T-cell Receptor Excision Circles Research Articles

Newborn screening for SCID: the very first prospective pilot study from Türkiye

PurposeThe measurement of T-cell receptor excision circle (TREC) is used for newborn screening (NBS) in dried blood spot (DBS) samples from Guthrie card for severe combined immunodeficiency (SCID). Here, we report the results of first newborn screening pilot program for SCID conducted in Türkiye.MethodsThe study was carried out together with Ankara University School of Medicine and The Ministry of Health, Public Health General Directorate, Pediatric and Adolescent Health Department. TREC measurements were performed in randomly selected Guthrie card samples obtained from 20253 babies born between October 2018 and October 2020. The TREC analyses were performed together with beta Actin (β-Actin) via RT-PCR (Real Time Polymerase Chain Reaction).ResultsTRECs found to be normal (≥15 copies/µl) in 98,6% of the newborns (n: 19975) but low (<15 copies/µl) in 1.4% (n:278) at the initial analyses. TRECs were retested in 278 suspected infants and found to be normal in 160 (0.8%) while low in 118 (0.58%). New DBS were obtained from the babies with low TRECs (new sample test). TRECs were normal in 108 (0.53%) of the new sample tests and low in 10 (0.049%). Two among 10 babies who had abnormal (undetectable) TRECs were diagnosed as SCID; ADA (P1) and RAG1 (P2) defects were confirmed respectively. They both received curative treatments [gene therapy (P1) and HSCT (P2)]. The remaining 6 of 8 newborns with abnormal TRECs were found normal after clinical and laboratory immune work-up, while medical records of other two revealed early postnatal death due to extreme prematurity.ConclusionIn the light of this study the incidence of SCID was detected at least 1/10000 live births in Türkiye. This study shows the feasibility and usefulness of initiating SCID screening in Türkiye.

Quantification of T-Cell Receptor Excision Circles (TRECs).

Aging is a natural process that compromises the immune system's functionality increasing the risk of infectious, tumors, and autoimmune diseases. The thymus involution is an age-dependent process characterized by decreased cellularity, peripheral lymphocyte infiltration into the perivascular space, and expansion of adipose tissue. All those modifications hamper the functionality of the organ and lead to a decline of naïve T-cell production with a shrinking of the T-cell repertoire. Thymus atrophy is described in several disorders including autoimmune diseases. The quantification of T-cell receptor excision circles (TRECs) in recent thymus emigrants is a standard procedure to investigate the thymic function. In this chapter, we discuss the methodology used to quantify this molecule in peripheral blood mononuclear cells and isolated CD4+ and CD8+ T cells.

РОЛЬ ИММУННОГО ОТВЕТА В РАЗВИТИИ ГИПОКСИЧЕСКИХ ОСЛОЖНЕНИЙ РЕСПИРАТОРНЫХ ИНФЕКЦИЙ НИЖНИХ ДЫХАТЕЛЬНЫХ ПУТЕЙ У ДЕТЕЙ

Pneumonia is one of the most common severe infectious diseases in pediatric practice. Hypoxia is common in children with pneumonia and is one of the main predictors of poor outcome. The role of the central link in the cellular response to hypoxia is played by the family of hypoxia-inducible factors (HIF). HIF-1α is the key regulator of both T- and B-lymphocyte functions. The processes of their maturation are accompanied by the formation of circular DNA fragments, TREC (T-cell Receptor Excision Circle) and KREC (Kappa-deleting Recombination Excision Circle). TREC and KREC are unable to replicate and serve as markers for naive lymphocyte production. In its turn, HIF-1α can be used as a marker for cellular hypoxia. The use of TREC and KREC in assessing the immune system in patients with various immunological and infectious pathologies has already been described. At the same time, HIF-1α has so far been little studied as yet. The purpose of this research was to analyze the scientific bibliographical data on the HIF-1α assessment as a marker for hypoxia as well as the quantitative level of TREC and KREC as markers for the adaptive immune response; to study the bibliographical sources on the effect of hypoxia on the immune response, including as part of lower respiratory tract infections. Methods used: bibliographical research in Scopus, PubMed and Google Scholar with keywords “TREC,” “KREC,” “HIF,” “hypoxia,” “pneumonia,” “immune response,” that was then followed by the search results’ systematization, analysis and synthesis. Results: the analysis of bibliographical sources had allowed the Authors to conclude that there is a possible close connection between acute hypoxia and the formation of an adequate immune response in infectious pathologies. Research in this area is still extremely scarce. At the same time, the prospects for this area determine the need for further fundamental and clinical research. Conclusion: it is of a great scientific and clinical interest to be able to non-invasively assess the degree of acute hypoxia during an ongoing infectious disease by examining urinary HIF-1α levels. Particularly promising is the assessment of the HIF-1α level not only as a regulator of tissue adaptation to acute hypoxia, but also as a modulator of the immune response. Taking into consideration the analyzed data, both TREC and KREC appear to be the optimal markers characterizing the functional state of the adaptive immune system.

