A Rare Entity of Accelerated Chronic Lymphocytic Leukemia: A Report of Two Cases and Review of Literature

Background: Accelerated-chronic lymphocytic leukemia (A-CLL) is a rare disease entity as it represents less than 1% of all reported cases of chronic lymphoid leukemia (CLL). Moreover, it is most likely an under diagnosed entity due to its rarity and the non-standardized practice of lymph node biopsy in CLL. Purpose: The aims of our work are to establish the diagnosis of A-CLL and to study the prognosis and treatment of this rare entity. Method: here, we report the clinical presentation and the follow up of two cases of A-CLL. Results: Distinguishing Richter transformation (RT) from A-CLL is important as it may result in a major change in disease management. The prognosis of A-CLL is intermediate between CLL and RT. The prognosis is mainly poor due to a predominance of poor prognostic markers including an increasing number of p53-positive cases. Conclusion: To this date, no prospective study has been led to define the best treatment for A-CLL. The shorter survival of A-CLL when compared to typical CLL implies the need of a more aggressive treatment.

Serum LDH level may predict outcome of chronic lymphocytic leukemia patients with a 17p deletion： a retrospective analysis of prognostic factors in China

Objective： This study aims to evaluate the natural history of patients with chronic lymphocytic leukemia （CLL） and a 17p deletion （17p-） and identify the predictive factors within this subgroup. Methods： The sample of patients with CLL were analyzed by fluorescence in situ hybridization for deletions in chromosome bands 1 lq22, 13q14 and 17p13; trisomy of bands 12q13; and translocation involving band 14q32. The data from 456 patients with or without a 17p- were retrospectively collected and analyzed. Results： The overall response rate （ORR） in patients with a 17p- was 56.9%, and patients with a high percentage of 17p- （defined as more than 25% of cells harbouring a 17p-） had a lower ORR. The median overall survival （OS） in patients with a 17p- was 78.0 months, which was significantly shorter than the OS in patients without this genetic abnormality （median 162.0 months, P〈0.001）. Within the subgroup with a 17p-, the progression-free survival was significantly shorter in patients at Binet stage B-C and patients with elevated lactate dehydrogenase （LDH）, B symptoms, unmutated IGHVand a high percentage of 17p-. Conclusions： These results indicated that patients with a 17p- CLL have a variable prognosis that might be predicted using simple clinical and laboratory characteristics.

Time-limited,Combined Regimen in Chronic Lymphocytic Leukemia:A Promising Strategy to Achieve a Drug Holiday

Chemoimmunotherapy(CIT)is defined as standard first line treatment for chronic lymphocytic leukemia(CLL)patients while patients with unfavorable biological characteristics such as unmutated immunoglobulin heavy chain(UM-IGHV)and TP53 aberration failed to benefit from it.The emergency of the small molecular targeted agents including Bruton’s tyrosine kinase(BTK)inhibitor(BTKi)leads to a brand-new era,from a CIT to a chemo-free era in CLL.However,the treatment of target agents is not enough to attain a deep remission and high rate of complete remission(CR),especially in patients with high risks.The long duration brought about problems,such as cost,drug resistance and toxicity.To benefit CLL in progression free survival(PFS)and long-term remission,exploration of time-limited therapies,mainly with BTKi plus CIT and BCL2i based combination therapy has become a mainstream in clinical trials.The time-limited combination therapy shed light on the promising potentiality to attain sustainable deep remission and partly overcame the risk factors,although long term follow-up is required to consolidate the conclusion.In this review,we intend to introduce key results of clinical trials with combination therapy,discuss the achievements and limitations and put forward future direction for clinical trial design in this field.

Synchronous diagnosis and treatment of acute myeloid leukemia and chronic lymphocytic leukemia:Two case reports

BACKGROUND The concurrence of acute myeloid leukemia(AML)and chronic lymphocytic leukemia(CLL)is rare.Previous reports of such cases have focused mainly on clinical diagnosis and characteristics,so the mechanism remains unclear,and therapy options have been poorly explored.CASE SUMMARY Here,we report two cases of synchronous AML and CLL.Flow cytometry revealed two distinct abnormal cell populations(myeloblasts and lymphoid cells)according to scatter characteristics.CD5-positive B cell lymphoma with myeloid leukemia invasion was observed on lymph node biopsy.Chemotherapy regimens indicated for both AML and CLL were used in our patients,and our patients achieved complete response after chemotherapy.Next-generation sequencing of 88 genes was performed.CONCLUSION We conclude that early mutation and dysregulation at the hematopoietic stem cell stage and the accumulation of multiple rearrangements may cause the concurrence of CLL and AML.The treatment of infection and combination therapy aimed at the CLL component are significant in the management of patients with concurrent CLL and AML.

