Prognostyczna wartość mutacji NOTCH1 i MYD88 u pacjentów z przewlekłą białaczką limfocytową

Introduction: Chronic lymphocytic leukemia (CLL) is one of the most common type of leukemia in adults with highly heterogenous clinical course of the disease. Currently available prognostic factors are not fully efficient in predicting the course of CLL. New molecular mutations like NOTCH1 and MYD88 could partly explain the CLL heterogeneity and help in identifying clinically relevant groups of patients. Material and methods: NOTCH1 c.7544_7545delCT (n=200) in PEST domain (exon 34) and MYD88 L265P (n=60) mutations was investigated by (ARMS) amplification refractory mutation system. Expression of MYD88 in CLL was assessed in (PB) peripheral blood (n=60) and (BM) bone marrow (n=92) CLL patients and 25 (HVs) healthy volunteers using qRT-PCR. Results: NOTCH1 mutation occurred in 18/200 (9.0%) CLL patients. Patients harboring NOTCH1 mutations prevalently belonged to aggressive cases, i.e. cases with an unmutated IGVH gene status, expression of CD38 and ZAP-70. MYD88 mutation occured in 2/60 (3.3%) CLL patients. MYD88 mutations were strikingly enriched among patients expressing mutated IGVH genes. Our study demonstrated significantly higher PB MYD88 expression than in HVs and relevantly higher PB MYD88 expression in comparison with BM (respectively pWstęp. Przewlekła białaczka limfocytowa (PBL) jest najczęstszą postacią białaczki u dorosłych. Chorzy z PBL stanowią heterogenną grupę kliniczną. Obecnie dostępne czynniki rokownicze nie są do końca skuteczne w prognozowaniu przebiegu PBL. Nowe mutacje molekularne, takie jak NOTCH1 i MYD88, mogą częściowo wyjaśnić heterogenność PBL i być pomocne w klasyfikacji grup chorych już przy rozpoznaniu. Materiał i metody. Mutację NOTCH1 c.7544_7545delCT (n = 200) w domenie PEST (exon 34) i MYD88 L265P (n = 60) oznaczano metodą ARMS. Ekspresję MYD88 we krwi obwodowej (n = 60) i szpiku kostnym (n = 92) pacjentów z PBL i 25 zdrowych ochotników oceniano przy użyciu metody qRT-PCR. Wyniki. Mutację w genie NOTCH1 wykryto u 18/200 (9%) pacjentów z PBL. Pacjenci z mutacją NOTCH1 należeli do grupy z bardziej agresywnym przebiegiem choroby: niezmutowanymi łańcuchami IGVH, ekspresją CD38 i ZAP-70. Mutację MYD88 wykryto u 2/60 (3.3%) pacjentów z PBL i należeli oni do grupy ze zmutowanymi genami IGVH. Mediana ekspresji MYD88 we krwi obwodowej pacjentów z PBL była istotnie wyższa niż u zdrowych ochotników, a także istotnie wyższa niż w szpiku kostnym (odpowiednio p < 0,0001 i p = 0,0015). Nie stwierdzono korelacji między ekspresją MYD88 we krwi obwodowej i szpiku kostnym a ekspresją ZAP-70, CD38 oraz stanem mutacji IGVH. Wnioski. Mutacje w genie NOTCH1 są znacznie częściej wykrywane u pacjentów z niekorzystnymi parametrami komórek białaczkowych i mogą stanowić czynnik predykcyjny gorszego rokowania u pacjentów z PBL. Niezbędne są dalsze badania nad rolą mutacji MYD88 jak czynnika prognostycznego i predykcyjnego u pacjentów z PBL

Ten years of Polish Myeloma Study Group – history and key achievements

Polish Myeloma Study Group (PMSG) was created in 2005 during XXI Congress of Polish Society of Haematology and Transfusion Medicine with the initiative of prof. Anna Dmoszynska as a forum for exchanging experiences and for clinical and scientific cooperation focused on the issue of multiple myeloma (MM). The main aims of PMSG include development of joint study projects, elaboration of unified diagnostic procedures and common treatment algorithms using new drugs. PMSG has established cooperation with international MM study group in Europe. During ten years of activity, the members of PMSG published many papers in peer-review Polish and international journals. In current article some important studies of PMSG are described