MS treatment de-escalation: review and commentary.

Almost all currently licensed disease-modifying therapies (DMTs) for MS treatment require prolonged if not lifelong administration. Yet, as people age, the immune system has increasingly reduced responsiveness, known as immunosenescence. Many MS DMTs reduce the responsiveness of the immune system, increasing the risks for infections and possibly cancers. As people with MS (pwMS) age, it is recognized that inflammatory MS activity declines. Several studies have addressed de-escalation of DMTs for relapsing MS under special circumstances. Here, we review evidence for de-escalating DMTs as a strategy that is particularly relevant to pwMS of older age. Treatment de-escalation can involve various strategies, such as extended or reduced dosing, switching from high-efficacy DMTs having higher risks to moderately effective DMTs with lesser risks, or treatment discontinuation. Studies have suggested that for natalizumab extended dosing maintained clinical efficacy while reducing the risk of PML. Extended interval dosing of ocrelizumab mitigated the decline of Ig levels. Retrospective and observational discontinuation studies demonstrate that age is an essential modifier of drug efficacy. Discontinuation of MS treatment in older patients has been associated with a stable disease course, while younger patients who discontinued treatment were more likely to experience new clinical activity. A recently completed 2-year randomized-controlled discontinuation study in 260 stable pwMS > 55years found stable clinical multiple sclerosis with only a small increased risk of new MRI activity upon discontinuation. DMT de-escalation or discontinuation in MS patients older than 55years may be non-inferior to continued treatment with immunosuppressive agents having higher health risks. However, despite several small studies, a definite conclusion about treatment de-escalation in older pwMS will require larger and longer studies. Ideally, comparison of de-escalation versus continuation versus discontinuation of DMTs should be done by prospective randomized-controlled trials enrolling sufficient numbers of subjects to allow comparisons for MS patients of both sexes within age groups, such as 55-59, 60-65, 66-69, etc. Optimally, such studies should be 3years or longer and should incorporate testing for specific markers of immunosenescence (such as T-cell receptor excision circles) to account for differential aging of individuals.

A Unique Comprehensive Model to Screen Newborns for Severe Combined Immunodeficiency-An Ontario Single-Centre Experience Spanning 2013-2023.

Severe combined immunodeficiency (SCID) is a life-threatening genetic disorder caused by critical defects of the immune system. Almost all cases are lethal if not treated within the first two years of life. Early diagnosis and intervention are thus essential for improving patient outcomes. In 2013, Ontario became the first Canadian province to perform newborn screening (NBS) for SCID by T cell receptor excision circles (TRECs) analysis, a surrogate marker of thymic function and lymphocyte maturation. This retrospective study reports on nearly 10 years of NBS for SCID at a quaternary referral centre. From August 2013 to April 2023, our centre's densely populated catchment area flagged 162 newborns with low TRECs levels, including 10 cases with SCID. Follow-up revealed other causes of low TRECs, including non-SCID T cell lymphopenia (secondary/reversible or idiopathic causes, and syndromic conditions) and prematurity. A small number of cases with normal repeat TRECs levels and/or T cell subsets were also flagged. Province-wide data from around this period revealed at least 24 diagnosed cases of SCID or Leaky SCID. This is the first report of NBS outcomes in a Canadian province describing the causative genetic defects, and the non-SCID causes of a positive NBS for SCID.