Concomitant Chronic Lymphocytic Leukemia and Multiple Myeloma: Proof of Common Clonal Origin

We describe a patient with concomitant B-cell chronic lymphocytic leukemia (CLL) and multiple myeloma (MM). CLL- and MM-cell were separated by preparative flourescence-activated cell sorting (FACS). DNA sequence analysis of the complementarity-determinining region III (CDR III) of the immunoglobulin heavy chain genes showed identical gene rearrangements in the CLL- and the MM-cell population. Our findings prove a common clonal tumor origin of both B-cell diseases in this patient.

Chronic lymphocytic leukemia/small lymphocytic lymphoma complicated with skin Langerhans cell sarcoma:A case report

BACKGROUND Langerhans cell sarcoma(LCS)is a rare malignancy with poor prognosis.LCS and chronic lymphocytic leukemia(CLL)/small lymphocytic lymphoma(SLL)can occur in the same diseased tissues,such as lymph nodes or skin.CASE SUMMARY A 48-year-old female Han Chinese patient was admitted for generalized lymph node enlargement for 6 years and abdominal distension for 1 wk.She was diagnosed with small B-cell lymphoma(stage IV)/CLL(Benet stage B)and received chemotherapy.She started oral ibrutinib in February 2019.She was hospitalized on June 11,2019,and a 1.5 cm×1.5 cm dark-red nodule with ulceration scalp lesion was found.Biopsy revealed LCS but without CLL/SLL.She was diagnosed with CLL/SLL(Binet stage C,Rai stage IV)accompanied by secondary histiocytic sarcomas and skin LCS and received cyclophosphamide,doxorubicin,vincristine,dexamethasone,and etoposide but developed severe cytopenia.She ultimately refused treatments and discharged spontaneously.She died on September 12,2019.The literature review showed that in patients with CLL/SLL,skin lesions of LCS are accompanied by CLL/SLL.This patient was different from the previously reported cases of skin LCS in patients with CLL/SLL.CONCLUSION In this patient,the skin lesion of LCS showed no concomitant CLL/SLL.

Massive ascites as a presenting manifestation of chronic lymphocytic leukemia

Ascites is not an uncommon manifestation of certain solid tumors like gastrointestinal malignancies, ovarian cancer and breast cancer. However, it is unusual to encounter ascites in patients with hematological malignancies especially chronic leukemia. The patient described here presented with massive ascites and blood lymphocytosis. Further studies confirmed the diagnosis of chronic lymphocytic leukemia with ascites. The ascitic fluid was exudative, consisting of mature-looking B-lymphocytes, which were morphologically and immunophenotypically similar to peripheral blood and bone marrow cells. The patient was treated with chemotherapy and achieved a good response and diminution of ascitic fluid accumulation.

Chronic Lymphocytic Leukemia of del 17p in a Young Subject: About a Case and Reviewed a Literature

Objective: To report a case of Chronic Lymphoid Leukemia in a 28-year-old young subject, with variable clinical features and a TP53 mutation, diagnosed and followed up in the Onco-Hematology department of the HNN. Observation: 28-year-old patient, having consulted for polyadenopathy and physical asthenia, whose clinical examination found a conscious patient, submaxillary, laterocervical, axillary and inguinal lymphadenopathy, bilateral, symmetrical, painless and non-compressive whose largest measures 3 cm in diameter. Hepato-splenomegaly and epistaxis. Predominantly lymphocyte hyperleukocytosis, immunophenotyping revealed low CD19+, CD5+, CD23+, CD20 monoclonal B lymphoid proliferation. The Matutes score was 4. A karyotype showed a three-chromosome translocation;the short arm of a chromosome 2, the long arm of a chromosome 11 and the long arm of a chromosome 13, and a translocation between the long arm of a chromosome 6 and the long arm of a chromosome 18. A FISH objectified a led 17p. The diagnosis of Binet Stage C CLL with positive del 17p and complex karyotype was retained. Despite the poor prognosis, the R-C (Rituximab-Chlorambucil) protocol was instituted with once-weekly transfusions. The patient is still alive in partial clinical and biological remission. Conclusion: Despite therapeutic progress, the presence of the deletion of chromosome 17p with TP53 mutation and the young age of the patient does not change the patient’s prognosis.

