Heterozygous RPA2 variant as a novel genetic cause of telomere biology disorders.

Premature telomere shortening or telomere instability is associated with a group of rare and heterogeneous diseases collectively known as telomere biology disorders (TBDs). Here we identified two unrelated individuals with clinical manifestations of TBDs and short telomeres associated with the identical monoallelic variant c.767A>G; Y256C in RPA2 Although the replication protein A2 (RPA2) mutant did not affect ssDNA binding and G-quadruplex-unfolding properties of RPA, the mutation reduced the affinity of RPA2 with the ubiquitin ligase RFWD3 and reduced RPA ubiquitination. Using engineered knock-in cell lines, we found an accumulation of RPA at telomeres that did not trigger ATR activation but caused short and dysfunctional telomeres. Finally, both patients acquired, in a subset of blood cells, somatic genetic rescue events in either POT1 genes or TERT promoters known to counteract the accelerated telomere shortening. Collectively, our study indicates that variants in RPA2 represent a novel genetic cause of TBDs. Our results further support the fundamental role of the RPA complex in regulating telomere length and stability in humans.

Haematological features of telomere biology disorders diagnosed in adulthood: A French nationwide study of 127 patients.

Data on haematological features of telomere biology disorders (TBD) remain scarce. We describe haematological, extra-haematological characteristics and prognosis of 127 genetically confirmed TBD patients diagnosed after the age of 15. Ninety-three index cases and 34 affected relatives were included. At diagnosis of TBD, 76.3% of index cases had haematological features, half pulmonary features and a third liver features. At diagnosis, bone marrow failure (BMF) was present in 59 (46.5%), myelodysplastic syndrome (MDS) in 22 (17.3%) and acute myeloid leukaemia (AML) in 2 (1.6%) while 13 (10.2%) developed or worsened bone marrow involvement during follow-up. At diagnosis, compared to MDS/AML patients, BMF patients were younger (median 23.1 years vs. 43.8, p = 0.007), and had a better outcome (4-year overall survival 76.3% vs. 31.8%, p < 0.001). While frequencies and burden of cytogenetical and somatic mutations increased significantly in myeloid malignancies, some abnormalities were also observed in patients with normal blood counts and BMF, notably somatic spliceosome variants. Solid cancers developed in 8.7% patients, mainly human papillomavirus-related cancers and hepatocellular carcinomas. TBD is a multiorgan progressive disease. While BMF is the main haematological disorder, high-risk myeloid malignancies are common, and are, together with age, the only factors associated with a worse outcome.

Alemtuzumab-based conditioning regimen before hematopoietic stem cell transplantation in patients with short telomere syndromes: a retrospective study of the SFGM-TC.

While HSCT is the only curative option for patients with short telomere syndromes (STSs) and severe bone marrow failure (BMF) or myeloid malignancies (MM), their increase sensitivity to conditioning regimen strongly affect outcomes. To minimize HSCT related mortality, alemtuzumab-based conditioning regimens have been proposed, but the number of patients transplanted with those regimens reported in the literature remains very low. We retrospectively analyzed outcome of adults and adolescents with STSs transplanted after an alemtuzumab, fludarabine and cyclophosphamide based regimen registered by the SFGM-TC. Seven patients were transplanted for a BMF and 5 for a MM (median age 34 years, (IQR [22-45])). The 2-year GRFS for patients with MM was 20% (95% CI [3;100]), and 57% (95% CI [30;100]) in others. In univariate (hazard ratio, HR = 6, 95% CI [1;31]) and multivariate analysis (HR = 26, 95% CI [2;414]) stem cell source was a predictive factor for GRFS. Three of the 5 patients with pre-transplant MM relapsed and 2 of them died at last follow up. The 2-year OS was 66% (95% CI [43;99]) in the whole cohort with a median follow up of 32 months (IQR [13-56]). In conclusion, Alemtuzumab-based conditioning regimen with bone marrow is an option for patients with STSs and BMF, but others modalities have to be explored for patients with MM.

A predictive classifier of poor prognosis in transplanted patients with juvenile myelomonocytic leukemia: a study on behalf of the Société Francophone de Greffe de Moelle et de Thérapie Cellulaire.

