Posted on Authorea 17 Sep 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.160037743.35386400 — This a preprint and has not been peer reviewed. Data may be preliminary. Valle Garc´ıa-Barber´an Martin Lorena cancer in predisposition involved potentially PTPRT Germline 05 mt ta. 03 al,21) otFCXcssrmi nxlie.I at CT comprises FCCTX fact, In unexplained. al., Segu´ı et remain 2014; cases al., FCCTX et Schulz most 2014; 2017), al., 2018; Valle, Woods, et 2013; & Nieminen al., Green, 2017; the et Evans, allowed al., Mart´ın-Morales 2016; Smith has et 2015; al., (NGS) 2015; 2011, et Sequencing al., Generation (Esteban-Jurado Next remains et cancers of Garre predisposition related arrival CRC other X the MMR-proficient new and Type Although stable, of CRC criteria Cancer 2009). to identification microsatellite Amsterdam Colorectal predisposition (Lindor, Familial with the their elucidated term families underlying fulfill be the basis HNPCC to that genetic reason, of families the this whom group For the for such of genes. tumors, designate half MMR to the almost emerged in that defects (FCCTX) estimated any (MSI) germline is instability present by microsatellite caused it not in is However, & do results and Mecklin, which Syndrome tumors. Watson, genes, Lynch Vasen, the as (MMR) 1991; known repair in Lynch, is mismatch families the & by these in Khan, defined of Mecklin, and inactivating fraction (Vasen, tumors important criteria An associated clinical other 1999). II and Lynch, (CRC) and cancer I colorectal Amsterdam charac- to condition the susceptibility inherited increased dominant autosomal an an by is terized (HNPCC) Cancer Colorectal Non-Polyposis Hereditary hereditary and susceptibility cancer Introduction with linked epigenetic progression is study. and mutation exhibit present PTPRT initiation carriers the germline cancer a of role the that relevance role PTPRT’s of time the causal Although first highlights a tumors family. the which with the is the consistent cancer, of this genes, phosphatase addition, predisposition studied, target second well cancer In downstream the been PTPRT the of has of in activity. fraction expression mutation PTPRT’s significant altered germline a an for this of and of essential allele loss is wild-type the shows the variant in which of germline results inactivation , This and development. family the many tumor in the of level promote in somatic that domain cancer at drivers mutated with as frequently cosegregation act be mutations compatible to these found a p.(Asp1364GlyfsTer24)]. predisposition that [c.4090dup, one increased proven suppressor PTPRT and tumor an been in cancer-affected a having with mutation two is cancers, germline families in and truncating phosphatase sequencing of receptor a Whole-exome group a revealed basis. encodes a family PTPRT genetic FCCTX describe unknown a to an of but used members cancers, term healthy related a other is and (FCCTX) colorectal X to Type Cancer Colorectal Familial Abstract 2020 17, September 6 5 4 3 2 1 optlciioSnCro.IdISSC Carlos. San clinico Hospital ICO-IDIBELL IDIBELL CIBERONC IdISCC, Carlos, Cl´ınico IdISSC San Hospital Carlos. San Clinico IdISCC Hospital Carlos. San Clinico Hospital 4 iuld aHoya la de Miguel , 1 itrLorca Victor , 3 ai us Gonzalez-Morales Luisa Maria , 2 ia Garre Pilar , 2 er P´erez-Segura Pedro , 6 n rnddCaldes Trinidad and , 1 2 aiBelhad Sami , 2 arcaLlovet Patricia , 2 4 are Capell´a Gabriel , 1 Vanesa , 5 Laura , Posted on Authorea 17 Sep 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.160037743.35386400 — This a preprint and has not been peer reviewed. 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All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.160037743.35386400 — This a preprint and has not been peer reviewed. Data may be preliminary. a efre sn obnto ftodffrn loihs(aSa n AK n h identified the and GATK) and average (VarScan 50x algorithms a different and two 101bp of combination of version calling Variant a library genome reads SAMtools. reference and using the paired-end human Picard-tools by performed the and with processing against was Technologies), by platform followed aligned software, Agilent BWA subsequently 2000 the and (51Mb, using capture HiSeq GRCh37/hg19 trimmed V3 were exome Illumina Reads Exon depth. 