NOTCH1 pathway activating mutations and clonal evolution in pediatric T-cell acute lymphoblastic leukemia (T-ALL).

Cancer Sci. 2018 Nov 02;:

Authors: Kimura S, Seki M, Yoshida K, Shiraishi Y, Akiyama M, Koh K, Imamura T, Manabe A, Hayashi Y, Kobayashi M, Oka A, Miyano S, Ogawa S, Takita J

Abstract
Molecular mechanisms involved in the relapse of T-cell acute lymphoblastic leukemia (T-ALL) are not fully understood, although activating NOTCH1 signaling due to NOTCH1/FBXW7 alterations is a major oncogenic driver. To unravel the relevance of NOTCH1/FBXW7 mutations associated with relapse, we performed whole exome sequencing in 30 pediatric T-ALL cases, of which 11 diagnosis-relapse paired cases were further investigated to track the clonal evolution of relapse using amplicon-based deep sequencing. NOTCH1/FBXW7 alterations were detected in 73.3% (diagnosis) and 72.7% (relapse). Single nucleotide variations in the heterodimerization domain were the most frequent (40.0%) at diagnosis, whereas PEST domain alterations were the most frequent at relapse (54.5%). Comparison between non-relapsed and relapsed cases at diagnosis showed a predominance of PEST alterations in relapsed cases (P=0.045) although we failed to validate in TARGET cohort. Based on the clonal analysis of diagnosis-relapse samples, we identified NOTCH1 "switching" characterized by different NOTCH1 mutations in a major clone between diagnosis and relapse samples in two out of eleven diagnosis-relapse paired cases analyzed. We found another NOTCH1 "switching" case in previously reported Berlin-Frankfurt-Münster cohort (n = 13), indicating NOTCH1 importance in both the development and progression of T-ALL. Despite the limitations of small sample size and non minimal residual disease-based protocol, our results suggest that the presence of NOTCH1 mutations might contribute to the disease relapse of T-ALL. This article is protected by copyright. All rights reserved.

PMID: 30387229 [PubMed - as supplied by publisher]

LRBA Deficiency in a Patient With a Novel Homozygous Mutation Due to Chromosome 4 Segmental Uniparental Isodisomy.

Front Immunol. 2018;9:2397

Authors: Soler-Palacín P, Garcia-Prat M, Martín-Nalda A, Franco-Jarava C, Rivière JG, Plaja A, Bezdan D, Bosio M, Martínez-Gallo M, Ossowski S, Colobran R

Abstract
LRBA deficiency was first described in 2012 as an autosomal recessive disorder caused by biallelic mutations in the LRBA gene (OMIM #614700). It was initially characterized as producing early-onset hypogammaglobulinemia, autoimmune manifestations, susceptibility to inflammatory bowel disease, and recurrent infection. However, further reports expanded this phenotype (including patients without hypogammaglobulinemia) and described LRBA deficiency as a clinically variable syndrome with a wide spectrum of clinical manifestations. We present the case of a female patient who presented with type 1 diabetes, psoriasis, oral thrush, and enlarged liver and spleen at the age of 8 months. She later experienced recurrent bacterial and viral infections, including pneumococcal meningitis and Epstein Barr viremia. She underwent two consecutive stem cell transplants at the age of 8 and 9 years, and ultimately died. Samples from the patient and her parents were subjected to whole exome sequencing, which revealed a homozygous 1-bp insertion in exon 23 of the patient's LRBA gene, resulting in frameshift and premature stop codon. The patient's healthy mother was heterozygous for the mutation and her father tested wild-type. This finding suggested that either one copy of the paternal chromosome 4 bore a deletion including the LRBA locus, or the patient inherited two copies of the mutant maternal LRBA allele. The patient's sequencing data showed a 1-Mb loss of heterozygosity region in chromosome 4, including the LRBA gene. Comparative genomic hybridization array of the patient's and father's genomic DNA yielded normal findings, ruling out genomic copy number abnormalities. Here, we present the first case of LRBA deficiency due to a uniparental disomy (UPD). In contrast to classical Mendelian inheritance, UPD involves inheritance of 2 copies of a chromosomal region from only 1 parent. Specifically, our patient carried a small segmental isodisomy of maternal origin affecting 1 Mb of chromosome 4.

PMID: 30386343 [PubMed - in process]

Novel missense mutation in PTPN22 in a Chinese pedigree with Hashimoto's thyroiditis.

