Bloom Syndrome Research Articles

Protocol for a high titer of BaEV-Rless pseudotyped lentiviral vector: Focus on syncytium formation and detachment

The development of hematopoietic stem cell (HSCs) gene therapy for DNA repair disorders, such as Fanconi anemia and Bloom syndrome, is challenging because of the induction of HSCs apoptosis by cytokine stimulation. Although the Baboon envelope pseudotyped lentiviral vector (BaEV-Rless-LV) has been reported as a non-stimulatory gene transfer tool, the virus titer of BaEV-Rless-LV is too low for use in clinical applications. Transfected 293 T cells with helper plasmids, including the BaEV-Rless plasmid, showed morphological changes, such as syncytium formation and detachment. To establish a novel protocol for producing a high titer of BaEV-Rless-LV, we optimized three aspects of a basic virus production protocol by focusing on modifying culture conditions and the use of reagents: the virus titer increased 3-fold when the amount of BaEV-Rless plasmid was increased 1.2-fold; the highest titer was obtained when the viral supernatant was harvested at 48-h post-transfection, despite complete syncytium formation and detachment of the 293 T cells; and the use of poly-L-lysine-coated culture plates to enhance the adhesion and proliferation of 293 T cells and prevent detachment doubled the titer. Collectively, our novel protocol resulted in a 10-fold titer increase compared to the basic protocol and may be useful in clinical applications for treating DNA repair disorders.

Intellectual disability and abnormal cortical neuron phenotypes in patients with Bloom syndrome.

Bloom syndrome (BS) is a rare autosomal recessive disorder characterized by genomic instability that leads to various complications, including cancer. Given the low prevalence of BS in Japan, we conducted a nationwide survey. We recruited eight patients with BS, three of whom exhibited intellectual disability. The 631delCAA mutation in the BLM gene was detected in 9 out of 16 alleles. To investigate neuronal development in patients with BS, we generated induced pluripotent stem cells derived from one of these patients (BS-iPSCs). We examined the phenotypes of the induced cortical neurons derived from the generated BS-iPSCs using a previously reported protocol; the generated BS-iPSCs showed an approximately 10-times higher frequency of sister-chromatid exchange (SCE) than the control iPSCs. Immunocytochemistry revealed shorter axons and higher proliferative potential in BS-iPSC-derived cortical neurons compared with control iPSCs. To our knowledge, our study is the first to clarify the abnormality of the cortical neuron phenotypes derived from patients with BS. Our findings may help identify the pathogenesis of neuronal differentiation in BS and aid in the development of novel therapeutic agents.

Germline Pathogenic Variants in Squamous Cell Carcinoma of the Head and Neck.

Head and neck squamous cell carcinoma (HNSCC) presents a significant global health problem with variable geographic distribution and risk factors, including tobacco and alcohol abuse, human papillomavirus infections, and genetic predisposition. While the majority of cases are sporadic, several well-defined hereditary syndromes have been associated with a higher risk of developing HNSCC including Li-Fraumeni syndrome, Fanconi anaemia, Bloom syndrome, familial atypical multiple mole melanoma, and dyskeratosis congenita. There is also evidence of familial clusters of HNSCC, suggesting a genetic component in the development of the disease. Germ-line genetic testing in HNSCC using next-generation sequencing has revealed a wide range of germline variants, some of which were not anticipated based on standard guidelines. These variants may influence treatment decisions and have the potential to be targeted with precision medicine in the future. Despite these advances, routine germline genetic testing for HNSCC is not currently recommended and remains reserved for HNSCC cases with early onset or strong family cancer history. However, the increasing availability of germline genetic testing warrants development of more comprehensive and standardized testing protocols. Germline genetic testing also has the potential to influence precision-guided treatment in HNSCC patients carrying germline pathogenic variants.

BLM promotes malignancy in PCa by inducing KRAS expression and RhoA suppression via its interaction with HDGF and activation of MAPK/ERK pathway.

