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Objective: 3′ tRNA‐derived fragments (3′ tRFs) are important epigenetic regulators in normal and pathological conditions. In this study, we aimed to explore the potential value of a 3′ tRF as a prognostic and/or screening biomarker for B‐cell chronic lymphocytic leukemia (B‐CLL). Methods: Publicly available next‐generation sequencing data from 20 B‐CLL cases were analyzed, followed by prediction of targets of the most abundantly and ubiquitously expressed 3′ tRFs, leading to selection of tRF‐LeuAAG/TAG. PBMCs were isolated from blood samples of 91 B‐CLL patients and 43 non‐leukemic donors, followed by total RNA extraction, in‐vitro polyadenylation, and first‐strand cDNA synthesis. Next, a real‐time quantitative PCR (qPCR) assay was developed for the accurate quantification of tRF‐LeuAAG/TAG and applied in all samples, prior to biostatistical analysis. Results: High tRF‐LeuAAG/TAG levels are associated with inferior overall survival (OS) of B‐CLL patients. The unfavorable significance of tRF‐LeuAAG/TAG was independent of established prognostic factors in B‐CLL. Stratified Kaplan‐Meier OS analysis uncovered the unfavorable prognostic role of high tRF‐LeuAAG/TAG levels for patients in Binet A or Rai I stage, negative CD38 expression, mutated, or unmutated IGHV genomic locus. Conclusion: Our approach revealed the independent prognostic value of a particular 3′ tRF, derived from tRNALeuAAG and tRNALeuTAG (tRF‐LeuAAG/TAG) in B‐CLL. [ABSTRACT FROM AUTHOR]

• A qPCR assay was developed for the quantification of a new mt-tRNA-derived fragment. • i-tRF-PheGAA can distinguish between CLL patients and normal controls. • An association was observed between i-tRF-PheGAA expression and CLL clinical stage. • Patients overexpressing i-tRF-PheGAA show significantly poorer overall survival. • The prognostic power of i-tRF-PheGAA is independent of established prognosticators. Chronic lymphocytic leukemia (CLL) is the most common leukemia among adults. The prognosis of CLL patients varies considerably. Transfer RNA-derived RNA fragments (tRFs) constitute a class of small non-coding RNA fragments excised from mature tRNAs and pre-tRNAs located in nuclei as well as in mitochondria. In this study, the clinical utility of i-tRF-PheGAA, a novel mitochondrial tRF, was investigated in CLL. Peripheral blood mononuclear cells (PBMCs) were isolated from 91 CLL patients and 43 non-leukemic controls. Total RNA was isolated from each sample, polyadenylated at the 3′ end and reversely transcribed. An in-house developed real-time quantitative PCR assay was developed and applied, and the results were biostatistically analyzed. For the normalization of the i-tRF-PheGAA expression levels, the expression of a small nucleolar RNA (RNU48) was used as reference. Mann-Whitney U test showed that i-tRF-PheGAA can distinguish between CLL samples and normal controls (p < 0.001). As determined by Kaplan-Meier survival analysis, overexpression of i-tRF-PheGAA was related to poor overall survival of the CLL patients (p < 0.001). Univariate bootstrap Cox regression analysis exhibited a higher hazard ratio of 7.95 (95% CI = 2.37–26.72, p < 0.001) for patients with positive i-tRF-PheGAA expression status. Multivariate bootstrap Cox regression analysis showed that the prognostic value of this tRF is independent of clinical stage, mutational status of the immunoglobulin heavy chain variable (IGHV) genetic locus, and CD38 expression status (p = 0.010). Our results show that i-tRF-PheGAA can serve as a molecular biomarker of poor prognosis in CLL, alongside with the existing factors for CLL prognosis. [ABSTRACT FROM AUTHOR]

MicroRNA-92a-3p (miR-92a-3p) derives from the oncogenic miR-17/92 cluster and its highly homologous miR-106a/363 cluster. miR-92a-3p regulates the expression of key transcription factors such as HIF1 and inhibits SOCS1 to enhance the anti-apoptotic STAT3/IL6 signaling pathway. In this study, we assessed the putative usefulness of miR-92a-3p as a prognostic and/or diagnostic biomarker in chronic lymphocytic leukemia (CLL). For this purpose, total RNA was extracted from mononuclear cells isolated from the peripheral blood of 88 CLL patients and 36 non-leukemic blood donors, was polyadenylated and reversely transcribed. miR-92a-3p expression was quantified using an accurate qPCR method. miR-92a-3p levels were significantly lower in peripheral blood mononuclear cells of CLL patients. Overall survival (OS) analysis revealed that high miR-92a-3p expression predicts significantly prolonged OS of CLL patients. Interestingly, miR-92a-3p overexpression remains a significant prognosticator in subgroups of CLL patients with distinct prognosis. In conclusion, miR-92a-3p overexpression is a potential surrogate biomarker of favorable outcome of CLL patients. [ABSTRACT FROM AUTHOR]