BMC Endocr Disord. 2018 Nov 01;18(1):76

Authors: Gong L, Liu B, Wang J, Pan H, Qi A, Zhang S, Wu J, Yang P, Wang B

Abstract
BACKGROUND: Hashimoto's thyroiditis is a complex autoimmune thyroid disease, the onset of which is associated with environmental exposures and specific susceptibility genes. Its incidence in females is higher than its incidence in males. Thus far, although some susceptibility loci have been elaborated, including PTPN22, FOXP3, and CD25, the aetiology and pathogenesis of Hashimoto's thyroiditis remains unclear.
METHODS: Four affected members from a Chinese family with Hashimoto's thyroiditis were selected for whole-exome sequencing. Missense, nonsense, frameshift, or splicing-site variants shared by all affected members were identified after frequency filtering against public and internal exome databases. Segregation analysis was performed by Sanger sequencing among all members with available DNA.
RESULTS: We identified a missense mutation in PTPN22 (NM_015967.5; c. 77A > G; p.Asn26Ser) using whole-exome sequencing. PTPN22 is a known susceptibility gene associated with increased risks of multiple autoimmune diseases. Cosegregation analysis confirmed that all patients in this family, all of whom were female, carried the mutation. All public and private databases showed that the missense mutation was extremely rare.
CONCLUSIONS: We found a missense mutation in PTPN22 in a Chinese HT pedigree using whole-exome sequencing. Our study, for the first time, linked a rare variant of PTPN22 to Hashimoto's thyroiditis, providing further evidence of the disease-causing or susceptibility role of PTPN22 in autoimmune thyroid disease. Functional studies regarding the effects of this variant on thyroid autoimmunity and thyroid function are warranted.

PMID: 30384852 [PubMed - in process]

Prognostic utility of six mutated genes for older patients with acute myeloid leukemia.

Int J Cancer. 2018 04 15;142(8):1664-1670

Authors: Wang J, Ma Z, Wang Q, Guo Q, Huang J, Yu W, Wang H, Huang J, Washington Shao Y, Chen S, Jin J

Abstract
Approximately 50% of older patients with acute myeloid leukemia (AML) do not obtain chromosomal abnormalities as an effective risk-stratification, and present cytogenetically normal AML (CN-AML). To develop a reliable prediction model for stratifying the risk of these elderly patients, we conducted a study with a discovery and validation design. As a result, we found the top 6 mutated genes in the discovery cohort of 26 case by the whole exome sequencing, and verified as recurrent mutations in the large cohort of 329 patients by Sanger sequencing. The top 6 genes were NPM1, FLT3-ITD, DNMT3A, CEBPA double allele, IDH1 and IDH2 mutations, and the frequency of each gene in the combining cohort was 36.8%, 19.8%, 20.1%, 5.8%, 14.9% and 22.5%, respectively. In addition, clinical variables such as age, white blood cell counts, genes of IDH1 and DNMT3A mutations, European LeukemiaNet genotype (NPM1 mutations and lacking FLT3-ITD or CEBPA double allele mutations) and treatment protocols were independent factors for predicting the probabilities of overall and event-free survival. The prediction nomograms based on these significant factors showed accurate discrimination. In conclusion, we developed a reliable prediction model for stratifying the risk of elderly patients with CN-AML.

PMID: 29193057 [PubMed - indexed for MEDLINE]

KDF1 is a novel candidate gene of non-syndromic tooth agenesis.

Arch Oral Biol. 2018 Oct 23;97:131-136

Authors: Zeng B, Lu H, Xiao X, Yu X, Li S, Zhu L, Yu D, Zhao W

Abstract
OBJECTIVE: Tooth agenesis (TA) is featured by congenital loss of teeth, and can be divided into two subtypes, non-syndromic TA (NSTA) and syndromic TA (STA). Although 12 candidate genes of NSTA have been revealed, the genetic basis of NSTA needs to be further studied. We noticed an overlap of candidate genes between NSTA and STA, and hypothesized that some candidate genes of STA may be new candidate genes of NSTA.
METHODS: Sanger sequencing, whole exome sequencing, bioinformatics analyses and immunohistochemical staining were performed to reveal the genetic basis of the patients in a family with NSTA.
RESULTS: No pathogenic mutation was found in the 12 candidate genes of NSTA. We screened the variants of 76 STA candidate genes and identified a novel pathogenic mutation c.G908C (p.R303 P) in Keratinocyte Differentiation Factor 1 (KDF1). This mutation was cosegregated with the disease in the family. Bioinformatics analyses predicted the mutation to be pathogenic. Immunohistochemical staining of kdf1 in developing tooth germs indicated that kdf1 expression is important for the development of teeth.
CONCLUSIONS: This study identified KDF1 as a novel candidate gene for NSTA. STA candidate genes may be a promising source of new NSTA genes.

PMID: 30384154 [PubMed - as supplied by publisher]

Landscape of genomic alterations in high-grade serous ovarian cancer from exceptional long- and short-term survivors.

Genome Med. 2018 Oct 31;10(1):81

Authors: Yang SYC, Lheureux S, Karakasis K, Burnier JV, Bruce JP, Clouthier DL, Danesh A, Quevedo R, Dowar M, Hanna Y, Li T, Lu L, Xu W, Clarke BA, Ohashi PS, Shaw PA, Pugh TJ, Oza AM

