Diffuse osteolytic lesions in leukemic transformation of myelofibrosis

Myelofibrosis is a clonal myeloproliferative disorder characterized by splenomegaly, abnormal deposition of reticulin and collagen in the bone marrow, extramedullary hematopoiesis, dacryocytosis and leukoerythroblastic blood smear. Development and sustainment of fibrosis are mediated by complex network of several cytokines. Osteosclerosis is the most frequently observed bone change in myelofibrosis. We present an atypical case of leukemic transformation in myelofibrosis associated with diffuse osteolytic lesions and extremely elevated lactate dehydrogenase in serum, which indicates high bone turnover during leukemic infiltration and bone destruction

Reversal of FLT3 Mutational Status and Sustained Expression of NPM1 Mutation in Paired Presentation, and Relapse Samples in a Patient with Acute Myeloid Leukemia

We report a case of de novo acute myeloid leukemia (AML) with unstable FLT3 gene mutations and stable NPM1 mutation. FLT3/D835 and NPM1 (Type A) mutations were detected upon diagnosis. During the relapse, the FLT3/D835 mutation changed to an FLT3/ITD mutation while the NPM1 (Type A) mutation was retained. Cytogenetic analyses showed the normal karyotype at diagnosis and relapse. Our findings raise interesting questions about the significance of these mutations in the leukemogenic process, about their stability during the evolution of the disease, and regarding the selection of appropriate molecular markers for the monitoring of minimal residual disease

Granulomatous rosacea: Like leukemid in a patient with acute myeloid leukemia

Introduction. Skin findings in leukemias may be divided into specific lesions (leukemia cutis) and non-specific lesions (leukemids) which may be found in up to 80% of all patients with leukemias. The leukemids vary clinically and they are usually a manifestation of bone marrow or immunologic impairment, but also Sweet syndrome, pyoderma gangrenosum, erythroderma, maculopapular exanthema, prurigo-like papules, generalized pigmentation, follicular mucinosis, generalized pruritus may be found during the course of leukemia. Case report. We report a 70-year-old male with a 3-month history of erythema, papules and pustules on the face, ears and neck and over a month history of refractory anemia, anorexia, weight loss, malaise, and fever. Physical examination revealed symmetric erythematous, violaceous papules, papulo-nodules and plaques with slate scale and sparse, small pustules on the face, earlobes and neck. Histopathologic findings of involved skin showed diffuse mixed inflammatory cell infiltrate with perifollicular accentuation and focal granulomatous inflammation in the papillary and upper reticular dermis. Extensive checkup revealed the presence of acute myeloid leukemia French- American-British (FAB) classification subtype M2, with signs of three-lineage dysplasia. The patient was treated by L6 protocol which led to complete remission, both in bone marrow and skin, but after seven months he had relapse of leukemia with the fatal outcome. Conclusion. This case indicates the importance of skin eruptions in the context of hematological malignancies

Association of Bax expression and Bcl2/Bax ratio with clinical and molecular prognostic markers in chronic lymphocytic leukemia