Mutation Analysis of IgVH Gene in B-Cell Chronic Lymphocytic Leukemia

Summary: The variable heavy chain region (VH) genes of 3 untreated patients with B cell chronic lymphocytic leukemia (B CLL) were cloned and analyzed. The VH family used was VH3 11, VH3 72 and VH3 33. More than 2 % difference from the corresponding germline gene was detected in all the 3 obtained potential functional genes (average 16.7). Mutation pattern analysis indicated evidence of antigen selective pressure observed in 1 of 3 cases. Our findings suggested that the tumor cells originate from post GC cells.

Cytogenetic characteristics of B cell chronic lymphocytic leukemia in 275 Chinese patients by fluorescence in situ hybridization： a multicenter study

Background Under conventional cytogenetic （CC） analysis, only 30%-50% of B cell chronic lymphocytic leukemia （B-CLL） cases show clonal aberrations. Using fluorescence in situ hybridization （FISH）, the percentage of patients with abnormalities rises to almost 80%, among them, the most frequent abnormalities were 13q14, 1 lq22, p53 deletions and trisomy 12. The aim of this study was to explore the incidence of cytogenetic changes in Chinese patients with B-CLL. Methods We used FISH methods to detect the cytogenetic features in 275 cases of B-CLL from 48 hospitals. The correlation between FISH abnormalities and clinical characteristics such as age, gender, white blood cell count, peripheral hemoglobin （Hb） level, peripheral platelet count （PLT）, lactate dehydrogenase （LDH） level, Rai stage, Binet stage, and overall survival was analyzed, and the relationship between them and overall survival was also analyzed to evaluate their prognostic implications. Results Of the 275 patients, genetic aberrations were found in 77.8% using FISH. The frequencies of abnormalities were as follows： 13q deletion （56.4%）, trisomy 12 （34.5%）, p53 deletion （33.5%） and 11q22 deletion （30.5%）. It was obvious that the patients with p53 deletion had lower level of Hb （P=0.001） and PLT （P=-0.003） when compared to patients without p53 deletion. Significant differences were obtained in the distribution of p53 deletion according to Rai and Binet classification systems （P=0.016 and 0.008 respectively）. Significant differences were also observed when the overall survival was correlated with p53 deletion （P=-0.043）, Rai stage （P=0.006）, Binet stage （P=0.013）, Hb level （P=-0.004） and PLT level （P=-0.010）. Conclusions Chinese CLL patients have the similar frequencies of del（13q）, trisomy 12, del（11q） and a higher frequency of del（17p） when compared to literatures. Del（17p） is associated with advanced stage and low levels of Hb and PLT. Patients with p53 deletion, or advanced stage probably have poor survival in China.

Chronic Lymphocytic Leukemia Prognostic Index： A New Inteorated Scorino System to Predict the Time to First Treatment in Chinese Patients with Chronic Lymphocytic Leukemia