Abstract
BACKGROUND: Patients diagnosed with high-grade serous ovarian cancer (HGSOC) who received initial debulking surgery followed by platinum-based chemotherapy can experience highly variable clinical responses. A small percentage of women experience exceptional long-term survival (long term (LT), 10+ years), while others develop primary resistance to therapy and succumb to disease in less than 2 years (short term (ST)). To improve clinical management of HGSOC, there is a need to better characterize clinical and molecular profiles to identify factors that underpin these disparate survival responses.
METHODS: To identify clinical and tumor molecular biomarkers associated with exceptional clinical response or resistance, we conducted an integrated clinical, exome, and transcriptome analysis of 41 primary tumors from LT (n = 20) and ST (n = 21) HGSOC patients.
RESULTS: Younger age at diagnosis, no residual disease post debulking surgery and low CA125 levels following surgery and chemotherapy were clinical characteristics of LT. Tumors from LT survivors had increased somatic mutation burden (median 1.62 vs. 1.22 non-synonymous mutations/Mbp), frequent BRCA1/2 biallelic inactivation through mutation and loss of heterozygosity, and enrichment of activated CD4+, CD8+ T cells, and effector memory CD4+ T cells. Characteristics of ST survival included focal copy number gain of CCNE1, lack of BRCA mutation signature, low homologous recombination deficiency scores, and the presence of ESR1-CCDC170 gene fusion.
CONCLUSIONS: Our findings suggest that exceptional long- or short-term survival is determined by a concert of clinical, molecular, and microenvironment factors.

PMID: 30382883 [PubMed - in process]

Recovery of T-cell receptor V(D)J recombination reads from lower grade glioma exome files correlates with reduced survival and advanced cancer grade.

J Neurooncol. 2018 Oct 31;:

Authors: Chobrutskiy BI, Zaman S, Tong WL, Diviney A, Blanck G

Abstract
BACKGROUND: The immune system plays an important role in cancer survival and disease progression, but the role of the immune system in lower grade glioma (LGG) is largely unknown METHODS: To investigate the relationship between lymphocyte infiltration into the LGG microenvironment and LGG survival, we used a genomics approach to recover productive V(D)J recombination sequences from primary tumor, whole exome sequence files available via the cancer genome atlas RESULTS: Increased T-cell receptor V(D)J read recovery, indicating increased T-lymphocyte infiltration into the primary tumor site, strongly correlated with decreased overall and disease-free survival; and with a more advanced cancer grade. In addition, this result was more significant than related results obtainable using RNASeq-based, T-cell biomarkers, similar to a recently reported case for pancreatic cancer, where the recovery of BCR recombination reads from WXS files clearly associated with reduced survival, despite the fact that no such association was demonstrable with B-cell based, RNASeq biomarkers CONCLUSIONS: Overall, the results presented here support V(D)J recombination read recovery, from whole exome files, as a uniquely useful biomarker for distinct LGG survival rates.

PMID: 30382482 [PubMed - as supplied by publisher]

"Infostery" analysis of short molecular dynamics simulations identifies highly sensitive residues and predicts deleterious mutations.

Sci Rep. 2018 Oct 31;8(1):16126

Authors: Karami Y, Bitard-Feildel T, Laine E, Carbone A

Abstract
Characterizing a protein mutational landscape is a very challenging problem in Biology. Many disease-associated mutations do not seem to produce any effect on the global shape nor motions of the protein. Here, we use relatively short all-atom biomolecular simulations to predict mutational outcomes and we quantitatively assess the predictions on several hundreds of mutants. We perform simulations of the wild type and 175 mutants of PSD95's third PDZ domain in complex with its cognate ligand. By recording residue displacements correlations and interactions, we identify "communication pathways" and quantify them to predict the severity of the mutations. Moreover, we show that by exploiting simulations of the wild type, one can detect 80% of the positions highly sensitive to mutations with a precision of 89%. Importantly, our analysis describes the role of these positions in the inter-residue communication and dynamical architecture of the complex. We assess our approach on three different systems using data from deep mutational scanning experiments and high-throughput exome sequencing. We refer to our analysis as "infostery", from "info" - information - and "steric" - arrangement of residues in space. We provide a fully automated tool, COMMA2 ( www.lcqb.upmc.fr/COMMA2 ), that can be used to guide medicinal research by selecting important positions/mutations.

PMID: 30382169 [PubMed - in process]

Recurrent CCND3 mutations in MLL-rearranged acute myeloid leukemia.

Blood Adv. 2018 Nov 13;2(21):2879-2889

Authors: Matsuo H, Yoshida K, Fukumura K, Nakatani K, Noguchi Y, Takasaki S, Noura M, Shiozawa Y, Shiraishi Y, Chiba K, Tanaka H, Okada A, Nannya Y, Takeda J, Ueno H, Shiba N, Yamato G, Handa H, Ono Y, Hiramoto N, Ishikawa T, Usuki K, Ishiyama K, Miyawaki S, Itonaga H, Miyazaki Y, Kawamura M, Yamaguchi H, Kiyokawa N, Tomizawa D, Taga T, Tawa A, Hayashi Y, Mano H, Miyano S, Kamikubo Y, Ogawa S, Adachi S