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Other Counseling 1999) the MMR Genetic fulfilled 1991, of the families al., expression at the et recruited of families (Vasen All FCCTX HNPCC Spain). 13 (Madrid, in Carlos studied Cl´ınico was San exome whole The a describes study population present Study The families. methods performed and HNPCC was Materials MMR-proficient sequencing the Amsterdam-positive whole-exome in FCCTX, 13 mutation underlying germline of cause truncating group genetic 2010). the 2017, a al., identifying in et of Zhao aim (Y. the pathway With PI3K-Akt the inhibiting hence Akt), and 2. 1. PTPRT μ eeta a dnie noeo hs families. these of one in identified was that gene -hc FEtsu etos n eaoyi n eosin- and hematoxylin a and sections, tissue FFPE m-thick 3 Posted on Authorea 17 Sep 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.160037743.35386400 — This a preprint and has not been peer reviewed. Data may be preliminary. hsasy hl omrilhg-ehltdadnnmtyae N a sda oto (CpGenome control a as used was DNA non-methylated following and and high-methylated from buffers commercial DNA and tissue while reagents assay, healthy Q96 or this PyroMark Tumor the instructions. of advantage manufacturer’s taking the Qiagen), (Biotage, of sequencer of measurement MD fraction the a 1 for targeting of PCR used conversion bisulfite control. also the a was purpose, as assay used pyrosequencing was A PCR tissue the breast and instructions, or electrophoresis. manufacturers’ gel colon Pyrosequencing the agarose healthy following by 2016) out from analyzed al., carried then obtained et were were (Peyser DNA two MSP products al. from and while et conversion DNA assay, Peyser Tumor bisulfite by this The S2). or previously for Table CRC) been (Supplementary (in used had carcinoma) 2013) was and neck al., DNA and et unmethylated (Laczmanska head or methylated (in al. of (Qiagen) detected et region Kit Laczmanska specifically promoter MSP by which EpiTect the primers, described the targeting of using primers pairs (MSP) specific 2 PCR of of specific sets 1 methylation different of two a conversion by and bisulfite in followed initial described (Qiagen), an are Kit of Bisulfite used consisted assay primers methylation the promoter All The alleles. the assay of methylation any Promoter germline of order and in height tumor compared peak analyses, were the S2. 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All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.160037743.35386400 — This a preprint and has not been peer reviewed. Data may be preliminary. fdffrn acrtpsi w ucsiegnrtos( Rs noera n ratcne) the cancer), was variant breast identified 1 The S3). and Table Supplementary endometrial and 1 1A CRCs, (Figure diagnosed old (3 members years 5 generations 28 with successive being II:6 criteria, onset two (II:5, II of in Amsterdam cc765 age the types earliest family fulfilled cancer family of different This relative relative. of cancer-free germline unaffected the a one in revealed and absent 1A) members Figure affected II:7, two and in sequencing Whole-exome and Portugal) (Caparica, VIDA 13). 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No reuse without permission. — https://doi.org/10.22541/au.160037743.35386400 — This a preprint and has not been peer reviewed. Data may be preliminary. te addt ainsi hsfml included family this in variants all candidate affects and family Other Genomes) the 1000 in ExAc, variants (gnomAD, candidate databases Other public the of any in reported of not is variant This iia C a hnpromdi re ocekteall-pcfi xrsino idtp n mutant and wild-type of expression allele-specific the check to order in performed then was member PCR of digital tissue A breast healthy the subtle, to more again compared Tumor once methylation Although increased 3C). 