CHEK2 rinta- sekä paksu- ja peräsuolisyövässä

Breast and colorectal cancers, are common types of cancer, with over two million newly diagnosed cases annually worldwide. Cancer is a genetic disease and defects in DNA integrity restoring functions make a significant contribution to cancer risk. CHEK2 is a checkpoint kinase functioning as a regulator of cell cycle checkpoints, apoptosis, and DNA repair in response to DNA double-strand breaks. The aim of this study was to evaluate the role of CHEK2 in breast cancer predisposition in Finnish breast cancer families and in breast cancer risk at the population level. We were interested in the clinical and biological characteristics of the breast tumors associated with the CHEK2 germline mutations or aberrant CHEK2 protein expression and the effect on survival of patients with these CHEK2 defects. We also assessed the role of CHEK2 mutations, namely 1100delC and I157T, in colorectal cancer susceptibility in Finland. CHEK2 I157T was found to be a low-penetrance breast cancer susceptibility allele, conferring a 1.4-fold risk for carriers. Reduced or absent CHEK2 protein expression was observed in one-fifth of breast tumors from patients unselected for family history, implying that defective CHEK2 signaling contributes to tumorigenesis. Reduction in CHEK2 expression was more common in tumors with larger diameter and ER expression, but with regard to other tumor characteristics and prognosis of a patient no association was observed. Results from comparison of CHEK2 1100delC carrier tumors with noncarrier tumors were in line with the findings from the CHEK2 expression study. Tumors from CHEK2 1100delC carriers were more often of higher grade than tumors from noncarriers, and they also tended to be ER-positive more often, although generally 1100delC status does not seem to radically affect the tumor characteristics. Our results suggest that CHEK2 1100delC may not be a susceptibility allele for CRC, although a very small effect cannot be excluded. Furthermore, CHEK2 1100delC is equally frequent in HBCC (hereditary breast and colorectal cancer) phenotype families and in breast cancer families. Over 1000 CRC cases were screened for CHEK2 I157T, and a significantly higher frequency of I157T was observed among both familial and sporadic CRC cases. The relation of CHEK2 I157T with familial CRC has not been studied previously. CHEK2 I157T seems to be a susceptibility allele for both familial and sporadic CRC, conferring a 1.5-fold risk for carriers of this variant. CHEK2 I157T has been proposed to have a role as a multiple cancer susceptibility allele, which is supported by our results since we observed a trend towards higher frequency of the variant among cases with multiple primary tumors or those with a family history of cancer. During the last five years CHEK2 has established its role as an important cancer susceptibility gene. It has become apparent that CHEK2 is a low-penetrance susceptibility gene for several cancer types, significantly contributing to familial cancer risk as well as to cancer risk at the population level.Rinta- sekä paksu- ja peräsuolisyövät ovat yleisiä syöpätyyppejä, joihin sairastuu vuosittain maailmassa yli 2 miljoonaa ihmistä. Syöpä on geneettinen sairaus ja häiriöt DNA:n kunnossapitomekanismeissa lisäävät syöpäriskiä. CHEK2 on solusyklin säätelykinaasi, joka aktivoiduttuaan DNA:n kaksoisjuosteen katkeamisen seurauksen säätelee solusyklin tarkastuspisteitä, ohjelmoitua solukuolemaa ja DNA:n korjausmekanismeja. Tutkimuksen tavoitteena oli arvioida CHEK2:n vaikutusta rintasyöpäalttiuteen suomalaisissa rintasyöpäperheissä sekä väestötasolla. Tutkimme CHEK2:n mutaatioihin ja epänormaaliin CHEK2-proteiinin ilmentymiseen liittyvistä rintasyöpäkasvaimien kliinisistä ja biologisista ominaisuuksista. Vertasimme myös rintasyöpäpotilaiden selviytymisestä suhteessa CHEK2:n vaurioihin. Lisäksi arviomme kahden CHEK2-mutaation, 1100delC ja I157T, osuutta paksu- ja peräsuolisyöpäriskiin Suomessa. CHEK2 I157T mutaation havaittiin olevan alhaisen ilmenemisyleisyyden rintasyöpäalttiusalleeli, joka kohottaa rintasyöpäriskin 1,4-kertaiseksi variantin kantajilla. Alentunut tai puuttuva CHEK2-proteiinin ilmentyminen havaittiin viidesosassa rintasyöpäkasvaimia, mikä viittaa häiriintyneen CHEK2-signaloinnin vaikuttavan kasvainten kehittymiseen. Alentunut CHEK2:n ilmentyminen oli yleisempää suuremmissa ja estrogeenireseptoripositiivisissa kasvaimissa, mutta muihin kasvainten ominaisuuksiin tai potilaan ennusteeseen ei havaittu yhteyttä. Tulokset CHEK2 1100delC kantajien kasvainten ja ei-kantajien kasvainten vertailusta tukivat CHEK2-proteiinin ilmentymistutkimuksen tuloksia. CHEK2 1100delC kantajien kasvaimet olivat useammin erilaistumattomampia ja estrogeenireseptoripositiivisia, vaikka yleisesti 1100delC ei näytä olennaisesti vaikuttavan kasvainten ominaisuuksiin. Suuren tutkimusaineiston perusteella CHEK2 1100delC ei näytä merkittävästi vaikuttavan paksu-ja peräsuolisyöpäriskiin. CHEK2 1100delC on myös yhtä yleinen HBCC (perinnöllinen rinta- sekä paksu- ja peräsuolisyöpä) -perheissä kuin rintasyöpäperheissä. Yli tuhat paksu- ja peräsuolisyöpätapausta tutkittuamme havaitsimme CHEK2 I157T-mutaation olevan merkittävästi yleisempi potilailla kuin väestössä riippumatta siitä, onko potilaan suvussa kyseisiä syöpiä. Tätä mutaatiota ei ole aikaisemmin tutkittu suvuttain esiintyvässä paksu- ja peräsuolisyövässä. CHEK2 I157T vaikuttaa olevan sekä satunnaisesti että suvuttain esiintyvän paksu- ja peräsuolisyövän alttiusalleeli kohottaen kantajien syöpäriskin 1,5-kertaiseksi. Tuloksemme tukevat aikaisempaa ehdotusta, jonka mukaan CHEK2 I157T olisi monisyöpäalttiusalleeli. Havaitsimme nimittäin enemmän CHEK2 I157T -kantajia potilaissa, joilla oli useampia primäärikasvaimia sekä potilaissa, joiden sukutaustassa esiintyi syöpää. Kuluneen viiden vuoden aikana CHEK2 on osoittautunut merkittäväksi syöpäalttiusgeeniksi. CHEK2 on alhaisen ilmenemisyleisyyden alttiusgeeni useissa syöpätyypeissä vaikuttaen syöpäriskiin sekä perhe- että väestötasolla