Uvod: Rezistencija na apoptozu koja karakteriše maligne B limfocite in vivo u hroničnoj limfocitnoj leukemiji (HLL) delimično je uzrokovana unutrašnjim poremecajima apoptotske mašinerije u ovim celijama. Ti poremecaji su rezultat genetičkih promena i aberantne ekspresije regulatora procesa apoptoze, među kojima ključnu ulogu imaju članovi Bcl2 familije. Cilj: Cilj ove studije je bio da se ispita udruženost nivoa ekspresije proapoptotskog Bax gena, kao i Bcl2/Bax odnosa, sa kliničkim karakteristikama bolesnika sa HLL kao i molekularnim prognostičkim markerima, i to mutacionim statusom rearanžiranih gena za teške lance imunoglobulina (IGHV) i ekspresijom gena za lipoproteinsku lipazu (LPL). Metode: Analizirana je ekspresija Bax iRNK i Bcl2/Bax iRNK odnos u mononuklearnim celijama periferne krvi 58 bolesnika sa HLL i 10 zdravih kontrola metodom reverzne transkripcije i lančane reakcije polimeraze u realnom vremenu (qRT-PCR). Rezultati: Detektovana je povišena ekspresija Bax gena u HLL uzorcima u odnosu na kontrolne uzorke (p=0,003), kao i povišen Bcl2/Bax odnos (p= lt 0,001). Kada je u pitanju udruženost sa prognostičkim markerima, Bcl2/Bax odnos je ispoljio negativnu korelaciju sa vremenom udvostručavanja broja limfocita (r=-0,307; p=0,0451), dok je visoka ekspresija Bax bila povezana sa LPL-pozitivnim statusom (p=0,035). I ekspresija Bax gena i Bcl2/Bax odnos su bili viši kod bolesnika sa nemutiranim u odnosu na bolesnike sa mutiranim IGHV genima, ali nije dostignuta statistička značajnost. Zaključak: Rezultati ove studije ukazuju na mogucu ulogu poremecene ekspresije Bcl2 i Bax gena, koja dovodi do visokog Bcl2/Bax odnosa u leukemijskim celijama, u patogenezi i kliničkom toku HLL.Background: In chronic lymphocytic leukemia (CLL), in vivo apoptotic resistance of malignant B lymphocytes results, in part, from the intrinsic defects of their apoptotic machinery. These include genetic alterations and aberrant expression of many apoptosis regulators, among which the Bcl2 family members play a central role. Aim: The aim of this study was to investigate the association of pro-apoptotic Bax gene expression and Bcl2/Bax ratio with the clinical features of CLL patients as well as with molecular prognostic markers, namely the mutational status of rearranged immunoglobulin heavy variable (IGHV) genes and lipoprotein lipase (LPL) gene expression. Methods: We analyzed the expression of Bax mRNA and Bcl2/Bax mRNA ratio in the peripheral blood mononuclear cells of 58 unselected CLL patients and 10 healthy controls by the quantitative reverse-transcriptase polymerase chain reaction. Results: We detected significant Bax gene overexpression in CLL samples compared to non-leukemic samples (p=0.003), as well as an elevated Bcl2/Bax ratio (p= lt 0.001). Regarding the association with prognostic markers, the Bcl2/Bax ratio showed a negative correlation to lymphocyte doubling time (r=-0.307; p=0.0451), while high-level Bax expression was associated with LPL-positive status (p=0.035). Both the expression of Bax and Bcl2/Bax ratio were higher in patients with unmutated vs. mutated IGHV rearrangements, but this difference did not reach statistical significance. Conclusions: Our results suggest that dysregulated expression of Bcl2 and Bax, which leads to a high Bcl2/Bax ratio in leukemic cells, contributes to the pathogenesis and clinical course of CLL

Мијелодиспластични синдром са приближном тетраплоидијом удруженом са мутацијом гена TP53 – редак случај

