors for HHV-6 reactivation: previous allo-HSCT, posttransplant cyclophosphamide (PT-Cy), and time-dependent steroids introduction. The use of PT-Cy and steroids were associated with clinically relevant infections, whereas higher CD3+ cell counts seemed to be protective. Interestingly, circulating HHV-6-specific T cells were significantly higher in patients with reactivated virus. Moreover, HHV-6-specific T-cell responses, quantified at >4 days after the first viremia detection, predicted clinically relevant infections (P < .0001), with higher specificity (93%) and sensitivity (79%) than polyclonal CD3+ cells per µL. Overall survival and transplant-related mortality were not affected by time-dependent HHV-6 reactivation, whereas a significant association was observed between clinically relevant infections and acute graft-versus-host disease. These results shed light on the role of HHV-6 in allo-HSCT and may affect HHV-6 monitoring and treatment.

Once-Daily Foscarnet Is Effective for Human Herpesvirus 6 Reactivation after Hematopoietic Stem Cell Transplantation.

Human herpesvirus 6 (HHV-6) reactivation is common after allogeneic hematopoietic stem cell transplantation (allo-HSCT) and is associated with higher mortality and increased transplantation-related complications. We hypothesized that preemptive treatment with a short course of foscarnet at a lower cutpoint of plasma HHV-6 viral load would be effective in treating early HHV-6 reactivation, preventing complications and precluding hospitalization of these patients. We reviewed outcomes of adult patients (age ≥18 years) who received preemptive treatment with once-daily foscarnet 60 to 90 mg/kg for 7 days for HHV-6 reactivation after allo-HSCT at our institution between May 2020 and November 2022. Plasma HHV-6 viral load was monitored by quantitative PCR twice monthly in the first 100 days post-transplantation and twice weekly after reactivation until resolution. Eleven patients with a median age of 46 years (range, 23 to 73 years) were included in the analysis. HSCT was performed with a haploidentical donor in 10 patients and with an HLA-matched related donor in 1 patient. The most common diagnosis was acute leukemia (9 patients). Myeloablative and reduced-intensity conditioning regimens were used in 4 and 7 patients, respectively. Ten of the 11 patients received post-transplantation cyclophosphamide-based graft-versus-host disease prophylaxis. The median follow-up was 440 days (range, 174 to 831 days), and the median time to HHV-6 reactivation was 22 days post-transplantation (range, 15 to 89 days). The median viral load at first reactivation was 3,100 copies/mL (range, 210 to 118,000 copies/mL), and the median peak viral load was 11,300 copies/mL (range, 600 to 983,000 copies/mL). All patients received a short course of foscarnet at either 90 mg/kg/day (n = 7) or 60 mg/kg/day (n = 4). In all patients, plasma HHV-6 DNA was undetectable at completion of 1 week of treatment. No HHV-6 encephalitis or pneumonitis occurred. All patients achieved neutrophil and platelet engraftment after a median of 16 days (range, 8 to 22 days) and 26 days (range, 14 to 168 days), respectively, with no secondary graft failure. No complications related to foscarnet administration were noted. One patient with very high HHV-6 viremia had recurrent reactivation and received a second course of foscarnet as an outpatient. A short course of once-daily foscarnet is effective in treating early HHV-6 reactivation post-transplantation and may reduce the incidence of HHV-6-related and treatment-related complications and preclude hospitalization in these patients.

Human herpesvirus-6 in hematopoietic stem cell transplant recipients: a prospective cohort study in Egypt.