Prostate cancer (PCa) has long been the leading cause of cancer-associated deaths among male worldwide. Our previous studies have shown that Bloom syndrome protein (BLM) plays a vital role in PCa proliferation, yet the underlying molecular mechanism remains largely obscure. Mechanistically, BLM directly interacted with hepatoma-derived growth factor (HDGF). Functionally, BLM and HDGF knockdown resulted in the higher impairment of PC3 proliferation, clonogenicity, migration and invasion than that their counterpart with either BLM or HDGF knockdown exclusively. Of note, HDGF overexpression expedited, whereas its knockdown suppressed, PC3 proliferation, clonogenicity, migration and invasion. Additionally, the potentiation or attenuation was partially antagonized upon BLM depletion or overexpression. In line with the vitro data, the impact of BLM and HDGF on tumor growth was investigated in mouse xenograft models. ChIP-seq, dual-luciferase reporter and western blotting assays were employed to expound the regulatory network in PC3 cells. The results unveiled that HDGF activated KRAS and suppressed RhoA transcription, and that the function of HDGF was mediated, in part, by interaction with BLM. Accordingly, the MAPK/ERK pathway was activated. Moreover, the regulation of HDGF on KRAS and RhoA had a signal crosstalk. To recapitulate, BLM and HDGF may serve as novel prognostic markers and potential therapeutic targets in PCa.

The MRN complex and topoisomerase IIIa–RMI1/2 synchronize DNA resection motor proteins

DNA resection-the nucleolytic processing of broken DNA ends-is the first step of homologous recombination. Resection is catalyzed by the resectosome, a multienzyme complex that includes bloom syndrome helicase (BLM), DNA2 or exonuclease 1 nucleases, and additional DNA-binding proteins. Although the molecular players have been known for over a decade, how the individual proteins work together to regulate DNA resection remains unknown. Using single-molecule imaging, we characterized the roles of the MRE11-RAD50-NBS1 complex (MRN) and topoisomerase IIIa (TOP3A)-RMI1/2 during long-range DNA resection. BLM partners with TOP3A-RMI1/2 to form the BTRR (BLM-TOP3A-RMI1/2) complex (or BLM dissolvasome). We determined that TOP3A-RMI1/2 aids BLM in initiating DNA unwinding, and along with MRN, stimulates DNA2-mediated resection. Furthermore, we found that MRN promotes the association between BTRR and DNA and synchronizes BLM and DNA2 translocation to prevent BLM from pausing during resection. Together, this work provides direct observation of how MRN and DNA2 harness the BTRR complex to resect DNA efficiently and how TOP3A-RMI1/2 regulates the helicase activity of BLM to promote efficient DNA repair.

ML216-Induced BLM Helicase Inhibition Sensitizes PCa Cells to the DNA-Crosslinking Agent Cisplatin.

Using standard DNA-damaging medicines with DNA repair inhibitors is a promising anticancer tool to achieve better therapeutic responses and reduce therapy-related side effects. Cell viability assay, neutral comet assay, western blotting (WB), and cell cycle and apoptosis analysis were used to determine the synergistic effect and mechanism of ML216, a Bloom syndrome protein (BLM) helicase inhibitor, and cisplatin (CDDP), a DNA-crosslinking agent, in PCa cells. Based on the online database research, our findings revealed that BLM was substantially expressed in PCa, which is associated with a bad prognosis for PCa patients. The combination of ML216 and CDDP improved the antiproliferative properties of three PCa cell lines. As indicated by the increased production of γH2AX and caspase-3 cleavage, ML216 significantly reduced the DNA damage-induced high expression of BLM, making PC3 more susceptible to apoptosis and DNA damage caused by CDDP. Furthermore, the combination of ML216 and CDDP increased p-Chk1 and p-Chk2 expression. The DNA damage may have triggered the ATR-Chk1 and ATM-Chk2 pathways simultaneously. Our results demonstrated that ML216 and CDDP combination therapy exhibited synergistic effects, and combination chemotherapy could be a novel anticancer tactic.

The BLM helicase is a new therapeutic target in multiple myeloma involved in replication stress survival and drug resistance.

