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Development of the target therapies of lung cancer was a rapid process which fundamentally changed the pathological diagnosis as well. Furthermore, molecular pathology became essential part of the routine diagnostics of lung cancer. These changes generated several practical problems and in underdeveloped countries or in those with reimbursement problems have been combined with further challenges. The central and eastern region of Europe are characterized by similar problems in this respect which promoted the foundation of NSCLC Working Group to provide up to date protocols or guidelines. This present paper is a summary of the molecular pathology and target therapy guidelines written with the notion that it has to be upgraded continuously according to the development of the field.

Primary genetic abnormalities of leukemia cells have important prognostic significance in childhood acute leukemia. In the last two years 30 newly diagnosed or recurrent childhood ALL bone marrow samples were analyzed for chromosomal abnormalities with conventional G-banding and interphase-fluorescence in situ hybridization (I-FISH) using probes to detect BCR/ABL fusions, cryptic TEL/AML1 and MLL rearrangements and p16(9p21) tumor suppressor gene deletions. G-banded karyotype analysis found clonal chromosomal aberrations in 50% of cases. With the use of complementary I-FISH techniques, ALL-specific structural and numerical changes could be identified in 70% of the patients. Nine cases (30%) had subtle chromosomal aberrations with prognostic importance that had not been detected in G-banded analysis. Conventional G-banding yielded additional information (rare and complex structural aberrations) in 19% of patients. The most common aberration (30%) was AML1 copy number increase present in G-banded hyperdiploid karyotype as a chromosome 21 tetrasomy in the majority of cases; one case displayed 5-6 copies and in another case amplification of AML1 gene on der(21) was combined with TEL/AML1 fusion of the homologue AML1 gene and deletion of the remaining TEL allele. High hiperdiploidy was detected in 6 cases, in one patient with normal G-banding karyotype. TEL/AML1 fusion signals were identified in four patients. Deletion of p16 locus was found in eight cases (23%), of which only two had cytogenetically visible rearrangements. G-banding in combination with I-FISH has produced major improvements in the sensitivity and accuracy of cytogenetic analysis of ALL patients and this method helps to achieve a more precise identification of different risk categories in order to choose the optimal treatment.

Introduction: Many new prognostic factors established in recent years in chronic lymphocytic leukemia. May help predicting survival., Aims: The goal of the present study was to determine the frequency and the correlation of these novel prognostic factors in samples of 419 leukemia patients., Methods: The mutation status of the IgH gene was evaluated in 160 cases., Results: In 62% of cases, non-mutated IgH gene was found, the heavy chain family usage was different in mutated and non-mutated cases. The CD38 expression demonstrated 78% concordance with the mutation status, the ZAP-70 expression failed to show any correlation. Cytogenetic abnormalities were seen in 76% of cases, the most frequent were del(13q) (57%), trisomy 12 (15%), del(11q) (12%) and del(17p) (6%). 95% of cases with del(11q) harbored non-mutated, 74% of cases with del(13q) as the sole anomaly demonstrated mutated IgH genes., Conclusions: The parameters analysed are not independent of each other, utilization of them in the clinical routine needs careful planning.

Cytogenetic syndrome involving bands 3q21 and 3q26, known as "3q21q26 syndrome" has been observed in adult patients with acute myelogenous leukemia (0.5-2%), chronic myelogenous leukemia in blast crisis (20%), myelodysplastic syndromes and myeloproliferative disorders. In the present study bone marrow samples from two boys (12 and 16 years), diagnosed with CML and AML respectively, were investigated using conventional cytogenetic methods, interphase "multipoint" fluorescence in situ hybridization (FISH), dual color-FISH and multiplex FISH. The "multipoint" FISH analysis identified in de novo childhood AML case an inv(3)(q21q26) and a complex 3q rearrangement including inversion and duplication in the CML case. The "3q21q26 syndrome" is associated with normal or elevated platelet counts with marked abnormalities of megakaryocytopoiesis, involvement of multiple hematopoietic lineages. The affected patients were resistant to conventional chemotherapy and had a short survival. This syndrome is very rare in de novo childhood AML, and simultaneous presence of 3q inversion and duplication, to our knowledge, has not yet been identified in hematological malignancies. The results of our study emphasize the importance of classical and modern cytogenetic analysis in the diagnosis of hematologic malignancies, because in the majority of cases they can provide additional diagnostic information for the clinicians in deciding the best therapeutic approach, precise classification and prognosis of the disease.