Quantitative indicators of TREC and KREC excision circles in malignancies: a prospective cohort study

Background. In oncology, of particular interest is the study of the T-cell receptor excision circles (TREC) and the κ-deletion B-cell receptor excision circles (KREC), which are extrachromosomal DNA structures. In many malignancies, the effectiveness of immune checkpoint inhibitors depends on the mutational load of the tumor, which correlates with the formation of specific antitumor immunity. Quantitative indicators of recombination excision circles reflect the occurrence of a different repertoire of T-cell receptors, an integral component in the formation of specific immunity. Understanding the change in quantitative values of TREC and KREC in cancer patients can improve the selection of patients for immunotherapy. Aim. To determine quantitative indicators of TREC and KREC for immunological evaluation of patients with malignancies. Materials and methods. The study included 55 healthy individuals and 180 patients with malignancies. Among healthy individuals, 49.1% (27/55) were males and 50.9% (28/55) females. Among patients with malignancies, 20.5% (37/180) were males and 79.5% (143/180) females. The median age in healthy individuals was 36 years [Q1–Q3: 26–58]. The median age in the group of patients with malignancies was 57 years [Q1–Q3: 47.5–67]. Results. In the general population of healthy individuals, the median TREC level was 60.1 [Q1-Q3: 31.3-188.9] and the median KREC level was 256 [Q1-Q3: 149.8-353]. In the general population of patients with malignancies, the median TREC rate was 4.6 [Q1-Q3: 0.9-17.3] and the median KREC was 111.9 [Q1-Q3: 29.3-339.28]. According to the results of the study, we noted statistically significant differences in TREC and KREC indices between all patients with malignancies and healthy individuals (p0.001, p=0.001). Analysis of TREC and KREC indices in patients with malignancies of various localizations (breast cancer, ovarian cancer, lung cancer, colorectal cancer, skin melanoma, lymphomas) in comparison with healthy individuals statistically significant differences in TREC level were noted (p=0.001, p0.001). When analyzing the KREC level in the studied groups, statistically significant differences in patients with ovarian malignancies (p0.001), lymphoma (p0.001), colorectal cancer (p=0.001) and melanoma (p=0.039) in comparison with healthy individuals were obtained. When comparing groups pairwise, it was found that TREC level in patients with malignancies in the age group of 25–44 years was significantly higher than in the age group of 45–60 years (p=0.03); TREC level in the age group of 25–44 years was significantly higher than in the age group of persons over 60 years (p0.001); TREC level in the age group of 45–60 years was significantly higher than in the age group over 60 years (p0.001). Statistically significant differences of KREC level in the studied patients with malignancies depending on the age group were not established (p=0.16), there were no age differences of groups by KREC level. Conclusion. The results demonstrate a significant decrease in TREC and KREC levels in patients with malignancies compared to healthy individuals. The study of TREC and KREC excision circles in peripheral blood is one of the promising approaches for the immunological evaluation of cancer patients.

Expert consensus on newborn screening of T-cell receptor excision circles and (or) Kappa-recombination excision circles (2024)

Expert consensus on newborn screening of T-cell receptor excision circles and (or) Kappa-recombination excision circles (2024)

T-cell receptor excision circles and postnatal cytomegalovirus infection in very low birth weight infants.

T-cell receptor excision circles and postnatal cytomegalovirus infection in very low birth weight infants.

Live Rotavirus Vaccination Appears Low-risk in Infants Born to Mothers With Inflammatory Bowel Disease on Biologics