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All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.160037743.35386400 — This a preprint and has not been peer reviewed. Data may be preliminary. CS6096)i soitdwt R codn oCSI Tt ta. 09.Ee huhterole the though Even 2019). al., et (Tate COSMIC to according CRC with associated pathogenic is a (COSV62009665) with compatible are that results discussed, both previously mentioned, As things the all development this For tumor of inhibiting 2017). role role al., important et an 2017). plays (Chu al., SOCS3 metastasis et that showed (Chu reduced also pathway that al. signaling STAT3 et and of Chu by activation study sustained same and The imbalance to of leads regulation IL-6/STAT3, negative of This feedback. of negative expression its the inactivating inhibiting while pathway of signaling STAT3’s hypermethylation that the 2013; proven Yoshimura, as been such & has Ito, it Kondo, and (Inagaki-Ohara, and 2017), pathway al., signaling et JAK-STAT compared Jiang the when tumors of breast regulators and feedback colon the contrast, both in In observed of was controls. 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Zhang matter the Wang genes D2’s by a target on (Z. out downstream As affecting effects half pointed 2017). mutation significant by also al., activity had missense was et enzyme’s domain a domain Zhao the catalytic just (Y. decrease second activity that to this PTPRT’s enough reported known for was one al. essential mutant, the be et our is to Wang (D1) protein D2 proven domain fact, the essential been phosphatase of of of has first activity phospho-tyrosine amount the and phosphatase substrate’s Although considerable activity the affects 2014). the this a for al., that surrounding and responsible et directly domain mutation be (Lui mutation residues to D2 frameshift this of the with a of (69.2%) lost is are 36.2% majority including D1364Gfs*24 the Actually, protein, protein, D2. residues. the the of as on acids known variant amino domain, the catalytic second of suppressor PTPRT’s tumor effects this of the inactivation the affects Regarding in mainly involved silencing hit epigenetic second decrease the a no that considered while supports be gene. individual, This could same allele. and the mutant allele the from fact, wild-type of colon In the healthy expression carriers. which the to the tumors), in compared from observed colon when tumors was of tumor and the expression colorectal (breast in the the tested allele of in that wild-type were showed reduced region the that assay of promoter expression carriers inactivation the allele-specific the of an mechanism Indeed, of different tumors 2016). a two al., be the could et in Peyser hypermethylated 2013; be to al., et (Laczmanska tumors PTPRT STAT3 PTPRT rmtrhsbe eetyrpre ob rqetyhprehltdi prdcCCadother and CRC sporadic in hypermethylated frequently be to reported recently been has promoter PTPRT SOCS3 r peuae Cue l,21) hsi huh oocrtruhdffrn mechanisms, different through occur to thought is This 2017). al., et (Chu upregulated are slne ohrdtr acr neetnl,asmtcmtto ffcigtesm codon same the affecting mutation somatic a Interestingly, cancer. hereditary to linked is SOCS3 PTPRT variant. xrsinwsdcesdi ohtmr.Nntees OSpoen r negative are proteins SOCS Nonetheless, tumors. both in decreased was expression PTPRT sepeso ffcstmrgnssadCCporsin rmtn rwhand growth promoting progression, CRC and tumorigenesis affects expression ’s ain a eivle ntecneso hsFCXfml,tetmr from tumors the family, FCCTX this of cancers the in involved be may variant SOCS3 sascainwt acrhsbe elsuid u hsi h rttime first the is this but studied, well been has cancer with association ’s eepooes loigiflmaoyctknsI- oactivate to IL-6 cytokines inflammatory allowing promoters, gene SOCS3 PTPRT 8 suulydwrgltdi R,ee when even CRC, in downregulated usually is 16Gs2 eut ntels ftels 97 last the of loss the in results D1364Gfs*24 e se per SOCS3 SOCS3 PTPRT 2i epnil o h euainof regulation the for responsible is D2 , Cue l,21) ihteproeof purpose the with 2017), al., et (Chu xrsin eitdb h activation the by mediated expression, ´ idtp leewssignificantly was allele wild-type s BCL-XL hc sa oncogenic an is which , BCL-XL PTPRT and a found was SOCS3 IL-6 Posted on Authorea 17 Sep 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.160037743.35386400 — This a preprint and has not been peer reviewed. Data may be preliminary. rmr3( 3 Primer ( ( ( per ( Condel author corresponding the from available resources are Web study current the request. during reasonable analyzed upon and generated datasets The statement availability interests. Data of conflicts no declare authors The (Madrid, Carlos statement San records. We interest Clinico study. pathology of Hospital and the of Conflict sections in and Biobank blocks, Millan part the Diaz FFPE taking and Isabel the for Diaque, assistance, for Paula families technical Spain) collaboration, the their Asso- his thank Case for for European Wang to Ravillah in the Zhenghe like Durgadevi Dr. lab, of acknowledge would ZhengheWang’s fellowship to authors Dr. travel like also The at a would Research. by LMM Cancer of supported (Spain) for was stay Institute ciation USA), short Health (Cleveland, A III University Funds. Carlos Reserve Development the Western from Regional PI-19/1366 European and the PI-16/01292 grants and by supported was work This this cancer stablished, in well role work. Acknowledgements its been this clarify of has relevance us progression a the help and that highlights will additional which initiation – time in families cancer first gene high-risk the in this other we is role in of PTPRT’s reason, even screening Although or The that – predisposition. location. For cohorts cancer variant family. cancer preferred germline colorectal carrier and familial the risk the of conferred role of penetrance, causal causality, susceptibility probable cancer a the this to propose of point in effect presented together, p.(Asp1364GlyfsTer24) the here or c.4090dup modifying independently results or contributing, candidate the family these be together, the that may Taken of possibility – susceptibility the factors cancer environmental out increased rule or cannot the genetic we to other causality, even its or support – report variants acts this pathway INVS in Wnt suppress Finally, presented canonical Although 2001). to results Nakamura, the 2010). reported inhibiting & al., pathway pathways, (Unoki et homozygous signaling signaling cycle (Bellavia and Wnt PTEN cell 2005), different the al., the the et of between of (Simons gastric inhibition switch mediator familial the molecular with a by a associated cells is as been cancer have ABTB1 of gene 2014). growth this al., the in this mutations et for germline (Gaston addition, prioritized In cancer were 2014). variants al., of candidate et regulation (Gaston three the another in risk, involved in variants is cancer this missense increased though two the even the that family: of However, out explanation pointed patient. be the CRC should of for early-onset it matter determined. least, an not a been in but not As Last domain has task. D1 variant this the this with of inside a help effect located as would functional variant involvement cohort its germline larger confirm missense to a rare of needed in screening are gene studies the this more fact, Studying undeniable, gene. is development susceptibility tumor cancer in protein this of www.omim.org www.ensembl.org www.cbioportal.org/mutation PTPRT primer3.ut.ee bbglab.irbbarcelona.org/fannsdb/ ;Ptcrs( Pathcards ); sanvlcne rdsoiingn.Hwvr oersac sncsayt ofimthe confirm to necessary is research more However, gene. predisposition cancer novel a as ;Gncrs( Genecards ); PTPRT PTPRT ;POEN( PROVEAN ); na needn aiileryostCCchr dnie n additional one identified cohort CRC familial/early-onset independent an in VEGF emievrati ikdwt acrssetblt n eeiaycancer, hereditary and susceptibility cancer with linked is variant germline pathcards..org PTPRT MAP3K6 www.genecards.org mapper INVS xrsinadhsas enrpre oata uo suppressor tumor a as act to reported been also has and expression 16Gs2 stebs addt ain o hsfml n the and family this for variant candidate best the is D1364Gfs*24 provean.jcvi.org/ and uain aebe soitdwt ueienephronophthisis juvenile with associated been have mutations ;MttoTse ( MutationTaster ); ;CSI ( COSMIC ); ABTB1 9 ;HF( HSF ); ;PlPe ( 2 PolyPhen ); n piergo ain in variant region splice a and , ;Ratm ( Reactome ); PTPRT www.umd.be/HSF/ www.mutationtaster.org cancer.sanger.ac.uk uainhdtehgetpotential highest the had mutation genetics.bwh.harvard.edu/pph2/ reactome.org PTPRT ;Mtto Map- Mutation ); ;SF ( SIFT ); INVS mutation. ;Ensembl ); ;OMIM ); MAP3K6 . sift.bii.a- PTPRT ); Posted on Authorea 17 Sep 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.160037743.35386400 — This a preprint and has not been peer reviewed. Data may be preliminary. edako