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Exp Hematol Oncol. 2016 Mar 1;5:7. doi: 10.1186/s40164-016-0036-3. eCollection 2015.

New CD20 alternative splice variants: molecular identification and differential expression within hematological B cell malignancies.

Author information

1
INSERM UMR1098, Établissement Français du Sang Bourgogne Franche Comté, Université de Franche-Comté, SFR FED4234, 25020 Besançon, France.
2
EA3181 et Service de Dermatologie, Université de Franche Comté, CHU de Besançon, Besançon, France.
3
EA3181 et Service de Dermatologie, Université de Franche Comté, CHU de Besançon, Besançon, France ; CHRU, Department of Rheumatology, Université de Franche-Comté EA 4266, INSERM CIC-1431, 25000 Besançon, France ; EA 4266, Université de Franche-Comté, Besançon, France.
4
INSERM UMR1098, Établissement Français du Sang Bourgogne Franche Comté, Université de Franche-Comté, SFR FED4234, 25020 Besançon, France ; EA3181 et Service de Dermatologie, Université de Franche Comté, CHU de Besançon, Besançon, France.
5
Inserm U1037, Université Toulouse 3-ERL CNRS, CHU Purpan, Toulouse, France.
6
Hématologie Clinique, CHU Estaing, 1 Place Lucie Aubrac, 63003 Clermont-Ferrand Cedex 1, France.
7
Hematologie, CHU Bretonneau, Tours, France.
8
INSERM UMR1098, Établissement Français du Sang Bourgogne Franche Comté, Université de Franche-Comté, SFR FED4234, 25020 Besançon, France ; Hematology Department, CHU Jean Minjoz, 25020 Besançon, France.
9
INSERM UMR1098, Établissement Français du Sang Bourgogne Franche Comté, Université de Franche-Comté, SFR FED4234, 25020 Besançon, France ; EA3181 et Service de Dermatologie, Université de Franche Comté, CHU de Besançon, Besançon, France ; Hematology Department, CHU Jean Minjoz, 25020 Besançon, France.
10
INSERM UMR1098, Établissement Français du Sang Bourgogne Franche Comté, Université de Franche-Comté, SFR FED4234, 25020 Besançon, France ; Laboratoire de Thérapeutique Immuno-Moléculaire et cellulaire des cancers, INSERM UMR1098, Etablissement Français du Sang-Bourgogne/Franche-Comté, 8, rue du Docteur Jean-François-Xavier Girod, 25020 Besançon Cedex, France.

Abstract

BACKGROUND:

CD20 is a B cell lineage-specific marker expressed by normal and leukemic B cells and targeted by several antibody immunotherapies. We have previously shown that the protein from a CD20 mRNA splice variant (D393-CD20) is expressed at various levels in leukemic B cells or lymphoma B cells but not in resting, sorted B cells from the peripheral blood of healthy donors.

RESULTS:

Western blot (WB) analysis of B malignancy primary samples showed additional CD20 signals. Deep molecular PCR analysis revealed four new sequences corresponding to in-frame CD20 splice variants (D657-CD20, D618-CD20, D480-CD20, and D177-CD20) matching the length of WB signals. We demonstrated that the cell spliceosome machinery can process ex vivo D480-, D657-, and D618-CD20 transcript variants by involving canonical sites associated with cryptic splice sites. Results of specific and quantitative RT-PCR assays showed that these CD20 splice variants are differentially expressed in B malignancies. Moreover, Epstein-Barr virus (EBV) transformation modified the CD20 splicing profile and mainly increased the D393-CD20 variant transcripts. Finally, investigation of three cohorts of chronic lymphocytic leukemia (CLL) patients showed that the total CD20 splice variant expression was higher in a stage B and C sample collection compared to routinely collected CLL samples or relapsed refractory stage A, B, or C CLL.

CONCLUSION:

The involvement of these newly discovered alternative CD20 transcript variants in EBV transformation makes them interesting molecular indicators, as does their association with oncogenesis rather than non-oncogenic B cell diseases, differential expression in B cell malignancies, and correlation with CLL stage and some predictive CLL markers. This potential should be investigated in further studies.

KEYWORDS:

Alternative splicing; B malignancies; CD20; CLL; EBV transformation

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