MicroRNA-20b-5p (miR-20b-5p) is part of the miR-106a/363 cluster and a member of the cancer-related miR-17 family. miR-20b-5p regulates important transcription factors, including hypoxia-inducible factor 1 (HIF1) and signal transducer and activator of transcription 3 (STAT3). Recently, the dysregulation of miR-20b-5p expression has been observed in many B-cell lymphomas and T-cell leukemias. In this research study, we examined the putative prognostic value of miR-20b-5p in CLL. Therefore, total RNA was isolated from peripheral blood mononuclear cells (PBMCs) collected from 88 CLL patients; next, total RNA was polyadenylated and first-strand cDNA was synthesized, using an oligo-dT–adapter primer. miR-20b-5p expression was quantified using an in-house–developed real-time quantitative PCR assay. Kaplan-Meier OS analysis and bootstrap univariate Cox regression showed that high miR-20b-5p expression predicts better OS for CLL patients ( p <  0.001). Interestingly, miR-20b-5p overexpression retains its favorable prognostic role in CLL patients of intermediate risk or stratified according to established prognostic factors [CD38 expression and mutational status of the immunoglobulin heavy chain variable ( IGHV ) region]. In conclusion, miR-20b-5p is a potential independent molecular biomarker of favorable prognosis in CLL. [ABSTRACT FROM AUTHOR]

MicroRNA-155-5p (miR-155-5p) is a proinflammatory, oncogenic miRNA, involved in various physiological processes, including hematopoiesis, immunity, inflammation, and cell lineage differentiation. It regulates important transcription factors, such as E2F2, hypoxia-inducible factor 1 (HIF1), and FOXO3. Recently, the dysregulation of miR-155-5p expression has been linked to chronic lymphocytic leukemia (CLL) pathogenesis. In this research study, we investigated the potential diagnostic and prognostic value of miR-155-5p in CLL. To achieve our goal, we isolated total RNA from peripheral blood mononuclear cells (PBMCs) collected from 88 CLL patients and 36 nonleukemic blood donors and performed polyadenylation of total RNA and reverse transcription. Next, we quantified miR-155-5p levels using an in-house-developed real-time quantitative PCR method, before proceeding to extensive biostatistical analysis. Thus, it appears that miR-155-5p is significantly overexpressed in PBMCs of CLL patients and can distinguish them from nonleukemic population. Kaplan-Meier OS analysis and bootstrap univariate Cox regression showed that high miR-155-5p expression predicts inferior OS for CLL patients (p<0.001). Interestingly, miR-155-5p overexpression retains its unfavorable prognostic role in CLL patients stratified according to established prognostic factors [CD38 expression and mutational status of the immunoglobulin heavy chain variable region (IGHV)]. Thus, miR-155-5p appears as a promising, independent molecular biomarker of unfavorable prognosis in CLL. [ABSTRACT FROM AUTHOR]

Accumulated data indicate a significant role of T cell dysfunction in the pathogenesis of chronic lymphocytic leukemia. In CLL, regulatory T cells are significantly higher and show lower apoptotic levels compared to healthy donors. We demonstrate that CLL derived CD4 + CD25 − CD127 − and CD4 + CD25 low CD127 − subpopulations share a common immunophenotypic profile with conventional Tregs and are associated with advanced stage disease. We further provide evidence that the increased number of Tregs contributes indirectly to the proliferation of the CLL clone, by suppressing the proliferation of Teffs which in turn suppress CLL cells. These data are further supported by our observations that CLL derived Tregs appear rather incapable of inducing apoptosis of both normal B cells and CLL cells, in contrast to normal Tregs, suggesting an immunoediting effect of CLL cells on Tregs which negatively affects the functionality of the latter and contributes to the failure of Tregs in CLL to efficiently eliminate the abnormal clone. [ABSTRACT FROM AUTHOR]

