5265d62c6edeebeafe31867aed569fb9 cancers-14-03088-v2.pdf 11bd74c3f1ff41e1ded9d573d667e8018029125d cancers-14-03088-v2.pdf 05d801fb5394b4784df5a4ab5809771839f4d800f9d8e5fac1d7ebf0605d126c cancers-14-03088-v2.pdf Title: Bone Marrow Stromal Cell Regeneration Profile in Treated B-Cell Precursor Acute Lymphoblastic Leukemia Patients: Association with MRD Status and Patient Outcome Subject: For the last two decades, measurable residual disease (MRD) has become one of the most powerful independent prognostic factors in B-cell precursor acute lymphoblastic leukemia (BCP-ALL). However, the effect of therapy on the bone marrow (BM) microenvironment and its potential relationship with the MRD status and disease free survival (DFS) still remain to be investigated. Here we analyzed the distribution of mesenchymal stem cells (MSC) and endothelial cells (EC) in the BM of treated BCP-ALL patients, and its relationship with the BM MRD status and patient outcome. For this purpose, the BM MRD status and EC/MSC regeneration profile were analyzed by multiparameter flow cytometry (MFC) in 16 control BM (10 children; 6 adults) and 1204 BM samples from 347 children and 100 adult BCP-ALL patients studied at diagnosis (129 children; 100 adults) and follow-up (824 childhood samples; 151 adult samples). Patients were grouped into a discovery cohort (116 pediatric BCP-ALL patients; 338 samples) and two validation cohorts (74 pediatric BCP-ALL, 211 samples; and 74 adult BCP-ALL patients; 134 samples). Stromal cells (i.e., EC and MSC) were detected at relatively low frequencies in all control BM (16/16; 100%) and in most BCP-ALL follow-up samples (874/975; 90%), while they were undetected in BCP-ALL BM at diagnosis. In control BM samples, the overall percentage of EC plus MSC was higher in children than adults (p = 0.011), but with a similar EC/MSC ratio in both groups. According to the MRD status similar frequencies of both types of BM stromal cells were detected in BCP-ALL BM studied at different time points during the follow-up. Univariate analysis (including all relevant prognostic factors together with the percentage of stromal cells) performed in the discovery cohort was used to select covariates for a multivariate Cox regression model for predicting patient DFS. Of note, an increased percentage of EC (>32%) within the BCP-ALL BM stromal cell compartment at day +78 of therapy emerged as an independent unfavorable prognostic factor for DFS in childhood BCP-ALL in the discovery cohort—hazard ratio (95% confidence interval) of 2.50 (1–9.66); p = 0.05—together with the BM MRD status (p = 0.031). Further investigation of the predictive value of the combination of these two variables (%EC within stromal cells and MRD status at day +78) allowed classification of BCP-ALL into three risk groups with median DFS of: 3.9, 3.1 and 1.1 years, respectively (p = 0.001). These results were confirmed in two validation cohorts of childhood BCP-ALL (n = 74) (p = 0.001) and adult BCP-ALL (n = 40) (p = 0.004) treated at different centers. In summary, our findings suggest that an imbalanced EC/MSC ratio in BM at day +78 of therapy is associated with a shorter DFS of BCP-ALL patients, independently of their MRD status. Further prospective studies are needed to better understand the pathogenic mechanisms involved. Keywords: B-cell precursor acute lymphoblastic leukemia; stromal cells; mesenchymal stem cells; endothelial cells; bone marrow microenvironment; multiparameter flow cytometry; disease free survival; measurable residual disease Author: Elen Oliveira, Elaine S. Costa, Juana Ciudad, Giuseppe Gaipa, Łukasz Sedek, Susana Barrena, Tomasz Szczepanski, Chiara