Praktyka kliniczna oceny minimalnej choroby resztkowej u chorych na szpiczaka plazmocytowego w Polsce: badanie ankietowe Polskiego Konsorcjum Szpiczakowego

Studies exploring the significance of minimal residual disease (MRD) in plasma cell myeloma (PCM) have proven its prognostic value, regardless of the type of administered treatment. In order to assess the current practice for evaluating MRD in Poland, we conducted a survey on the methods for assessing MRD and on the MRD testing time points at Polish hematological centres. Seven out of 15 institutions surveyed use of the flow cytometry (FC) method for MRD assessment. The FC-MRD assessment is performed uniformly only in those patients achieving complete remission(CR). However, the specific indications and assessment time points differed at the tested centres including: testing MRD only after autologous hematopoietic stem cell transplantation (auto-HSCT), after auto-HSCT and consolidation, after completion of first line chemotherapy or after obtaining CR in any line of treatment. The study also showed considerable heterogeneity in the FC-MRD methodology, which affects test sensitivity (from 10–3 to 10–5). None of the surveyed centres uses molecular techniques for MRD assessment. In 8 of the 15 institutions, patients are monitored by imaging techniques. Our survey may thus be useful for developing guidelines and standardization of MRD assessment in PCM in Poland.W badaniach nad znaczeniem minimalnej choroby resztkowej (MRD) w szpiczaku plazmocytowym (PCM) dowiedziono, że status MRD ma wartość prognostyczną niezależnie od zastosowanego leczenia. W celu poznania zasad monitorowania MRD u chorych na PCM w polskich ośrodkach hematologicznych przeprowadzono badanie ankietowe. W ankiecie zadano pytania dotyczące stosowanych metod wykrywania MRD oraz punktów czasowych, w których badania są wykonywane. W 7 z 15 ośrodków objętych badaniem ankietowym oznaczenia MRD w PCM wykonuje się w aspiratach szpiku kostnego metodą cytometrii przepływowej (FC). We wszystkich ośrodkach oznaczenia FC-MRD są wykonywane jedynie u chorych w całkowitej remisji (CR), jednak w różnych punktach czasowych — tylko po autologicznym przeszczepieniu krwiotwórczych komórek macierzystych (allo-HSCT), po allo-HSCT i konsolidacji, po zakończeniu leczenia pierwszej linii lub, w przypadku uzyskania CR, po dowolnej linii leczenia. Stwierdzono ponadto znaczne różnice w sposobie wykonywania badania FC-MRD wpływające na osiąganą czułość detekcji MRD (od 10–3 do 10–5). W żadnym z ankietowanych ośrodków nie ocenia się MRD w szpiku kostnym technikami molekularnymi. Monitorowanie choroby resztkowej metodami obrazowymi stosuje personel 8 z 15 ośrodków. Wyniki przeprowadzonej ankiety mogą posłużyć wypracowaniu wspólnych wytycznych i standaryzacji oceny MRD w PCM w Polsce

Peptide vaccination induces profound changes in the immune system in patients with B-cell chronic lymphocytic leukemia

Although the immune status of chronic lymphocytic leukemia (CLL) patients is mostly characterized by immunosuppression, there is an accumulation of in vivo (graft-versus-leukemia effect) and <i>in vitro</i> (spontaneous remissions after infections) data that indicates that CLL might be effectively targeted by T-cell based immunotherapy. Recently, we characterized receptor for hyaluronic acid mediated motility (RHAMM) as a preferential target for immunotherapy of CLL. We also completed a RHAMM-derived peptide vaccination phase I/II clinical trial in CLL. Here, we present a detailed immunological analysis of six CLL patients vaccinated with HLA-A2 restricted RHAMM-derived epitope R3 (ILSLELMKL). Beside effective induction of R3-specific cytotoxic T-cells, peptide vaccination caused profound changes in different T-cell subsets as well as cytokines. We present longitudinal analyses of Th17, CD8<sup>+</sup>CD103<sup>+</sup>, CD8<sup>+</sup>CD137<sup>+</sup> and IL-17 producing CD8<sup>+</sup> T cells (CD8<sup>+</sup>IL- -17<sup>+</sup>) as well as important cytokines involved in regulation of immune response such as TGF-β, IL-10, IL-2 and TNF throughout the peptide vaccination period. <i>(Folia Histochemica et Cytobiologica 2011, Vol. 49, No. 1, 161–167