Anti-CD20-based chemo-immunotherapeutic regimens have been suggested to assist in the management of Epstein-Barr virus (EBV)-induced hemophagocytic lymphohistiocytosis (HLH) and EBV-associated post-transplant lymphoproliferative disorders (EBV-PTLD), by reducing EBV viral load and EBV-induced inflammation. Herein we report a fatal EBV-related HLH in the context of Hodgkin lymphoma (HL)-like Richter’s transformation of B chronic lymphocytic leukemia (B-CLL), two months after rituximab treatment. The complex balance between EBV driven T-cell stimulation and immunosuppressive therapy in the context of multiple immune deficits is discussed. [ABSTRACT FROM AUTHOR]

Simple Summary: The involvement of miRNAs in physiological cellular processes has been well documented. The development of B cells, which is dictated by a miRNA-transcription factor regulatory network, suggests a typical process partly orchestrated by miRNAs. Besides their contribution in normal hematopoiesis, miRNAs have been severally reported to be implicated in hematological malignancies, a typical example of which is B-cell chronic lymphocytic leukemia (B-CLL). Numerous studies have attempted to highlight the regulatory role of miRNAs in B-CLL or establish some of them as molecular biomarkers or therapeutic targets. Thus, a critical review summarizing the current knowledge concerning the multifaceted role of miRNAs in normal B-cell development and B-CLL progression, prognosis, and therapy, is urgent. Moreover, this review aims to highlight important miRNAs in both normal B-cell development and B-CLL and discuss future perspectives concerning their regulatory potential and establishment in clinical practice. MicroRNAs (miRNAs) represent a class of small non-coding RNAs bearing regulatory potency. The implication of miRNAs in physiological cellular processes has been well documented so far. A typical process orchestrated by miRNAs is the normal B-cell development. A stage-specific expression pattern of miRNAs has been reported in the developmental procedure, as well as interactions with transcription factors that dictate B-cell development. Besides their involvement in normal hematopoiesis, miRNAs are severally implicated in hematological malignancies, a typical paradigm of which is B-cell chronic lymphocytic leukemia (B-CLL). B-CLL is a highly heterogeneous disease characterized by the accumulation of abnormal B cells in blood, bone marrow, lymph nodes, and spleen. Therefore, timely, specific, and sensitive assessment of the malignancy is vital. Several studies have attempted to highlight the remarkable significance of miRNAs as regulators of gene expression, biomarkers for diagnosis, prognosis, progression, and therapy response prediction, as well as molecules with potential therapeutic utility. This review seeks to outline the linkage between miRNA function in normal and malignant hematopoiesis by demonstrating the main benchmarks of the implication of miRNAs in the regulation of normal B-cell development, and to summarize the key findings about their value as regulators, biomarkers, or therapeutic targets in B-CLL. [ABSTRACT FROM AUTHOR]

• An innovative qPCR method for quantification of i-tRF-GlyCCC levels was developed. • High i-tRF-GlyCCC expression predicts poor overall survival in CLL patients. • Prognostic significance of i-tRF-GlyCCC is independent of other prognostic factors. • i-tRF-GlyCCC is an unfavorable prognosticator in distinct subgroups of CLL patients Chronic lymphocytic leukemia (CLL) is one of the most common types of leukemia in adults. Several studies have identified various prognostic biomarkers in CLL. In this study, we investigated the potential value of an internal fragment of the tRNAs bearing the Glycine anticodon CCC (i-tRF-GlyCCC), which is a small non-coding RNA, as a prognostic and screening biomarker in CLL. For this purpose, blood samples were collected from 90 CLL patients and 43 non-leukemic blood donors. Peripheral blood mononuclear cells (PBMCs) were isolated, total RNA was extracted and in-vitro polyadenylated, and first-strand cDNA was synthesized using an oligo-dT–adaptor primer. A real-time quantitative PCR assay was developed and applied for the quantification of i-tRF-GlyCCC in our samples. The biostatistical analysis revealed that i-tRF-GlyCCC levels are significantly lower in PBMCs of CLL patients, compared to PBMCs of non-leukemic controls, and that i-tRF-GlyCCC could be considered as a screening biomarker. Kaplan-Meier overall survival (OS) analysis revealed reduced OS for CLL patients with positive i-tRF-GlyCCC expression (P = 0.001). Multivariate Cox regression confirmed its independent unfavorable prognostic power with regard to OS. In conclusion, i-tRF-GlyCCC may constitute a promising molecular biomarker in CLL, for screening and prognostic purposes. [ABSTRACT FROM AUTHOR]