Mutational Signatures Associate With Survival in Gastrointestinal Carcinomas

Background/Aim: Mutational signatures reflect common patterns based on the counts of mutations and their sequence context. The prognostic value of these signatures, mirroring various carcinogenetic processes of cancers, are unexplored in gastrointestinal cancers. Our aim was to evaluate possible prognostic relevance of mutational signatures in gastrointestinal carcinomas after adjusting with the traditional prognostic factors. Materials and Methods: We used publicly available data from The Cancer Genome Atlas and Pan-Cancer Analysis of Whole Genomes to evaluate the associations between survival endpoints and activity of mutational signatures in seven types of gastrointestinal cancers. Results: Most strikingly, the high activity of age -related single-base substitution 5 (SBS5) and SBS40 signatures were in rectal adenocarcinomas associated with both improved overall survival (OS) [for SBS5 hazard ratio (HR) 0.130; 95% CI=0.03-0.56, for SBS40 HR=0.072; 95% CI=0.012-0.44, respectively] and similarly also to rectal cancer-specific survival. In patients with left-sided (but not right-sided) colon adenocarcinoma, the high activity of SBS2 signatures, formed due to APOBEC activity, predicted shortened OS. In pancreatic cancer, the high activity of SBS10b, caused by polymerase epsilon exonuclease proofreading defects, was associated both with longer OS (HR=0.44; 95% CI=0.205-0.96) and pancreatic cancer-specific survival (HR=0.32; 95% CI=0.112-0.91). Conclusion: Several mutational signatures seem to have clinically meaningful, cancer-specific associations with prognosis among gastrointestinal cancers.Peer reviewe

Whole-exome sequencing identifies novel candidate predisposition genes for familial polycythemia vera

Background: Polycythemia vera (PV), characterized by massive production of erythrocytes, is one of the myeloproliferative neoplasms. Most patients carry a somatic gain-of-function mutation in JAK2, c.1849G > T (p.Val617Phe), leading to constitutive activation of JAK-STAT signaling pathway. Familial clustering is also observed occasionally, but high-penetrance predisposition genes to PV have remained unidentified. Results: We studied the predisposition to PV by exome sequencing (three cases) in a Finnish PV family with four patients. The 12 shared variants (maximum allowed minor allele frequency G (p.Phe418Leu) in ZXDC, c.1931C > G (p.Pro644Arg) in ATN1, and c.701G > A (p.Arg234Gln) in LRRC3. We also observed a rare, predicted benign germline variant c.2912C > G (p.Ala971Gly) in BCORL1 in all four patients. Somatic mutations in BCORL1 have been reported in myeloid malignancies. We further screened the variants in eight PV patients in six other Finnish families, but no other carriers were found. Conclusions: Exome sequencing provides a powerful tool for the identification of novel variants, and understanding the familial predisposition of diseases. This is the first report on Finnish familial PV cases, and we identified three novel candidate variants that may predispose to the disease.Peer reviewe