Introduction. Chromosomal numerical aberrations are very common in hematological malignancies, but near-tetraploidy (80-104 chromosomes) is rare in myeloid lineage malignancies, with only a few cases reported in myelodysplastic syndrome (MDS). Due to a small number of cases with this rare cytogenetic abnormality, clinicopathological significance of near-tetraploidy in MDS is still unknown. In this case report we present a case of de novo MDS patient with near-tetraploidy in association with TP53 mutation, and we aimed to elucidate the prognostic significance of this rare genetic feature. Case Outline. In August of 2018 a 71-year-old male presented with severe anemia, thrombocytopenia, and leucopenia and enlarged spleen. Laboratory data were as follows: hemoglobin (Hb) 93 g/L, white blood cells (WBC) 2.8×109/L and platelets 23x109/L. The bone marrow aspirate was hypercellular, megakaryocytes were not found, granulocytic cells were 15% with signs of dysplasia, with 16% of blast cells without Auer rods. The finding was in correlation with diagnosis of MDS, type RAEB2 which was also confirmed by immunophenotyping. Cytogenetic finding was near-tetraploidy (48,XY+mar[10]/92,XXYY[10]), and TP53 mutational analysis showed the presence of mutation in exon 8 (p.D281A; c.842 A>C). The patient received from time-to-time packed red blood cells and platelets, and died four months after initial diagnosis. Conclusion. Near-tetraploidy associated with TP53 mutation has been described only in few MDS cases. Results of these reports including ours suggest that the association of TP53 mutation and near-tetra polyploidy is a poor prognostic factor.Нумеричке аберације хромозома су веома честе код хематолошких малигнитета, али су приближне тетраплои- дије (80–104 хромозома) ретке у малигнитетима мијелоидне лозе, са само неколико случајева пријављених у мијелоди- спластичком синдрому (МДС). Због малог броја случајева са овом ретком цитогенетском абнормалношћу, клиничко- -патолошки значај приближне тетраплоидије у МДС-у је још увек непознат. Овим приказом de novo болесника са МДС-ом, са приближном тетраплоидијом и мутацијом у гену TP53, циљ нам је био да расветлимо прогностички значај ове рет- ке генетске карактеристикe. Приказ болесника Приказан је 71-годишњи болесник који је у августу 2018. године развио симптоме тешке анемије, тромбоцитопеније, леукопеније и увећане слезине. Лабо- раторијске анализе су показале следеће: хемоглобин 93 g/L, леукоцити 2,8 × 109 /L и тромбоцити 23 × 109 /L. Аспират коштане сржи је био хиперћелијски, мегакариоцити нису на- ђени, 15% гранулоцитa је било са знацима дисплазије, 16% бластa без Ауерових штапића. Налаз је одговарао дијагнози МДС-а, типа рефракторне анемије са вишком бласта 2, што је потврђено и имунолошком фенотипизацијом. Цитогенет- ском анализом утврђено је присуство приближне тетрапло- идије (48,XY+mar10/92,XXYY[10]), а анализа мутација у гену TP53 показала је присуство мутације у егзону 8 (p.D281A; c.842 A > C). Болесник је по потреби примао трансфузију еритроцита и тромбоците, а умро је четири месеца након почетне дијагнозе. Закључак Присуство приближне тетраплоидије удружене са мутацијама у гену TP53 описано је само у неколико слу- чајева МДС-а. Резултати ових случајева, као и наши резул- тати, указују на то да приближна тетраплоидија повезана са присуством мутација у гену TP53 представља фактор лоше прогнозе

Expression of the Bcl2 gene in chronic lymphocytic leukaemia patients

Hronična limfocitna leukemija (HLL) se manifestuje kao klonska ekspanzija zrelih B limfocita, čija se akumulacija pripisuje prvenstveno poremećajima procesa apoptoze. U HLL su uočene genetičke promene i aberantna ekspresija različitih članova Bcl2 genske familije, koji imaju ključnu ulogu u regulaciji unutrašnjeg, mitohondrijskog puta aktivacije apoptoze. U ovom radu je analizirana ekspresija anti-apoptotskog Bcl2 gena u grupi od 58 pacijenata obolelih od HLL. Metodom kvantitativnog RT-PCRa detektovana je povišena ekspresija Bcl2 mRNA u HLL uzorcima u odnosu na kontrolne uzorke (p= lt 0.001). 'Receiver operating characteristic' (ROC) analiza je pokazala da nivo ekspresije Bcl2 ima visoku moć diskriminacije između pacijenata i zdravih kontrola (A=0.98, 95% CI=0.95-1.009, p lt 0.0001).Chronic lymphocytic leukaemia (CLL) manifests as clonal expansion of mature B lymphocytes, whose accumulation is primarily attributed to the dysregulation of apoptosis. Aberrant expression, as well as genetic alterations within various Bcl2 family members and central regulators of the intrinsic, mitochondriamediated apoptotic pathway all hasve been observed in CLL. Here, we report the expression analysis of the anti-apoptotic Bcl2 gene in a cohort of 58 CLL patients. Quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR) analysis revealed a significant overexpression of Bcl2 mRNA in CLL samples compared to control samples (p= lt 0.001). Receiver operating characteristic (ROC) analysis showed that the level of Bcl2 expression exerts a high discriminatory power between patients and healthy subjects (A=0.98, 95% CI=0.95-1.009, p lt 0.0001)