Multiple myeloma (MM) is a hematologic cancer characterized by accumulation of malignant plasma cells in the bone marrow. To date, no definitive cure exists for MM and resistance to current treatments is one of the major challenges of this disease. The DNA helicase BLM, whose depletion or mutation causes the cancer-prone Bloom's syndrome (BS), is a central factor of DNA damage repair by homologous recombination (HR) and genomic stability maintenance. Using independent cohorts of MM patients, we identified that high expression of BLM is associated with a poor outcome with a significant enrichment in replication stress signature. We provide evidence that chemical inhibition of BLM by the small molecule ML216 in HMCLs (human myeloma cell lines) leads to cell cycle arrest and increases apoptosis, likely by accumulation of DNA damage. BLM inhibition synergizes with the alkylating agent melphalan to efficiently inhibit growth and promote cell death in HMCLs. Moreover, ML216 treatment re-sensitizes melphalan-resistant cell lines to this conventional therapeutic agent. Altogether, these data suggest that inhibition of BLM in combination with DNA damaging agents could be of therapeutic interest in the treatment of MM, especially in those patients with high BLM expression and/or resistance to melphalan.

N6‑methyladenosine‑induced long non‑coding RNA PVT1 regulates the miR‑27b‑3p/BLM axis to promote prostate cancer progression.

Prostate cancer (PCa) is one of the most fundamental causes of cancer‑related mortality and morbidity among males. However, the underlying mechanisms have not yet been fully clarified. The present study aimed to investigate the effects of plasmacytoma variant translocation 1 (PVT1) on the malignant behaviors of PCa cells and to explore the possible molecular mechanisms involved. The expression levels of PVT1 and microRNA (miRNA/miR)‑27b‑3p in PCa tissues and cell lines were measured using reverse‑transcritpion‑quantitative polymerase chain reaction. Methyltransferase 3 (METTL3)‑mediated PVT1 N6‑methyladenosine (m6A) modifications were detected using RNA immunoprecipitation (RIP) and RNA pull‑down assays. Bioinformatics analysis was used to predict the interactions of miR‑27b‑3p with PVT1 and bloom syndrome protein (BLM), and these interactions were validated using RIP, dual‑luciferase reporter and biotin pull‑down assays. The functional importance of miR‑27b‑3p, PVT1 and BLM within PCa cells was assessed through the invitro utilization of Cell Counting Kit‑8, Transwell, wound healing and colony formation assays, and the invivo use of a mouse xenograft model. The results revealed the high expression level of PVT1 in PCa tissues and cells, and epigenetic analyses revealed the upregulation of PVT1 expression following METTL3‑mediated m6A modification. PVT1 overexpression induced PCa cells to become more proliferative, migratory and invasive, whereas PVT1 knockdown led to the opposite phenotype. Furthermore, miR‑27b‑3p was found to target both PVT1 and BLM, and PVT1 functioned to sequester miR‑27b‑3p within cells, thereby indirectly promoting the BLM expression level. BLM overexpression reversed the adverse effects of PVT1 knockdown on the migratory, proliferative and invasive capabilities of PCa cells invitro and invivo. The overexpression of PVT1 contributed to the aggressive phenotype of PCa cells by regulating the miR‑27b‑3p/BLM axis. On the whole, the findings of the present study may provide novel potential targets for the treatment of PCa.

Design, synthesis and evaluation of N3-substituted quinazolinone derivatives as potential Bloom's Syndrome protein (BLM) helicase inhibitor for sensitization treatment of colorectal cancer

The homologous recombination repair (HRR) pathway is critical for repairing double-strand breaks (DSB). Inhibition of the HRR pathway is usually considered a promising strategy for anticancer therapy. The Bloom's Syndrome Protein (BLM), a DNA helicase, is essential for promoting the HRR pathway. Previously, we discovered quinazolinone derivative 9h as a potential BLM inhibitor, which suppressed the proliferation of colorectal cancer (CRC) cell HCT116. Herein, a new series of quinazolinone derivatives with N3-substitution was designed and synthesized to improve the anticancer activity and explore the structure-activity relationship (SAR). After evaluating their BLM inhibitory activity, the SAR was discussed, leading to identifying compound 21 as a promising BLM inhibitor. 21 exhibited the potent BLM-dependent cytotoxicity against the CRC cells but weak against normal cells. Further evaluation revealed that 21 could disrupt the HRR level while inhibiting BLM located on the DSB site and trigger DNA damage in the CRC cells. This compound effectively suppressed the proliferation and invasion of CRC cells, along with cell cycle arrest and apoptosis. Consequently, 21 might be a promising candidate for treating CRC, and the BLM might be a new potential therapeutic target for CRC.