Abstract
In acute myeloid leukemia (AML), MLL (KMT2A) rearrangements are among the most frequent chromosomal abnormalities; however, knowledge of the genetic landscape of MLL-rearranged AML is limited. In this study, we performed whole-exome sequencing (n = 9) and targeted sequencing (n = 56) of samples from pediatric MLL-rearranged AML patients enrolled in the Japanese Pediatric Leukemia/Lymphoma Study Group AML-05 study. Additionally, we analyzed 105 pediatric t(8;21) AML samples and 30 adult MLL-rearranged AML samples. RNA-sequencing data from 31 patients published in a previous study were also reanalyzed. As a result, we identified 115 mutations in pediatric MLL-rearranged AML patients (2.1 mutations/patient), with mutations in signaling pathway genes being the most frequently detected (60.7%). Mutations in genes associated with epigenetic regulation (21.4%), transcription factors (16.1%), and the cohesin complex (8.9%) were also commonly detected. Novel CCND3 mutations were identified in 5 pediatric MLL-rearranged AML patients (8.9%) and 2 adult MLL-rearranged AML patients (3.3%). Recurrent mutations of CCND1 (n = 3, 2.9%) and CCND2 (n = 8, 7.6%) were found in pediatric t(8;21) AML patients, whereas no CCND3 mutations were found, suggesting that D-type cyclins exhibit a subtype-specific mutation pattern in AML. Treatment of MLL-rearranged AML cell lines with CDK4/6 inhibitors (abemaciclib and palbociclib) blocked G1 to S phase cell-cycle progression and impaired proliferation. Pediatric MLL-MLLT3-rearranged AML patients with coexisting mutations (n = 16) had significantly reduced relapse-free survival and overall survival compared with those without coexisting mutations (n = 9) (P = .048 and .046, respectively). These data provide insights into the genetics of MLL-rearranged AML and suggest therapeutic strategies.

PMID: 30381403 [PubMed - in process]

Heterozygous STUB1 mutation causes familial ataxia with cognitive affective syndrome (SCA48).

Neurology. 2018 Oct 31;:

Authors: Genis D, Ortega-Cubero S, San Nicolás H, Corral J, Gardenyes J, de Jorge L, López E, Campos B, Lorenzo E, Tonda R, Beltran S, Negre M, Obón M, Beltran B, Fàbregas L, Alemany B, Márquez F, Ramió-Torrentà L, Gich J, Volpini V, Pastor P

Abstract
OBJECTIVE: To describe a new spinocerebellar ataxia (SCA48) characterized by early cerebellar cognitive-affective syndrome (CCAS) and late-onset SCA.
METHODS: This is a descriptive study of a family that has been followed for more than a decade with periodic neurologic and neuropsychological examinations, MRI, brain SPECT perfusion, and genetic analysis. Whole exome sequencing was performed in 3 affected and 1 unaffected family member and subsequently validated by linkage analysis of chromosome 16p13.3.
RESULTS: Six patients fully developed cognitive-affective and complete motor cerebellar syndrome associated with vermian and hemispheric cerebellar atrophy, suggesting a continuum from a dysexecutive syndrome slowly evolving to a complete and severe CCAS with late truncal ataxia. Three presymptomatic patients showed focal cerebellar atrophy in the vermian, paravermian, and the medial part of cerebellar lobes VI and VII, suggesting that cerebellar atrophy preceded the ataxia, and that the neurodegeneration begins in cerebellar areas related to cognition and emotion, spreading later to the whole cerebellum. Among the candidate variants, only the frameshift heterozygous c.823_824delCT STUB1 (p.L275Dfs*16) pathogenic variant cosegregated with the disease. The p.L275Dfs*16 heterozygous STUB1 pathogenic variant leads to neurodegeneration and atrophy in cognition- and emotion-related cerebellar areas and reinforces the importance of STUB1 in maintaining cognitive cerebellar function.
CONCLUSIONS: We report a heterozygous STUB1 pathogenic genetic variant causing dominant cerebellar ataxia. Since recessive mutations in STUB1 gene have been previously associated with SCAR16, these findings suggest a previously undescribed SCA locus (SCA48; MIM# 618093).

PMID: 30381368 [PubMed - as supplied by publisher]

TIRAP p.R81C is a novel lymphoma risk variant which enhances cell proliferation via NF-κB mediated signaling in B-cells.

Haematologica. 2018 Oct 31;:

Authors: Burkhard R, Keller I, Arambasic M, Juskevicius D, Tzankov A, Lundberg P, Bruggman R, Dirnhofer S, Radpour R, Novak U

Abstract
Diffuse large B-cell lymphoma is the most common malignant lymphoma in adults. By gene-expression profiling, this lymphoma is divided in three cell-of-origin subtypes with distinct molecular and clinical features. Most lymphomas arise sporadically, yet familial clustering is known, suggesting a genetic contribution to disease risk. Familial lymphoma cases are a valuable tool to investigate risk genes. We studied a Swiss/Japanese family with two sisters affected by a primary mediastinal B-cell lymphoma and a non-germinal center diffuse large B-cell lymphoma not otherwise specified, respectively. The somatic landscape of both lym-phomas was marked by alterations affecting multiple components of the JAK-STAT pathway. Consequently, this pathway was constitutively activated as evidenced by high pJAK2 as well as increased nuclear pSTAT3 and pSTAT6 in malignant cells. Potential lymphoma risk vari-ants were identified by whole-exome sequencing of the germline DNA derived from siblings and unaffected family members. This analysis revealed a pathogenic variant in TIRAP, an upstream regulator of NF-κB, in both affected siblings and their mother. We observed in-creased B-cell proliferation in family members harboring the TIRAP p.R81C variant. B-cell proliferation correlated with TIRAP and NF-κB target gene expression, suggesting enhanced NF-κB pathway activity in TIRAP p.R81C individuals. TIRAP knockdown reduced B-cell sur-vival and NF-κB target gene expression, particularly in individuals with TIRAP p.R81C. Func-tional studies revealed significantly increased NF-κB activity and resistance to stress-induced cell-death by TIRAP p.R81C. The identification of an inherited TIRAP variant pro-vides evidence for a novel link between genetic alterations affecting the NF-κB pathway and lymphomagenesis.