yoieSgaigi acngnssadIsSgicnei acrTreatment. Cancer in Negative SOCS-Mediated Significance of Its Dysregulation (2017). and J. Carcinogenesis Yu, & in T., Signaling Liu, Immunology W., Cytokine Yu, P., of Liu, Feedback W., Zhang, M., Jiang, cancer. and inflammation, SOCS, (2013). A. Yoshimura, & , M., Ito, T., Kondo, K. K., Inagaki-Ohara, Bedard, counting. and . years . . Thirty cancer. phosphorylation: C., 21 Tyrosine gastric Macgillivray, (2009). familial H., T. with Hunter, Pinheiro, associated M., are https://doi.org/10.1371/journal.pgen.1004669 gene: Nightingale, MAP3K6 e1004669. BRCA2 C., in (10), (2015). mutations Oliveira, Cald´es, T. Germline X. S., (2014). type . Hansford, . (2011). . cancer D., T. I., colorectal Gaston, Bando, Caldes, familial A., in . Tosar, . testing . Mis- A., clinical https://doi.org/10.1111/cge.12427 J., from Romero, for Sanz, Patients J., candidate M., in Sanz, A Hoya, Mart´ın, L., MUTYH P., la and Garre, de OGG1, NUDT1, J., (MSS-HNPCC). Genes B. Families mutations Repair HNPCC Doyle, https://doi.org/10.1158/1078-0432.CCR-10-2491 pathogenic Proficient Damage M., for Repair Oxidative R. BMPR1a match the Xicola, of of V., Newfoundland. Screening Analysis Briceno, from (2018). P., O. families Garre, M. X Woods, type & cancer S., https://doi.org/10.1007/s10689-017-0016-8 colorectal J. familial Green, pre- in R., germline D. for Evans, spotlight the in pathway repair cancer. ps://doi.org/10.1038/ejhg.2016.44 damage Cas- colorectal therapy. DNA to . anemia . L´opez-Cer´on, cancer disposition . Fanconi S., M., The Carballal, in (2016). Oca˜na, T., tellv´ı-Bel, S. Mu˜noz, J., Stat3 Franch-Exp´osito, S., C., Validating Esteban-Jurado, its and (2005). SOCS3 of E. significance https://doi.org/10.1038/nm0605-595 Prognostic (2017). J. W. Zhang, Darnell, & C., cancer. E. Liu, colorectal J. H., in Wang, Darnell, function L., & biological He, C., D., Albanese, Shen, G., Q., R. Chu, Pestell, Y., oncogene. Zhao, an of G., as Devgan, substrates Stat3 H., Intracellular (1999). M. Wrzeszczynska, (2006). F., A. J. Rotter, Bromberg, & (RPTPrho/PTPRT). A., rho Frostholm, phosphatase https://doi.org/10.1016/j.brainres.2006.07.122 R., tyrosine 50–57. Huijsduijnen, protein van receptor Hooft brain-enriched A., J. Besco, Association Transplant pathway. and Wnt/beta- Association Dialysis mutation European Renal the homozygous European the of A - of reactivity (2010). Publication abnormal Y. Official Transplantation: Pirson, with Dialysis, & nephronophthisis phrology, O., juvenile (2019). Devuyst, causing J.-P., L. Cosyns, INVS Valle, S., in Terryn, . K., . . Dahan, in S., M., phenotype, Bellavia, multi-tumor Navarro, a H., or M. adenomas. polyposis serrated Alonso, colorectal cancer, of M., colorectal absence P. cause Munoz-Torres, seldom P., mutations biallelic Mur, NTHL1 I., Quintana, S., Belhadj, References del.expasy.org star.edu.sg 2 2,1016 https://doi.org/10.1016/j.ceb.2009.01.028 140–146. (2), 3,e45.https://doi.org/10.4161/jkst.24053 e24053. (3), ;SAT( SMART ); , ;Uirt( UniProt ); 8 https://doi.org/10.3389/fimmu.2017.00070 . smart.embl-heidelberg.de www..org Cell , 25 cetfi Reports Scientific uoenJunlo ua eeis EJHG Genetics: Human of Journal European , 1) 0740.https://doi.org/10.1093/ndt/gfq519 4097–4102. (12), 98 3,295–303. (3), Gene ). , 627 ;SRN ( STRING ); , 9 1–2.https://doi.org/10.1016/j.gene.2017.06.013 114–122. , 10 1,92.https://doi.org/10.1038/s41598-019-45281-1 9020. (1), lnclCne Research Cancer Clinical string-db.org aueMedicine Nature aiilCancer Familial lnclGenetics Clinical urn pno nCl Biology Cell in Opinion Current ;SISMDL( SWISS-MODEL ); ri Research Brain , 24 1) 5110.htt- 1501–1505. (10), , LSGenetics PLoS , , 17 11 , 17 87 7,1701–1712. (7), 6,595–596. (6), 2,205–208. (2), 6,582–587. (6), rnir in Frontiers JAK-STAT , 1116 swissmo- , Ne- (1), 10 , Posted on Authorea 17 Sep 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.160037743.35386400 — This a preprint and has not been peer reviewed. Data may be preliminary. ae .G,Bmod . ub .C,Snk,Z,Bae .M,Bna,N,...Fre,S .