Folliculotropic mycosis fungoides: clinicopathological analysis of 17 patients.

BACKGROUND: Folliculotropic mycosis fungoides (FMF) represents a variant of MF characterized by hair follicle invasion of mature, CD4-positive small lymphoid cells with cerebriform nuclei. The disease displays resistance to standard treatment modalities and has an unfavourable course. OBJECTIVE: Clinical analysis of 17 patients with FMF collected between 2005 and 2012, investigation of tumour cells and involved hair follicle. METHODS: Re-evaluation of clinical data, wide panel immunohistochemistry investigation on paraffin-embedded biopsy material, T-cell receptor gene rearrangement analysis of the samples. RESULTS: Male and older age group predominance, frequent head-neck involvement, acneiform lesions, keratotic plugs, cysts, nodules, follicular papules, alopecia and classic mycosis fungoides-like plaques represented the main clinical characteristics. Treatment response showed a wide range from transient complete response to therapy resistance and death due to the disease. The pathological alterations: folliculotropism, mild epidermotropism, follicular plugging, mucinous degeneration of hair follicle, basaloid hyperplasia, syringotropism were similar to those observed previously. The first case of a CD8-positive folliculotropic mycosis fungoides - with unusual clinical presentation - is reported here. Nestin overexpression of mesenchymal cells of the isthmic and suprabulbar regions of hair follicle and the reappearance of dermal nestin-expressing cells were observed in association with immature dendritic cell hyperplasia. Altered CK19 expression was detected suggesting a potential role of follicular keratinocytes in the disease process. It was found that a proportion of neoplastic T cells constantly express programmed death-1 receptor in our patients contrary to classic mycosis fungoides. CONCLUSION: The spectrum of the clinical manifestation and the course of folliculotropic mycosis fungoides are broad and differ from classic mycosis fungoides. Folliculotropic neoplastic T-cell proliferation is associated with activation of inflammatory reactive T- and B-lymphoid cells, mesenchymal cells and changes in the hair follicle.

Richter's and prolymphocytic transformation of chronic lymphocytic leukemia are associated with high mRNA expression of activation-induced cytidine deaminase and aberrant somatic hypermutation.

Chronic lymphocytic leukemia (CLL) is an indolent B-cell non-Hodgkin's lymphoma that may transform into higher-grade lymphoma. The transformation involves an increased number of prolymphocytic cells, termed prolymphocytic transformation (PLT) or the development of diffuse large B-cell lymphoma (DLBL), also referred to as Richter's transformation (RT). To analyze whether activation-induced cytidine deaminase (AID), which is essential for somatic hypermutation (SHM) of normal B-cells, and malfunction of SHM termed aberrant somatic hypermutation (ASHM) are associated with higher-grade transformation of CLL, AID mRNA expression and the mutation pattern of c-MYC, PAX-5 and RhoH genes were analyzed in eight cases of CLL without transformation and in 21 cases that showed RT or PLT. Chronic lymphocytic leukemia cases, which showed no transformation or eventually transformed into higher-grade lymphoma, showed low levels of AID mRNA expression and low frequency of mutations of c-MYC, PAX-5 and RhoH genes. In both RT and PLT, high-levels of AID mRNA expression and high-frequency mutations of c-MYC, PAX-5 and RhoH genes were detected. These results indicate that AID expression and ASHM are associated with higher-grade transformation of CLL and provide further evidences that AID expression and ASHM may be activated during the clonal history of B-cell lymphomas.

Clonal selection in the bone marrow involvement of follicular lymphoma.