Juvenile myelomonocytic leukemia (JMML) is an aggressive pediatric myeloproliferative neoplasm requiring hematopoietic stem cell transplantation (HSCT) in most cases. We retrospectively analyzed 119 JMML patients who underwent first allogeneic HSCT between 2002 and 2021. The majority (97%) carried a RAS-pathway mutation, and 62% exhibited karyotypic alterations or additional mutations in SETBP1, ASXL1, JAK3 and/or the RAS pathway. Relapse was the primary cause of death, with a 5-year cumulative incidence of 24.6% (95% CI: 17.1-32.9). Toxic deaths occurred in 12 patients, resulting in treatment-related mortality (TRM) of 9.0% (95% CI: 4.6-15.3). The 5-year overall (OS) and event-free survival were 73.6% (95% CI: 65.7-82.4) and 66.4% (95% CI: 58.2-75.8), respectively. Four independent adverse prognostic factors for OS were identified: age at diagnosis >2 years, time from diagnosis to HSCT ≥6 months, monocyte count at diagnosis >7.2x109/L, and the presence of additional genetic alterations. Based on these factors, we proposed a predictive classifier. Patients with 3 or more predictors (21% of the cohort) had a 5-year OS of 34.2%, whereas those with none (7%) had a 5-year OS of 100%. Our study demonstrates improved transplant outcomes compared to prior published data, which can be attributed to the synergistic impacts of a low TRM and a reduced, yet still substantial, relapse incidence. By integrating genetic information with clinical and hematologic features, we have devised a predictive classifier. This classifier effectively identifies a subgroup of patients who are at a heightened risk of unfavorable post-transplant outcomes who would benefit from novel therapeutic agents and post-transplant strategies.

Characterization of novel mutations in the TEL-patch domain of the telomeric factor TPP1 associated with telomere biology disorders.

Telomeres are nucleoprotein structures that protect the chromosome ends from degradation and fusion. Telomerase is a ribonucleoprotein complex essential to maintain the length of telomeres. Germline defects that lead to short and/or dysfunctional telomeres cause telomere biology disorders (TBDs), a group of rare and heterogeneous Mendelian diseases including pulmonary fibrosis, dyskeratosis congenita, and Høyeraal-Hreidarsson syndrome. TPP1, a telomeric factor encoded by the gene ACD, recruits telomerase at telomere and stimulates its activity via its TEL-patch domain that directly interacts with TERT, the catalytic subunit of telomerase. TBDs due to TPP1 deficiency have been reported only in 11 individuals. We here report four unrelated individuals with a wide spectrum of TBD manifestations carrying either heterozygous or homozygous ACD variants consisting in the recurrent and previously described in-frame deletion of K170 (K170∆) and three novel missense mutations G179D, L184R, and E215V. Structural and functional analyses demonstrated that the four variants affect the TEL-patch domain of TPP1 and impair telomerase activity. In addition, we identified in the ACD gene several motifs associated with small deletion hotspots that could explain the recurrence of the K170∆ mutation. Finally, we detected in a subset of blood cells from one patient, a somatic TERT promoter-activating mutation that likely provides a selective advantage over non-modified cells, a phenomenon known as indirect somatic genetic rescue. Together, our results broaden the genetic and clinical spectrum of TPP1 deficiency and specify new residues in the TEL-patch domain that are crucial for length maintenance and stability of human telomeres in vivo.

Liver disease in germline mutations of telomere-related genes: Prevalence, clinical, radiological, pathological features, outcome, and risk factors.

