Efstathios Papageorgiou

The role of microRNAs in normal and malignant hematopoiesis

MicroRNAs are small non‐coding RNAs that act at the post‐transcriptional level, regulating protein expression by repressing translation or destabilizing mRNA target. Because of their discovery, microRNAs have been associated with almost every normal cell function, including proliferation, differentiation and apoptosis. Several lines of evidence suggest that they have an important role in normal hematopoiesis as exemplified by the role of mir‐155 and mir‐150 in the differentiation of B and T lymphocytes, the suppressive role of mir‐221 and mir‐222 in erythroid differentiation, the inhibitory effect of mir‐181 on hematopoietic differentiation and the induction of myeloid differentiation by mir‐223. Moreover, they play a role both as oncogenes, probably by a variety of mechanisms, namely through elimination of tumor suppressor proteins, or as tumor suppressor genes by targeting oncogenic mRNAs. Their aberrant expression has been associated with solid tumors and hematopoietic malignancies as suggested by the frequent deletion of mir‐15a and mir‐16‐1 in chronic lymphocytic leukemia, the increased levels of mir‐155 in diffuse large B‐cell lymphomas and the increased levels of mir‐181 in acute myeloid leukemia M1 and M2. The purpose of this review is to summarize current knowledge on the role of microRNAs in normal hematopoiesis and hematopoietic malignancies and, moreover, to highlight their role as potential therapeutic tools.

Thrombotic thrombocytopenic purpura/hemolytic uremic syndrome (TTP/HUS): treatment outcome, relapses, prognostic factors. A single-center experience of 48 cases.

Abstract The thrombotic thrombocytopenic purpura/hemolytic uremic syndrome (TTP/HUS) is a rare disorder characterized by microangiopathic hemolysis and thrombocytopenia. We have undertaken a retrospective analysis of the clinical characteristics, treatment outcome, and prognosis of 48 patients diagnosed and treated in our institution during a 13-year period. Among our patients 22 (46%) had fever, 35 (73%) neurological abnormalities, and 22 (46%) renal impairment at presentation of the syndrome. All patients were treated with a multimodality regimen including plasma exchange, steroids, antiplatelet agents, and IgG infusion. Of the 48 patients, 41 achieved complete remission, two had a partial response, and five had no response and died of progressive disease. Within a median follow-up period of 40 months, ten of the 41 patients who had achieved remission relapsed, most of them within the first 2 years, and nine of these responded promptly to plasma exchange therapy. Eight deaths were observed, seven of refractory disease and one in fourth relapse. The analysis of prognostic factors revealed advanced age and severe renal impairment (creatinine levels above 2 mg%) as the only parameters associated with treatment failure and poor outcome. However, none of the pretreatment characteristics proved to be of prognostic value regarding the probability of relapse. In conclusion, TTP/HUS represent a syndrome of variable clinical expression and aggressiveness. The use of a multimodality regimen in our series produced a high response rate. Nevertheless, the early identification, based on clinical characteristics, of poor-prognosis cases that probably need more or alternative forms of treatment is an issue that remains to be elucidated in prospective trials.

Myelodysplastic syndromes: Analysis of 131 cases according to the FAB classification

The clinical and haematological findings in 131 patients with myelodysplastic syndromes (MDS), none of which had previously received chemotherapy or radiotherapy, classified according to the FAB criteria, were analysed. The distribution among the 5 subgroups was: RA 31 patients, RAS 19, RAEB 23, CMML 29 and RAEBT 29 patients. There were difficulties in the classification of 24 patients. These included, first, 8 cases with myeloid hyperplasia of the bone marrow (BM) but without mono‐cytosis or excess of blasts of the BM. They were classified as RA. Second, 8 cases with sideroblastosis but with monocytosis or excess of blasts of the BM were classified 3 as RAEB, 2 as CMML and 3 as RAEBT. Finally, 8 cases with absolute monocytosis and BM blasts 15–30% were classified as CMML. 37 of 82 dead patients (45.1%) had transformed to acute non‐lymphoblastic leukaemia (ANLL). The incidence of evolution to ANLL was low for RA and RAS (6.30% and 12.5% respectively), while it was 37.5% for RAEB, 57.1% for CMML and 77.2% for RAEBT. The median survival for each subgroup was: RA 18 months; RAS 25; RAEB 13; CMML 14 and RAEBT 10 months. It is concluded that the FAB classification with some modifications recognises group of MDS with different prognosis.