Background： The established clinical staging systems （Rai/Binet） of chronic lymphocytic leukemia （CLL） cannot accurately predict the appropriate treatment of patients in the earlier stages. In the past two decades, several prognostic factors have been identified to predict the outcome of patients with CLL, but only a few studies investigated more markers together, To predict the time to first treatment （TTFT） in patients of early stages, we evaluated the prognostic role of conventional markers as well as cytogenetic abnormalities and combined them together in a new prognostic scoring system, the CLL prognostic index （CLL-PI）. Methods： Taking advantage of a population of 406 untreated Chinese patients with CLL at early and advanced stage of disease, we identified the strongest prognostic markers of TTFT and, subsequently, in a cohort of 173 patients who had complete data for all 3 variables, we integrated the data of traditional staging system, cytogenetic aberrations, and mutational status of immunoglobulin heavy chain variable region （1GI：tV） in CLL-PI. The median follow-up time was 45 months and the end point was TTFT. Results： The median TTFT was 38 months and the 5-year overall survival was 80%. According to univariate analysis, patients of advanced Rai stages （P 〈 0.001） or with 11q- （P = 0.002）, 17p- （P 〈 0.001）, unmutated IGHV （P 〈 0.001）, negative 13q- （P = 0.007） and elevated lactate dehydrogenase levels （P = 0.001 ） tended to have a significantly shorter TTFT. And subsequently, based on multivariate Cox regression analysis, three independent factors for TTFT were identified： advanced clinical stage （P = 0.002）, 17p- （P = 0.050） and unmutated 1GHV （P = 0.049）. Applying weighted grading of these independent factors, a CLL-PI was constructed based on regression parameters, which could categorize tbur different risk groups （low risk [score 0], intermediate low [score 1], intermediate high [score 2] and high risk [score 3-6]） with significantly different TTFT （median TTFT of not reached （NR）, 65.0 months, 36.0 months and 19.0 months, respectively, P 〈 0.001 ）. Conclusions： This study developed a weighted, integrated CLL-PI prognostic system of CLL patients which combines the critical genetic prognostic markers with traditional clinical stage. This novel modified PI system could be used to discriminate among groups and may help predict the TTFT and prognosis of patients with CLL.

TOSO interacts with SYK and enhances BCR pathway activation in chronic lymphocytic leukemia

Background:TOSO,also named Fas inhibitory molecule 3(FAIM3),has recently been identified as an immunoglobulin M(IgM)Fc receptor(FcμR).Previous studies have shown that TOSO is specifically over-expressed in chronic lymphocytic leukemia(CLL).However,the functions of TOSO in CLL remain unknown.The B-cell receptor(BCR)signaling pathway has been reported to be constitutively activated in CLL.Here,we aimed to investigate the functions of TOSO in the BCR signaling pathway and the pathogenesis of CLL.Methods:We over-expressed TOSO in B-cell lymphoma cell lines(Granta-519 and Z138)by lentiviral transduction and knocked down TOSO by siRNA in primary CLL cells.The over-expression and knockdown of TOSO were confirmed at the RNA level by polymerase chain reaction and protein level by Western blotting.Co-immunoprecipitation with TOSO antibody followed by liquid chromatography coupled with tandem mass spectrometry(IP/LCMS)was used to identify TOSO interacting proteins.Western blotting was performed to detect the activation status of BCR signaling pathways as well as B-cell lymphoma 2(BCL-2).Flow cytometry was used to examine the apoptosis of TOSO-over-expressing B lymphoma cell lines and TOSO-down-regulated CLL cells via the staining of Annexin V and 7-AAD.One-way analyses of variance were used for intergroup comparisons,while independent samples t tests were used for two-sample comparisons.Results:From IP/LCMS,we identified spleen tyrosine kinase(SYK)as a crucial candidate of TOSO-interacting protein and confirmed it by co-immunoprecipitation.After stimulation with anti-IgM,TOSO over-expression increased the phosphorylation of SYK,and subsequently activated the BCR signaling pathway,which could be reversed by a SYK inhibitor.TOSO knockdown in primary CLL cells resulted in reduced SYK phosphorylation as well as attenuated BCR signaling pathway.The apoptosis rates of the Granta-519 and Z138 cells expressing TOSO were(8.46±2.90)%and(4.20±1.21)%,respectively,significantly lower than the rates of the control groups,which were(25.20±4.60)%and(19.72±1.10)%,respectively(P<0.05 for both).The apoptosis rate was reduced after knocking down TOSO in the primary CLL cells.In addition,we also found that TOSO down-regulation in primary cells from CLL patients led to decreased expression of BCL-2 as well as lower apoptosis,and vice versa in the cell line.Conclusions:TOSO might be involved in the pathogenesis of CLL by interacting with SYK,enhancing the BCR signaling pathway,and inducing apoptosis resistance.

Role of signaling pathways and miRNAs in chronic lymphocytic leukemia

Objective To summarize the recent findings of dysregulation of signaling pathways and miRNAs in chronic lymphocytic leukemia （CLL）. Data sources We searched PubMed database with the keywords ＂chronic lymphocytic leukemia＂, ＂signal pathway＂, or ＂miRNA＂ for relevant articles in recent years.