Biologic therapies in the context of inflammatory bowel disease and pregnancy lead to improved maternal and fetal outcomes. Placental transfer results in detectable drug concentrations in infants. Rotavirus infection results in diarrheal related hospitalizations; however, the live oral vaccine is not currently recommended in biologic exposed infants. The aim of this study was to assess the effect of maternal biologic therapies on the infant immune system and safety of live rotavirus vaccination in biologic-exposed infants. Biologic-exposed infants underwent standardized clinical assessments, drug concentration, and immune function testing (complete blood count, differential, immunoglobulin levels, extended B-cell and T-cell subset enumeration, Recent Thymic Emigrants, regulatory T-cell numbers, mitogen stimulation assays, and review of T-cell Receptor Excision Circles in the newborn screen). Rotavirus vaccine-specific adverse effects following immunizations up to 42 days post the last dose of the vaccine series were recorded. There were 57 infants born to 52 mothers with inflammatory bowel disease exposed to infliximab (n= 21), adalimumab (n= 19), vedolizumab (n= 10), and ustekinumab (n= 7) in the third trimester for a median of 39 weeks (interquartile range, 38-39 weeks) at delivery. Immunologic assessments validated for age were normal in all infants despite median infliximab concentrations of 6.1 ug/mL (range, 0.4-28.8 ug/mL), adalimumab concentrations of 1.7ug/mL (range, 0.7-7.9 ug/mL), ustekinumab concentrations of 0.6 ug/mL (range, 0-1.1), and undetectable for vedolizumab at 10.7 weeks (interquartile range, 9.4-12.4) of age. The live oral rotavirus vaccine series was provided to 50 infants with the first dose given at a median of 13weeks of age. No adverse effects following immunization were reported. Immune function testing was normal, and administration of live rotavirus vaccination appeared low-risk in biologic-exposed infants irrespective of circulating drug levels.

Peripheral T Cell Development and Immunophenotyping of Twins with Heterozygous FOXN1 Mutations.

The transcription factor FOXN1 plays an established role in thymic epithelial development to mediate selection of maturing thymocytes. Patients with heterozygous loss-of-function FOXN1 variants are associated with T cell lymphopenia at birth and low TCR excision circles that can ultimately recover. Although CD4+ T cell reconstitution in these patients is not completely understood, a lower proportion of naive T cells in adults has suggested a role for homeostatic proliferation. In this study, we present an immunophenotyping study of fraternal twins with low TCR excision circles at birth. Targeted primary immunodeficiency testing revealed a heterozygous variant of uncertain significance in FOXN1 (c.1205del, p.Pro402Leufs*148). We present the immune phenotypes of these two patients, as well as their father who carries the same FOXN1 variant, to demonstrate an evolving immune environment over time. While FOXN1 haploinsufficiency may contribute to thymic defects and T cell lymphopenia, we characterized the transcriptional activity and DNA binding of the heterozygous FOXN1 variant in 293T cells and found the FOXN1 variant to have different effects across several target genes. These data suggest multiple mechanisms for similar FOXN1 variants pathogenicity that may be mutation specific. Increased understanding of how these variants drive transcriptional regulation to impact immune cell populations will guide the potential need for therapeutics, risk for infection or autoimmunity over time, and help inform clinical decisions for other variants that might arise.

Comprehensive Flow Cytometric, Immunohistologic, and Molecular Assessment of Thymus Function in Rhesus Macaques.

The critical importance of the thymus for generating new naive T cells that protect against novel infections and are tolerant to self-antigens has led to a recent revival of interest in monitoring thymic function in species other than humans and mice. Nonhuman primates such as rhesus macaques (Macaca mulatta) provide particularly useful animal models for translational research in immunology. In this study, we tested the performance of a 15-marker multicolor Ab panel for flow cytometric phenotyping of lymphocyte subsets directly from rhesus whole blood, with validation by thymectomy and T cell depletion. Immunohistochemical and multiplex RNA expression analysis of thymus tissue biopsies and molecular assays on PBMCs were used to further validate thymus function. Results identify Ab panels that can accurately classify rhesus naive T cells (CD3+CD45RA+CD197+ or CD3+CD28+CD95-) and recent thymic emigrants (CD8+CD28+CD95-CD103+CD197+) using just 100 µl of whole blood and commercially available fluorescent Abs. An immunohistochemical panel reactive with pan-cytokeratin (CK), CK14, CD3, Ki-67, CCL21, and TdT provides histologic evidence of thymopoiesis from formalin-fixed, paraffin-embedded thymus tissues. Identification of mRNAs characteristic of both functioning thymic epithelial cells and developing thymocytes and/or molecular detection of products of TCR gene rearrangement provide additional complementary methods to evaluate thymopoiesis, without requiring specific Abs. Combinations of multiparameter flow cytometry, immunohistochemistry, multiplex gene expression, and TCR excision circle assays can comprehensively evaluate thymus function in rhesus macaques while requiring only minimal amounts of peripheral blood or biopsied thymus tissue.