Buracchi, Daniela Silvestri, Patrícia F. R. Siqueira, Fabiana V. Mello, Rafael C. Torres, Leonardo M. R. Oliveira, Isabelle V. C. Fay-Neves, Edwin Sonneveld, Vincent H. J. van der Velden, Esther Mejstrikova, Josep-Maria Ribera, Valentino Conter, Martin Schrappe, Jacques J. M. van Dongen, Marcelo G. P. Land and Alberto Orfao Creator: LaTeX with hyperref Producer: pdfTeX-1.40.21 CreationDate: Mon Jun 27 04:59:54 2022 CEST ModDate: Mon Jun 27 09:20:45 2022 CEST Tagged: no UserProperties: no Suspects: no Form: none JavaScript: no Pages: 18 Encrypted: no Page size: 595.276 x 841.89 pts (A4) Page rot: 0 File size: 18221810 bytes Optimized: no PDF version: 1.7 name type encoding emb sub uni object ID ------------------------------------ ----------------- ---------------- --- --- --- --------- WLENKM+VnURWPalladioL-Bold Type 1 Custom yes yes no 10 0 VJAUGW+VnURWPalladioL Type 1 Custom yes yes no 15 0 HZFBWV+URWPalladioL-Bold Type 1 Custom yes yes no 19 0 PATWLQ+URWPalladioL-Roma Type 1 Custom yes yes no 24 0 FAKNYV+URWPalladioL-Ital Type 1 Custom yes yes no 28 0 LCJEVJ+TimesNewRomanPS-BoldItalicMT Type 1C WinAnsi yes yes no 47 0 KOSUXD+CMSY10 Type 1 Builtin yes yes no 75 0 CBHVLJ+EURM10 Type 1 Builtin yes yes no 79 0 MPUDFL+PazoMath Type 1 Builtin yes yes no 100 0 WBVKDJ+URWPalladioL-BoldItal Type 1 Custom yes yes no 104 0 DFFMPE+Symbol Type 1C Custom yes yes yes 111 0 DFFPPG+CordiaNew,Bold TrueType WinAnsi yes yes no 116 0 DFFHLO+PalatinoLinotype,Bold TrueType WinAnsi yes yes no 119 0 DFFHLP+PalatinoLinotype CID TrueType Identity-H yes yes yes 122 0 DFFHMA+PalatinoLinotype TrueType WinAnsi yes yes no 128 0 DFFHMC+PalatinoLinotype,Italic TrueType WinAnsi yes yes no 131 0 DFFPPE+PalatinoLinotype,BoldItalic TrueType WinAnsi yes yes no 134 0 Jhove (Rel. 1.22.1, 2019-04-17) Date: 2023-07-01 02:41:37 CEST RepresentationInformation: cancers-14-03088-v2.pdf ReportingModule: PDF-hul, Rel. 1.12.1 (2019-04-17) LastModified: 2023-02-10 18:19:48 CET Size: 18221810 Format: PDF Version: 1.7 Status: Well-Formed and valid SignatureMatches: PDF-hul MIMEtype: application/pdf PDFMetadata: Objects: 439 FreeObjects: 7 IncrementalUpdates: 0 DocumentCatalog: PageLayout: SinglePage PageMode: UseNone Outlines: Item: Title: Introduction Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@44c8afef Item: Title: Materials and Methods Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@7b69c6ba Children: Item: Title: Patients, Samples, and Controls Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@46daef40 Item: Title: Immunophenotypic Studies Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@12f41634 Item: Title: Statistical Methods Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@13c27452 Item: Title: Results Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@262b2c86 Children: Item: Title: Immunophenotypic Identification and Characterization of Stromal Cells in Bone Marrow Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@371a67ec Item: Title: Distribution of MSC and EC in Normal vs. BCP-ALL Bone Marrow Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@5ed828d Item: Title: MRD Status in Follow-Up BM of Childhood and Adult BCP-ALL and Its Relationship with the Distribution of Stromal Cells Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@50d0686 Item: Title: Relationship between the Distribution of MSC and EC in BM at Day +78 and Both the MRD Status and Outcome of BCP-ALL Patients Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@7a3d45bd Item: Title: Discussion Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@1e7c7811 Item: Title: Conclusions Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@77ec78b9 Item: Title: References Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@1a3869f4 Info: Title: Bone Marrow Stromal Cell Regeneration Profile in Treated B-Cell Precursor Acute Lymphoblastic Leukemia Patients: Association with MRD Status and Patient Outcome Author: Elen Oliveira, Elaine S. Costa, Juana Ciudad, Giuseppe Gaipa, Łukasz Sedek, Susana Barrena, Tomasz Szczepanski, Chiara Buracchi, Daniela Silvestri, Patrícia F. R. Siqueira, Fabiana V. Mello, Rafael C. Torres, Leonardo M. R. Oliveira, Isabelle V. C. Fay-Neves, Edwin Sonneveld, Vincent H. J. van der Velden, Esther Mejstrikova, Josep-Maria Ribera, Valentino Conter, Martin Schrappe, Jacques J. M. van Dongen, Marcelo G. P. Land and Alberto Orfao Subject: For the last two decades, measurable residual disease (MRD) has become one of the most powerful independent prognostic factors in B-cell precursor acute lymphoblastic leukemia (BCP-ALL). However, the effect of therapy on the bone marrow (BM) microenvironment and its potential relationship with the MRD status and disease free survival (DFS) still remain to be investigated. Here we analyzed the distribution of mesenchymal stem cells (MSC) and endothelial cells (EC) in the BM of treated BCP-ALL patients, and its relationship with the BM MRD status and patient outcome. For this purpose, the BM MRD status and EC/MSC regeneration profile were analyzed by multiparameter flow cytometry (MFC) in 16 control BM (10 children; 6 adults) and 1204 BM samples from 347 children and 100 adult BCP-ALL patients studied at diagnosis (129 children; 100 adults) and follow-up (824 childhood samples; 151 adult samples). Patients were grouped into a discovery cohort (116 pediatric BCP-ALL patients; 338 samples) and two validation cohorts (74 pediatric BCP-ALL, 211 samples; and 74 adult BCP-ALL patients; 134 samples). Stromal cells (i.e., EC and MSC) were detected at relatively low frequencies in all control BM (16/16; 100%) and in most BCP-ALL follow-up samples (874/975; 90%), while they were undetected in BCP-ALL BM at diagnosis. In control BM samples, the overall percentage of EC plus MSC was higher in children than adults (p = 0.011), but with a similar EC/MSC ratio in both groups. According to the MRD status similar frequencies of both types of BM stromal cells were detected in BCP-ALL BM studied at different time points during the follow-up. Univariate analysis (including all relevant prognostic factors together with the percentage of stromal cells) performed in the discovery cohort was used to select covariates for a multivariate Cox regression model for predicting patient DFS. Of note, an increased percentage of EC (>32%) within the BCP-ALL BM stromal cell compartment at day +78 of therapy emerged as an independent unfavorable prognostic factor for DFS in childhood BCP-ALL in the discovery cohort hazard ratio (95% confidence interval) of 2.50 (1 9.66); p = 0.05 together with the BM MRD status (p = 0.031). Further investigation of the predictive value of the combination of these two variables (%EC within stromal cells and MRD status at day +78) allowed classification of BCP-ALL into three risk groups with median DFS of: 3.9, 3.1 and 1.1 years, respectively (p = 0.001). These results were confirmed in two validation cohorts of childhood BCP-ALL (n = 74) (p = 0.001) and adult BCP-ALL (n = 40) (p = 0.004) treated at different centers. In summary, our findings suggest that an imbalanced EC/MSC ratio in BM at day +78 of therapy is associated with a shorter DFS of BCP-ALL patients, independently of their MRD status. Further prospective studies are needed to better understand the pathogenic mechanisms involved. 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