Acute Promyelocytic Leukemia With Torque Teno Mini Virus::RARA Fusion: An Approach to Screening and Diagnosis.

Acute promyelocytic leukemia (APL) with variant RARA translocation is linked to over 15 partner genes. Recent publications encompassing 6 cases have expanded the spectrum of RARA partners to torque teno mini virus (TTMV). This entity is likely underrecognized due to the lack of clinician and pathologist familiarity, inability to detect the fusion using routine testing modalities, and informatic challenges in its recognition within next-generation sequencing (NGS) data. We describe a clinicopathologic approach and provide the necessary tools to screen and diagnose APL with TTMV::RARA using existing clinical DNA- or RNA-based NGS assays, which led to the identification of 4 cases, all without other known cytogenetic/molecular drivers. One was identified prospectively and 3 retrospectively, including 2 from custom automated screening of multiple data sets (50,257 cases of hematopoietic malignancy, including 4809 acute myeloid leukemia/myeloid sarcoma/APL cases). Two cases presented as myeloid sarcoma, including 1 with multiple relapses after acute myeloid leukemia-type chemotherapy and hematopoietic stem cell transplant. Two cases presented as leukemia, had a poor response to induction chemotherapy, but achieved remission upon reinduction (including all-trans retinoic acid in 1 case) and subsequent hematopoietic stem cell transplant. Neoplastic cells demonstrated features of APL including frequent azurophilic granules and dim/absent CD34 and HLA-DR expression. RARA rearrangement was not detected by karyotype or fluorescent in situ hybridization. Custom analysis of NGS fusion panel data identified TTMV::RARA rearrangements and, in the prospectively identified case, facilitated monitoring in sequential bone marrow samples. APL with TTMV::RARA is a rare leukemia with a high rate of treatment failure in described cases. The diagnosis should be considered in leukemias with features of APL that lack detectable RARA fusions and other drivers, and may be confirmed by appropriate NGS tests with custom informatics. Incorporation of all-trans retinoic acid may have a role in treatment but requires accurate recognition of the fusion for appropriate classification as APL.

Detection of circulating tumor human papillomavirus DNA before diagnosis of HPV-positive head and neck cancer.

Human papillomavirus (HPV), most commonly HPV16, causes a growing subset of head and neck squamous cell carcinomas (HNSCCs), including the overwhelming majority of oropharynx squamous cell carcinomas in many developed countries. Circulating biomarkers for HPV-positive HNSCC may allow for earlier diagnosis, with potential to decrease morbidity and mortality. This case-control study evaluated whether circulating tumor HPV DNA (ctHPVDNA) is detectable in prediagnostic plasma from individuals later diagnosed with HPV-positive HNSCC. Cases were participants in a hospital-based research biobank with archived plasma collected ≥6 months before HNSCC diagnosis, and available archival tumor tissue for HPV testing. Controls were biobank participants without cancer or HPV-related diagnoses, matched 10:1 to cases by sex, race, age and year of plasma collection. HPV DNA was detected in plasma and tumor tissue using a previously validated digital droplet PCR-based assay that quantifies tumor-tissue-modified viral (TTMV) HPV DNA. Twelve HNSCC patients with median age of 68.5 years (range, 51-87 years) were included. Ten (83.3%) had HPV16 DNA-positive tumors. ctHPV16DNA was detected in prediagnostic plasma from 3 of 10 (30%) patients with HPV16-positive tumors, including 3 of 7 (43%) patients with HPV16-positive oropharynx tumors. The timing of the plasma collection was 19, 34 and 43 months before cancer diagnosis. None of the 100 matched controls had detectable ctHPV16DNA. This is the first report that ctHPV16 DNA is detectable at least several years before diagnosis of HPV16-positive HNSCC for a subset of patients. Further investigation of ctHPV16DNA as a biomarker for early diagnosis of HPV16-positive HNSCC is warranted.

Prevalence of human anelloviruses in Romanian healthy subjects and patients with common pathologies.