To characterize the pathways of bone marrow (BM) involvement of follicular lymphoma (FL), we performed morphological and immunophenotypical analysis of tumor cells from lymph nodes (LNs) and corresponding BMs in 21 patients with FL. In three cases, genealogical trees were constructed based on the immunoglobulin variable region heavy chain (IgV(H)) gene sequences of tumor clones from LNs and BMs. Results showed that FLs within the BMs display identical or lower cytological grades than in the LNs. In the majority of cases, different proportions of tumor cells expressed bcl-2, CD10 and Ki67 in LNs and BMs. Tumor cells in the BM showed ongoing somatic hypermutation of the IgV(H) genes; the distribution of these mutations was highly consistent with antigen selection. The topology of the genealogical trees revealed that different subclones populate the LN and BM and BM infiltration may occur at different points of the clonal evolution of FL. Early descendants of the original tumor clone and derivatives of diversified tumor clones may invade the BM. These results suggest that the BM involvement of FL is associated with intensive clonal selection of tumor cells, and the BM provides a microenvironment similar to the germinal centers of LNs, where tumor cells retain their biological nature.

Prognostic significance and detection of the internal tandem duplication of the FLT3 gene in acute myeloid leukemia.

The FMS-like tyrosine kinase-3 (FLT3), which belongs to the class III receptor tyrosine kinase family, expressed by immature hematopoietic cells, plays an important role in the proliferation, differentiation and survival of stem cells. The activating mutations of FLT3 gene have been reported to be of prognostic significance. The most common somatic alteration of the FLT3 gene is the Internal Tandem Duplication (FLT3/ITD), which is caused by the elongation of the juxtamembrane (JM) domain of FLT3. The duplicated fragment size varies from 3 to more than 400 base pair, always occurs in multiples of three while the reading frame is preserved. The elongated segment of DNA can be amplified by polymerase chain reaction (PCR), and the products are separated by gel electrophoresis. The FLT3/ITD is found in 20-40% of adult AML patients and is the most frequent mutation in leukemia. Using native peripheral blood and bone marrow from AML and non-AML patients (total of 19 samples), and samples from the RNA bank (total of eight samples), the authors purpose was to work out a method for FLT3/ITD detection, which can be used in routine diagnostics. All samples produced detectable PCR products, which proofs that this procedure can be used for the detection of FLT3/ITD mutations in daily clinical practice.

Microsatellite instability and hMLH1 promoter hypermethylation in Richter's transformation of chronic lymphocytic leukemia.

Chronic lymphocytic leukemia (CLL) is an indolent B cell non-Hodgkin lymphoma (NHL) that may transform into diffuse large B cell lymphoma (DLBL). This transformation is referred to as Richter's syndrome or transformation. To analyze whether microsatellite instability (MSI) and DNA mismatch repair defects are associated with Richter's transformation, we have performed microsatellite analysis, mutational analysis of hMLH1 and hMSH2 genes and methylation status analysis of CpG island of the hMLH1 promoter on serial biopsy specimens from 19 patients with CLL. Ten cases of CLL showed no histologic alteration in the second biopsy, and nine cases of CLL underwent morphologic transformation to DLBL in the second biopsy. Using eight microsatellite loci, high level of MSI was associated with Richter's transformation in four cases of CLL, but none of the CLLs displayed this level of MSI without transformation. Mutations of the hMLH1 or hMSH2 genes were not detected in any of the lymphoma samples. In five cases of Richter's transformation the hMLH1 promoter was hypermethylated in both CLL and DLBL samples. Hypermethylation of the hMLH1 promoter associated with high-level of MSI in four cases, and low-level of MSI in one case. These results suggest that in certain cases of Richter's transformation the DNA mismatch-repair defect-initiated genetic instability may play a role in tumor progression.

Relationship between the mutational status of VH genes and pathogenesis of diffuse large B-cell lymphoma in Richter's syndrome.