BACKGROUND: Immunocompromised patients face reactivation of latent viruses that increase the risk of morbidity. AIM: The study aimed to detect human herpes virus 6 (HHV-6) reactivation among allogeneic (allo) and autologous (auto) hematopoietic stem cell transplant (HSCT) recipients and to correlate potentially attributed clinical manifestations to HHV-6 DNA plasma level. METHODS: A prospective study included all (forty) patients undergoing allo and auto-HSCT from Jan 2020 till June 2022. Plasma samples were collected for HHV-6 serology, and for HHV-6 quantitative PCR at post-transplantation weeks 2, 4, 6. Demographic and clinical data were recorded. RESULTS: Out of 40 peripheral blood stem cell transplant (PBSCT) recipients, 34 (85%) were HHV-6 IgG positive pre-HSCT. Of which, fourteen patients (14/34, 41.2%) showed positive HHV-6 DNaemia. HHV-6 DNAemia (15/40, 37.5%) was significantly higher among allo (8/12, 66.7%) versus auto (7/28, 25%) HSCT recipients (p = 0.030). Patients with HHV-6 DNAemia developed fever, delayed engraftment and bone marrow suppression in 6/15, 40%, thrombocytopenia (5/15, 33.3%), rash and pneumonitis (2/15, 13.3%), acute GVHD (aGVHD) (1/15, 6.7%). HHV-6 DNAemia ranged from 101 to 102,000 copies/mL. Univariate analysis identified conditioning with busulfan-cyclophosphamide as a significant risk (p = 0.043), while receiving BEAM protocol was a protective factor (p = 0.045). In multivariate analysis, receiving BEAM protocol retained significance (p = 0.040). CONCLUSION: Frequent HHV-6 reactivation was detected after HSCT, especially in allo-HSCT recipients with clinical manifestations which could not be otherwise explained. To our best knowledge this is the first study of HHV6 reactivation in HSCT recipients from Egypt. Raising awareness for HHV-6 reactivation manifestations and screening in HSCT recipients could be lifesaving.

Studying the Interactions of U24 from HHV-6 in Order to Further Elucidate Its Potential Role in MS.

A number of studies have suggested that human herpesvirus 6A (HHV-6A) may play a role in multiple sclerosis (MS). Three possible hypotheses have been investigated: (1) U24 from HHV-6A (U24-6A) mimics myelin basic protein (MBP) through analogous phosphorylation and interaction with Fyn-SH3; (2) U24-6A affects endocytic recycling by binding human neural precursor cell (NPC) expressed developmentally down-regulated protein 4-like WW3* domain (hNedd4L-WW3*); and (3) MS patients who express Killer Cell Immunoglobulin Like Receptor 2DL2 (KIR2DL2) on natural killer (NK) cells are more susceptible to HHV-6 infection. In this contribution, we examined the validity of these propositions by investigating the interactions of U24 from HHV-6B (U24-6B), a variant less commonly linked to MS, with Fyn-SH3 and hNedd4L-WW3* using heteronuclear single quantum coherence (HSQC) nuclear magnetic resonance (NMR) titrations and isothermal titration calorimetry (ITC). In addition, the importance of phosphorylation and the specific role of U24 in NK cell activation in MS patients were examined. Overall, the findings allowed us to shed light into the models linking HHV-6 to MS and the involvement of U24.

ATF1 Restricts Human Herpesvirus 6A Replication via Beta Interferon Induction.

The stimulus-induced cAMP response element (CRE)-binding protein (CREB) family of transcription factors bind to CREs to regulate diverse cellular responses, including proliferation, survival, and differentiation. Human herpesvirus 6A (HHV-6A), which belongs to the Betaherpesvirinae subfamily, is a lymphotropic herpesvirus frequently found in patients with neuroinflammatory diseases. Previous reports implicated the importance of CREs in the HHV-6A life cycle, although the effects of the binding of transcription factors to CREs in viral replication have not been fully elucidated. In this study, we analyzed the role of the CREB family of transcription factors during HHV-6A replication. We found that HHV-6A infection enhanced phosphorylation of the CREB family members CREB1 and activating transcription factor 1 (ATF1). Knockout (KO) of CREB1 or ATF1 enhanced viral gene expression and viral replication. The increase in viral yields in supernatants from ATF1-KO cells was greater than that in supernatants from CREB1-KO cells. Transcriptome sequencing (RNA-seq) analysis showed that sensors of the innate immune system were downregulated in ATF1-KO cells, and mRNAs of beta interferon (IFN-ß) and IFN-regulated genes were reduced in these cells infected with HHV-6A. IFN-ß treatment of ATF1-KO cells reduced progeny viral yields significantly, suggesting that the enhancement of viral replication was caused by a reduction of IFN-ß. Taken together, our results suggest that ATF1 is activated during HHV-6A infection and restricts viral replication via IFN-ß induction. IMPORTANCE Human herpesvirus 6A (HHV-6A) is a ubiquitous herpesvirus implicated in Alzheimer's disease, although its role in its pathogenesis has not been confirmed. Here, we showed that the transcription factor ATF1 restricts HHV-6A replication, mediated by IFN-ß induction. Our study provides new insights into the role of ATF1 in innate viral immunity and reveals the importance of IFN-ß for regulation of HHV-6A replication, which possibly impairs HHV-6A pathogenesis.