Acute Myeloid Leukemia Associated With Near-Tetraploid Karyotype and Mutations in the FLT3 Gene

Tetraploidy and near-tetraploidy are rare in acute myeloid leukemia (AML), contrary to other hematological disease. In this paper we describe a case of a 52-year-old male with AML associated with tetraploidy, mutation in tyrosine kinase receptor FLT3, and very short survival. At presentation maculopapular rash with crustae, lymphadenopathy, and hepatosplenomegaly was diagnosed. The blasts comprised 80% of marrow nucleated cells (POX negative and PAS finely granular positive). Immunophenotyping done on marrow cells was (CD34, HLA DR, CD14, CD64, CD33, CD11b, and CD15) and correlated with the acute monoblastic leukemia. Detection of FLT3 mutation was done by polymerase chain reaction (PCR). Cytogenetic analysis show: 85-93. XXYY,inc(cp5)/46,XY. Based on these considerations, we suggest the detection of FLT3 mutations as a diagnostic procedure for all AML patients

The role of FasR/FasL system in pathogenesis of myeloprolyferative neoplasms

Mijeloproliferativne neoplazije (MPN) su hematološki maligniteti koji se karakterišu nekontrolisanom ćelijskom proliferacijom i poremećajem u procesu apoptoze. Sistem FasR/FasL je uključen u kontrolu apoptoze u različitim tipovima ćelija. U ovom radu je izučavana uloga sistema FasR/FasL u patogenezi mijeloproliferativnih neoplazija. Upoređena je ekspresija FasR i FasL između pacijenata sa MPN (24) i zdravih kontrola korišćenjem metode 'real-time' PCR. Detektovana je povećana ekspresija FasR kod pacijenata sa MPN. Nije utvrđena razlika u ekspresiji FasL. Mutacija B617F u JAK2 genu, karakteristična za MPN, je nađena kod 13 od 24 pacijenta. Pokazano je da ekspresija FasR i FasL nije povezana sa prisustvom B617F JAK2 mutacije.Myeloproliferative neoplasms (MPN) are hematological malignancies characterized by uncontrolled cell proliferation and impaired apoptosis. The FasR/FasL system is involved in the control of apoptosis in different cell types. Here we have investigated the role of FasR/FasL in the pathogenesis of MPNs. We compared FasR/FasL expression between MPN patients (24) and healthy individuals using the real-time PCR assay. We found an increase of FasR expression in MPN patients. No difference was detected in FasL expression. Mutation V617F in the JAK2 gene, a hallmark of MPN, was detected in 13/24 patients. We found that neither FasR nor FasL expression were related to the presence of JAK2 V617F mutation

JAK2-V617F mutation in patients with myeloproliferative neoplasms: Association with FLT3-ITD mutation