BACKGROUND AND AIM: Germline mutations of telomere-related genes (TRG) induce multiorgan dysfunction, and liver-specific manifestations have not been clearly outlined. We aimed to describe TRG mutations-associated liver diseases. APPROACH AND RESULTS: Retrospective multicenter analysis of liver disease (transaminases > 30 IU/L and/or abnormal liver imaging) in patients with TRG mutations. Main measurements were characteristics, outcomes, and risk factors of liver disease in a TRG mutations cohort. The prevalence of liver disease was compared to a community-based control group (n = 1190) stratified for age and matched 1:3 for known risk factors of liver disease. Among 132 patients with TRG mutations, 95 (72%) had liver disease, with associated lung, blood, skin, rheumatological, and ophthalmological TRG diseases in 82%, 77%, 55%, 39%, and 30% of cases, respectively. Liver biopsy was performed in 52/95 patients, identifying porto-sinusoidal vascular disease in 48% and advanced fibrosis/cirrhosis in 15%. After a follow-up of 21 months (12-54), ascites, hepato-pulmonary syndrome, variceal bleeding, and HCC occurred in 14%, 13%, 13%, and 2% of cases, respectively. Five-year liver transplantation-free survival was 69%. A FIB-4 score ≥ 3·25 and ≥1 risk factor for cirrhosis were associated with poor liver transplantation-free survival. Liver disease was more frequent in patients with TRG mutations than in the paired control group [80/396, (20%)], OR 12.9 (CI 95%: 7.8-21.3, p < 0.001). CONCLUSIONS: TRG mutations significantly increase the risk of developing liver disease. Although symptoms may be mild, they may be associated with severe disease. Porto-sinusoidal vascular disease and cirrhosis were the most frequent lesions, suggesting that the mechanism of action is multifactorial.

[Juvenile myelomonocytic leukemia and pediatric myelodysplastic syndromes]. / Leucémies myélo-monocytaires juvéniles et syndromes myélodysplasiques de l'enfant.

Juvenile myelomonocytic leukemia (JMML) and myelodysplastic syndromes (MDS) of children are rare and aggressive diseases. They both have the particularity of being very frequently associated with an underlying predisposition syndrome, which must be systematically investigated by meticulous clinical exam completed by molecular analysis on fibroblasts, in order to guarantee the best therapeutic management. New generation sequencing techniques have made it possible to better define the landscape of constitutional predisposing pathologies, to understand the clonal evolution that leads to the development of hematological malignancies and to identify new prognostic markers. In these two diseases, the only curative treatment is allogeneic hematopoietic stem cell transplantation, for which the appropriate timeframe, the type of donor and the conditioning must be decided in consultation with the expert teams in each entity.

De novo NUF2 variant in a novel inherited bone marrow failure syndrome including microcephaly and renal hypoplasia.

In a patient with severe microcephaly, congenital bone marrow failure, growth retardation, and renal hypoplasia, we identified a likely pathogenic variant in NUF2 that impairs the cell's ability to properly complete mitosis. Interestingly, these clinical features as well as the observed cellular alterations are highly reminiscent of what is reported in Fanconi Anaemia supporting a unifying causal role of the variant in the disease. This case provides the first evidence that a kinetochore defect, previously associated with microcephaly, can be responsible for an inherited bone marrow failure syndrome, highlighting the unique pathological link between neurogenesis and haematopoiesis.

Interstitial lung diseases associated with mutations of poly(A)-specific ribonuclease: A multicentre retrospective study.

BACKGROUND AND OBJECTIVE: Poly(A)-specific ribonuclease (PARN) mutations have been associated with familial pulmonary fibrosis. This study aims to describe the phenotype of patients with interstitial lung disease (ILD) and heterozygous PARN mutations. METHODS: We performed a retrospective, observational, non-interventional study of patients with an ILD diagnosis and a pathogenic heterozygous PARN mutation followed up in a centre of the OrphaLung network. RESULTS: We included 31 patients (29 from 16 kindreds and two sporadic patients). The median age at ILD diagnosis was 59 years (range 54 to 63). In total, 23 (74%) patients had a smoking history and/or fibrogenic exposure. The pulmonary phenotypes were heterogenous, but the most frequent diagnosis was idiopathic pulmonary fibrosis (n = 12, 39%). Haematological abnormalities were identified in three patients and liver disease in two. In total, 21 patients received a specific treatment for ILD: steroids (n = 13), antifibrotic agents (n = 11), immunosuppressants (n = 5) and N-acetyl cysteine (n = 2). The median forced vital capacity decline for the whole sample was 256 ml/year (range -363 to -148). After a median follow-up of 32 months (range 18 to 66), 10 patients had died and six had undergone lung transplantation. The median transplantation-free survival was 54 months (95% CI 29 to ∞). Extra-pulmonary features were less frequent with PARN mutation than telomerase reverse transcriptase (TERT) or telomerase RNA component (TERC) mutation. CONCLUSION: IPF is common among individuals with PARN mutation, but other ILD subtypes may be observed.