Bloom Syndrome Patients Display Accelerated Epigenetic Aging in Blood Lymphocytes

Bloom Syndrome (BSyn) is an autosomal recessive disorder caused by variants in the BLM genes at locus 15q26.1. The BLM gene encodes a protein that is part of the DNA RECQ helicase family, which has a role in homologous recombination, DNA structural integrity, DNA repair responses, and telomere maintenance. Patients with BSyn present with poor growth, sun sensitivity, mild immunodeficiency, diabetes, and increased risk of cancer, most commonly leukemias. Additionally, patients with BSyn develop myelodysplasia and life-threatening side effects from standard chemotherapy and radiation, often due to bone marrow toxicity. Cells from patients with BSyn display increased sister chromatid exchange, telomere defects, and telomere-dependent chromosome fusions. Despite these aging-associated findings, patients with BSyn do not appear to have typical premature aging characteristics such as graying hair, osteoporosis, and early-onset cardiovascular disease. We hypothesized that patients with BSyn may have increased epigenetic aging in blood, which may be related to their increased risk of hematologic dysfunction and malignancies. In this study, we aimed to use DNA methylation in order to identify an aging signature that could be a potential biomarker of disease risk. In partnership with the Bloom Syndrome Registry (BSR) as well as JScreen and our own clinical practice, we extracted 49 DNA samples from blood lymphocytes of patients with BSyn as well as carriers. The samples were then processed per standard Illumina protocol for the Infinium MethylationEPIC BeadChip kit. We utilized the online DNA Methylation Age Calculator (https://dnamage.genetics.ucla.edu/) to calculate multiple measures of methylation aging for each patient, including the Horvath pan-tissue epigenetic clock DNAmAge, the Hannum clock, the GrimAge lifespan predictor, and the PhenoAge phenotypic age estimate. Simple linear regression was used to determine statistical significance. Of the 49 total samples, 18 samples were from patients with BSyn (ages 1-38 years) and 31 samples were from carriers with one variant in BLM (ages 23-69 years). We discovered that BSyn patients had increased DNAm age compared to carriers measured by the pan-tissue clock (p=0.0054). Similarly, epigenetic clocks that were designed to predict human mortality risk (GrimAge and PhenoAge) indicate significant age acceleration as well, p<0.0088 and p<0.0001 respectively. Finally, DNA methylation-based estimators of telomere length show significantly shorter telomere lengths in BSyn patients compared to carriers, p<0.0001. Epigenetic age can be a better predictor of health and disease over biologic or chronologic age. In this study, we show for the first time that patients with BSyn have evidence of epigenetic aging across several measures in blood lymphocytes. Future studies are ongoing to investigate underlying mechanisms for this and whether epigenetic aging in these patients can predict hematologic dysfunction and cancer risk.

The Impact of Next Generation Sequencing in Diagnosis and Management of Rare Diseases: Bloom Syndrome

Bloom syndrome is an exceptionally rare autosomal recessive disorder characterized by a considerable genomic instability due to the defective DNA damage repair machine. It is caused by biallelic pathogenic variants in the gene encoding for one of the five human RecQ helicases, RECQL3/BLM. The disorder manifests clinically as growth deficiency, skin anomalies, immunodeficiencies, insulin resistance, and a high predisposition to cancers. Less than 300 patients have been reported so far. In this paper, we report on the first Romanian patient of bi-ethnic origin, molecularly diagnosed with Bloom syndrome. As the most severe complications of the disorder are the malignancies, developing even in childhood, an early diagnosis is essential for further surveillance and therapeutic approach of Bloom patients.

Beta Human Papillomavirus 8 E6 Induces Micronucleus Formation and Promotes Chromothripsis.