(2019). A. S. Forbes, . . . N., Bindal, M., D. Beare, Z., Sondka, Cancer. C., In Mutations H. Somatic Jubb, Of Catalogue S., The Resequen- Bamford, COSMIC: Exome (2013). G., P. J. J. Cheadle, Tate, Cancer. . Colorectal . . to N., Predispose Li, that H., West, 34 Genes J., Tumor-Suppressor Colley, Potential R., Identifies Harris, cing M., Implications Naven, Phosphorylation: (2017). G., K. C. K. Smith, Singh, & K., B. Roy, R., Cancer. Prasad, R., in Kumar, M., signaling Ram, Wnt V., between Singh, switch molecular a as functions Kr II, type M., nephronophthisis pathways. Bashkurov, in A., mutated Bullerkotte, product gene A., the Ganner, J., Gloy, Repair. M., DNA Simons, Germline Impairing (2015). by L. Valle, Germline Cancer https://doi.org/10.1053/j.gastro.2015.05.056 Colorectal (2014). . . 563–566. . Hereditary H. (3), M., Cause Navarro, Sill, FAN1 T., in Pons, . . R., Mutations . Sanz-Pamplona, L´azaro, C., W., B., X. L. Renner, type Mina, Segu´ı, C. P., cancer N., Ulz, colorectal B. familial R., Koehler, to A. https://doi.org/10.1038/ncomms6191 predispose Janecke, 5191. . gene . SEMA4A . S., the Holzapfel, A., in P., variants Heller, cancer. Klampfl, K., colorectal E., Schulz, Duglova, in P., driver Radhakrishnan, oncogenic an M., https://doi.org/10.1038/cddis.2016.233 (2011). is Salou, E. Bcl-xL G., Campo, (2016). Gdynia, leukaemia. . . A.-L., . lymphocytic N., Scherr, chronic Villamor, Ord´o˜nez, in R., L., mutations https://doi.org/10.1038/nature10113 G. Conde, 101–105. recurrent Frequent (7354), V., (2016). identifies Quesada, R. sequencing M., J. Pinyol, Whole-genome in Grandis, S., inhibition & STAT3 X. E., to Puente, D. sensitivity Johnson, and H., cancer. activation Li, neck STAT3 and Y., increases head Zeng, PTPRT L., of Wang, hypermethylation M., promoter Freilino, Peltom D., . N. . . Hereditary Peyser, D., to A. Predisposition Deficiency. Paterson, Causes Repair Protein, W., Mismatch https://doi.org/10.1053/j.gastro.2014.06.009 Ribosomal S. DNA 595-598.e5. Without a Scherer, Carcinoma Encoding H., Colorectal RPS20, Lohi, Nonpolyposis of Y., Mutation Wu, Germline M.-F., (2014). O’Donohue, T., T. X. Nieminen, type dominant cancer SETD6 colorectal (2017). familial D. https://doi.org/10.1093/hmg/ddx336 Frequent Levy, in (2014). & Cald´es, R. mutation T., P., J. Garre, negative Grandis, Z., Vershinin, nonpoly- . M., . Feldman, . Mart´ın-Morales, hereditary L., Y., Lu, of Y., head Zeng, in https://doi.org/10.1073/pnas.1319551111 half hyperactivation J., STAT3 1114–1119. for Hritz, mechanism other K.-S., a cancer. P. provides neck phosphatases Ng, and The tyrosine D., protein N. receptor X: of Peyser, mutation Y., type W. V. cancer Lui, colorectal syndrome. Familial cancer ps://doi.org/10.1016/j.soc.2009.07.003 (2009). colon M. posis N. can- (2013). colorectal Lindor, M. sporadic M. in Sasiadek, hypermethylated & frequently E., are Szmida, genes D., cer. Ramsey, receptor-like T., phosphatase Sedziak, tyrosine M., Protein Bebenek, P., phosphatome. Karpinski, tyrosine I., cancer Laczmanska, human the Inside (2011). L. Cancer M. Tremblay, Reviews. & Nature S., Hardy, Dub´e, N., G., S. Julien, 7,12–04 https://doi.org/10.1002/humu.22333 1026–1034. (7), ora fHmnGenetics Human of Journal aueGenetics Nature h rti Journal Protein The rceig fteNtoa cdm fSine fteUie ttso America of States United the of Sciences of Academy National the of Proceedings Oncogene , 11 , 1,3–9 https://doi.org/10.1038/nrc2980 35–49. (1), 37 , 5,5753 https://doi.org/10.1038/ng1552 537–543. (5), ugclOclg lnc fNrhAmerica North of Clinics Oncology Surgical , , 36 58 35 1,16 https://doi.org/10.1007/s10930-017-9696-z 1–6. (1), 1,1–5 https://doi.org/10.1038/jhg.2012.119 11–15. (1), 9,16–19 https://doi.org/10.1038/onc.2015.171 1163–1169. (9), 11 ua oeua Genetics Molecular Human uli cd Research Acids Nucleic elDah&Disease & Death Cell ng . az .(05.Inversin, (2005). G. Walz, . . . C., ¨ onig, aueCommunications Nature Gastroenterology , 18 , Gastroenterology , 47 , 26 7 4,6765 htt- 637–645. (4), ua Mutation Human D) D941–D947. (D1), 8,e2342–e2342. (8), 2) 4481–4493. (22), Nature , , 147 111 k,P. ¨ aki, , , , (3), (3), 149 475 5 , , Posted on Authorea 17 Sep 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.160037743.35386400 — This a preprint and has not been peer reviewed. Data may be preliminary. iuelegends Figure https://doi.org/10.1073/pnas.0914884107 in involved genes two EGR2, and BPOZ of effects pathway. Growth-suppressive signaling (2001). PTEN and R Y. the syndrome) Nakamura, A., & (Lynch Chapelle, M., cancer la Unoki, colorectal de nonpolyposis S., hereditary Syngal, for P., instability. Guidelines microsatellite J. Terdiman, Bethesda R., Revised C. (2004). Boland, A., Umar, ho . hn,X,Gd,K,Lwec,E,Sn . aaae . ag .