Paving the way in implementation of SCID newborn screening in developing nations: feasibility study and strategies to move forward in Malaysia.

Early diagnosis and effective management of Primary immunodeficiency diseases (PIDs), particularly severe combined immunodeficiency (SCID), play a crucial role in minimizing associated morbidities and mortality. Newborn screening (NBS) serves as a valuable tool in facilitating these efforts. Timely detection and diagnosis are essential for swiftly implementing isolation measures and ensuring prompt referral for definitive treatment, such as allogeneic hematopoietic stem cell transplantation. The utilization of comprehensive protocols and screening assays, including T cell receptor excision circles (TREC) and kappa-deleting recombination excision circles (KREC), is essential in facilitating early diagnosis of SCID and other PIDs, but their successful application requires clinical expertise and proper implementation strategy. Unfortunately, a notable challenge arises from insufficient funding for the treatment of PIDs. To address these issues, a collaborative approach is imperative, involving advancements in technology, a well-functioning healthcare system, and active engagement from stakeholders. The integration of these elements is essential for overcoming the existing challenges in NBS for PIDs. By fostering synergy between technology providers, healthcare professionals, and governmental stakeholders, we can enhance the efficiency and effectiveness of early diagnosis and intervention, ultimately improving outcomes for individuals with PIDs.

BIOM-04. ALTERED IMMUNE ACTIVATION AND REDUCED LYMPHOPOIESIS IN TREATMENT NAÏVE PEDIATRIC BRAIN TUMOUR PATIENTS

Abstract Pediatric central nervous system (CNS) tumors are the most common solid tumor in children. Despite improvements in diagnostic and therapeutic approaches over time, treatment effectiveness remains unsatisfactory. Optimal immune function is critical to protect against development and dissemination of cancer and to offer the ideal conditions for the success of anticancer treatment. Here we analyzed the peripheral immunological features of patients affected by pediatric brain tumors, the process of de novo T- and B-cell development and its association with the lymphopenia often observed in these patients. Eighty-six steroids-naïve pediatric patients with CNS tumors at the onset of the disease were included in the study. Brain tumors were classified into low-grade gliomas (LGG: n=44, 51%), pediatric-type diffuse high-grade gliomas (pHGG: n=16, 19%), medulloblastoma (n =9, 10%), and other CNS tumors (n =17, 20%). Patient’s peripheral immunological status was analyzed by flow-cytometry and by evaluating T-cell receptor excision circles (TRECs) and kappa-deleting excision circles (KREC) to quantify T- and B-cell neogenesis. Although all patients showed tendencies towards low lymphocyte counts, this effect was particularly evident in pHGG patients. T-cells of pHGG patients displayed a shift from naïve to effector memory phenotype compared to age-matched controls. On the other hand, LGG patients were characterized by an expansion of the T-central memory pool and higher expression of T-cell activation markers. Both patient cohorts exhibited decreased frequency of B-cells with activated phenotype. Interestingly, pHGG patients exhibited reduced thymic function as revealed by reduced levels of the recent thymic emigrant population and TRECs. In addition, LGG and pHGG patients were characterized by lower KREC values, suggesting suboptimal B-cell formation. At diagnosis, pediatric patients with brain tumor show alterations in their immunological status and reduced thymic and medullary lymphopoiesis, particularly pronounced in pHGG, which may contribute to explain the systemic lymphopenia characterizing these patients.

Premature thymic functional senescence is a hallmark of childhood acute lymphoblastic leukemia survivorship