Patients with chronic lymphocytic leukemia (CLL) may develop diffuse large B-cell lymphoma (DLBL), also known as Richter's syndrome. Mutational status of immunoglobulin (Ig) heavy-chain variable region (VH) genes have prognostic impact in CLL. Patients with mutated VH genes have a stable disease, whereas patients with unmutated VH gene have more aggressive disease. The mutational status of CLLs that transform to DLBL is unknown. To reveal whether Richter's syndrome occurs in CLLs with mutated or unmutated VH genes, we have performed mutational analysis on serial specimens from eight patients. CLL and DLBL tumorclones were identical in five cases and they were different in three cases. Six CLLs expressed unmutated and two cases expressed mutated VH genes. In five of the six unmutated CLLs, the DLBL clones evolved from CLL tumorclones and the VH genes expressed by DLBLs were also unmutated. In one unmutated and two mutated CLLs, the DLBLs expressed mutated VH genes, but in these three cases the DLBL tumorclones developed as independent secondary neoplasm. These results suggest that Richter's syndrome may develop in both mutated or unmutated CLLs, but clonal transformation of CLL to DLBL occur only in the unmutated subgroup of CLL.

Genetic instability is associated with histological transformation of follicle center lymphoma.

Follicle center lymphoma (FCL) is an indolent B cell non-Hodgkin's lymphoma (NHL) characterized genetically by the t(14;18) translocation. Histological transformation and clinical progression of FCLs are frequently associated with secondary genetic alterations at both nucleic acid and chromosomal levels. To determine the type and pattern of genomic instability occurring in histological transformation of FCLs and the role of DNA mismatch repair defects in this procedure, we have performed microsatellite analysis, comparative genomic hybridization (CGH) and mutational analysis of hMLH1 and hMSH2 genes on serial biopsy specimens from patients with FCL transformed to diffuse large cell lymphoma (DLCL). Paired biopsy samples of eight patients were analyzed for microsatellite instability and structural alterations for hMLH1 and hMSH2 genes, and tumor samples of five patients were subjected to CGH analysis. A high level of microsatellite instability was associated with histological transformation of two cases of FCL, but no mutations of the hMLH1 and hMSH2 genes were detected in any of the lymphoma samples. In the five cases subjected to CGH analysis, the histological transformation of FCLs was associated with genomic imbalances at 21 chromosomal regions. The genomic abnormalities found were rather heterogeneous and none of the genetic changes were overrepresented in the transformed DLCLs. These data suggest that histological transformation of FCLs to DLCL is frequently associated with genome wide instability at both nucleic acid and chromosomal levels, although mutations of the hMSH1 and hMLH2 genes are not involved in this process.

Immunoglobulin V(H) gene mutational analysis suggests that blastic variant of mantle cell lymphoma derives from different stages of B-cell maturation.

To characterise the nature of the cellular origin of the blastic variant of mantle cell lymphoma (MCL-BV), we analysed the immunoglobulin (Ig) heavy chain variable region (V(H)) genes in four cases of MCL-BV. The rearranged V(H)-D J(H) genes were PCR-amplified, cloned and sequenced. In one case, the comparison of the rearranged V(H) gene sequence to known germline V(H) gene templates showed no somatic mutations suggesting a pre-germinal centre B-cell origin for tumour cells. In the other three cases, the V(H) gene sequences showed varied number of point mutations relative to the putative germline V(H) gene sequences but the point mutations were not associated with intraclonal diversification. In one of the mutated cases, the distribution and type of the mutations indicated that tumour cells had been selected by an antigen. Since somatically mutated Ig genes are expressed by B-cells that have reached a germinal centre/post-germinal centre stage of development, these findings suggest that the MCL-BV cell of origin may also be a germinal centre or a post-germinal centre B-cell. Taken together, our findings suggest that the development of MCL-BC may not be restricted to one stage of B-cell differentiation and that they may represent transformants of B-cells at different stages of ontogeny.

Morphologic and flow cytometric analysis of circulating megakaryoblasts in chronic myeloid leukaemia.