Epstein-Barr Virus and Human Herpesvirus-6 Reactivation in Acute COVID-19 Patients.

Beyond their pulmonary disease, many COVID-19 patients experience a complex constellation of characteristics, including hyperinflammatory responses, autoimmune disorders, and coagulopathies. However, the pathogenesis of these aspects of COVID-19 is obscure. More than 90% of people are latently infected with the lymphotropic herpesviruses Epstein-Barr Virus (EBV) and/or Human Herpesvirus-6 (HHV-6). Some of the inflammatory features of COVID-19 resemble clinical syndromes seen during EBV and HHV-6 infection, and these latent viruses can be reactivated by inflammatory mediators. We hypothesized that EBV and HHV-6 reactivation might be a common feature of early COVID-19, particularly in patients with more inflammation. We tested for EBV and HHV-6 reactivation in 67 patients acutely hospitalized with COVID-19 using previously validated quantitative PCR assays on the plasma. In our cohort, we found that 15/67 (22.4%) patients had detectable EBV and 3/67 (4.5%) had detectable HHV-6. This frequency of activation is somewhat more than the frequency reported for some healthy cohorts, such as blood donors and other healthy control cohorts. There was no association between EBV or HHV-6 and markers indicative of more inflammatory disease. We conclude that EBV and HHV-6 activation at about day 7 of hospitalization occurred in a modest fraction of our cohort of COVID-19 patients and was not associated with high levels of inflammation. In the modest fraction of patients, EBV and HHV-6 reactivation could contribute to some features of acute disease and pre-disposition to post-acute sequelae in a subset of patients.

Differential Impacts of HHV-6A versus HHV-6B Infection in Differentiated Human Neural Stem Cells.

Within the family Herpesviridae, sub-family ß-herpesvirinae, and genus Roseolovirus, there are only three human herpesviruses that have been described: HHV-6A, HHV-6B, and HHV-7. Initially, HHV-6A and HHV-6B were considered as two variants of the same virus (i.e., HHV6). Despite high overall genetic sequence identity (~90%), HHV-6A and HHV-6B are now recognized as two distinct viruses. Sequence divergence (e.g., >30%) in key coding regions and significant differences in physiological and biochemical profiles (e.g., use of different receptors for viral entry) underscore the conclusion that HHV-6A and HHV-6B are distinct viruses of the ß-herpesvirinae. Despite these viruses being implicated as causative agents in several nervous system disorders (e.g., multiple sclerosis, epilepsy, and chronic fatigue syndrome), the mechanisms of action and relative contributions of each virus to neurological dysfunction are unclear. Unresolved questions regarding differences in cell tropism, receptor use and binding affinity (i.e., CD46 versus CD134), host neuro-immunological responses, and relative virulence between HHV-6A versus HHV-6B prevent a complete characterization. Although it has been shown that both HHV-6A and HHV-6B can infect glia (and, recently, cerebellar Purkinje cells), cell tropism of HHV-6A versus HHV-6B for different nerve cell types remains vague. In this study, we show that both viruses can infect different nerve cell types (i.e., glia versus neurons) and different neurotransmitter phenotypes derived from differentiated human neural stem cells. As demonstrated by immunofluorescence, HHV-6A and HHV-6B productively infect VGluT1-containing cells (i.e., glutamatergic neurons) and dopamine-containing cells (i.e., dopaminergic neurons). However, neither virus appears to infect GAD67-containing cells (i.e., GABAergic neurons). As determined by qPCR, expression of immunological factors (e.g., cytokines) in cells infected with HHV-6A versus HHV6-B also differs. These data along with morphometric and image analyses of infected differentiated neural stem cell cultures indicate that while HHV-6B may have greater opportunity for transmission, HHV-6A induces more severe cytopathic effects (e.g., syncytia) at the same post-infection end points. Cumulatively, results suggest that HHV-6A is more virulent than HHV-6B in susceptible cells, while neither virus productively infects GABAergic cells. Consistency between these in vitro data and in vivo experiments would provide new insights into potential mechanisms for HHV6-induced epileptogenesis.