PMID: 30381301 [PubMed - as supplied by publisher]

Blastic plasmacytoid dendritic cell neoplasm: genomics mark epigenetic dysregulation as a primary therapeutic target.

Haematologica. 2018 Oct 31;:

Authors: Sapienza MR, Abate F, Melle F, Orecchioni S, Fuligni F, Etebari M, Tabanelli V, Laginestra MA, Pileri A, Motta G, Rossi M, Agostinelli C, Sabattini E, Pimpinelli N, Truni M, Falini B, Cerroni L, Talarico G, Piccioni R, Amente S, Indio V, Tarantino G, Brundu F, Paulli M, Berti E, Facchetti F, Dellino GI, Bertolini F, Tripodo C, Rabadan R, Pileri SA

Abstract
Blastic Plasmacytoid Dendritic Cell Neoplasm is a rare and aggressive hematological malignancy currently lacking an effective therapy. To possibly identify genetic alterations useful for a new treatment design, we analyzed by whole-exome sequencing fourteen Blastic Plasmacytoid Dendritic Cell Neoplasm patients and the patient-derived CAL-1 cell line. The functional enrichment analysis of mutational data reported the epigenetic regulatory program as the most significantly undermined (P<.0001). In particular, twenty-five epigenetic-modifiers were found mutated (e.g., ASXL1, TET2, SUZ12, ARID1A, PHF2, CHD8); ASXL1 was the most frequently affected (28.6% of cases). To evaluate the impact of the identified epigenetic mutations at the gene-expression and Histone H3 lysine 27 trimethylation/acetylation levels, we performed additional RNA and Pathology tissue-chromatin immunoprecipitation sequencing experiments; the patients displayed enrichment in gene-signatures regulated by methylation and modifiable by Decitabine administration, shared common H3K27-acetylated regions and featured a set of cell-cycle genes aberrantly up-regulated and marked by promoter acetylation. Collectively, the integration of sequencing data showed the potential of a therapy based on epigenetic agents. Through the adoption of a preclinical Blastic Plasmacytoid Dendritic Cell Neoplasm mouse model, established by the CAL-1 cell line xenografting, we demonstrated the efficacy of the combination of the epigenetic drugs 5'-Azacytidine and Decitabine in controlling the disease progression in vivo.

PMID: 30381297 [PubMed - as supplied by publisher]

Pitfalls of immunotherapy: lessons from a patient with CTLA-4 haploinsufficiency.

Allergy Asthma Clin Immunol. 2018;14:65

Authors: Watson LR, Slade CA, Ojaimi S, Barnes S, Fedele P, Smith P, Marum J, Lunke S, Stark Z, Hunter MF, Bryant VL, Low MSY

Abstract
Background: Daclizumab is a humanized monoclonal antibody that blocks CD25, the high affinity alpha subunit of the interleukin-2 receptor. Daclizumab therapy targets T regulatory cell and activated effector T cell proliferation to suppress autoimmune disease activity, in inflammatory conditions like relapsing and remitting multiple sclerosis. Here, we present the first report of agranulocytosis with daclizumab therapy in a patient with relapsing and remitting multiple sclerosis.
Case presentation: Our patient was a 24-year-old Australian female with a clinical history of atopy, lymphocytic enteritis complicated by B12 deficiency, relapsing and remitting multiple sclerosis, recurrent lower respiratory tract infections, vulval/cervical intraepithelial neoplasia and melanoma. She was commenced on daclizumab therapy after failing several lines of treatment for relapsing and remitting multiple sclerosis. During a hospital admission for lymphocytic enteritis, she was incidentally diagnosed with combined immunodeficiency with hypogammaglobulinaemia and declined proposed regular intravenous immunoglobulin infusions. Following six months of daclizumab therapy, our patient presented to hospital with febrile neutropenia. No clear infective cause was found, despite numerous investigations. However, bone marrow biopsy revealed agranulocytosis with an apparent maturation block at the myeloblasts stage. Neustrophil recovery occurred following cessation of daclizumab and the initiation of T cell immunosuppressive agents including systemic corticosteroids and methotrexate. The patient was further investigated for combined immunodeficiency and whole exome sequencing revealed a novel heterozygous missense variant in cytotoxic T lymphocyte antigen 4 (CTLA4), leading to a diagnosis of CTLA-4 haploinsufficiency with autoimmune infiltration (CHAI).
Conclusion: This case demonstrates that autoimmune disease may be the presenting feature of primary immunodeficiency and should be appropriately investigated prior to the commencement of immunotherapy. Genetic clarification of underlying primary immunodeficiency may provide critical clinical information that alters the safety of the proposed treatment strategy.

PMID: 30377434 [PubMed]

Immune Escape of Relapsed AML Cells after Allogeneic Transplantation.