The immunophenotype (a), ultrastructural features (b) and cell kinetics (c) of circulating megakaryoblasts have been studied in two cases of pure megakaryoblastic and one case of mixed (myeloblastic, megakaryoblastic) cell proliferation in chronic myeloid leukaemia (CML). (a) The blast cells showed early megakaryocyte differentiation antigen (HLA-DR), platelet specific GpIIIa (CD61) and GpIIb-IIIa (CD41) antigens in different percentages. (b) The megakaryoblasts were recognized by the presence of platelet GpIIIa (CD61) demonstrated by an immunoelectron microscopic method. The labelled cells were "lymphocyte-like" megakaryoblasts and cells with features of cytoplasmic maturation (demarcation membranes, alpha granules and vacuoles). (c) Cellular DNA content of the megakaryoblasts was measured by propidium iodide (PI) staining of cells expressing platelet GpIIIa (CD61). Flow cytometric (FC) DNA analysis revealed no aneuploidy and high ploidy (greater than 4N) cell population. In the two cases of pure megakaryoblastic proliferation a high percentage of the megakaryoblasts were in the S-phase, while the non-megakaryoblastic cell fraction showed no elevated S-phase compartment. It is concluded that in CML the circulating megakaryoblasts (1) have a nuclear maturation arrest and accumulation at the level of tetraploid DNA content, (2) surface antigen expression and cytoplasmic organelles show a tendency to mature and (3) in pure megakaryoblastic proliferation the myeloid cells are not in the cell compartment showing high proliferation.

Phenotypic and genotypic analyses of blastic cell population suggest that pure B-lymphoblastic leukemia may arise from myelodysplastic syndrome.

The case history of a 70-year-old man with myelodysplastic syndrome terminated into acute leukemia in 22 months is presented. The leukemic cells exhibited multifocal acid phosphatase positivity and expressed TdT, CD45, CD34 and HLA-DR but not myeloid, monocytic or megakaryocytic differentiation antigenes. The genotypic analysis revealed clonal immunoglobulin heavy chain gene rearrangement. These phenotypic and genotypic analyses of the blastic cell population suggest that myelodysplastic syndrome may transform to pure acute lymphoblastic leukemia of B-cell origin.

Different clonal origin of B-cell populations of chronic lymphocytic leukemia and large-cell lymphoma in Richter's syndrome.

Richter's syndrome is defined as the morphologic transformation of chronic lymphocytic leukemia (CLL) into diffuse large-cell lymphoma (DLL). To determine the clonal nature of the two malignancies, we microdissected the CLL and DLL cells from a lymph node of Richter's syndrome and analyzed the sequences of the rearranged Ig VH-D-JH genes of the two lymphomas. Using the Ig VH-D-JH sequence as a marker of clonality, we delineated the clonal relationship of the CLL and DLL cells. The microdissected CLL and DLL cells productively rearranged different VH, D, and JH genes, suggesting that these DLL B cells emerge as discrete elements independent of the CLL B-cell population. The productively rearranged Ig V gene sequence of the CLL clone was 100% identical to the VH6 germline gene, but the rearranged Ig VH gene of the DLL clone was somatically point-mutated based on comparison of its sequence with those of reported germline genes. In the DLL clone, the random distribution and nature of the somatic point-mutations suggests a lack of antigen selection; the identity of the somatic point-mutations in multiple independent isolates of the same B-cell clone suggests a lack of intraclonal diversity. Thus, Richter's syndrome DLL B cells are monoclonal and can emerge as discrete elements independent of the preexisting CLL cells; antigen selection and clonal diversification are not necessarily associated with the events leading to this aggressive neoplastic transformation.

High-grade transformation of low-grade non-Hodgkin's lymphomas: mechanisms of tumor progression.

In the natural history of low-grade non-Hodgkin's lymphomas (NHL) a prolonged indolent phase of the disease may be followed by clinical progression toward intermediate and high-grade disease. The abrupt appearance of diffuse large cell lymphoma (DLL) in patients with low-grade NHL is usually associated with an accelerated clinical course and shorter time of survival. The histologic transformation has been described for chronic lymphocytic leukemia or small lymphocytic lymphoma (CLL/SLL), follicular lymphoma (FL), mantle cell lymphoma (MCL) and lymphoma of mucosa-associated lymphoid tissue (MALT). Although the histological transformation of low-grade lymphomas are relatively frequent, the clonal relationship between the two neoplasms and pathogenetic mechanisms underlying the progression of the disease are widely debated. In this review, we will focus on the possible relationship between the low-grade and the transformed high-grade NHLs and genetic lesions that may be associated with the histologic transformation and clinical progression of the disease.