Identification of homozygous deletion in ACAN and other candidate variants in familial classical Hodgkin lymphoma by exome sequencing

Tutkimuksessamme tarkastelimme Lähi-idästä lähtöisin olevaa perhettä, jossa kolmella viidestä lapsesta on todettu nuorellä iällä klassinen Hodgkinin lymfooma (cHL). Perinnöllinen alttius cHL:lle tunnetaan huonosti, eikä taudille mahdollisesti altistavia geenimuutoksia ole aiemmin raportoitui kuin yksi kappale. Geenimuutosten selvittämiseksi eksomisekvensoimme kolmen sairastuneen lapsen verinäytteestä eristetyn DNA:n ja poimimme joukosta kaikkien kolmen jakamat muutokset. Suodatimme lasten jakamien DNA-muutosten joukosta pois omissa vertailujoukoissamme ja useissa julkisissa tietokannoissa esiintyvät geneettiset muutokset ja arvioimme jäljellejääneiden muutosten haitallisuutta kahdella laskennallisella priorisaatioalgoritmilla. Näin saimme järjestettyä jäljelle jääneet 35 jaettua muutosta laskennalliseen haitallisuusjärjestykseen. Jaetuista muutoksista merkittävimmäksi nousi ACAN-geenissä oleva homotsygoottinen 57 emäksen pituinen deleetio c.2836_2892del, jota ei ole aiemmin liitytty cHL-fenotyyppiin

A germline exome analysis reveals harmful POT1 variants in multiple myeloma patients and families

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Detection of subclonal L1 transductions in colorectal cancer by long-distance inverse-PCR and Nanopore sequencing

Long interspersed nuclear elements-1 (L1s) are a large family of retrotransposons. Retrotransposons are repetitive sequences that are capable of autonomous mobility via a copy-and-paste mechanism. In most copy events, only the L1 sequence is inserted, however, they can also mobilize the flanking non-repetitive region by a process known as 3' transduction. L1 insertions can contribute to genome plasticity and cause potentially tumorigenic genomic instability. However, detecting the activity of a particular source L1 and identifying new insertions stemming from it is a challenging task with current methodological approaches. We developed a long-distance inverse PCR (LDI-PCR) based approach to monitor the mobility of active L1 elements based on their 3' transduction activity. LDI-PCR requires no prior knowledge of the insertion target region. By applying LDI-PCR in conjunction with Nanopore sequencing (Oxford Nanopore Technologies) on one L1 reported to be particularly active in human cancer genomes, we detected 14 out of 15 3' transductions previously identified by whole genome sequencing in two different colorectal tumour samples. In addition we discovered 25 novel highly subclonal insertions. Furthermore, the long sequencing reads produced by LDI-PCR/Nanopore sequencing enabled the identification of both the 5' and 3' junctions and revealed detailed insertion sequence information.Peer reviewe

Enrichment of cancer-predisposing germline variants in adult and pediatric patients with acute lymphoblastic leukemia

Despite recent progress in acute lymphoblastic leukemia (ALL) therapies, a significant subset of adult and pediatric ALL patients has a dismal prognosis. Better understanding of leukemogenesis and recognition of germline genetic changes may provide new tools for treating patients. Given that hematopoietic stem cell transplantation, often from a family member, is a major form of treatment in ALL, acknowledging the possibility of hereditary predisposition is of special importance. Reports of comprehensive germline analyses performed in adult ALL patients are scarce. Aiming at fulfilling this gap of knowledge, we investigated variants in 93 genes predisposing to hematologic malignancies and 70 other cancer-predisposing genes from exome data obtained from 61 adult and 87 pediatric ALL patients. Our results show that pathogenic (P) or likely pathogenic (LP) germline variants in genes associated with predisposition to ALL or other cancers are prevalent in ALL patients: 8% of adults and 11% of children. Comparison of P/LP germline variants in patients to population-matched controls (gnomAD Finns) revealed a 2.6-fold enrichment in ALL cases (CI 95% 1.5-4.2, p = 0.00071). Acknowledging inherited factors is crucial, especially when considering hematopoietic stem cell transplantation and planning post-therapy follow-up. Harmful germline variants may also predispose patients to excessive toxicity potentially compromising the outcome. We propose integrating germline genetics into precise ALL patient care and providing families genetic counseling.Peer reviewe