Introduction: In the Western world the second most common type of non-Hodgkin's lymphomas is follicular lymphoma (FL) comprising 30-35% of all cases. According to the data of the National Cancer Registry and our institute, this ratio is lower in Hungary and is about 15-20%, but the occurrence shows an increasing tendency., Aims: Our aim was to survey and revise FLs that had been diagnosed at the National Institute of Oncology between 1990-1995. We studied the diagnostic relevance of histology, immunohistochemistry and the detection of immunoglobulin heavy chain (IgH) and bcl-2 gene rearrangements., Materials and Methods: We surveyed 53 cases that were previously diagnosed as follicular or centrocytic, centroblastic lymphoma. Following histological re-examination, immunohistochemistry (CD20, CD3, bcl-2, CD10, bcl-6, CD5, p53, cyclin D1 and Ki-67) was performed on each case. We also studied the IgH and bcl-2 (major breakpoint region=MBR) gene rearrangement on paraffin embedded samples with conventional PCR methods. The classification was made according to the new WHO classification., Results: After the revision of the 53 cases we found 37 follicular, 11 diffuse large B-cell, 1 mantle cell and 1 marginal zone lymphomas, 1 nodular lymphocyte predominant Hodgkin's lymphoma and 2 follicular hyperplasias. The grade of the FLs correlated with the expression of different antigens. CD10, bcl-2 expression and the proliferation index with Ki-67 showed good correlation with the grade of FLs. We could detect bcl-2 gene rearrangement in 55% of the FLs., Conclusion: Considering the diagnostic relevance of the different methods we can conclude that histology alone is not sufficient to make a correct diagnosis. Ninety percent of our cases were solvable with the help of immunohistochemistry and in 10% of the cases the diagnosis was partly based on the molecular pathological results.

Diagnosis of cutaneous T-cell lymphomas, especially of mycosis fungoides is often difficult in the early stage of the disease, as well as their differentiation from other reactive inflammatory dermatoses. The diagnostic exactness could be improved by the simultaneous histological, immunophenotypic and gene rearrangement analysis of the skin biopsy specimen, and these results together with the clinical findings could help to establish the early diagnosis. Using this complex analysis effective treatment can be started in time and repeated skin biopsies are not always necessary. Authors performed this complex investigation in skin biopsy specimens and peripheral blood of 49 patients with cutaneous T-cell lymphoma and other T-cell lymphoproliferative diseases and estimated the diagnostic value of the T-cell receptor gamma gene rearrangement analysis. Based on the results of the clinical, histological, immunophenotypic and gene rearrangement studies authors could made a clear distinction between the benign dermatoses (19 cases) and malignant cutaneous lymphomas (17 cases). However less than one third of the patients (13 cases) remained unclassified, so definite diagnosis of these cases could be based on the long term follow up and on the results of the repeated skin biopsies.