Primary extranodal non-Hodgkin's lymphoma of the head and neck

Primary extranodal NHL of the head and neck (HN-NHL) accounts for 10-20% of all cases of NHL. Despite their frequency, the natural history and biological behaviour of these lymphomas is poorly understood. In this study we analysed the data 116 cases of HN-NHL. There were 65 males and 51 females with a median age of 56 years. The distribution among different anatomical sites was: tonsils 56 cases (48.3%), nasopharynx 15 (12.9%), mandible/gingiva 9 (7.8%), hard palate 7 (6%), parotis 6 (5.2%), nasal cavity 6 (5.2%), hypopharynx/larynx 6 (5.2%), thyroid 5 (4.3%), ocular adnexa 4 (3.5%), paranasal sinuses 2 (1.7%). The patients were treated with radiotherapy alone (14 cases), combined chemotherapy (52 cases) and combined modality (50 cases). According to the WF histological classification 73 cases (62.9%) had intermediate, 32 (27.6%) high and 11 (9.5%) low grade. Patients were separated in two groups: Tonsillar NHL (56 cases) and NHL of all other sites (non-tonsillar group-60 cases). A comparison between the two groups showed that there was no statistically significant difference with respect to age, sex, and histological subtypes. Also treatment response was similar (82.1% for the tonsillar vs 83.3% for the non-tonsillar). The two groups differed in stage distribution, survival and pattern of relapse. Stage I was more frequent in the non-tonsillar NHL (60%) in contrast to tonsillar NHL where stage II was more prominent (51.8%). Median survival was 86 months for the tonsillar while it has not been reached yet for the non-tonsillar patients. Patients in stage I and stage II of the non-tonsillar group had better survival compared to stages I and II of the tonsillar patients. Finally GI tract was a common site of relapse in the tonsillar group while a considerable number in CNS relapses were observed in the non-tonsillar group. We concluded that HN-NHL constitutes a heterogeneous group of patients. Tonsillar lymphomas represent a distinct group with some special clinicopathological findings.

Massive hemothorax due to intrathoracic extramedullary hematopoiesis in a patient with hereditary spherocytosis

Abstract Extramedullary hematopoiesis (EMH) is a rare disorder, characterized by the appearance of hematopoietic elements outside of the bone marrow, which occurs in patients with chronic myeloproliferative disorders or congenital hemolytic anemias. We report on a 64-year-old man with hereditary spherocytosis, who presented with anemia, jaundice, intrathoracic EMH, and massive hemothorax. The diagnosis of EMH was established after computer tomography (CT)-guided punctuation of the paravertebral mass. The patient underwent splenectomy and thoracic drainage. After 1 year, the patient is in good health, with normal hemoglobin values, and hemothorax has not recurred.

Primary Extranodal Non-Hodgkin's Lymphoma in Adults: Clinicopathological and Survival Characteristics

Among 318 cases of non-Hodgkins lymphoma (NHL) treated in our unit, 145 (45.6%) had primary extranodal NHL (PE-NHL). The stomach was the most common site (42.1%), followed by the PE-NHL of the head and neck region. Histologically aggressive histologies (65.5% intermediate and 20.7% high grade) predominated. 89.6% of the cases were localized (stage IE, 51% and stage II, 38.6%) but 28% had B symptoms. CR was achieved in 82.1% of the cases. 5-years disease free survival and overall survival were both 65%. Factors that influence prognosis were stage and high grade histology. Among various primary sites the Waldeyers ring, small intestine and testes had the worse prognosis. Compared to nodal NHL, the PE-NHL were more frequently localized, belonged more often to aggressive histologies and had more often distal extranodal relapses. CR rates and disease free and overall survival were significantly better for PE-NHL. The survival rates, however, listed according to stage and histology for nodal and PE-NHL were not different. We conclude that although PE-NHL differed from nodal NHL in several respects, prognosis is mainly a factor of stage and histology rather than of the primary localization per se.

Extensive apoptosis of bone marrow cells as evaluated by the in situ end-labelling (ISEL) technique may be the basis for ineffective haematopoiesis in patients with myelodysplastic syndromes

Abstract: Apoptosis is a gene‐directed cellular self‐destruction which begins with internucleosomal cleavage of DNA and ends eventually with fragmentation of the nucleus. We have shown that the technique of ISEL of fragmented DNA appears to be an accurate and reliable measurement of the early stages of apoptosis. The present study was undertaken in order to define the incidence of programmed cell death in bone marrow (BM) haematopoietic and stromal cells of myelodysplastic syndromes (MDS). The ISEL technique was employed in 21 BM biopsies of MDS patients. The analysis showed that in 11/21 patients, >70% cells (high score) were undergoing programmed cell death while 5 patients showed up to 1/3 of the biopsy containing apoptotic cells and 2 patients had only few occasional ISEL positive cells. Stromal cells including fat cells, endothelial cells and fibroblasts were frequently in apoptosis in large clusters. Our results indicate that extensive apoptosis of haematopoietic cells documented in BM biopsies of MDS patients may be the explanation for the ineffective haematopoiesis which is the hallmark of these disorders.