Prognostic impact of NOTCH1 and MYD88 mutations in chronic lymphocytic leukemia patients

Background. Chronic lymphocytic leukemia (CLL) is one of the most common types of leukemia in adults with highly heterogeneous clinical course of the disease. Currently available prognostic factors are not fully efficient in predicting the course of CLL. New molecular mutations such as NOTCH1 and MYD88 could partly explain the CLL heterogeneity and help in identifying clinically relevant groups of patients. Material and methods. NOTCH1 c.7544_7545delCT (n = 200) in PEST domain (exon 34) and MYD88 L265P (n = 60) mutations was investigated by amplification refractory mutation system (ARMS). Expression of MYD88 in CLL was assessed in peripheral blood (PB) (n = 60) and bone marrow (BM) (n = 92) of CLL patients and 25 healthy volunteers (HVs) using qRT-PCR. Results. NOTCH1 mutation occurred in 18/200 (9.0%) CLL patients. Patients harboring NOTCH1 mutations prevalently belonged to aggressive cases, i.e. cases with an unmutated IGVH gene status, expression of CD38 and ZAP-70. MYD88 mutation occurred in 2/60 (3.3%) CLL patients. MYD88 mutations were strikingly enriched among patients expressing mutated IGVH genes. Our study demonstrated significantly higher PB MYD88 expression than in HVs and relevantly higher PB MYD88 expression in comparison with BM (respectively p < 0.0001 and p = 0.0015). There was no correlation between MYD88 expression in PB and BM and expression of ZAP-70, CD38 and IGVH mutational status. Conclusions. NOTCH1 mutations are more frequently detected in cases with unfavorable biological markers and seem to be independent predictive markers for worse outcome in CLL patients. Further collaborative studies in CLL are obligate to study the prognostic and predictive relevance of MYD88 mutations and expression

Differential Function of a Novel Population of the CD19+CD24hiCD38hi Bregs in Psoriasis and Multiple Myeloma

Psoriasis (Ps), an autoimmune disease, and multiple myeloma (MM), a blood neoplasm, are characterized by immune dysregulation resulting from the imbalance between the effector and regulatory cells, including B regulatory (Breg) lymphocytes. Peripheral blood samples from 80 Ps patients, 17 relapsed/refractory MM patients before and after daratumumab (anti-CD38 monoclonal antibody) treatment, 23 healthy volunteers (HVs), and bone marrow samples from 59 MM patients were used in the study. Bregs were determined by flow cytometry using CD19, CD24, and CD38. Intracellular production of interleukin-10 (IL-10) was assessed by flow cytometry after CD40L, LPS, and CpG stimulation. IL-10 serum or plasma concentrations were tested using ELISA method. The percentage of CD19+CD24hiCD38hi Bregs was not different whereas the production of IL-10 in Bregs was significantly higher in Ps patients in comparison with HVs. The percentage of CD19+CD24hiCD38hi Bregs in MM patients was significantly higher than in HVs (p < 0.0001). The percentage of CD19+CD24hiCD38hi Bregs was significantly higher in MM patients with the ISS stage I (p = 0.0233) while IL-10 production in Bregs was significantly higher in ISS stage III (p = 0.0165). IL-10 serum or plasma concentration was significantly higher in Ps and MM patients when compared to HVs (p < 0.0001). Following the treatment with daratumumab the percentages of CD19+CD24hiCD38hi Bregs significantly decreased (p < 0.0003). Here, in the two opposite immune conditions, despite the differences in percentages of Bregs in Ps and MM we have identified some similarities in the IL-10 producing Bregs. Effective treatment of daratumumab besides the anti-myeloma effect was accompanied by the eradication of Bregs