Uvod. Stečena somatska mutacija V617F u genu za Janus kinazu 2 (JAK2) uzrok je nekontrolisane proliferacije ćelija kod bolesnika s mijeloproliferativnim neoplazijama. Poznato je da do proliferacije ćelija mijeloidne loze dolazi i pod uticajem mutacija u drugim genima, kao što su mutacije u genu za tirozin-kinazni receptor FLT3, koji je najčešći mutirani gen u akutnim mijeloidnim leukemijama. Od posebnog je značaja to što mutirani protein FLT3 koristi isti signalni put kao protein JAK2. To je signalni put preko proteina STAT5, čija je aktivacija važna za samoobnavljanje matičnih ćelija hematopoeze. Cilj rada. Cilj istraživanja je bio da se otkrije mutacija V617F u genu za JAK2 kod bolesnika s mijeloproliferativnim neoplazijama. Ispitano je i istovremeno prisustvo mutacije FLT3-ITD kod ovih bolesnika radi rasvetljavanja hipoteze o sličnoj ulozi ova dva molekularnogenetička markera u hematološkim malignitetima. Metode rada. Metodom alel-specifičnog PCR (engl. polymerase chain reaction) analiziran je 61 bolesnik sa potvrđenom dijagnozom mijeloproliferativne neoplazije na mutaciju V617F u genu za JAK2 ili sumnjom na ovu dijagnozu. Kod bolesnika sa mutacijom u genu JAK2 je zatim ispitano postojanje mutacije FLT3-ITD metodom PCR. Rezultati. Kod 18 ispitanika je otkrivena mutacija V617F u genu za JAK2. Među njima je kod osam bolesnika dijagnostikovana policitemija vera, a kod deset esencijalna trombocitemija. Ni kod jednog ispitanika s mutacijom V617F u genu za JAK2 nije otkrivena mutacija FLT3-ITD. Zaključak. Rezultati ovog istraživanja podržavaju hipotezu da je za malignu transformaciju matične ćelije hematopoeze dovoljna jedna mutacija koja izaziva poremećaj proliferacije ćelije.Introduction. An acquired somatic mutation V617F in Janus kinase 2 gene (JAK2) is the cause of uncontrolled proliferation in patients with myeloproliferative neoplasms. It is known that uncontrolled myeloid cell proliferation is also provoked by alteration in other genes, e.g. mutations in receptor tyrosine kinase FLT3 gene. FLT3 represents the most frequently mutated gene in acute myeloid leukaemia. Interestingly, mutated FLT3- ITD (internal tandem duplication) protein is a member of the same signalling pathway as JAK2 protein, the STAT5 signalling pathway. STAT5 activation is recognized as important for selfrenewal of haematopoetic stem cells. Objective. The aim of this study was the detection of JAK2- V617F mutation in patients with myeloproliferative neoplasms. Additionally, we investigated the presence of FLT3-ITD mutation in JAK2-V617F-positive patients in order to shed the light on the hypothesis of a similar role of these two molecular markers in haematological malignancies. Methods. Using allele-specific PCR, 61 patients with known or suspected diagnosis of myeloproliferative neoplasms were tested for the presence of JAK2-V617F mutation. Samples that were positive for JAK2 mutation were subsequently tested for the presence of FLT3-ITD mutation by PCR. Results. Eighteen of 61 analysed patients were positive for JAK2-V617F mutation. Among them, 8/18 samples were diagnosed as polycythaemia vera, and 10/18 as essential thrombocythaemia. None of JAK2-V617F-positive patient was positive for FLT3-ITD mutation. Conclusion. This study suggests that one activating mutation is sufficient for aberrant cell proliferation leading to malignant transformation of haematopoetic stem cell

Drug-induced agranulocytosis: Case series

Introduction: Agranulocytosis is a life-threatening disorder characterized by disappearance of granulocytes from peripheral blood or decrease of peripheral neutrophil count below 0.5x109 /l. It frequently occurs as idiosyncratic adverse drug reaction. It occurs especially in association with the use of antithyroid drugs, metamizole, aminopyrine and clozapine. The patients usually present with severe acute infections and sepsis. With modern management of broad-spectrum antibiotics and hematopoietic growth factors (G-CSF) the prognosis has been significantly improved. Methods and results: We present 15 patients with drug-induced agranulocytosis, discuss the criteria for diagnosis, management and prognosis. Median patients age was 36 years (ranging from 25 to 61 years). The male/female ratio was 2/13. Underlying diseases were found in 10/15 (66.6%) patients, consisting of hyperthyreosis in 7 (46.6%) patients, mental disorders in 2 patients (13.3%) and in 1 (6.6%) patient suffering from perianalfistula. Nadir median neutrophil count was 0.3x109 /l, range 0-0.5x109 /l, hemoglobin was between 104 and 128 g/L, anaemia was present in 6 (54.5%) patients, platelet count was between 170 and 230x109 /l. The treatment consisted of broad spectrum antibiotics, hematopoietic growth factor in 10, and antimycotic fluconazole in 3 patients. The outcome was favorable in 14 (93.3%) patients and 1 patient who had thyreotoxic crisis and agranulocytosis died. After 4 months, 1 (6.6%) patient, who had a complete recovery from agranulocytosis, developed an acute myeloid leukemia and died. Conclusions: Drug-induced agranulocytosis is a rare haematological complication, with stable incidence and mortality rate of 5% to 10%. The responsible drug for agranulocytosis must be identified, discontinued and permanently contraindicated