Cutaneous beta genus human papillomaviruses (β-HPVs) are suspected to promote the development of nonmelanoma skin cancer (NMSC) by destabilizing the host genome. Multiple studies have established the genome destabilizing capacities of β-HPV proteins E6 and E7 as a cofactor with UV. However, the E6 protein from β-HPV8 (HPV8 E6) induces tumors in mice without UV exposure. Here, we examined a UV-independent mechanism of HPV8 E6-induced genome destabilization. We showed that HPV8 E6 reduced the abundance of anaphase bridge resolving helicase, Bloom syndrome protein (BLM). The diminished BLM was associated with increased segregation errors and micronuclei. These HPV8 E6-induced micronuclei had disordered micronuclear envelopes but retained replication and transcription competence. HPV8 E6 decreased antiproliferative responses to micronuclei and time-lapse imaging revealed HPV8 E6 promoted cells with micronuclei to complete mitosis. Finally, whole-genome sequencing revealed that HPV8 E6 induced chromothripsis in nine chromosomes. These data provide insight into mechanisms by which HPV8 E6 induces genome instability independent of UV exposure. IMPORTANCE Some beta genus human papillomaviruses (β-HPVs) may promote skin carcinogenesis by inducing mutations in the host genome. Supporting this, the E6 protein from β-HPV8 (8 E6) promotes skin cancer in mice with or without UV exposure. Many mechanisms by which 8 E6 increases mutations caused by UV have been elucidated, but less is known about how 8 E6 induces mutations without UV. We address that knowledge gap by showing that 8 E6 causes mutations stemming from mitotic errors. Specifically, 8 E6 reduces the abundance of BLM, a helicase that resolves and prevents anaphase bridges. This hinders anaphase bridge resolution and increases their frequency. 8 E6 makes the micronuclei that can result from anaphase bridges more common. These micronuclei often have disrupted envelopes yet retain localization of nuclear-trafficked proteins. 8 E6 promotes the growth of cells with micronuclei and causes chromothripsis, a mutagenic process where hundreds to thousands of mutations occur in a chromosome.

Bloom Syndrome Helicase Compresses Single-Stranded DNA into Phase-Separated Condensates.

Bloom syndrome protein (BLM) is a conserved RecQ family helicase involved in the maintenance of genome stability. BLM has been widely recognized as a genome "caretaker" that processes structured DNA. In contrast, our knowledge of how BLM behaves on single-stranded (ss) DNA is still limited. Here, we demonstrate that BLM possesses the intrinsic ability for phase separation and can co-phase separate with ssDNA to form dynamically arrested protein/ssDNA co-condensates. The introduction of ATP potentiates the capability of BLM to condense on ssDNA, which further promotes the compression of ssDNA against a resistive force of up to 60 piconewtons. Moreover, BLM is also capable of condensing replication protein A (RPA)- or RAD51-coated ssDNA, before which it generates naked ssDNA by dismantling these ssDNA-binding proteins. Overall, our findings identify an unexpected characteristic of a DNA helicase and provide a new angle of protein/ssDNA co-condensation for understanding the genomic instability caused by BLM overexpression under diseased conditions.

Bloom Syndrome Helicase Compresses Single‐Stranded DNA into Phase‐Separated Condensates

Bloom syndrome protein (BLM) is a conserved RecQ family helicase involved in the maintenance of genome stability. BLM has been widely recognized as a genome “caretaker” that processes structured DNA. In contrast, our knowledge of how BLM behaves on single-stranded (ss) DNA is still limited. Here, we demonstrate that BLM possesses the intrinsic ability for phase separation and can co-phase separate with ssDNA to form dynamically arrested protein/ssDNA co-condensates. The introduction of ATP potentiates the capability of BLM to condense on ssDNA, which further promotes the compression of ssDNA against a resistive force of up to 60 piconewtons. Moreover, BLM is also capable of condensing replication protein A (RPA)- or RAD51-coated ssDNA, before which it generates naked ssDNA by dismantling these ssDNA-binding proteins. Overall, our findings identify an unexpected characteristic of a DNA helicase and provide a new angle of protein/ssDNA co-condensation for understanding the genomic instability caused by BLM overexpression under diseased conditions.

Downregulation of BLM RecQ helicase inhibits proliferation, promotes the apoptosis and enhances the sensitivity of bladder cancer cells to cisplatin.

Bloom syndrome protein (BLM) is known to maintain genomic integrity including DNA repair, recombination, replication and transcription. Its dysregulation affects the genomic instability of cells, which results in a high risk of developing various types of cancer and even Bloom syndrome. However, to date, to the best of our knowledge, no association has been made between human BLM and bladder cancer. Thus, the aim of the present study was to investigate the role of BLM in human bladder cancer. The expression pattern of BLM in bladder cancer tissue was detected by immunohistochemistry. The viability, proliferation, cell cycle and apoptosis of bladder cancer cell lines were determined by Cell Counting Kit-8, EdU and flow cytometry following transfection of BLM small interfering RNA. Finally, the effect of BLM on sensitivity of bladder cancer cell lines to cisplatin was investigated by reverse transcription-quantitative PCR and western blot. It was demonstrated that the expression of BLM in human bladder cancer was increased compared with adjacent healthy bladder tissues. In addition, silencing of BLM inhibited the proliferation and promoted the apoptosis of bladder cancer cells and it also enhanced the sensitivity of bladder cancer cells to cisplatin. Together, the findings of the present study demonstrated that the regulation of BLM activity may have potential for use as a novel therapeutic target and a predictor for the prognosis of bladder cancer.