Markers of Epstein-Barr virus and Human Herpesvirus-6 infection and multiple sclerosis clinical progression.

BACKGROUND: Infections with Epstein-Barr virus (EBV) and human herpesvirus-6 (HHV-6) have been implicated in multiple sclerosis (MS) onset but little work has studied their relationships in early disease. OBJECTIVE: Evaluate associations between markers of EBV and HHV-6 infection/reactivation and MS conversion, relapse and EDSS/MSSS amongst 205 CIS participants with EBV/HHV-6 data followed over 5 years. METHOD: Baseline serological and viral load measures of EBV and HHV-6 exposure/reactivation were measured and infectious mononucleosis (IM) history recorded. Conversion to MS and relapses were assessed annually, and EDSS/MSSS measured at 5-year review. Determinants of MS conversion and relapse assessed by Cox regression, and disability progression by linear regression. RESULTS: IM history showed a strong positive trend with higher relapse risk (aHR=1.45,95%CI=0.97-2.16) but was not associated with MS conversion (aHR=0.92,95%CI=0.57-1.48). Anti-HHV-6 IgG titre>40 also showed strong positive trends with higher relapse (aHR=1.61,95%CI=0.99-2.63) and MS conversion risks (aHR=1.48,95%CI=0.89-2.46). Anti-HHV-6 IgG titre≥640 was significantly associated with higher MSSS (0.15(95%CI=0.00,0.30) and also showed a strong positive trend with higher EDSS 0.10(95%CI=-0.02,0.21). HHV-6 DNA detection showed strong positive trends with 83%(95%CI=-6-357) and 77%(95%CI=-4-328) higher MS conversion and relapse risk. Anti-EBV-EA-D IgG titre was associated with a lower annualised disability progression by EDSS (ptrend=0.037) and also showed strong positive trend with higher MSSS (ptrend=0.053). No associations were seen for other serological or viral load markers. CONCLUSION: Overall, our data provides evidence that higher HHV-6 IgG was associated with increased risk of MS conversion and relapse but of borderline significance, and greater annualised disability progression, while that for EBV was more limited.

The Spontaneous Course of Human Herpesvirus 6 DNA-Associated Myocarditis and the Effect of Immunosuppressive Intervention.

INTRODUCTION: This study investigated the spontaneous clinical course of patients with endomyocardial biopsy (EMB)-proven lymphocytic myocarditis and cardiac human herpesvirus 6 (HHV6) DNA presence, and the effectiveness of steroid-based intervention in HHV6-positive patients. RESULTS: 756 heart failure (HF) patients underwent an EMB procedure to determine the underlying cause of unexplained HF. Low levels of HHV6 DNA, detectable by nested PCR only, were found in 10.4% of the cases (n = 79) of which 62% (n = 49) showed myocardial inflammation. The spontaneous course of patients with EMB-proven HHV6 DNA-associated lymphocytic myocarditis (n = 26) showed significant improvements in the left ventricular ejection fraction (LVEF) and clinical symptoms, respectively, in 15/26 (60%) patients, 3-12 months after disease onset. EMB mRNA expression of components of the NLRP3 inflammasome pathway and protein analysis of cardiac remodeling markers, analyzed by real-time PCR and MALDI mass spectrometry, respectively, did not differ between HHV6-positive and -negative patients. In another cohort of patients with ongoing symptoms related to lymphocytic myocarditis associated with cardiac levels of HHV6-DNA copy numbers <500 copies/µg cardiac DNA, quantified by real-time PCR, the efficacy and safety of steroid-based immunosuppression for six months was investigated. Steroid-based immunosuppression improved the LVEF (≥5%) in 8/10 patients and reduced cardiac inflammation in 7/10 patients, without an increase in cardiac HHV6 DNA levels in follow-up EMBs. CONCLUSION: Low HHV6 DNA levels are frequently detected in the myocardium, independent of inflammation. In patients with lymphocytic myocarditis with low levels of HHV6 DNA, the spontaneous clinical improvement is nearby 60%. In selected symptomatic patients with cardiac HHV6 DNA copy numbers less than 500 copies/µg cardiac DNA and without signs of an active systemic HHV6 infection, steroid-based therapy was found to be effective and safe. This finding needs to be further confirmed in large, randomized trials.