JAK2 V617F positive essential thrombocythemia developing in a patient with CD5-chronic lymphocytic leukemia

Coexistence of chronic lymphocytic leukemia （CLL） and essential thrombocythemia （ET） in a patient is extremely rare, with only 10 cases reported thus far in literature. This paper describes a 94-year-old male having atypical B-CLL with CD5- （CD5-） phenotype and ET. In this patient, we performed interphase fluorescence in situ hybridization （FISH） analysis which revealed 13q14.3 deletion in 31% of B-lymphocyte nuclei and RB1 deletion in 27% of B-lymphocyte nuclei, but not in neutrophils and T-lymphocytes. Furthermore, we identified JAK2 V617F mutation in the peripheral blood nucleated cells and neutrophils, but not in the B- and T-lymphocyte populations. Therefore, it was concluded that the occurrence of CD5- B-CLL and ET in this patient was pathogenically independent.

Overexpression of c-Myc-dependent heterogeneous nuclear ribonucleoprotein A1 promotes proliferation and inhibits apoptosis in NOTCH1-mutated chronic lymphocytic leukemia cells

Background: NOTCH1 mutation is an essential molecular biologic aberration in chronic lymphocytic leukemia (CLL). CLL patients withNOTCH1 mutation have shown an unfavorable survival and a poor response to chemoimmunotherapy. This study aims to present the mechanisms of adverse prognosis caused byNOTCH1 mutation from the perspective of the splicing factor heterogeneous nuclear ribonucleoprotein A1 (hnRNPA1).Methods: The microarray data in Gene Expression Omnibus datasets were analyzed by bioinformatics and the function of hnRNPA1 was checked by testing the proliferation and apoptosis of CLL-like cell lines. Afterward, quantitative reverse transcription-polymerase chain reaction and Western blotting were applied to explore the relationship among NOTCH1, c-Myc, and hnRNPA1.Results: RNA splicing was found to play a vital part inNOTCH1-mutated CLL cells;hence, hnRNPA1 was selected as the focus of this study. Higher expression of hnRNPA1 validated in primaryNOTCH1-mutated CLL samples could promote proliferation and inhibit apoptosis in CLL. The expression of hnRNPA1 increased when NOTCH1 signaling was activated by transfection with NOTCH1 intracellular domain (NICD)-overexpressed adenovirus vector and declined after NOTCH1 signaling was inhibited by NOTCH1-shRNA. Higher expression of c-Myc was observed in NICD-overexpressed cells and hnRNPA1 expression was downregulated after applying c-Myc inhibitor 10058-F4. Moreover, in NICD-overexpressed cells, hnRNPA1 expression decreased through c-Myc inhibition.Conclusion: Overexpression of c-Myc-dependent hnRNPA1 could promote proliferation and inhibit apoptosis inNOTCH1- mutated CLL cells, which might partly account for the poor prognosis of patients withNOTCH1 mutation.

Stationary distribution and extinction for a stochastic two-compartment model of B-cell chronic lymphocytic leukemia

In this paper,we study the dynamical behavior of a stochastic two-compartment model of B-cell chronic lymphocytic leukemia,which is perturbed by white noise.Firstly,by constructing suitable Lyapunov functions,we establish sufficient conditions for the existence of a unique ergodic stationary distribution.Then,conditions for extinction of the disease are derived.Furthermore,numerical simulations are presented for supporting the theoretical results.Our results show that large noise intensity may contribute to extinction of the disease.

THE ULTRASTRUCTURAL STUDY OF NON－HODGKIN＇S LYMPHOMA CELL，CHRONIC LYMPHOCYTIC AND HAIRY CELL LEUKEMIA

Non-Hodgkin’s lymphoma cell leukemia (NHLCL),chronic lymphocytic leukemia (CLL) and hairy cell leukemia (HLC) are the diseases very similar to each other. The differential diagnosis is very difficult,especially when there are small lymphoid cells in Periphcral blood and bone marrow under light microscope. We have observed 34 cases with electron microscope. The studies were correlated with clinical manifestation, cytology, pathology and immunologic histochemistry. Ultrastructural features strongly indicated the difference in three various diseases, although all the immunologic markers showed B-cell type.It is concluded that electron microscopic examination is of a definite significance in the diaguosis and successful treatment.