N Engl J Med. 2018 Oct 31;:

Authors: Christopher MJ, Petti AA, Rettig MP, Miller CA, Chendamarai E, Duncavage EJ, Klco JM, Helton NM, O'Laughlin M, Fronick CC, Fulton RS, Wilson RK, Wartman LD, Welch JS, Heath SE, Baty JD, Payton JE, Graubert TA, Link DC, Walter MJ, Westervelt P, Ley TJ, DiPersio JF

Abstract
BACKGROUND: As consolidation therapy for acute myeloid leukemia (AML), allogeneic hematopoietic stem-cell transplantation provides a benefit in part by means of an immune-mediated graft-versus-leukemia effect. We hypothesized that the immune-mediated selective pressure imposed by allogeneic transplantation may cause distinct patterns of tumor evolution in relapsed disease.
METHODS: We performed enhanced exome sequencing on paired samples obtained at initial presentation with AML and at relapse from 15 patients who had a relapse after hematopoietic stem-cell transplantation (with transplants from an HLA-matched sibling, HLA-matched unrelated donor, or HLA-mismatched unrelated donor) and from 20 patients who had a relapse after chemotherapy. We performed RNA sequencing and flow cytometry on a subgroup of these samples and on additional samples for validation.
RESULTS: On exome sequencing, the spectrum of gained and lost mutations observed with relapse after transplantation was similar to the spectrum observed with relapse after chemotherapy. Specifically, relapse after transplantation was not associated with the acquisition of previously unknown AML-specific mutations or structural variations in immune-related genes. In contrast, RNA sequencing of samples obtained at relapse after transplantation revealed dysregulation of pathways involved in adaptive and innate immunity, including down-regulation of major histocompatibility complex (MHC) class II genes ( HLA-DPA1, HLA-DPB1, HLA-DQB1, and HLA-DRB1) to levels that were 3 to 12 times lower than the levels seen in paired samples obtained at presentation. Flow cytometry and immunohistochemical analysis confirmed decreased expression of MHC class II at relapse in 17 of 34 patients who had a relapse after transplantation. Evidence suggested that interferon-γ treatment could rapidly reverse this phenotype in AML blasts in vitro.
CONCLUSIONS: AML relapse after transplantation was not associated with the acquisition of relapse-specific mutations in immune-related genes. However, it was associated with dysregulation of pathways that may influence immune function, including down-regulation of MHC class II genes, which are involved in antigen presentation. These epigenetic changes may be reversible with appropriate therapy. (Funded by the National Cancer Institute and others.).

PMID: 30380364 [PubMed - as supplied by publisher]

Exome sequencing in large, multiplex bipolar disorder families from Cuba.

PLoS One. 2018;13(10):e0205895

Authors: Maaser A, Forstner AJ, Strohmaier J, Hecker J, Ludwig KU, Sivalingam S, Streit F, Degenhardt F, Witt SH, Reinbold CS, Koller AC, Raff R, Heilmann-Heimbach S, Fischer SB, Bipolar Disorder Working Group of the Psychiatric Genomics Consortium, Herms S, Hoffmann P, Thiele H, Nürnberg P, Löhlein Fier H, Orozco-Díaz G, Carmenate-Naranjo D, Proenza-Barzaga N, Auburger GWJ, Andlauer TFM, Cichon S, Marcheco-Teruel B, Mors O, Rietschel M, Nöthen MM

Abstract
Bipolar disorder (BD) is a major psychiatric illness affecting around 1% of the global population. BD is characterized by recurrent manic and depressive episodes, and has an estimated heritability of around 70%. Research has identified the first BD susceptibility genes. However, the underlying pathways and regulatory networks remain largely unknown. Research suggests that the cumulative impact of common alleles with small effects explains only around 25-38% of the phenotypic variance for BD. A plausible hypothesis therefore is that rare, high penetrance variants may contribute to BD risk. The present study investigated the role of rare, nonsynonymous, and potentially functional variants via whole exome sequencing in 15 BD cases from two large, multiply affected families from Cuba. The high prevalence of BD in these pedigrees renders them promising in terms of the identification of genetic risk variants with large effect sizes. In addition, SNP array data were used to calculate polygenic risk scores for affected and unaffected family members. After correction for multiple testing, no significant increase in polygenic risk scores for common, BD-associated genetic variants was found in BD cases compared to healthy relatives. Exome sequencing identified a total of 17 rare and potentially damaging variants in 17 genes. The identified variants were shared by all investigated BD cases in the respective pedigree. The most promising variant was located in the gene SERPING1 (p.L349F), which has been reported previously as a genome-wide significant risk gene for schizophrenia. The present data suggest novel candidate genes for BD susceptibility, and may facilitate the discovery of disease-relevant pathways and regulatory networks.

PMID: 30379966 [PubMed - in process]

Biallelic sequence and structural variants in RAX2 are a novel cause for autosomal recessive inherited retinal disease.