Childhood acute lymphoblastic leukemia (cALL) survivors suffer early-onset chronic diseases classically associated with aging. Normal aging is accompanied by organ dysfunctions, including immunological ones. We hypothesize that thymic immunosenescence occurs in cALL survivors and that its severity may correlate with early-onset chronic diseases. The PETALE study is a cALL survivor cohort with an extensive cardiovascular and metabolic evaluation. The thymic immunosenescence biomarker, signal joint T-cell receptor excision circles (TREC), was evaluated and was highly correlated with age in healthy participants (n = 281) and cALL survivors (n = 248). We observed a systematic thymic immunoage accentuation in each cALL survivor compared to controls ranging from 5.9 to 88.3 years. The immunoage gain was independent of age at diagnosis and treatment modalities and was more severe for females. Thymic aging was associated with several pathophysiological parameters, was greater in survivors suffering from metabolic syndrome, but there was no significant association with global physical condition. The decrease in TREC was independent from blood cell counts, which were normal, suggesting a segmental aging of the thymic compartment. Indeed, increased plasmatic T cell regulatory cytokines IL-6, IL-7 and GM-CSF accompanied high immunoage gain. Our data reveal that cALL or its treatment trigger a rapid immunoage gain followed by further gradual thymic immunosenescence, similar to normal aging. This leads to an enduring shift in accentuated immunoage compared to chronological age. Thus, accentuated thymic immunosenescence is a hallmark of cALL survivorship and TREC levels could be useful immunosenescence biomarkers to help monitoring the health of cancer survivors.

Newborn screening for inborn errors of immunity: The status worldwide

BackgroundNewborn screening (NBS) for the early detection of inborn errors of immunity (IEI) has been implemented in a few countries. The objective of this study was to verify the situation and define obstacles to the implementation of NBS worldwide. MethodsA questionnaire was developed by the Inborn Errors of Immunity Committee of the World Allergy Organization (WAO) with 17 questions regarding NBS for IEI in the physician's workplace, NBS test type, problems hindering NBS implementation, reimbursement for IEI therapy, presence of a national IEI registry, referral centers, molecular diagnosis, hematopoietic stem cell transplantation centers, gene therapy, and immunoglobulin replacement therapy. The survey was sent by email once a week to doctors and others associated with WAO and the main immunology societies worldwide as a Google Form™ to be completed during September and October 2021. ResultsTwo hundred twenty-nine questionnaires were completed, of which 216 (94.3%) were completed by physicians. One hundred seventy-six (76.8%) physicians were both allergists and immunologists. The agreement between allergists/immunologists and non-allergists/non-immunologists for the question “Is there NBS for IEI in the country you work in?” was good (κ = 0,64: 95% CI 0.55–0.69). Ninety-eight (42.8%) participants were from Latin America, 35 (15.3%) from North America, 29 (12.6%) from Europe, 18 (7.9%) from Africa, 44 (19.2%) from Asia, and 5 (2.2%) from Oceania. More than half the participants (n = 124, 54.2%) regularly treated patients with IEI, followed by occasional treatment (n = 77, 33.6%), or never (n = 28, 12.2%). Of the respondents, 14.8% reported that their countries performed NBS for IEI, whereas 42.2% reported their countries did not. T-cell receptor excision circles was the most widely used technique in some countries, with 75 (59.9%) for the diagnosis of NBS for IEI, followed by combined use with kappa deleting-recombination excision circles. Only 13 participants (10.3%) underwent neonatal exon screening in their respective countries. Financial and technical issues were among the major obstacles to the implementation of NBS for IEI. ConclusionsThis pilot study showed that few countries have implemented NBS for IEI, despite the presence of immunology referral centers and the availability of hematopoietic stem cell transplantation and intravenous immunoglobulin replacement therapy. The findings highlight the difficulties, mainly financial and technical, hindering wide application of NBS. Sharing experiences, technologies, and resources at the international level can help overcome these difficulties.

Determining the relationship of p16INK4a and additional molecular markers of aging with clinical frailty in hematologic malignancy