Inherited human Apollo deficiency causes severe bone marrow failure and developmental defects.

Inherited bone marrow failure syndromes (IBMFSs) are a group of disorders typified by impaired production of 1 or several blood cell types. The telomere biology disorders dyskeratosis congenita (DC) and its severe variant, Høyeraal-Hreidarsson (HH) syndrome, are rare IBMFSs characterized by bone marrow failure, developmental defects, and various premature aging complications associated with critically short telomeres. We identified biallelic variants in the gene encoding the 5'-to-3' DNA exonuclease Apollo/SNM1B in 3 unrelated patients presenting with a DC/HH phenotype consisting of early-onset hypocellular bone marrow failure, B and NK lymphopenia, developmental anomalies, microcephaly, and/or intrauterine growth retardation. All 3 patients carry a homozygous or compound heterozygous (in combination with a null allele) missense variant affecting the same residue L142 (L142F or L142S) located in the catalytic domain of Apollo. Apollo-deficient cells from patients exhibited spontaneous chromosome instability and impaired DNA repair that was complemented by CRISPR/Cas9-mediated gene correction. Furthermore, patients' cells showed signs of telomere fragility that were not associated with global reduction of telomere length. Unlike patients' cells, human Apollo KO HT1080 cell lines showed strong telomere dysfunction accompanied by excessive telomere shortening, suggesting that the L142S and L142F Apollo variants are hypomorphic. Collectively, these findings define human Apollo as a genome caretaker and identify biallelic Apollo variants as a genetic cause of a hitherto unrecognized severe IBMFS that combines clinical hallmarks of DC/HH with normal telomere length.

Genotype-Phenotype Relationships in Inheritable Idiopathic Pulmonary Fibrosis: A Greek National Cohort Study.

BACKGROUND: Monogenic and polygenic inheritances are evidenced for idiopathic pulmonary fibrosis (IPF). Pathogenic variations in surfactant protein-related genes, telomere-related genes (TRGs), and a single-nucleotide polymorphism in the promoter of MUC5B gene encoding mucin 5B (rs35705950 T risk allele) are reported. This French-Greek collaborative study, Gen-Phen-Re-GreekS in inheritable IPF (iIPF), aimed to investigate genetic components and patients' characteristics in the Greek national IPF cohort with suspected heritability. PATIENTS AND METHODS: 150 patients with familial PF, personal-family extrapulmonary disease suggesting short telomere syndrome, and/or young age IPF were analyzed. RESULTS: MUC5B rs35705950 T risk allele was detected in 103 patients (90 heterozygous, 13 homozygous, allelic frequency of 39%), monoallelic TRG pathogenic variations in 19 patients (8 TERT, 5 TERC, 2 RTEL1, 2 PARN, 1 NOP10, and 1 NHP2), and biallelic ABCA3 pathogenic variations in 3. Overlapping MUC5B rs35705950 T risk allele and TRG pathogenic variations were shown in 11 patients (5 TERT, 3 TERC, 1 PARN, 1 NOP10, and 1 NHP2), MUC5B rs35705950 T risk allele, and biallelic ABCA3 pathogenic variations in 2. In 38 patients, neither MUC5B rs35705950 T risk allele nor TRG pathogenic variations were detectable. Kaplan-Meier curves showed differences in time-to-death (p = 0.025) where patients with MUC5B rs35705950 T risk allele alone or in combination with TRG pathogenic variations presented better prognosis. CONCLUSION: The Gen-Phen-Re-GreekS in iIPF identified multiple and overlapping genetic components including the rarest, underlying disease's genetic "richesse," complexity and heterogeneity. Time-to-death differences may relate to diverse IPF pathogenetic mechanisms implicating "personalized" medical care driven by genotypes in the near future.