Genet Med. 2018 Oct 31;:

Authors: Van de Sompele S, Smith C, Karali M, Corton M, Van Schil K, Peelman F, Cherry T, Rosseel T, Verdin H, Derolez J, Van Laethem T, Khan KN, McKibbin M, Toomes C, Ali M, Torella A, Testa F, Jimenez B, Simonelli F, De Zaeytijd J, Van den Ende J, Leroy BP, Coppieters F, Ayuso C, Inglehearn CF, Banfi S, De Baere E

Abstract
PURPOSE: RAX2 encodes a homeobox-containing transcription factor, in which four monoallelic pathogenic variants have been described in autosomal dominant cone-dominated retinal disease.
METHODS: Exome sequencing in a European cohort with inherited retinal disease (IRD) (n = 2086) was combined with protein structure modeling of RAX2 missense variants, bioinformatics analysis of deletion breakpoints, haplotyping of RAX2 variant c.335dup, and clinical assessment of biallelic RAX2-positive cases and carrier family members.
RESULTS: Biallelic RAX2 sequence and structural variants were found in five unrelated European index cases, displaying nonsyndromic autosomal recessive retinitis pigmentosa (ARRP) with an age of onset ranging from childhood to the mid-40s (average mid-30s). Protein structure modeling points to loss of function of the novel recessive missense variants and to a dominant-negative effect of the reported dominant RAX2 alleles. Structural variants were fine-mapped to disentangle their underlying mechanisms. Haplotyping of c.335dup in two cases suggests a common ancestry.
CONCLUSION: This study supports a role for RAX2 as a novel disease gene for recessive IRD, broadening the mutation spectrum from sequence to structural variants and revealing a founder effect. The identification of biallelic RAX2 pathogenic variants in five unrelated families shows that RAX2 loss of function may be a nonnegligible cause of IRD in unsolved ARRP cases.

PMID: 30377383 [PubMed - as supplied by publisher]

Improved diagnostics by exome sequencing following raw data reevaluation by clinical geneticists involved in the medical care of the individuals tested.

Genet Med. 2018 Oct 31;:

Authors: Salmon LB, Orenstein N, Markus-Bustani K, Ruhrman-Shahar N, Kilim Y, Magal N, Hubshman MW, Bazak L

Abstract
PURPOSE: Reanalysis of exome sequencing data when results are negative may yield additional diagnoses. We sought to estimate the contribution of clinical geneticists to the interpretation of sequencing data of their patients.
METHODS: The cohort included 84 probands attending a tertiary genetics institute (2015-2018) with a nondiagnostic result on clinical exome sequencing performed in one of five external laboratories. The raw data were uploaded to the Emedgene bioinformatics and interpretation platform for reanalysis by a team of two clinical geneticists, the geneticist directly involved in the patient's care, and a bioinformatician.
RESULTS: In ten probands (11.9%), a new definitive diagnosis was reached based on genes that were known to be associated with the phenotype at the time the original report was issued. The main reasons for a negative exome result were incorrect interpretation of the clinical context and absence of OMIM entry. Pathogenic variants in genes with previously unknown gene-disease associations were discovered to be causative in three probands. In total, new diagnoses were established in 13/84 individuals (15.5%).
CONCLUSION: Direct access to complete clinical data and shortening of time to including gene-phenotype associations in databases can assist the analytics team and reduce the need for additional unnecessary tests.

PMID: 30377382 [PubMed - as supplied by publisher]

Characterization of Squamous Cell Lung Cancers from Appalachian Kentucky.

Cancer Epidemiol Biomarkers Prev. 2018 Oct 30;:

Authors: Liu J, Murali T, Yu T, Liu C, Sivakumaran TA, Moseley HNB, Zhulin IB, Weiss HL, Durbin EB, Ellingson SR, Liu J, Huang B, Hallahan BJ, Horbinski CM, Hodges K, Napier DL, Bocklage T, Mueller J, Vanderford N, Fardo DW, Wang C, Arnold SM

Abstract
BACKGROUND: Lung cancer is the leading cause of cancer mortality in the United States (US). Squamous cell carcinoma (SQCC) represents 22.6% of all lung cancers nationally, and 26.4% in Appalachian Kentucky (AppKY), where death from lung cancer is exceptionally high. The Cancer Genome Atlas (TCGA) characterized genetic alterations in lung SQCC, but this cohort did not focus on AppKY residents.
METHODS: Whole exome sequencing was performed on tumor and normal DNA samples from 51 lung SQCC subjects from AppKY. Somatic genomic alterations were compared between the AppKY and TCGA SQCC cohorts.
RESULTS: From this AppKY cohort, we identified an average of 237 non-silent mutations per patient and, in comparison to TCGA, we found that PCMTD1 (18%) and IDH1 (12%) were more commonly altered in AppKY versus TCGA. Using IDH1 as a starting point, we identified a mutually exclusive mutational pattern (IDH1, KDM6A, KDM4E, JMJD1C) involving functionally-related genes. We also found actionable mutations (10%) and/or intermediate or high tumor mutation burden (65%), indicating potential therapeutic targets in 65% of subjects.
CONCLUSIONS: This study has identified an increased percentage of IDH1 and PCMTD1 mutations in SQCC arising in the AppKY residents versus TCGA, with population-specific implications for the personalized treatment of this disease.
IMPACT: Our study is the first report to characterize genomic alterations in lung SQCC from AppKY. These findings suggest population differences in the genetics of lung SQCC between AppKY and US populations, highlighting the importance of the relevant population when developing personalized treatment approaches for this disease.

PMID: 30377206 [PubMed - as supplied by publisher]

Whole Exome Sequencing Is the Preferred Strategy to Identify the Genetic Defect in Patients With a Probable or Possible Mitochondrial Cause.