PurposeOlder adults with hematologic malignancies (HM) have unique challenges due to age and fitness. The primary aim of this pilot study was to benchmark the ability of multiple biomarkers of aging (p16, epigenetic clocks, T cell gene expression profiles, and T cell receptor excision circles (TREC) to identify frailty as measured by a clinical impairment index (I2) in patients with HM.Methods70 patients newly diagnosed with HM had peripheral blood T lymphocytes (PBTL) analyzed for p16INK4a expression using the OSU_Senescence Nanostring CodeSet. PBTL epigenetic age was measured using 7 epigenetic clocks, and TREC were quantified by qRT-PCR. A composite clinical impairment index (I2) was generated by combining values from 11 geriatric metrics (Independent Activities of Daily Living (iADL), physical health score, Short Physical Performance Battery (SPPB), Body Mass Index (BMI), Eastern Cooperative Oncology Group (ECOG) performance status, self-reported KPS, Blessed Orientation Memory Concentration (BOMC), polypharmacy, Mental Health Inventory (MHI)-17, Medical Outcomes Study (MOS) subscales). Clinical frailty was defined as a score of 7 or greater on the I2.ResultsAge-adjusted p16INK4a was similar in newly diagnosed patients and healthy controls (p > 0.1). PBTL p16INK4a levels correlated positively with the Hannum [r = 0.35, 95% CI (0.09–0.75); p adj. = 0.04] and PhenoAge [r = 0.37, 95% CI (0.11–0.59); p adj. = 0.04] epigenetic clocks. The discrimination ability of the I2 model was calculated using the area under the receiver operating characteristic curve (AUC). After adjusting for chronologic age and disease group, baseline p16INK4a [AUC = 0.76, 95% CI (0.56–0.98); p = 0.01], Hannum [AUC = 0.70, 95% CI (0.54–0.85); p = 0.01], PhenoAge [AUC = 0.71, 95% CI (0.55–0.86); p = 0.01], and DunedinPACE [AUC = 0.73, 95% CI (0.57–0.88); p = < 0.01] measures showed the greatest potential to identify clinical frailty using the I2.ConclusionsOur pilot data suggest that multiple blood-based aging biomarkers have potential to identify frailty in older adults with HM.Implications for Cancer SurvivorsWe developed the I2 index to quantify impairments across geriatric domains and discovered that PBTL p16, Hannum, PhenoAge, and DunedinPACE are promising indicators of frailty in HM.

Newborn Screening for Severe T and B Cell Lymphopenia Using TREC/KREC Detection: A Large-Scale Pilot Study of 202,908 Newborns.

Newborn screening (NBS) for severe inborn errors of immunity (IEI), affecting T lymphocytes, and implementing measurements of T cell receptor excision circles (TREC) has been shown to be effective in early diagnosis and improved prognosis of patients with these genetic disorders. Few studies conducted on smaller groups of newborns report results of NBS that also include measurement of kappa-deleting recombination excision circles (KREC) for IEI affecting B lymphocytes. A pilot NBS study utilizing TREC/KREC detection was conducted on 202,908 infants born in 8 regions of Russia over a 14-month period. One hundred thirty-four newborns (0.66‰) were NBS positive after the first test and subsequent retest, 41% of whom were born preterm. After lymphocyte subsets were assessed via flow cytometry, samples of 18 infants (0.09‰) were sent for whole exome sequencing. Confirmed genetic defects were consistent with autosomal recessive agammaglobulinemia in 1/18, severe combined immunodeficiency - in 7/18, 22q11.2DS syndrome - in 4/18, combined immunodeficiency - in 1/18 and trisomy 21 syndrome - in 1/18. Two patients in whom no genetic defect was found met criteria of (severe) combined immunodeficiency with syndromic features. Three patients appeared to have transient lymphopenia. Our findings demonstrate the value of implementing combined TREC/KREC NBS screening and inform the development of policies and guidelines for its integration into routine newborn screening programs.

Expanded Newborn Screening for Inborn Errors of Immunity: The Experience of Tuscany

Inborn errors of immunity (IEIs) include 485 inherited disorders characterized by an increased susceptibility to life threatening infectious diseases, autoimmunity and malignant diseases with a high mortality rate in the first years of life. Severe Combined Immunodeficiency is the most severe of the IEIs and its detection should be a primary goal in a newborn screening (NBS) program. The term "actionable" has recently been used for all IEIs with outcomes that can be demonstrably improved through early specialized intervention. to evaluate the results of the expanded NBS strategy for IEIs in Tuscany Region (Italy), based on TREC (T-cell Receptor Excision Circles), KREC (Kappa Recombining Excision Circles) and Tandem Mass-based assays. This is a retrospective study collecting data from all infants born in Tuscany from October 10, 2018, to October 10, 2022. Tandem mass assay to identify Adenosine deaminase (ADA) and purine nucleoside phosphorylase (PNP) deficiency, together with TREC and KREC molecular analysis were conducted on dried blood spot (DBS) from the newborns' Guthrie Cards. A new DBS and evaluation by an immunologist were carried out when the results of the first test were outside the diagnostic cut-offs. 94,319 newborns were evaluated. Referral rates for TREC (0.031%) and KREC (0.074%) in this study are in line with the data available in literature. The results from the expanded NBS strategy revealed an incidence rate of 1/9,431 affected newborns. This work represents the first description of a sustainable and real-life based expanded NBS program for IEIs with a high diagnostic incidence facilitating prompt management of identified patients.