BACKGROUND: Human anelloviruses (TTV, TTMDV and TTMV) are at high prevalence all across the globe, having also a controversial disease-inducing potential. This study aimed to estimate the prevalence of anelloviral DNA in the Romanian human population and to investigate the association of infections with common pathologies in Romanian population. METHODS: After informed consent, blood samples were collected from 2000 subjects represented by: clinically healthy individuals (n = 701) and a group of patients with pathologies linked to low grade inflammation or alteration of carbohydrate metabolism (n = 1299). All samples were analysed for the presence of TTV, TTMDV and TTMV DNA by hemi-nested PCR. RESULTS: The prevalence of TTV, TTMDV and TTMV in the studied population was 68.2, 54.4%, respectively 40.1%, lower than the recent reports from other geographic regions. The three viral species were significantly more frequent in the group of patients compared to the healthy subjects and were associated with type 2 diabetes mellitus. The presence of anelloviral DNA was also associated with medical procedures (e.g. haemodialysis/transfusions, surgical procedures) and previous hepatitis A virus infection. Lifestyle choices related to alcohol consumption, smoking, physical activity and living environment were not associated with differences in distribution of the three viruses. CONCLUSION: Further evidence is needed to establish a correlation between infection with human anelloviruses and a pathology or group of pathologies.

Detection of a new species of torque teno mini virus from the gingival epithelium of patients with periodontitis.

We describe a novel species of torque teno mini virus called TTMV-204, which was isolated from the gingival epithelium of patients with periodontitis and characterized using viral metagenomics. The sequence of the full genome is 2824 nt in length. Phylogenetic analysis and genetic analyses show classic Betatorquevirus species organization with less than 40% amino acid similarity in ORF1. The prevalence of TTMV-204 in the periodontitis patient population was 18.75% (15/80), which was higher than in periodontally healthy individuals (10.00%, 10/80). However, the difference of the TTMV-204 prevalence between two groups was not statistically significant (p = 0.115). Further investigation is required to determine whether this new virus is associated with inflammation.

Case report of pediatric TTMV-related acute promyelocytic leukemia with central nervous system infiltration and rapid accumulation of RARA-LBD mutations.

TTMV::RARA is a recently reported fusion gene associated with acute promyelocytic leukemia (APL), caused by the integration of torque teno mini virus (TTMV) genomic fragments into the second intron of the RARA gene. Currently, there have been only six documented cases, with clinical presentations showing significant variability. Although initial responses to all-trans retinoic acid (ATRA) treatment may be observed in patients with TTMV::RARA-APL, the overall prognosis remains unfavorable among infrequent reported cases. This article presents a pediatric case that manifested as PML::RARA-negative APL with central nervous system involvement at onset. The patient experienced both intramedullary and extramedullary relapse one year after undergoing allogeneic hematopoietic stem cell transplantation. Upon identification as TTMV::RARA-APL and subsequent administration of two rounds of ATRA-based treatment, the patient rapidly developed multiple RARA ligand-binding domain mutations and demonstrated extensive resistance to ATRA and various other therapeutic interventions. Additionally, the patient experienced ARID1A mutant clone expansion and progressed MYC-targeted gene activation. This case represents the first documentation of extramedullary involvement at both the initial diagnosis and relapse stages, emphasizing the intricate clinical features and challenges associated with the rapid accumulation of multiple ATRA-resistant mutations in TTMV::RARA-APL, characterizing it as a distinct and complex sub-entity of atypical APL.

Over-Representation of Torque Teno Mini Virus 9 in a Subgroup of Patients with Myalgic Encephalomyelitis/Chronic Fatigue Syndrome: A Pilot Study.

Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a chronic disorder classified by the WHO as postviral fatigue syndrome (ICD-11 8E49 code). Diagnosing ME/CFS, often overlapping with fibromyalgia (FM), is challenging due to nonspecific symptoms and lack of biomarkers. The etiology of ME/CFS and FM is poorly understood, but evidence suggests viral infections play a critical role. This study employs microarray technology to quantitate viral RNA levels in immune cells from ME/CFS, FM, or co-diagnosed cases, and healthy controls. The results show significant overexpression of the Torque Teno Mini Virus 9 (TTMV9) in a subgroup of ME/CFS patients which correlate with abnormal HERV and immunological profiles. Increased levels of TTMV9 transcripts accurately discriminate this subgroup of ME/CFS patients from the other study groups, showcasing its potential as biomarker for patient stratification and the need for further research into its role in the disease. Validation of the findings seems granted in extended cohorts by continuation studies.

The Impact of First-Time SARS-CoV-2 Infection on Human Anelloviruses.