CD46 Isoforms Influence the Mode of Entry by Human Herpesvirus 6A/B in T Cells.

CD46 is a receptor for human herpesvirus 6A (HHV-6A) and is in some cells also important for infection with HHV-6B. CD46 has several isoforms of which the most commonly expressed can be distinguished by expression of a BC domain or a C domain in a serine-threonine-proline-rich (STP) extracellular region. Using a SupT1 CD46 CRISPR-Cas9 knockout model system reconstituted with specific CD46 isoforms, we demonstrated that HHV-6A infection was more efficient when BC isoforms were expressed as opposed to C isoforms, measured by higher levels of intracellular viral transcripts and recovery of more progeny virus. Although the B domain contains several O-glycosylations, mutations of Ser and Thr residues did not prevent infection with HHV-6A. The HHV-6A infection was blocked by inhibitors of clathrin-mediated endocytosis. In contrast, infection with HHV-6B was preferentially promoted by C isoforms mediating fusion-from-without, and this infection was less affected by inhibitors of clathrin-mediated endocytosis. Taken together, HHV-6A preferred BC isoforms, mediating endocytosis, whereas HHV-6B preferred C isoforms, mediating fusion-from-without. This demonstrates that the STP region of CD46 is important for regulating the mode of infection in SupT1 cells and suggests an epigenetic regulation of the host susceptibility to HHV-6A and HHV-6B infection. IMPORTANCE CD46 is the receptor used by human herpesvirus 6A (HHV-6A) during infection of T cells, but it is also involved in infection of certain T cells by HHV-6B. The gene for CD46 allows expression of several variants of CD46, known as isoforms, but whether the isoforms matter for infection of T cells is unknown. We used a genetic approach to delete CD46 from T cells and reconstituted them with separate isoforms to study them individually. We expressed the isoforms known as BC and C, which are distinguished by the potential inclusion of a B domain in the CD46 molecule. We demonstrate that HHV-6A prefers the BC isoform to infect T cells, and this occurs predominantly by clathrin-mediated endocytosis. In contrast, HHV-6B prefers the C isoform and infects predominantly by fusion-from-without. Thus, CD46 isoforms may affect susceptibility of T cells to infection with HHV-6A and HHV-6B.

Unique Challenges to Diagnosing Human Herpesvirus-6 (HHV-6) Encephalitis Following Post-Hematopoietic Stem Cell Transplant: A Case and Brief Review.

A patient with an ultimate diagnosis of human herpesvirus-6 (HHV-6) encephalitis developed central nervous system (CNS) symptoms 13 days after undergoing myeloablative haploidentical allogeneic hematopoietic stem cell transplant (HSCT). Due to the patient's body habitus, magnetic resonance (MR) imaging was not obtained until the onset of retrograde amnesia on day +24. MR imaging and other clinical findings eliminated all skepticism of HHV-6 encephalitis and HHV-6 antivirals were initiated on day +28, leading to gradual recovery. This case demonstrates some of the factors that may complicate the diagnosis of post-alloHSCT HHV-6 encephalitis. Because HHV-6 encephalitis and viremia can occur without warning, a single negative study should not exclude future development, especially if CNS symptoms are present. Acute graft-versus-host disease and cord blood transplantation are both significant risk factors for HHV-6 encephalitis. Human leukocyte antigen (HLA) mismatch, engraftment complications, or certain HLA alleles have also been associated with HHV-6 encephalitis. Chromosomally integrated HHV-6 must also be ruled out to prevent inappropriate and potentially harmful administration of antivirals. Due to the severe short- and long-term sequelae of HHV-6 encephalitis, appropriate treatment should be administered as soon as possible.