EFFECT OF ORAL VITAMIN D3 ON THYMIC INDEX IN MALNOURISHED CHILDREN

Objectives: To compare the thymic Index, vitamin D3 level, and T cell receptor excision circle (TREC) of healthy and malnourished infants. To evaluate the effect of oral vitamin D3 supplementation on the Thymic Index and TREC in malnourished children. Material and Methods: This quasi-experimental study (non-equivalent control group design) included 44 infants within the age range of 2 to 6 months. The cases were 22 malnourished infants, and 22 were healthy infants (controls) based on WHO Z scores classification (weight for length). Weight, age, thymic Index (TI), Vitamin D levels, and TRECs were recorded for cases and controls(pre-test). The cases were given Vitamin D supplementation for 8 weeks. The weight for length (z-score), thymic Index (TI), vitamin D levels, and TRECs were checked in these malnourished children on follow-up (post-test). Thymic Index (TI) was checked by ultrasonography (length multiplied by breadth). ELISA was performed to check Vitamin D levels and RT-PCR was done to compare TRECs. Results: The malnourished infants had a mean thymic index of 8.015 ±3.2192 as compared to controls of 11.667 ±3.2730. Post Vit D supplementation TI increased (8.459 ±3.2191) in cases. Weight, length, and Z scores were found to be directly related to TI values. Pre and Post supplementation TI were strongly correlated to each other (r= 0.985, p= &lt;0.001). The thymic index increased significantly (p= 0.001) after intervention. There was a statistically non-significant rise in TRECS (809 vs 4242) (p= 0.075) after vitamin D supplementation. The Pearson Correlation showed no significant correlation of TRECS to any of the variables including the Thymic index. Conclusion: TI (thymic size) and vitamin D levels increase with vitamin D supplementation in malnourished children. However, Vitamin D supplementation has no significant effect on TRECS. Keywords: Malnourished, Vitamin D3, T cell receptor excision circles, Thymic Index, TRECs.

Increased mortality in infants with abnormal T-cell receptor excision circles.

T-Cell Receptor Excision Circles based newborn screening (TREC-NBS) allows for early detection of T-cell lymphopenia in infants with primary immunodeficiency disorders (PIDD). The utility of abnormal TREC-NBS in infants without PIDD is not well studied. We sought to evaluate the association of abnormal TREC-NBS with mortality. 365,207 TREC-NBS from October 2011 to December 2014 were reviewed. 467 newborns had abnormal screens and did not meet the criteria for a PIDD diagnosis. Cases were matched to controls (1:3) based on gestational age, birth weight, neonatal intensive care unit status (NICU), and race. Data were obtained through NBS, birth and death certificates records from the Michigan Department of Health and Human Services (MDHHS) databases. Infants with abnormal TREC-NBS had higher mortality even when PIDD was ruled-out. Transient abnormal TREC-NBS was not associated with higher mortality, but unresolved or late abnormal TREC-NBS was associated with higher mortality. Infants with late abnormal TREC-NBS had severe prematurity, lower birth weight, lower Apgar scores, and higher percentage of congenital anomalies. Infants with abnormal TREC-NBS may be at a higher risk of morbidity and mortality and should be carefully followed, especially if discharged home before a repeat screen can be completed. This study explores the risk factors and mortality for newborns with secondary T-cell lymphopenia captured on T-Cell Receptor Excision Circles based newborn screening (TREC-NBS). Abnormal TREC-NBS allows for prompt life-saving interventions for primary immunological conditions such as Severe Combined Immunodeficiency (SCID), but can also be associated with non-immunologic conditions. Unresolved and late abnormal TREC-NBS is associated with higher mortality even without primary immunodeficiency, likely detected in infants with more severe prematurity, lower birth weight, and congenital anomalies. TREC-NBS positive infants with secondary T-cell lymphopenia require special attention and close monitoring.