(00.Identi- (2010). Z. Wang, receptor phosphatase tyrosine . . . protein T., of Watanabe, target Q., a Sun, as paxillin E., Regulation of Lawrence, (2017). characterization K., T. Z. functional Guda, Wang, and X., fication . Zhang, . . tumorigenesis. Y., S., colon Zhao, Somasundaram, impacts PTPRT X., and Feng, https://doi.org/10.18632/oncotarget.10654 Src Y., human 48782–48793. by Hao, in (30), axis P., phosphatases signaling Zhang, tyrosine paxillin-p130-PI3K-AKT protein A., of of Scott, alterations Y., Genetic Zhao, of (2015). Identification Z. (2007). Wang, J. & Z. cancers. D., Wang, Sedwick, S., . America T. . Zhao, . of phosphatase W., States tyrosine Zheng, United protein Y., the receptor Chen, of phospho- of Sciences A., substrate between Possemato, a J., Cross-talk Yu, as (2011). STAT3 A., Z. Guo, Wang, X., & Zhang, X., Zhang, https://doi.org/10.1158/1541-7786.MCR-11-0147 D., 1418–1428. Sedwick, (10), X., PLC Zhu, X. and Y., STAT3 Type Zhao, Cancer P., Colorectal Familial Zhang, (2017). L. M. Bisgaard, https://doi.org/10.2174/1389202918666170307161643 & B., Mutational D. (2004). Zetner, E. V. Velculescu, . . . cancers. S., colorectal Szabo, https://doi.org/10.1126/science.1096096 in J., phosphatome Genome-wide Sager, tyrosine A., (2013). the Bardelli, of Q. W., analysis Wei, D. Parsons, . D., . Shen, . cancer. Z., Wang, T., lung in A. capacity Lee, repair S.-F., DNA Weng, of Y., determinants , genetic Qiao, novel J., reveals Ying, study nonpo- association Y., hereditary on O. for group Gorlova, criteria Collaborative L.-E., International clinical Wang, New the by (1999). T. proposed H. syndrome) Lynch, Lynch HNPCC. & (HNPCC, P., cancer Group J. colorectal Collaborative Mecklin, lyposis P., International Watson, The F., (1991). H. T. Vasen, H. Lynch, & (ICG-HNPCC). M., Cancer P. 424–425. Colorectal Khan, Polyposis. Non-Polyposis P., and Hereditary J. Cancer on Mecklin, Colorectal F., Familial H. of Vasen, Genetics the in Hepatology Discoveries and Gastroenterology Recent (2017). L. Valle, disease. to function, to genes, (2019). From L. phosphatases: Valle, tyrosine Biology Protein . Cell prevalence . Molecular . (2006). the of J., K. assessment N. Brunet, and Tonks, M., genes polyposes. Navarro, predisposing MSH3-associated G., proposed and recently Aiza, NTHL1- of of S., polyposis Belhadj, colonic to M., Contribution P. Munoz-Torres, M., Terradas, https://doi.org/10.1093/nar/gky1015 73 rceig fteNtoa cdm fSine fteUie ttso America of States United the of Sciences of Academy National the of Proceedings 1,2624 https://doi.org/10.1158/0008-5472.CAN-12-1915 256–264. (1), Oncogene Gastroenterology γ ly rtclrl nclrca tumorigenesis. colorectal in role critical a plays 1 , 34 3) 8539.https://doi.org/10.1038/onc.2014.326 3885–3894. (30), , 7 ora fteNtoa acrInstitute Cancer National the of Journal 1) 3–4.https://doi.org/10.1038/nrm2039 833–846. (11), , 116 Oncogene 6,1453–1456. (6), , 15 6,8989 https://doi.org/10.1016/j.cgh.2016.09.148 809–819. (6), , 20 , 104 3) 4746.https://doi.org/10.1038/sj.onc.1204608 4457–4465. (33), ua Mutation Human 1) 0046.https://doi.org/10.1073/pnas.0611665104 4060–4064. (10), 12 cec NwYr,N.Y.) York, (New Science https://doi.org/10.1002/humu.23853 . iesso h oo n Rectum and Colon the of Diseases , rceig fteNtoa cdm of Academy National the of Proceedings 96 oeua acrRsac:MCR Research: Cancer Molecular 4,261–268. (4), shff . rvsaa S. Srivastava, . . . J., uschoff, ¨ urn Genomics Current , 304 , 107 57) 1164–1166. (5674), acrResearch Cancer aueReviews. Nature 6,2592–2597. (6), Oncotarget , Clinical , 18 34 (4). (5), , , 8 9 Posted on Authorea 17 Sep 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.160037743.35386400 — This a preprint and has not been peer reviewed. Data may be preliminary. aiBelhad Sami Gonz´alez-MoralesMar´ıa Luisa Garc´ıa-Barber´anVanesa 0000-0002-2531-0203 0000-0003-0190-2105 Llovet Patricia P´erez-Segura 0000-0001-5049-7199 Pedro 0000-0002-3681-4319 Lorca Victor Mart´ın-Morales 0000-0002-1970-8872 Lorena