Rare Association of Tuberous sclerosis with Acute Lymphoblastic Leukemia: Case Report with Review of Literature

AbstractAcute lymphoblastic leukemia (ALL) is the most common leukemia in children in which 85% of all cases are of B-cell ALL and approximately 15% cases are of T-cell ALL (T-ALL). Recent revolution in next-generation sequencing has uncovered many novel somatic mutations and rearrangements in ALL cells, which have prognostic and therapeutic implications, and it has also led to recognition of germline variants in the same genes with somatic mutations commonly associated with ALL. Apart from increasing the risk of developing ALL, germline variants may influence diagnostic testing, genetic counseling, and response to antileukemic treatment. This emphasizes importance of identification of new germline variants, or association of inherited syndromes with ALL or other malignancies. Down's syndrome, Shwachman's syndrome, Fanconi anemia, Bloom's syndrome, neurofibromatosis, and ataxia telangiectasia are well-recognized conditions associated with ALL. In this communication, we report a rare association of T-ALL with tuberous sclerosis (TS). This is the first reported case, showing association of T cell leukemia and TS with confirmatory genetic work-up.

BLM helicase inhibition synergizes with PARP inhibition to improve the radiosensitivity of olaparib resistant non-small cell lung cancer cells by inhibiting homologous recombination repair.

Objective:We aimed to investigate the radiosensitizing efficacy of the poly-ADP-ribose polymerase (PARP) inhibitor, olaparib, and the Bloom syndrome protein (BLM) helicase inhibitor, ML216, in non-small cell lung cancer (NSCLC) cells.Methods:Radiosensitization of NSCLC cells was assessed by colony formation and tumor growth assays. Mechanistically, the effects of ML216, olaparib, and radiation on cell and tumor proliferation, DNA damage, cell cycle, apoptosis, homologous recombination (HR) repair, and non-homologous end joining (NHEJ) repair activity were determined.Results:Both olaparib and ML216 enhanced the radiosensitivities of olaparib-sensitive H460 and H1299 cells, which was seen as decreased surviving fractions and Rad51 foci, increased total DNA damage, and γH2AX and 53BP1 foci (P < 0.05). The expressions of HR repair proteins were remarkably decreased in olaparib-treated H460 and H1299 cells after irradiation (P < 0.05), while olaparib combined with ML216 exerted a synergistic radiosensitization effect on olaparib-resistant A549 cells. In addition to increases of double strand break (DSB) damage and decreases of Rad51 foci, olaparib combined with ML216 also increased pDNA-PKcs (S2056) foci, abrogated G2 cell cycle arrest, and induced apoptosis in A549 lung cancer after irradiation in vitro and in vivo (P < 0.05). Moreover, Western blot showed that olaparib combined with ML216 and irradiation inhibited HR repair, promoted NHEJ repair, and inactivated cell cycle checkpoint signals both in vitro and in vivo (P < 0.05).Conclusions:Taken together, these results showed the efficacy of PARP and BLM helicase inhibitors for radiosensitizing NSCLC cells, and supported the model that BLM inhibition sensitizes cells to PARP inhibitor-mediated radiosensitization, as well as providing the basis for the potential clinical development of this combination for tumors intrinsically resistant to PARP inhibitors and radiotherapy.