Clinically relevant germline variants in allogeneic hematopoietic stem cell transplant recipients

Allogeneic hematopoietic stem cell transplantation (HSCT) provides patients with severe hematologic disease a well-established potential for curation. Incorporation of germline analyses in the workup of HSCT patients is not a common practice. Recognizing rare harmful germline variants may however affect patients' pre-transplantation care, choice of the stem cell donor, and complication risks. We analyzed a population-based series of germline exome data of 432 patients who had undergone HSCT. Our aim was to identify clinically relevant variants that may challenge the outcome of the HSCT. We focused on genes predisposing to hematological diseases, or solid tumors, and genes included in the American College of Medical Genetics secondary findings list v3.0. As population-specific controls, we used GnomAD non-cancer Finns (n = 10,816). We identified in our population-based analysis rare harmful germline variants in disease-predisposing or actionable toxicity-increasing genes in 17.8% of adult and pediatric patients that have undergone HSCT (15.1% and 22.9%, respectively). More than half of the patients with a family member as a donor had not received genetic diagnosis prior to the HSCT. Our results encourage clinicians to incorporate germline genetic testing in the HSCT protocol in the future in order to reach optimal long-term outcome for the patients.Peer reviewe

Comprehensive analysis of NuMA variation in breast cancer

<p>Abstract</p> <p>Background</p> <p>A recent genome wide case-control association study identified <it>NuMA </it>region on 11q13 as a candidate locus for breast cancer susceptibility. Specifically, the variant Ala794Gly was suggested to be associated with increased risk of breast cancer.</p> <p>Methods</p> <p>In order to evaluate the <it>NuMa </it>gene for breast cancer susceptibility, we have here screened the entire coding region and exon-intron boundaries of <it>NuMa </it>in 92 familial breast cancer patients and constructed haplotypes of the identified variants. Five missense variants were further screened in 341 breast cancer cases with a positive family history and 368 controls. We examined the frequency of Ala794Gly in an extensive series of familial (n = 910) and unselected (n = 884) breast cancer cases and controls (n = 906), with a high power to detect the suggested breast cancer risk. We also tested if the variant is associated with histopathologic features of breast tumors.</p> <p>Results</p> <p>Screening of <it>NuMA </it>resulted in identification of 11 exonic variants and 12 variants in introns or untranslated regions. Five missense variants that were further screened in breast cancer cases with a positive family history and controls, were each carried on a unique haplotype. None of the variants, or the haplotypes represented by them, was associated with breast cancer risk although due to low power in this analysis, very low risk alleles may go unrecognized. The <it>NuMA </it>Ala794Gly showed no difference in frequency in the unselected breast cancer case series or familial case series compared to control cases. Furthermore, Ala794Gly did not show any significant association with histopathologic characteristics of the tumors, though Ala794Gly was slightly more frequent among unselected cases with lymph node involvement.</p> <p>Conclusion</p> <p>Our results do not support the role of <it>NuMA </it>variants as breast cancer susceptibility alleles.</p

Germline ERCC excision repair 6 like 2 (ERCC6L2) mutations lead to impaired erythropoiesis and reshaping of the bone marrow microenvironment

Despite the inclusion of inherited myeloid malignancies as a separate entity in the World Health Organization Classification, many established predisposing loci continue to lack functional characterization. While germline mutations in the DNA repair factor ERCC excision repair 6 like 2 (ERCC6L2) give rise to bone marrow failure and acute myeloid leukaemia, their consequences on normal haematopoiesis remain unclear. To functionally characterise the dual impact of germline ERCC6L2 loss on human primary haematopoietic stem/progenitor cells (HSPCs) and mesenchymal stromal cells (MSCs), we challenged ERCC6L2-silenced and patient-derived cells ex vivo. Here, we show for the first time that ERCC6L2-deficiency in HSPCs significantly impedes their clonogenic potential and leads to delayed erythroid differentiation. This observation was confirmed by CIBERSORTx RNA-sequencing deconvolution performed on ERCC6L2-silenced erythroid-committed cells, which demonstrated higher proportions of polychromatic erythroblasts and reduced orthochromatic erythroblasts versus controls. In parallel, we demonstrate that the consequences of ERCC6L2-deficiency are not limited to HSPCs, as we observe a striking phenotype in patient-derived and ERCC6L2-silenced MSCs, which exhibit enhanced osteogenesis and suppressed adipogenesis. Altogether, our study introduces a valuable surrogate model to study the impact of inherited myeloid mutations and highlights the importance of accounting for the influence of germline mutations in HSPCs and their microenvironment.Peer reviewe