On the basis of data of the Human Genome Project, it was embraced the newest information about the gene content of the human genome, the disease genes, the parasitic DNA, the single nucleotide polymorphisms, the repeat sequences, the cytoskeletons, the regulation of cell proliferation, and their medical consequences. The applicability of the acquaintance with the human genome in pathology is presented with a few examples of our own. The significance of the single nucleotide polymorphisms in susceptibility to, or protection from, a host of disease is illustrated by the example of the allele variation of Apo-E gene. The copy number of the N-myc gene in neuroblastomas and HER2/neu gene in breast carcinomas was determined with quantitative PCR techniques. The monoclonally increased abnormal p53 protein expression was found in small cell lung cancer (in 90% frequency), in oro-pharyngeal carcinomas (82%), in esophageal squamous cell carcinomas (59%) in stomach cancer (33%), in colon carcinomas (27%) and in soft tissue sarcomas (13%). These data advert to the fact that the mutation of the p53 gene is much more frequent in those tumors in which the basic tissue is directly exposed to with the environmental carcinogens. It is now known, that near the repetitive sequences, gene rearrangement can more easily be evolve. Finally, we have determined the conditions of the accomplishment of the molecular pathological diagnosis: (1) It is applicable, when the classic morphology does not eventuate a conclusive result. (2) Well known and validated gene alterations are admissible to diagnostic purpose. (3) Only standard methods are applicable along with positive and negative controls. (4) The result has to correlate with the morphological picture, the immunohistochemical profile and the clinical data. (5) It is necessary to be able to appropriately interpret the molecular biological result, which is then incorporated in the pathological report. (6) The ethical, legal and social consequences must be considered.

Follicular lymphoma is one of the most common indolent nodal non-Hodgkin's lymphomas. The follicular lymphoma should be differentiated from other nodal non-germinal centre derived lymphomas. Since bcl-2 gene translocation can be detected in 80-90% of the cases, it could be used for the confirmation of the diagnosis, measure of the treatment efficacy as well as for the evaluation of the prognosis and follow-up of the disease. There is no uniform therapy employed for follicular lymphoma patients. The treatment depends on the histological grade, stage, age and condition of the patients. Since disease cannot be cured by conventional dosage therapy in the vast majority of the cases, a treatment--adjusted to the condition of the patients as well as the stage and grade of the disease--ensuring the longest period and best quality of life should be selected. The authors review the questions of diagnosis and treatment, including the results of radiotherapy and chemotherapy and discuss the role of the interferon-alpha, purine analogues, monoclonal antibodies and high-dose therapy.

Introduction: Anaplastic large cell lymphoma belongs to a relatively good prognostic group of high grade non-Hodgkin lymphomas. Molecular genetic methods can provide great help in the differential diagnosis of the disease., Aims: Sixteen frozen lymph node samples of CD30 positive lymphomas with anaplastic morphology and 1 bone marrow aspiration were analysed with polymerase chain reaction according to T-cell receptor gene and immunglobulin heavy chain gene rearrangement. T(2;5)(p23;q35), t(1;2)(q25;p23) chromosomal translocation was also investigated with reverse transcriptase polymerase chain reaction., Results: Seven cases showed null-, 3 T-, 4 B- and 2 T/B hibrid genotype. Two cases of t(2;5), whereas no t(1;2) translocation was demonstrated., Conclusions: Molecular genetic results correlated with the results of immunhistochemistry and can give further help to divide the cases into smaller, new prognostic subgroups.

Unlabelled: Hepatitis C virus (HCV) has cytopathogenic effect not only on hepatocytes, however on salivary glands, monocytes of peripheral blood and lymphoid cells, may explain the systemic manifestations of the infection. HCV activates B and T-cells, modifies the immune response, causes lymphoproliferation, leading the development of B-cell non-Hodgkin's lymphoma (NHL). In the majority of B-cell NHLs immunoglobulin heavy chain (IgH) and light chain (IgL) genes are rearranged and expressed on cell surface in the early stage of the ontogenesis. The analyses of IgH rearrangement prove the clonality of lymphoproliferative disorders giving a powerful approach to the B-cell origin identification of cell proliferation., Patients and Methods: IgH gene rearrangements were examined from the sera of 57 chronic HCV infected patients and 11 HCV-positive cryoglobulinemic patients as well., Results: Cryoglobulinemia was detected in 20% of all chronic hepatitis C virus infected patients and IgH rearrangement was observed in 10.29% (7/68), 4/7 patients (57.14%) suffered from cryoglobulinemia., Conclusions: These results support the hypothesis that IgH gene rearrangement in HCV positive patients can indicate the lymphoproliferative disorder in the HCV infection released B-cell proliferation and lymphoma development.