Primary effusional lymphoma: A new non-Hodgkin s lymphoma entity.

A distinct non-Hodgkin s lymphoma (NHL) entity that grow in the body cavities as lymphomatous effusions in the absence of clinically identifiable tumor masses has been defined as primary effusional lymphoma (PEL). This lymphoma characterized by distinetive morphology, immunophenotype, genotype and association with Kaposi s sarcoma-associated herpesvirus (KHSV)/human herpesvirus-8 (HHV-8) infection. In this minireview, the clinico-pathological and biological characteristics of PELs are summarized.

In vivo effects of CHOP protocol on onco and suppressor gene expression: follow-up study.

In vivo investigation of onco/suppressor gene effects may provide new findings concerning chemical carcinogenesis. In earlier studies we pointed out the carcinogenic potential of COPP and ABVD chemotherapeutical protocols in "long-term" experiments. In another follow up study we proved the connection between the early gene expression changes and the late consequences of COPP and ABVD treatment during, a one year latency period. CHOP protocol is containing both proved carcinogenic cyclophosphamide and highly mutagenic doxorubicyn. CHOP protocol in "short-term" experiments shows strong effect on Ha-ras oncogene expression.

[The role of genetic studies in the diagnosis and classification of non-Hodgkin lymphoma]. / Molekuláris genetikai vizsgálatok jelentösége a non-Hodgkin-lymphomák diagnostikájában és osztályozásában.

The revised European-American Classification of Lymphoid Neoplasms (REAL), proposed by the International Lymphoma Study Group, contains molecular genetic data which may characterize a given lymphoma entity. In this recent review, the molecular genetic alterations integrated into the diagnostic procedure of lymphomas are discussed. Based on our recent knowledge the rearrangements of bcl-1, bcl-2, bcl-6, anaplastic lymphoma kinase and c-myc genes are associated with mantle cell, diffuse large B-cell, anaplastic large cell and Burkitt's lymphomas, respectively. The integration of the relevant molecular data in the evaluation of objective diagnoses and therefore specific and successful therapies. Furthermore, the knowledge of the molecular event associated with lymphomas helps to better understand tumor development and biology.

[Diagnosis of chronic myeloproliferative diseases based on bone marrow biopsy]. / Krónikus myeloproliferatív betegségek diagnózisa csontvelöbiopsziás minták alapján.

The bone marrow biopsies and laboratory data of 248 patients with untreated chronic myeloproliferative disorders have been evaluated according to the Hannover Classification. 47.6 per cent of the cases were classed as primary or basic diseases, including chronic myeloid leukaemia common type, chronic myeloid leukaemia megakaryocytic type, chronic megakaryocytic-granulocytic myelosis, polycythaemia vera and essential thrombocythaemia. In 52.4 per cent of the biopsies the advanced stages of the primary diseases like increase of fibers and loss of differentiation were noted; the increase of fibers in myeloid leukaemia megakaryocytic type and in chronic megakaryocytic-granulocytic myelosis, and loss of differentiation in chronic myeloid leukaemia common type were frequently noted. In 14.1 per cent of the cases the advanced myelofibrosis and blast cell accumulations obscured the histological features of the primary disease, therefore these cases were placed in the unclassifiable group. The cases without increase of fibers and loss of differentiation accounted for only 4 per cent of the unclassifiable category. Leucocyte and platelet count as well as haematocrit values showed considerable overlapping and scattering and were generally lower in cases which developed myelofibrosis.

[Clonality analysis of B-cell lymphoproliferative disorders by means of immunoglobulin heavy chain polymerase reaction]. / B-sejtes lymphoproliferativ kórképek klonalitásának elemzése immunglobulin nehézlánc polimeráz láncreakció segítségével.