Regulation and function of angiogenic factors in chronic lymphocytic leukemia

Progression of chronic lymphocytic leukemia(CLL)is determined by the localization of malignant cells in lymphoid tissues,where they receive growth and survival signals.CLL cells produce angiogenic factors that are regulated by internal and external stimuli and whose levels vary according to the clinical stage of the disease.Stromal cellular and molecular components in CLL niches disturb the balance of pro-and antiangiogenic molecules in CLL cells and induce an angiogenic switch.Additionally,CLL cells also influence the behavior of microenvironmental cells,inducing endothelial cell proliferation and increasing the angiogenic capacity of macrophages,neutrophils,and other cells present in CLL niches.As a result of these reciprocal functional interactions,bone marrow angiogenesis is frequently increased in CLL and has been proposed as a prognostic marker in early disease.Besides their role in regulating angiogenesis,angiogenic factors are also involved in CLL cell migration and survival,all contributing to disease progression.Angiogenic factors,particularly vascular endothelial growth factor,have therefore been attractive therapeutic targets in CLL and many clinical trials were established in the past years.However,the results of these trials reveal that anti-angiogenic therapies alone are not as efficient as expected and should rather be used in combination with other treatments.

Promises and pitfalls of targeted agents in chronic lymphocytic leukemia

Targeted agents have significantly improved outcomes for patients with chronic lymphocytic leukemia,particularly high-risk subgroups for whom chemoimmunotherapy previously offered limited efficacy.Two classes of agent in particular,the Bruton tyrosine kinase inhibitors(e.g.,ibrutinib)and the B-cell lymphoma 2 inhibitor,venetoclax,induce high response rates and durable remissions in the relapsed/refractory and frontline settings.However,maturing clinical data have revealed promises and pitfalls for both agents.These drugs induce remissions and disease control in the majority of patients,often in situations where modest efficacy would be expected with traditional chemoimmunotherapy approaches.Unfortunately,in the relapsed and refractory setting,both agents appear to be associated with an inevitable risk of disease relapse and progression.Emerging patterns of resistance are being described for both agents but a common theme appears to be multiple sub-clonal drivers of disease progression.Understanding these mechanisms and developing effective and safe methods to circumvent the emergence of resistance will determine the longer-term utility of these agents to improve patients’quality and length of life.Rational drug combinations,optimised scheduling and sequencing of therapy will likely hold the key to achieving these important goals.

Construction,Expression and In Vitro Biological Behaviors of Ig scFv Fragment in Patients with Chronic B Cell Leukemia

The expression vector of SmIg scFv fragment was constructed in patient with B cell chronic lymphocyte leukemia （B-CLL） and expressed in E. coli to obtain scFv fragment, and the effect of the protein on the proliferation of stimulated peripheral blood mononuclear cells （PBMC） was investigated in vitro. Two pairs of primers were designed, and variable region genes of light chain and heavy chain were amplified by PCR respectively from the pGEM-T vectors previously constructed in our laboratory which containing light chain gene or Fd fragment of heavy chain gene. The PCR product was digested, purified and inserted into pHEN2 vector to construct the soluble expression vector pHEN2-scFv. After the induction by IPTG, the scFv protein was identified by SDS- PAGE electrophoresis and purified by Ni-NTA-Chromatography. MTT was used to determine the effect of purified protein on the proliferation of stimulated PBMC in vitro. Plasmid PCR and restriction enzyme digestion of pHEN2-scFv revealed the pHEN2-scFv vector was constructed successfully. Id-scFv protein was expressed in positive clone after induced by IPTG. SDS-PAGE analysis showed that the relative molecular weight of fusion protein was about 30 kD （1 kD= 0. 9921 ku）, which was consistent with the theoretically predicted value. Proliferation of PBMC could be induced by purified Id-scFv. It was suggested that the expression vector of SmIg scFv fragment was constructed successfully, and scFv protein was expressed and secreted from E. coil, which could induce proliferation of PBMC. This may lay an experimental foundation for further research of Id- HSP complex vaccine for B-CLL.