Novel TOP3A Variant Associated With Mitochondrial Disease

ObjectivesTopoisomerase III alpha plays a key role in the dissolution of double Holliday junctions and is required for mitochondrial DNA (mtDNA) replication and maintenance. Sequence variants in the TOP3A gene have been associated with the Bloom syndrome–like disorder and described in an adult patient with progressive external ophthalmoplegia. The purpose of this report is to expand the clinical phenotype of the TOP3A-related diseases and clarify the role of this gene in primary mitochondrial disorders.MethodsA 44-year-old woman was referred to our hospital because of exercise intolerance and creatine kinase increase. Muscle biopsy and a targeted next-generation sequencing (NGS) analysis were performed.ResultsA histopathologic assessment documented a mitochondrial myopathy, and a molecular analysis revealed a novel homozygous variant in the TOP3A gene associated with multiple mtDNA deletions.DiscussionThis case suggests that TOP3A is one of the several nuclear genes associated with mtDNA maintenance disorder and expands the spectrum of its associated phenotypes, ranging from a clinical condition defined Bloom syndrome–like disorder to canonical mitochondrial syndromes.

Genomic characterization of lymphomas in patients with inborn errors of immunity

AbstractPatients with inborn errors of immunity (IEI) have a higher risk of developing cancer, especially lymphoma. However, the molecular basis for IEI-related lymphoma is complex and remains elusive. Here, we perform an in-depth analysis of lymphoma genomes derived from 23 IEI patients. We identified and validated disease-causing or -associated germline mutations in 14 of 23 patients involving ATM, BACH2, BLM, CD70, G6PD, NBN, PIK3CD, PTEN, and TNFRSF13B. Furthermore, we profiled somatic mutations in the lymphoma genome and identified 8 genes that were mutated at a significantly higher level in IEI-associated diffuse large B-cell lymphomas (DLBCLs) than in non-IEI DLBCLs, such as BRCA2, NCOR1, KLF2, FAS, CCND3, and BRWD3. The latter, BRWD3, is furthermore preferentially mutated in tumors of a subgroup of activated phosphoinositide 3-kinase δ syndrome patients. We also identified 5 genomic mutational signatures, including 2 DNA repair deficiency-related signatures, in IEI-associated lymphomas and a strikingly high number of inter- and intrachromosomal structural variants in the tumor genome of a Bloom syndrome patient. In summary, our comprehensive genomic characterization of lymphomas derived from patients with rare genetic disorders expands our understanding of lymphomagenesis and provides new insights for targeted therapy.

OTHR-25 Germline Mutations in Pediatric Brain Tumor

Abstract BACKGROUND: The exploration of germline mutations in pediatric brain tumor is helpful to improve the understanding of tumor genesis, guide the treatment, and provide genetic counseling for patients and their families. METHODS: In this study, children with primary brain tumor aged ≤18 years old in Guangdong Sanjiu Brain Hospital were enrolled．Tumor tissue or cerebrospinal fluid (CSF) next-generation sequencing (NGS) tests were required for all included patients. The genetic mutations in these children were analyzed and patients with germline mutations were screened. RESULTS: 135 pediatric patients with NGS detection were included in this patients. 63 patients were medulloblastoma, 54 were glioma. The rest were 5 ependymoma, 2other embryonal tumors, 2 pineoblastoma, 2 germ cell tumors, 1 pituitary adenoma, 1 neuroepithelial tumor, 1 Schwannoma, 1 neuronal tumor, 1 craniopharyngioma and 1 Rosai-Dorfman disease. NGS was performed on tissues of 122 patients and CSF of 13 patients. 7 patients underwent whole exome sequencing and the remaining 115 patients underwent panel detection. 6.67% (9/135) patients took germline mutations, which included 4 gliomas, 3 medulloblastomas, 1 schwannoma and 1 ependymoma. The rate of germline mutations were 7.41% (4/54) and 4.76% (3/63) in gliomas and medulloblastoma, respectively. The germinal mutations included TP53 missense mutation, BLM nonsense mutation, MSH2 nonsense mutation, MSH2 frameshift mutation, SUFU frameshift mutation, FANCI frameshift mutation, SBDS splicing site mutation, NF2 nonsense mutation and DIS3L2 nonsense mutation. These mutations are closely related to with Ligemeni syndrome, Bloom syndrome, Lynch syndrome, nevus basal cell carcinoma syndrome, Fanconi anemia, Shwachman-Diamond syndrome, neurofibromatosis type II and Perlman syndrome, respectively. 2 glioma patients had a family history of cancer. CONCLUSION: Pediatric patients with glioma, medulloblastoma, schwannoma and ependymoma are easily harboring germline mutations. For these patients, a detailed genetic history of family members is required and genetic counseling is recommended.