Diagnosis of cutaneous lymphoma is based on the clinical picture and the histology. Several, different clinical entities are the members of this group with various clinical and histological features, with different course and prognosis. The clinical picture resembles that of a wide variety of benign, inflammatory dermatosis. The histological characteristics in the early lesions are not pathognomic, therefore immunohistochemistry and gene rearrangement studies are necessary to establish the diagnosis. These processes are evaluated in the article.

Despite B-cell counts and serum immunoglobulin levels usually normalized by one year, humoral immunity and the incidence of bacterial infections continue to be abnormal even after years following bone marrow transplantation. This immunodeficiency could be partially caused by B-cell repertoire restriction similar to that observed early in ontogeny. Immune reconstitution after haematopoietic stem cell transplantation really follows many established ontogenetic patterns relating to the appearance of particular membrane markers, immunoglobulin classes and subclasses, and onset of antigen receptor rearrangements. The sequence of events that occur during successful bone marrow transplantation can be regarded as a blueprint for immune reconstitution in other clinical settings as well. However, the repertoire does not resemble a fetal one, because it displays adult-size IgH CDR3s, adult-type immunoglobulin gene utilization and no evidence of bias towards any particular VH-gen family. Therefore, in the description and interpretation of these events, it is important to realize that immune reconstitution does not appear to recapitulate human fetal ontogeny. In terms of B lymphocyte diversity, the inadequacy of the recovering immune system is more likely to be explained by a combination of other factors--such as clonal dominance and the delayed occurrence of somatic hypermutation.

Results of morphological, cytochemical and immunological studies performed in adult acute leukaemias have been compared. Thirty one cases proved to be acute myeloid leukaemia, while 25 cases were shown to be acute lymphoid leukaemia. Based on our results we conclude that immunophenotyping with monoclonal antibodies does not help in distinguishing the subtypes of AML. For purposes of clinical diagnosis cytochemical methods are valuable. On the other hand the monoclonal antibodies are essential in distinguishing the very immature myeloid and lymphoid leukaemias and this is of great importance from the clinical point of view, in determining therapy. Moreover, the diagnosis of acute lymphoid leukaemias is not possible without the specific monoclonal antibodies. Their application is first of all in haematological centers caring for leukaemia patients nowadays already obligatory. Gene rearrangement studies make the diagnosis more accurate and help in the diagnosis of leukaemias of unknown immunological origin.

'Essential' cryoglobulinemias (EC) are characterized by the precipitation of immunoglobulins (Igs) from the sera of patients at temperatures below 25 degrees C. The cryoprecipitate of type I EC patients is comprised of a monoclonal Ig while that of patients with type II or 'mixed' EC contains monoclonal auto-antibodies with rheumatoid factor activity. In order to define if the high cryoglobulin production rate is related to a clonal B-cell expansion the rearrangement of Ig genes was investigated by Southern blot analysis of DNA extracted from peripheral blood lymphocytes of EC patients. Clonal expansion of B cells could be detected using Ig light and heavy chain specific DNA probes in 3/4 patients with type I EC and 4/12 patients with type II EC. In the group of patients with clonal Ig gene rearrangements, in two cases with type I EC and one case with type II EC, alterations of the c-myc locus was also noted. Demonstration of clonal B-cell expansions in EC patients shows that the clonal type of Ig gene rearrangements is not a unique marker of malignant lymphomas. Since malignant B-cell lymphomas can develop in a small number of EC cases, the follow-up of these patients should be pursued indefinitely.

In this report we summarize our experiences based on the gene rearrangement study of 111 leukaemic patients of different kind. The lymphocyte DNA of the patients was studied for rearrangement of the immunoglobulin light chain constant-, the heavy chain joining- and the T cell receptor beta chain constant region. Our data have well supplemented the results of the monoclonal antibody experiments. In 33 cases the DNA study was in good agreement with the immunological data. In 42 our data helped in gave different results, immunological results. In 11 cases evaluating the DNA and immunological data indicating the necessity of further investigation. The results were inconclusive in 25 cases. As a conclusion we consider the gene rearrangement study to be useful for diagnostic purposes.