Anti-Human Herpesvirus 6 A/B Antibodies Titers Correlate With Multiple Sclerosis-Associated Retrovirus Envelope Expression.

Human endogenous retrovirus W family envelope proteins (pHERV-W ENV/syncytin-1) have been repeatedly associated with multiple sclerosis (MS). Here, we have focused on the study of pHERV-W ENV/syncytin-1 expression levels in MS patients (relapsing and progressive forms) and in healthy donors (HD) and on exploring their possible relationship with Epstein-Barr virus (EBV) and human herpesvirus-6A/B (HHV-6A/B). We included blood samples from 101 MS patients and 37 HD to analyze antiviral antibody titers by ELISA and pHERV-W ENV/syncytin-1 expression levels by flow cytometry as well as by qPCR. Patients with relapsing MS forms showed significantly higher pHERV-W ENV/syncytin-1 protein and gene expression levels than HD. Progressive MS patients also showed significantly higher protein and gene expression levels than both HD and relapsing MS patients. Regarding antiviral antibodies titers, anti-HHV-6A/B IgM levels were positively correlated with pHERV-W ENV/syncytin-1 protein expression levels in patients with relapsing MS, while in the progressive forms patients this correlation was found with anti-HHVA/B IgG levels. Therefore, pHERV-W ENV could be involved in MS pathogenesis, playing a role in relapsing and progressive forms. Besides, anti-HHV-6A/B antibodies positively correlated with pHERV-W ENV expression. Further studies are needed to better understand this possible relationship.

The HHV-6A Proteins U20 and U21 Target NKG2D Ligands to Escape Immune Recognition.

The coevolution of the human immune system and herpesviruses led to the emergence and diversification of both cellular danger molecules recognized by immune cells on the one hand and viral countermeasures that prevent the expression of these proteins on infected cells on the other. There are eight ligands for the activating receptor NKG2D in humans - MICA, MICB, ULBP1-6. Several of them are induced and surface-expressed on herpesvirus-infected cells to serve as danger signals to activate the immune system. Therefore, these ligands are frequently targeted for suppression by viral immune evasion mechanisms. Mechanisms to downregulate NKG2D ligands and thereby escape immune recognition have been identified in all other human herpesviruses (HHV), except for HHV-6A. In this study, we identify two HHV-6A encoded immunoevasins, U20 and U21, which suppress the expression of the NKG2D ligands ULBP1 and ULBP3, respectively, during infection. Additionally, MICB is targeted by a so far unexplored viral protein. Due to the diminished NKG2D ligand surface expression on infected cells, recognition of HHV-6A infected cells by innate immune cells is impaired. Importantly, our study indicates that immune escape mechanisms between the related herpesviruses HHV-6A and HHV-6B are evolutionary conserved as the same NKG2D ligands are targeted. Our data contribute an additional piece of evidence for the importance of the NKG2D receptor - NKG2D ligand axis during human herpesvirus infections and sheds light on immune evasion mechanisms of HHV-6A.

Human Herpesviruses 6A and 6B in Reproductive Diseases.

Human herpesviruses 6A (HHV-6A) and human herpesvirus 6B (HHV-6B)-collectively, HHV-6A/B-are recently-discovered but ancient human viruses. The vast majority of people acquire one or both viruses, typically very early in life, producing an ineradicable lifelong infection. The viruses have been linked to several neurological, pulmonary and hematological diseases. In early human history, the viruses on multiple occasions infected a germ cell, and integrated their DNA into a human chromosome. As a result, about 1% of humans are born with the full viral genome present in every cell, with uncertain consequences for health. HHV-6A may play a role in 43% of cases of primary unexplained infertility. Both the inherited and acquired viruses may occasionally trigger several of the factors that are important in the pathogenesis of preeclampsia. Transplacental infection occurs in 1-2% of pregnancies, with some evidence suggesting adverse health consequences for the child. While emerging knowledge about these viruses in reproductive diseases is not sufficient to suggest any changes in current practice, we write this review to indicate the need for further research that could prove practice-changing.