The majority of B-cell non-Hodgkin's lymphomas (NHL) exhibit a highly specific immunoglobulin heavy chain (IgH) gene rearrangement as a result of sequential assembly of their Ig variable (VH), diversity (D) and joining (JH) region segments. The analyses of Ig gene rearrangements in B cells may help to differentiate reactive lymphoproliferations from NHLs, and to identify of their B-cell origin. The aim of this study was to reveal the usefulness of polymerase chain reaction analysis of the Ig gene rearrangement in the diagnosis of B-cell NHLs, using native and formol-paraffin embedded samples. The authors analysed 67 biopsy samples of immunohistochemically characterized lymph nodes diagnosed at the Department of Pathology. University Medical School of Pécs, between 1993 and 1995, using IgH gene polymerase chain reaction. The 67 samples included 10 reactive lymphoproliferations, 47 B-cell, 5 T-cell NHLs and 5 Hodgkin's diseases. In 54 cases, fresh, unfixed, in 13 cases, formalin-fixed samples have been used. The polymerase chain reaction amplification of the Ig heavy chain third complementary determining region (CDR 3) was performed by IgVH specific sense and JH specific antisense primer pairs. The polymerase chain reaction products were evaluated by agarose gel electrophoresis containing ethidium bromide. Sixty-four % of fresh, unfixed and 54% of formol-paraffin fixed B-cell NHLs samples showed clonal Ig gene rearrangement. The applied polymerase chain reaction technique did not show clonal amplification in reactive lymphoproliferations, T-cell NHLs or Hodgkin's disease. The polymerase chain reaction amplification of the IgH gene can be a powerful tool in the diagnosis of monoclonal B-cell lymphoproliferative disorders.

[Incidence of internal carotid artery thrombosis and disseminated intravascular coagulopathy in the early stages of acute myeloid leukemia]. / Arteria carotis interna thrombosis és disseminált intravascularis coagulopathia (DIC) elöfordulása akut myeloid leukaemia kezdetén.

The case history of a 15 year old boy in whom thrombosis of the internal carotid artery was associated with severe disseminated intravascular thrombosis (DIC) is described. Both peripheral blood smear and bone marrow aspirate revealed acute myelogenous leukemia FAB M-2 type as the cause of the disease. Consumption coagulopathy is common sign of hemostasis disturbances in leukemia. It is frequently observed in acute promyelocytic leukemia, but rarely it may be seen in the other forms of hemoblastosis, too.

[Detection of t(14;18) chromosome translocation in follicular lymphoma by polymerase chain reaction]. / A t(14;18) kromoszóma transzlokáció kimutatása follicularis lymphomában polimeráz láncreakció segítségével.

In most cases of centroblastic/centrocytic follicular lymphomas the bcl-2 proto-oncogene (18q21) is translocated to the immunoglobulin JH region of chromosome 14 (14q32). About three quarters of the translocations are concentrated on the 3' nontranslated, a few hundred basepare-long region of bcl-2, the so called major breakpoint region (mbr), the remaining 20-25% is located about 30 kilobases downstream of bcl-2 coding sequences in the minor cluster region (mcr). The majority of the immunoglobulin breakpoints can be found in JH6-4 genes. The polymerase chain reaction method can detect the translocation already in a very few number of cells (> 10(3)). This very sensitive technique makes it possible to detect the translocation in lymphoid/lymphoma of peripheral blood and bone marrow that are missed by other diagnostic methods. This way one can perform a quick, early diagnosis, examine the result of treatments as well as detect the remissions and the possible relapses right at the beginning. All the advantages of this method contribute to a more successful treatment of follicular lymphoma. This present work describes a polymerase chain reaction technique which is capable of a detection of the t(14;18) translocation in a patient of centroblastic/centrocytic lymphoma, moreover shows how this translocation disappears after 4 week of radiotherapy of the patient.