Extracorporeal photopheresis in combination with bexarotene in the treatment of mycosis fungoides and Sézary syndrome

were histologically compatible with MDE. Elastica staining showed band-like and sharply demarcated elastolysis in the upper reticular dermis with conservation of the fine vertically orientated elastic fibres of the papillary dermis. In addition, focal areas of fragmented and absent elastic fibres were observed in the mid-reticular dermis (Fig. 2c). Only a scant perivascular lymphohistiocytic infiltrate was seen in the upper dermis. On closer examination, isolated and small clusters of CD68 (PGM1)-positive multinucleated giant cells were detected in the mid-reticular dermis showing elastophagocytosis. This case documents the transition of AEGCG into a noninflammatory end stage, clinically and histologically compatible with MDE. The pathogenesis of MDE is still unknown. Exposure to natural or artificial UV radiation frequently precedes the onset of MDE suggesting a pathogenic UV-mediated immunological mechanism. However, MDE lesions are not usually confined to sun-exposed areas, so that other cofactors are likely to play a role. Extensive sun exposure was denied by our patient. An association of MDE with nicotine abuse and oral contraceptives as in our case has repeatedly been reported. It is still unclear if this is mere coincidence or if these substances represent relevant trigger factors. MDE has been described after resolution of urticaria, erythematous skin changes and lesions resembling granuloma annulare. In the majority of cases, however, MDE develops without a clinically obvious inflammatory prodromal stage. The clinical course in our patient suggests that AEGCG and MDE might represent different stages in the clinical spectrum of dermal elastolysis. After the inflammatory prodromal stage of AEGCG and phagocytosis of elastic fibres by multinucleate giant cells the noninflammatory ‘burned-out’ end stage of MDE could result in a loss of elastic fibres in the mid-dermis. As elastophagocytosis by macrophages is, to a much lesser extent, also a feature of early inflammatory MDE lesions, AEGCG might represent a severe inflammatory variant of the same entity.

A clinicopathological study of B-cell differentiation markers and transcription factors in classical Hodgkin’s lymphoma: a potential prognostic role of MUM1/IRF4

Background and Objectives Although most patients with classical Hodgkin’s lymphoma (CHL) are cured, a significant minority are refractory to treatment. The investigation of biological markers could improve the predictive capacity of clinical staging systems. The aim of our study was to detect B-cell differentiation markers and transcription factors in CHL in order to define subgroups with different histogeneses and prognoses. Design and Methods We evaluated 107 cases of CHL for BCL6, CD79a, MUM1/IRF4 and B-cell transcription factors BOB.1, OCT.2 expression by immunohistochemistry. Statistical analysis was performed using Fisher’s exact test, the Mann-Whitney test, the Kaplan-Meier method and the log rank test. Univariate and multivariate regression analyses were performed to identify variables with a significant effect on survival. Results CD79a was expressed in 5.8%, BCL6 in 14.7%, MUM1/IRF4 in 92.3%, BOB.1 in 53.4% and OCT.2 in 12.6% of cases. There was no significant association between CD79a or BCL6 expression and clinical characteristics. Univariate analysis showed that age of 45 or more, stage III and IV disease and MUM/IRF4 negative status were associated with significantly shorter time to progression (TTP) and overall survival (OS). On multivariate analysis the lack of MUM/IRF4 expression was associated with significantly shorter TTP while age of 45 or more and the presence of extralymphatic sites of disease were associated with significantly shorter OS. Interpretation and Conclusions Our study has confirmed that MUM1/IRF4 is expressed in most cases of CHL and shows that lack of this expression in a minority of cases may be a potential adverse prognostic factor.

The Novel Member of the BCL2 Gene Family, BCL2L12, Is Substantially Elevated in Chronic Lymphocytic Leukemia Patients, Supporting Its Value As a Significant Biomarker

BCL2L12 is a recently identified gene belonging to the BCL2 family, members of which are implicated in hematologic malignancies, including chronic lymphocytic leukemia (CLL). The aim of this study was to analyze the mRNA expression of the novel apoptosis-related gene BCL2L12 in patients with CLL and to examine its prognostic and predictive value and potential clinical application as a novel molecular biomarker for CLL. For this purpose, total RNA was isolated from peripheral blood of 65 CLL patients and 23 healthy donors. An ultrasensitive quantitative real-time polymerase chain reaction methodology for BCL2L12 and BCL2 mRNA quantification was developed using SYBR Green chemistry. After preparing cDNA by reverse transcription, relative quantification analysis was performed using the comparative C(T) (2(-ΔΔCT)) method. Furthermore, analysis of IGHV mutational status, CD38 expression, and detection of early apoptosis by double staining with Annexin V-FITC and propidium iodide were performed. According to our findings, BCL2L12 mRNA expression is significantly higher in CLL patients than in healthy donors. Receiver operating characteristic analysis demonstrated that BCL2L12 expression had significant discriminatory value, distinguishing very efficiently CLL patients from the non-leukemic population. Moreover, BCL2L12 expression predicts the presence of CLL, as demonstrated by both univariate and multivariate logistic regression analyses. Finally, high BCL2L12 mRNA levels are associated with advanced clinical stage and predict shorter overall survival in CLL patients.