co-authors repre- of bars IDs Data error ORCiD the respectively. experiments; cancer, Appendix: independent breast 2 and represent deviation. colorectal and standard with replicates the affected technical sent (B+D), three from II:4 collected and was two (A+C) of II:6 tumors members the family in genes of target Quantification STAT3 of Expression B) where 5. Target/Total, line). Figure as intervals. solid confidence presented with the (circled PCR to allele digital correspond wild-type by mutant bars the the obtained mutant tissue detects detects the expression dye dye colon FAM is VIC allele-specific ”Target” healthy The the the the II:6. while of in member line), family Quantification performed dashed cc765 assay with of expression (circled (bottom) (CRC) allele allele-specific cancer the colorectal of and (top) visualization D1364Gfs*24 PCR controls. and Digital commercial wild-type unmethylated (A) of and expression methylated com- Allele-specific a with and together 4. II:4 A, member Figure in colon of healthy analyzed breast a healthy samples and vs same II:6 cancer (C) the member breast for of the sample. of tissue breast status colon healthy methylation healthy the vs mercial shows CRC B the Panel of status pool. methylation the shows A Panel the (A+B) between differ that residues carriers. the tion 2013. residues all Those al. are mutation. et 3. Underlined Lui the Figure by from grey. produced Modified in PTPRT. effect marked SWISS-MODEL. of the by are and forms with obtained two pTyr (left) domain were wild-type the D2 models PTPRT’s of surrounding 3D of region directly truncation. sequence cytoplasmatic the acid the the of and Amino of effect mutation C) representation the studied showing the 3D PTPRT, of B) (right) location mutant (below). the with protein domains, mutant protein resulting PTPRT of representation Schematic (A) included inserted. is of is members guanine Effect healthy the the 2. of where of age Figure point sequence mutant current the and or show wild-type the diagnosis arrows of cancer at The Electropherogram (WT). endometrial age B) non-carriers gene. right: The years). were top (in (BC). three CRC, individual cancer while each right: beneath (+), breast bottom carriers corner; left: cosegregation were black the bottom individuals a and Four (EC), with box), marked grey III:5. are a and (in members II:4 II:7 Cancer-affected and II:3, II:6 II:2, II:5, in members analyzed family in done where was cc765, sequencing family Whole-exome II Amsterdam carrying of Pedigree family (A) the of Pedigree 1. Figure PTPRT PTPRT rmtrmtyainotie yMPo w ieetpooe ie rvosydescribed. previously sites promoter different two of MSP by obtained methylation promoter BCL-XL rmtri yemtyae nteclnadbes acr ftomuta- two of cancers breast and colon the in hypermethylated is promoter PTPRT PTPRT AB and (A+B) 16Gs2 ntepoenstructure. protein the on D1364Gfs*24 lee aawscletdfo w needn xeiet,adteerror the and experiments, independent two from collected was Data allele. PTPRT SOCS3 rmtrmtyainsau % bandb pyrosequencing by obtained (%) status methylation promoter CD xrsinotie yqC ntetmr fcc765 of tumors the in qPCR by obtained expression (C+D) PTPRT PTPRT 13 D1364Gfs*24. .00u .Ap34lfTr4,wsidentified. was p.(Asp1364GlyfsTer24), c.4090dup PTPRT PTPRT uaincarriers. mutation . PTPRT Posted on Authorea 17 Sep 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.160037743.35386400 — This a preprint and has not been peer reviewed. Data may be preliminary. rnddCald´es 0000-0002-1038-5392 Trinidad 0000-0001-8285-4138 Garre Pilar 0000-0002-8113-1410 Hoya la de Miguel 0000-0003-0371-0844 Valle Laura Capell´a 0000-0002-2159-1351 Gabriel 14 Posted on Authorea 17 Sep 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.160037743.35386400 — This a preprint and has not been peer reviewed. Data may be preliminary. 15 Posted on Authorea 17 Sep 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.160037743.35386400 — This a preprint and has not been peer reviewed. Data may be preliminary. 16 Posted on Authorea 17 Sep 2020 — The copyright holder is the author/funder. All rights reserved. No reuse without permission. — https://doi.org/10.22541/au.160037743.35386400 — This a preprint and has not been peer reviewed. Data may be preliminary. 17