Olivera Tarabar

Hypermethylation of p15 Gene in Diffuse – Large B‐Cell Lymphoma: Association with Less Aggressiveness of the Disease

In this study, methylation‐specific polymerase chain reaction was used to investigate the potential prognostic significance of the methylation status of p15, p16, MGMT, and DAPK genes in 51 specimens of diffuse large B‐cell lymphoma (DLBCL). Hypermethylation of p15 gene was significantly more prevalent in patients without relapse (p = 0.001) and there was a trend toward more frequent presence of p15 methylation in patients without death outcome within 5‐year follow‐up period (p = 0.086) Also, there was a trend toward accumulation of p15 methylation with favorable clinicopathological parameters including: age ≤ 60 years (p = 0.091), normal levels of lactate dehydrogenase (p = 0.090), Eastern Cooperative Oncology Group performance status < 2 (p = 0.095), and low/intermediate low International Prognostic Index (p = 0.076). In the female group and group of the patients without bulky tumor mass, treated with chemotherapeutic regimens including rituximab, methylation of p15 was significantly related to longer overall survival (p = 0.036 and 0.027, respectively). Our results suggest that promoter methylation of p15 gene could have prognostic value in DLBCL patients treated with rituximab when used in combination with gender and tumor size.

Genetic alterations in B-cell non-Hodgkin's lymphoma

BACKGROUND Although the patients with diagnosed B-NHL are classified into the same disease stage on the basis of clinical, histopathological, and immunological parameters, they respond significantly different to the applied treatment. This points out the possibility that within the same group of lymphoma there are different diseases at molecular level. For that reason many studies deal with the detection of gene alterations in lymphomas to provide a better framework for diagnosis and treatment of these hematological malignancies. AIM To define genetic alterations in the B-NHL with highest possibilites for diagnostic purposes and molecular detection of MRD. METHODS Formalin fixed and paraffin embedded lymph node tissues from 45 patients were examined by different PCR techniques for the presence of IgH and TCR gamma gene rearrangement; K-ras and H-ras mutations; c-myc amplification and bcl-2 translocation. There were 34 cases of B-cell non-Hodgkins lymphoma (B-NHL), 5 cases of T-cell non-Hodgkins lymphoma (T-NHL) and 6 cases of chronic lymphadenitis (CL). The mononuclear cell fraction of the peripheral blood of 12 patients with B-NHL was analyzed for the presence of monoclonality at the time of diagnosis and in 3 to 6 months time intervals after an autologous bone marrow transplantation (BMT). RESULTS The monoclonality of B-lymphocytes, as evidenced by DNA fragment length homogeneity, was detected in 88 % (30/34) of B-NHL, but never in CL, T-NHL, or in normal PBL. Bcl-2 translocation was detected in 7/31 (22.6%) B-NHL specimens, c-myc amplification 9/31 (29%, all were more than doubled), K-ras mutations in 1/31 (3.23%) and H-ras mutations in 2/31 (6.45%) of the examined B-NHL samples. In the case of LC and normal PBL, however, these gene alterations were not detected. All the patients (12) with B-NHL had dominant clone of B-lymphocyte in the peripheral blood at the time of diagnosis while only in 2 of 12 patients MRD was detected 3 or 6 months after BMT. CONCLUSION Because it is quic and simple, PCR analysis of clonal IgH rearrangements is very useful when diagnostic assistance is required. This technique is also very effecient for tracking minimal residual disease in lymphomas and leukemias and for monitoring clonal evolution in acute and chronic lymphoblastic leukemias and lymphomas. The presence of other genetic alterations, which we detected, should serve as an additional prognostic or predictive factor in the patients with B-NHL.

Primary Cutaneous Diffuse Large B-Cell Lymphoma – a Case Report

Abstract In 2005, the World Health Organization - European Organization for Research and Treatment of Cancer (WHOEORTC) classified cutaneous B-cell lymphomas into 4 categories: primary cutaneous marginal zone B-cell lymphoma (PCMZL), primary cutaneous follicle center lymphoma (PCFCL), primary cutaneous diffuse large B-cell lymphoma, leg type (PCDLBCL-LT), and primary cutaneous diffuse large B-cell lymphoma, other (PCDLBCL-O). The absence of evident extra-cutaneous disease is a necessary condition for the diagnosis of primary cutaneous B-cell lymphomas, because they have a completely different clinical behavior and prognosis from their nodal counterparts. PCDLBCL-O basically represents a morphological variation, lacking the typical features of PCDLBCLLT, neither confirming the definition of PCFCCL, but on the clinical ground, its behavior seems at least to partially overlap the indolent course of PCFCCL. In fact, the present WHO lymphoma classification from 2008 overcame the previous WHO-EORTC classification, including at least a part of PCDLBCL-O within the spectrum of PCFCCL. However, owing to the rarity and heterogeneity of the PCDLBCL-O, the precise clinicopathological characteristics have not been well characterized and the optimal treatment for this group of lymphomas is yet to be defined. Nevertheless, dermatologists and pathologists should be aware of this entity in order to avoid unnecessary aggressive treatment. We present a case of a 46-year-old Caucasian male with one large round-shaped tumor and a few scattered nodules localized on the back. The histopathological features of the lesion corresponded to PCDLBCL-O. The patient follow-up showed that he was disease-free three months after surgical excision of the lesions and adjuvant local radiotherapy. No additional therapy was introduced, including chemotherapy with rituximab, cyclophosphamide, doxorubicin hydrochloride, oncovin, prednisolone (R-CHOP).

PCR-based clonality assessment in patients with lymphocytic leukaemias: a single-institution experience

PCR-based clonality testing can be performed in all lymphoproliferations by analysing gene rearrangements of antigen receptors, rearrangements that are unique for each kind of lymphocyte. Reactive lymphoproliferations have polyclonally rearranged Ig/TCR genes, whereas malignant proliferations (leukaemias and lymphomas) show clonal rearrangements. The aim of this study was to assess the clinical benefits of clonality testing with previously evaluated consensus primers in leukaemia patients. The study included peripheral blood and bone marrow samples of 67 leukaemia patients (32 B-CLL, 24 B-ALL and 11 T-ALL). Clonality testing was based on PCR amplification of rearranged IgH and TCR genes. During diagnosis, monoclonal pattern was found in all analysed B-CLL and T-ALL samples. Testing in B-ALL patients showed positive results in all bone marrow and one peripheral blood samples. Results of clonality testing in B-CLL patients during follow-up were concordant between peripheral blood and bone marrow. Obtained results corresponded to clinical course in all but one patient. In B-ALL group, results of molecular testing in peripheral blood and bone marrow confirmed remission estimated according to clinical criteria in all except one patient. Before any clinical sign of relapse, monoclonal pattern was found in six/seven patients by bone marrow and in three/seven patients by peripheral blood analysis, respectively. Results of molecular monitoring in T-ALL patients did not confirme clinical evaluation in two patients. Obtained results indicate high accuracy of re-evaluated primers for clonality assessment in ALL and CLL patients at the time of diagnosis. Results of clonality testing in B-ALL patients indicate that bone marrow analysis has higher sensitivity compared to analysis of peripheral blood.