Front Genet. 2018;9:400

Authors: Theunissen TEJ, Nguyen M, Kamps R, Hendrickx AT, Sallevelt SCEH, Gottschalk RWH, Calis CM, Stassen APM, de Koning B, Mulder-Den Hartog ENM, Schoonderwoerd K, Fuchs SA, Hilhorst-Hofstee Y, de Visser M, Vanoevelen J, Szklarczyk R, Gerards M, de Coo IFM, Hellebrekers DMEI, Smeets HJM

Abstract
Mitochondrial disorders, characterized by clinical symptoms and/or OXPHOS deficiencies, are caused by pathogenic variants in mitochondrial genes. However, pathogenic variants in some of these genes can lead to clinical manifestations which overlap with other neuromuscular diseases, which can be caused by pathogenic variants in non-mitochondrial genes as well. Mitochondrial pathogenic variants can be found in the mitochondrial DNA (mtDNA) or in any of the 1,500 nuclear genes with a mitochondrial function. We have performed a two-step next-generation sequencing approach in a cohort of 117 patients, mostly children, in whom a mitochondrial disease-cause could likely or possibly explain the phenotype. A total of 86 patients had a mitochondrial disorder, according to established clinical and biochemical criteria. The other 31 patients had neuromuscular symptoms, where in a minority a mitochondrial genetic cause is present, but a non-mitochondrial genetic cause is more likely. All patients were screened for pathogenic variants in the mtDNA and, if excluded, analyzed by whole exome sequencing (WES). Variants were filtered for being pathogenic and compatible with an autosomal or X-linked recessive mode of inheritance in families with multiple affected siblings and/or consanguineous parents. Non-consanguineous families with a single patient were additionally screened for autosomal and X-linked dominant mutations in a predefined gene-set. We identified causative pathogenic variants in the mtDNA in 20% of the patient-cohort, and in nuclear genes in 49%, implying an overall yield of 68%. We identified pathogenic variants in mitochondrial and non-mitochondrial genes in both groups with, obviously, a higher number of mitochondrial genes affected in mitochondrial disease patients. Furthermore, we show that 31% of the disease-causing genes in the mitochondrial patient group were not included in the MitoCarta database, and therefore would have been missed with MitoCarta based gene-panels. We conclude that WES is preferable to panel-based approaches for both groups of patients, as the mitochondrial gene-list is not complete and mitochondrial symptoms can be secondary. Also, clinically and genetically heterogeneous disorders would require sequential use of multiple different gene panels. We conclude that WES is a comprehensive and unbiased approach to establish a genetic diagnosis in these patients, able to resolve multi-genic disease-causes.

PMID: 30369941 [PubMed]

Spontaneous Posterior Segment Vascular Disease Phenotype of a Mouse Model, rnv3, Is Dependent on the Crb1rd8 Allele.

Invest Ophthalmol Vis Sci. 2018 Oct 01;59(12):5127-5139

Authors: Chang B, FitzMaurice B, Wang J, Low BE, Wiles MV, Nishina PM

Abstract
Purpose: To determine the molecular basis of lesion development in a murine model of spontaneous retinal vascularization, rnv3 (retinal vascularization 3, aka JR5558).
Methods: Disease progression of rnv3 was examined in longitudinal studies by clinical evaluation, electroretinography (ERG) and light microscopy analyses. The chromosomal position for the recessive rnv3 mutation was determined by DNA pooling and genome-wide linkage analysis. The causative mutation was discovered by comparison of whole exome sequences of rnv3 mutant and wild-type (WT) controls. In order to confirm the causative mutation, transcription activator-like effector nuclease (TALEN)-mediated oligonucleotide directed repair (ODR) was utilized to correct the mutant allele. Phenotypic correction was assessed by fundus imaging and optical coherence tomography of live mice.
Results: rnv3 exhibits early-onset, multifocal depigmented retinal lesions observable by fundus examination starting at 18 days of age. The retinal lesions are associated with fluorescein leakage around 25 days of age, with peak leakage at about 4 weeks of age. ERG responses deteriorate as rnv3 mutants age, concomitant with progressive photoreceptor disruption and loss that is observable by histology. Genetic analysis localized rnv3 to mouse chromosome (Chr) 1. By high throughput sequencing of a whole exome capture library of a rnv3/rnv3 mutant and subsequent sequence analysis, a single base deletion (del) in the Crb1 [crumbs family member 1] gene, which was previously reported to cause retinal degeneration 8, was identified. The TALEN-mediated ODR rescued the posterior segment vascularization phenotype; heterozygous Crb1rd8+em1Boc/Crb1rd8 and homozygous Crb1rd8+em1Boc/Crb1rd8+em1Boc mice showed a normal retinal phenotype. Additionally, six novel disruptions of Crb1 that were generated through aberrant non-homologous end joining induced by TALEN exhibited variable levels of vascularization, suggesting allelic effects.
Conclusions: The rnv3 model and the models of six novel disruptions of Crb1 are all reliable, novel mouse models for the study of both early and late events associated with posterior segment vascularization and can also be used to test the effects of pharmacological targets for treating human ocular vascular disorders. Further study of these models may provide a greater understanding about how different Crb1 alleles result in aberrant angiogenesis.

PMID: 30372741 [PubMed - in process]