Members of the Anelloviridae family dominate the blood virome, emerging early in life. The anellome, representing the variety of anelloviruses within an individual, stabilizes by adulthood. Despite their supposedly commensal nature, elevated anellovirus concentrations under immunosuppressive treatment indicate an equilibrium controlled by immunity. Here, we investigated whether anelloviruses are sensitive to the immune activation that accompanies a secondary infection. As a model, we investigated 19 health care workers (HCWs) with initial SARS-CoV-2 infection, with blood sampling performed pre and post infection every 4 weeks in a 3-month-follow-up during the early 2020 COVID-19 pandemic. A concurrently followed control group (n = 27) remained SARS-CoV-2-negative. Serum anellovirus loads were measured using qPCR. A significant decrease in anellovirus load was found in the first weeks after SARS-CoV-2 infection, whereas anellovirus concentrations remained stable in the uninfected control group. A restored anellovirus load was seen approximately 10 weeks after SARS-CoV-2 infection. For five subjects, an in-time anellome analysis via Illumina sequencing could be performed. In three of the five HCWs, the anellome visibly changed during SARS-CoV-2 infection and returned to baseline in two of these cases. In conclusion, anellovirus loads in blood can temporarily decrease upon an acute secondary infection.

TTMV::RARA-positive acute promyelocytic leukemia with marrow necrosis and central nervous system involvement at disease recurrence.

Since the identification of the TTMV::RARA fusion in pediatric cases resembling acute promyelocytic leukemia (APL) by Astolfi et al. in 2021, several similar cases have been reported worldwide. In this report, we present a case of relapsed APL in an adolescent patient, who exhibited the TTMV::RARA fusion gene. This patient exhibited extensive central nervous system involvement and experienced bone marrow necrosis during disease recurrence. Despite achieving complete remission after re-induction chemotherapy, the patient experienced a rapid second relapse, highlighting the extremely aggressive nature of this subtype. These clinical manifestations contribute to the growing recognition of this rare disease.

The landscape of circulating tumor HPV DNA and TTMV-HPVDNA for surveillance of HPV-oropharyngeal carcinoma: systematic review and meta-analysis.

BACKGROUND: Human papilloma virus (HPV) related cancers of the oropharynx are rapidly increasing in incidence and may soon represent the majority of all head and neck cancers. Improved monitoring and surveillance methods are thus an urgent need in public health. MAIN TEXT: The goal is to highlight the current potential and limitations of liquid biopsy through a meta analytic study on ctHPVDNA and TTMV-HPVDNA. It was performed a Literature search on articles published until December 2023 using three different databases: MEDLINE, Embase, and Cochrane Library. Studies that evaluated post-treatment ctHPVDNA and TTMV-HPVDNA in patients with HPV + OPSCC, studies reporting complete data on the diagnostic accuracy in recurrence, or in which the number of true positives, false positives, true negatives, and false negatives was extractable, and methods of detection of viral DNA clearly defined. The meta-analysis was conducted following the Meta-analysis Of Observational Studies in Epidemiology (MOOSE) reporting guidelines. The aim of this meta-analysis was to evaluate the sensitivity, specificity, and accuracy of ctHPVDNA and TTMV by ddPCR to define its efficacy in clinical setting for the follow up of HPV-OPSCC. CONCLUSION: The 12 studies included in the meta-analysis provided a total of 1311 patients for the analysis (398 valuated with ctHPVDNA and 913 with TTMV-HPVDNA). Pooled sensitivity and specificity were 86% (95% CI: 78%-91%) and 96% (95% CI: 91%-99%), respectively; negative and positive likelihood ratios were 0.072 (95% CI: 0.057-0.093) and 24.7 (95% CI: 6.5-93.2), respectively; pooled DOR was 371.66 (95% CI: 179.1-918). The area under the curve (AUC) was 0.81 (95% CI, 0.67-0.91). Liquid biopsy for the identification of cell free DNA might identify earlier recurrence in HPV + OPSCC patients. At the present time, liquid biopsy protocol needs to be standardized and liquid biopsy cannot yet be used in clinical setting. In the future, a multidimensional integrated approach which links multiple clinical, radiological, and laboratory data will contribute to obtain the best follow-up strategies for the follow-up of HPV-OPSCC.

Rapid detection of human torque teno viruses using high-resolution melting analysis.

Torque teno viruses (TTVs) are recently discovered DNA viruses, with heterogeneous genomes, highly prevalent in populations worldwide. The species that infect humans are Torque teno virus (TTV), Torque teno midi virus (TTMDV) and Torque teno mini virus (TTMV). High-resolution melting analysis (HRMA) is a sensitive and effective method for genotyping and mutation scanning. Up to now, HRMA has not been utilized for detection of TTVs. The aim of this study was to asses if HRMA is suitable for detecting TTVs variants. DNA was extracted from the blood and saliva of 13 healthy subjects for method optimization. Additionally, saliva samples from 100 healthy individuals were collected for estimating the TTVs' prevalence. Viral DNA was amplified by heminested polymerase chain reaction (PCR). Second round amplicons were used for the HRMA. The samples were analyzed using two fluorescent dyes, SYBR (®) Green I and EvaGreen®. The prevalence values for TTV, TTMDV and TTMV were 71.0, 31.0 and 54.0%, respectively. The three major melting curve patterns corresponding to TTV, TTMDV and TTMV on HRMA can be easily distinguished regardless of kit used. Our results showed that HRMA is a rapid and efficient method of detecting human TTVs.

