High-throughput mutagenesis identifies mutations and RNA-binding proteins controlling CD19 splicing and CART-19 therapy resistance.
High-throughput mutagenesis identifies mutations and RNA-binding proteins controlling CD19 splicing and CART-19 therapy resistance.
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DOI：
10.1038/s41467-022-31818-y
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发表时间：
2022-09-22
影响因子：
16.6
通讯作者：
König J
中科院分区：
文献类型：
--
作者：
Cortés-López M;Schulz L;Enculescu M;Paret C;Spiekermann B;Quesnel-Vallières M;Torres-Diz M;Unic S;Busch A;Orekhova A;Kuban M;Mesitov M;Mulorz MM;Shraim R;Kielisch F;Faber J;Barash Y;Thomas-Tikhonenko A;Zarnack K;Legewie S;König J
在对B细胞急性淋巴细胞白血病（B - ALL）进行CART - 19免疫治疗后，许多患者由于同源CD19表位缺失而复发。由于表位缺失可能由异常的CD19外显子2加工过程导致，我们在此研究控制CD19剪接的调控密码。我们将高通量诱变与数学建模相结合，以定量地梳理包含CD19外显子1 - 3区域内所有突变的影响。于是，我们确定了约200个单点突变，这些突变改变了CD19剪接，因此可能使B - ALL患者易于产生CART - 19耐药性。此外，我们报道了近100种先前未知的剪接异构体，它们由隐蔽剪接位点产生，可能编码无功能的CD19蛋白。我们进一步确定了控制CD19剪接的顺式调控元件和反式作用RNA结合蛋白（例如，PTBP1和SF3B4），并验证了这些因子的缺失会导致广泛的CD19错误剪接。我们的数据集是用于识别CART - 19治疗预测性生物标志物的综合资源。
CD19 mRNA中的多种可变剪接事件与B细胞恶性肿瘤患者对CD19 CAR - T治疗的耐药性/复发相关。在此，通过结合患者数据和高通量诱变筛选，作者确定了能够控制CD19剪接并与CD19 CAR - T治疗耐药性相关的单点突变和RNA结合蛋白。
Following CART-19 immunotherapy for B-cell acute lymphoblastic leukaemia (B-ALL), many patients relapse due to loss of the cognate CD19 epitope. Since epitope loss can be caused by aberrant CD19 exon 2 processing, we herein investigate the regulatory code that controls CD19 splicing. We combine high-throughput mutagenesis with mathematical modelling to quantitatively disentangle the effects of all mutations in the region comprising CD19 exons 1-3. Thereupon, we identify ~200 single point mutations that alter CD19 splicing and thus could predispose B-ALL patients to developing CART-19 resistance. Furthermore, we report almost 100 previously unknown splice isoforms that emerge from cryptic splice sites and likely encode non-functional CD19 proteins. We further identify cis-regulatory elements and trans-acting RNA-binding proteins that control CD19 splicing (e.g., PTBP1 and SF3B4) and validate that loss of these factors leads to pervasive CD19 mis-splicing. Our dataset represents a comprehensive resource for identifying predictive biomarkers for CART-19 therapy. Multiple alternative splicing events in CD19 mRNA have been associated with resistance/relapse to CD19 CAR-T therapy in patients with B cell malignancies. Here, by combining patient data and a high-throughput mutagenesis screen, the authors identify single point mutations and RNA-binding proteins that can control CD19 splicing and be associated with CD19 CAR-T therapy resistance.
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影响因子：
20.3
作者：
Gardner, Rebecca A.;Finney, Olivia;Jensen, Michael C.
通讯作者：
Jensen, Michael C.
影响因子：
13.6
作者：
El Marabti E;Abdel-Wahab O
通讯作者：
Abdel-Wahab O
影响因子：
14.9
作者：
Frankish A;Diekhans M;Ferreira AM;Johnson R;Jungreis I;Loveland J;Mudge JM;Sisu C;Wright J;Armstrong J;Barnes I;Berry A;Bignell A;Carbonell Sala S;Chrast J;Cunningham F;Di Domenico T;Donaldson S;Fiddes IT;García Girón C;Gonzalez JM;Grego T;Hardy M;Hourlier T;Hunt T;Izuogu OG;Lagarde J;Martin FJ;Martínez L;Mohanan S;Muir P;Navarro FCP;Parker A;Pei B;Pozo F;Ruffier M;Schmitt BM;Stapleton E;Suner MM;Sycheva I;Uszczynska-Ratajczak B;Xu J;Yates A;Zerbino D;Zhang Y;Aken B;Choudhary JS;Gerstein M;Guigó R;Hubbard TJP;Kellis M;Paten B;Reymond A;Tress ML;Flicek P
通讯作者：
Flicek P
影响因子：
5.3
作者：
Bagashev A;Sotillo E;Tang CH;Black KL;Perazzelli J;Seeholzer SH;Argon Y;Barrett DM;Grupp SA;Hu CC;Thomas-Tikhonenko A
通讯作者：
Thomas-Tikhonenko A
影响因子：
64.5
作者：
Baeza-Centurion, Pablo;Minana, Belen;Lehner, Ben
通讯作者：
Lehner, Ben