High incidence of Epstein-Barr virus, cytomegalovirus, and human-herpes virus-6 reactivations in critically ill patients with COVID-19.

BACKGROUND: Systemic reactivation of herpesviruses may occur in intensive care unit (ICU) patients and is associated with morbidity and mortality. Data on severe Coronavirus disease-19 (COVID-19) and concomitant reactivation of herpesviruses are lacking. METHODS: We selected patients admitted to ICU for confirmed COVID-19 who underwent systematic testing for Epstein-Barr virus (EBV), cytomegalovirus (CMV) and human-herpes virus-6 (HHV-6) DNAemia while in the ICU. We retrospectively analysed frequency, timing, duration and co-occurrence of viral DNAemia. RESULTS: Thirty-four patients were included. Viremia with EBV, CMV, and HHV-6 was detected in 28 (82%), 5 (15%), and 7 (22%) patients, respectively. EBV reactivation occurred early after ICU admission and was associated with longer ICU length-of-stay. CONCLUSIONS: While in the ICU, critically ill patients with COVID-19 are prone to develop reactivations due to various types of herpesviruses.

IRAK4 Deficiency Presenting with Anti-NMDAR Encephalitis and HHV6 Reactivation.

IRAK4 deficiency is an inborn error of immunity predisposing patients to invasive pyogenic infections. Currently, there is no established simple assay that enables precise characterization of IRAK4 mutant alleles in isolation. Anti-N-methyl-D-aspartate receptor (NMDAR) encephalitis is an autoimmune condition that is characterized by psychiatric symptoms, involuntary movement, seizures, autonomic dysfunction, and central hypoventilation. It typically occurs in adult females associated with tumors. Only a few infantile cases with anti-NMDAR encephalitis have been so far reported. We identified a 10-month-old boy with IRAK4 deficiency presenting with anti-NMDAR encephalitis and human herpes virus 6 (HHV6) reactivation. The diagnosis of IRAK4 deficiency was confirmed by the identification of compound heterozygous mutations c.29_30delAT (p.Y10Cfs*9) and c.35G>C (p.R12P) in the IRAK4 gene, low levels of IRAK4 protein expression in peripheral blood, and defective fibroblastic cell responses to TLR and IL-1 (TIR) agonist. We established a novel NF-κB reporter assay using IRAK4-null HEK293T, which enabled the precise evaluation of IRAK4 mutations. Using this system, we confirmed that both novel mutations identified in the patient are deleterious. Our study provides a new simple and reliable method to analyze IRAK4 mutant alleles. It also suggests the possible link between inborn errors of immunity and early onset anti-NMDAR encephalitis.

The Presence of Human Herpesvirus 6 in the Brain in Health and Disease.

The human herpesvirus 6 (HHV-6) -A and -B are two dsDNA beta-herpesviruses infectingalmost the entire worldwide population. These viruses have been implicated in multipleneurological conditions in individuals of various ages and immunological status, includingencephalitis, epilepsy, and febrile seizures. HHV-6s have also been suggested as playing a role inthe etiology of neurodegenerative diseases such as multiple sclerosis and Alzheimer's disease. Theapparent robustness of these suggested associations is contingent on the accuracy of HHV-6detection in the nervous system. The effort of more than three decades of researching HHV-6 in thebrain has yielded numerous observations, albeit using variable technical approaches in terms oftissue preservation, detection techniques, sample sizes, brain regions, and comorbidities. In thisreview, we aimed to summarize current knowledge about the entry routes and direct presence ofHHV-6 in the brain parenchyma at the level of DNA, RNA, proteins, and specific cell types, inhealthy subjects and in those with neurological conditions. We also discuss recent findings relatedto the presence of HHV-6 in the brains of patients with Alzheimer's disease in light of availableevidence.