TTV and other anelloviruses: The astonishingly wide spread of a viral infection.

The broad family of viruses known as anelloviruses (AV) infects both humans and numerous animal species. They have a tiny, covalently closed single-stranded DNA genome and the astonishing capacity to infect a very high percentage of healthy and ill people with chronic infections that could last a lifetime. AV, and particularly the prototype Torquetenovirus, have established a successful interaction with the host's immune system and the rate at which they replicate is a gauge to measure overall immune function, even though many aspects of their life cycle and pathogenesis are still poorly understood.

Torque teno mini virus driven childhood acute promyelocytic leukemia: The third case report and sequence analysis.

In this manuscript, we report torque teno mini virus (TTMV) as a cause of acute promyelocytic leukemia (APL) lacking PML::RARA in a 3-year-old boy. Astolfi et al. firstly identified partial integration of the TTMV genome into RARA intron 2, which resulted in in-frame TTMV::RARA fusion in two APL-like pediatric cases without PML::RARA in November 2021. This fascinating report identified an unexpected exogenous genetic cause of APL and could be of great importance for diagnosing and managing APL. Here we report the third childhood APL-like case caused by TTMV integration and investigate the location and structure of the integrated TTMV sequence. These findings suggest TTMV::RARA is a recurrent cause of APL lacking PML::RARA. Considering the widespread prevalence of TTMV in the population, more TTMV::RARA positive APL-like cases might remain to be identified. Establishing a bioinformatic analysis strategy optimized for the highly variable TTMV genome sequence may facilitate the identification of TTMV::RARA by whole transcript sequencing. An effective PCR protocol to identify TTMV::RARA based on a profound analysis of the conservation of TTMV segments in the fusion transcript is also expected. Also, further investigation is needed to elucidate the oncogenic mechanisms of TTMV integration and the clinical features of TTMV::RARA positive patients.

Identification of a Torque Teno Mini Virus (TTMV) in Hodgkin's Lymphoma Patients.

At least 12% of human cancers are caused by virus infection. To understand whether other viruses are associated with human cancers, a viral metagenomics approach was used to analyze the composition of the viral communities of the serum of the patients with Hodgkin's lymphoma (HL) and non-Hodgkin lymphoma. In this report, a human anellovirus TTMV named TTMV-SH was discovered from three patients with HL. The complete genome of TTMV-SH is 2812nt in length. Phylogenetic analysis based on ORF1 indicated that TTMV-SH of the 11 isolates cluster with TTMV strain TLMV-CBD231 sharing only 60.3-62% sequence similarity, and the sequences divergence is 41.5-43.1%, which indicates that TTMV-SH is a novel species. The TTMV-SH prevalence in HL group, especially in nodular sclerosing Hodgkin's lymphomas (NSHL), was significantly higher than in the healthy group implicated that the TTMV-SH may be associated with HL, especially NSHL.

Detection of Small Anellovirus DNA from Blood Products / 대한수혈학회지

BACKGROUND: The small anellovirus (SAV) is a new member of the genus Anellovirus infecting humans. SAV can be transmissible by transfusion. However there are no reports on SAV infections in Korea. The aim of this study was to determine the prevalence of SAV in blood products. METHODS: A total of 90 plasma samples from blood products (each 30 units of Red blood cell, whole blood, and platelet concentrate) and 30 serum samples from non-A to C hepatitis patients were tested. SAV DNA was detected using nested polymerase chain reaction (PCR). At the same time, TTV and TTMV DNA were detected using nested PCR. RESULTS: SAV DNA was detected in 34% (31/90) of blood products. TTV and TTMV DNA were detected in 66% (54/90) and 29% (26/90) of blood products, respectively. One of the three anelloviruses (SAV, TTV, TTMV) was detected in a total of 77 blood products (86%). SAV DNA was detected in 40% (12/30) of hepatitis patients. TTV and TTMV DNA were detected in 73% (22/30) and 33% (10/30) of those patients, respectively. One of the three anelloviruses (SAV, TTV, TTMV) was detected in 97% (29/30) of hepatitis patients. CONCLUSION: Blood products are frequently infected with SAV and (or) other anelloviruses (TTV/TTMV) in Korea, and can be transmissible with a high probability.