A recombinant, fully human monoclonal antibody with antitumor activity constructed from phage-displayed antibody fragments.

A single-chain Fv antibody fragment specific for the tumor-associated Ep-CAM molecule was isolated from a semisynthetic phage display library and converted into an intact, fully human IgG1 monoclonal antibody (huMab). The purified huMab had an affinity of 5 nM and effectively mediated tumor cell killing in in vitro and in vivo assays. These experiments show that nonimmunized phage antibody display libraries can be used to obtain high-affinity, functional, and clinically applicable huMabs directed against a tumor-associated antigen.

[Three patients with differing manifestations of uterine sarcoma]. / Drie patiënten met verschillende manifestaties van uterussarcoom.

A 70-year-old woman with postmenopausal blood loss proved to have a stage-IV high-grade endometrial stromal sarcoma; 9 months after resection the patient was well. In a 53-year-old woman with symptoms of neurological deficit and weight loss accompanying an increase in abdominal girth and postmenopausal vaginal blood loss a high-grade leiomyosarcoma at stage IV was diagnosed. Despite treatment the neurological symptoms worsened and the patient died within 2 months of diagnosis. Another woman, aged 53, with abdominal pain but no blood loss proved to have a high-grade leiomyosarcoma at stage 1. Nine months after resection and radiotherapy the patient was well. The incidence of carcinomas of the uterus in The Netherlands is on average 113 women per year. They manifest themselves in different ways which can sometimes be misleading. The most common symptom is vaginal bleeding, in combination with abdominal pain or a pelvic mass. The only curative therapy is surgical excision. The 5-year survival rate is 50% in tumours confined to the uterus as opposed to 20% in those that spread further.

Multiple breakpoints within the BCL-1 locus in B-cell lymphoma: rearrangements of the cyclin D1 gene.

Centrocytic lymphoma (CC) and intermediately differentiated lymphocytic lymphoma (IDL) are B-cell non-Hodgkin's lymphomas composed of lymphocytes presumably derived from follicle mantle cells. In these lymphomas, a specific chromosomal translocation, t(11;14)(q13;q32), has been described. Previous studies suggested an association between t(11;14) chromosomal translocations and BCL-1 rearrangements. To evaluate the association between BCL-1 rearrangements and CC/IDL, Southern blot analysis was performed on a panel of 20 cases of CC/IDL, 22 cases of morphologically similar non-Hodgkin's lymphomas, 11 cases of chronic B-cell leukemias, and 2 cases of myelomas. We used various probes covering a considerable proportion of the 120-kilobase BCL-1 locus, and rearrangements in 50% of CC/IDL (10 of 20) were detected. In CC, all 4 breakpoints were located at the major translocation cluster (MTC). In contrast, in IDL, rearrangements were detected in 3 different cluster regions: 2 cases in the MTC, 2 cases with a breakpoint 24 kilobases outside the MTC, and 2 additional cases with breakpoints found 3 kilobases 5' of the first exon of the PRAD1/CCND1 gene, which is located 120 kilobases outside the MTC. In addition, one leukemia showed a breakpoint 63 kilobases outside the MTC. In all cases, there was comigration of the rearranged 11q13 fragment and the immunoglobulin heavy chain-joining gene complex, indicating a t(11;14)(q13;q32) chromosomal rearrangement. Our results show that Southern blot analysis is helpful to identify CC/IDL, but multiple breakpoints are present over a large region, and therefore, many probes are necessary to cover all breakpoints.

Mantle cell lymphoma. A morphological, immunohistochemical and molecular genetic study.

In order to identify helpful markers in the classification of mantle cell lymphoma, a morphological, immunohistochemical and molecular genetic analysis of 41 cases of NHL, originally referred to us as CC, ILL or IDL, was performed. We revised these lymphomas using the strict morphological criteria described in the updated Kiel classification and the more recently described criteria for MCL. The term MCL was used to designate the small lymphocytic B-cell NHL, previously referred to as CC or ILL/ IDL. This revision yielded 20 MCL, 8 CLL, 3 Cb/Cc, 1 CB, 6 IC and 3 MALT lymphomas. The presence of scattered histiocytes was seen in 90% of MCL and 5% of the other cases. No other morphological parameter, besides the used criteria, differentiated between MCL and similar small lymphocytic B cell lymphomas. Helpful immunohistochemical markers to distinguish MCL from similar small lymphocytic lymphomas were CD5+, CD10-, CD23- and Alkaline Phosphatase+. Large fields of dendritic reticulum cells, often in a loose and disrupted arrangement were seen in 82% of MCL and in 19% of the other lymphomas. Analysis with Southern blotting showed a rearrangement in the BCL-1 locus in 12/20 cases of MCL but not in the other 21 lymphomas. Although very specific for MCL, Southern blotting to detect BCL-1 rearrangements is, due to the large number of probes necessary, not of great help in daily practice for routine diagnostic purposes. We conclude that using strict morphological criteria, the diagnosis MCL can be made reliably and that immunophenotyping is helpful in supporting the diagnosis.

Urinary symptoms due to leukemic infiltration of the prostate. A case report.

A patient with chronic lymphocytic leukemia is presented who suffered from urinary obstructive symptoms due to leukemic infiltration of the prostate gland. He was treated with local radiotherapy, which resulted in complete relief of symptoms. Our report indicates that leukemic infiltration of the prostate should be considered in the differential diagnosis in patients with CLL presenting with obstructive symptoms of the urinary tract.

Immunohistochemical analysis of decalcified paraffin-embedded human bone marrow biopsies with emphasis on MHC class I and CD34 expression.

AIMS: To identify the stromal structures and haematopoietic cell lineages in normal bone marrow. The optimal conditions were studied for the reactivity of a panel of antibodies, applicable to paraffin sections of decalcified trephine biopsies using antigen retrieval methods. METHODS AND RESULTS: Two methods of antigen retrieval (pepsin and acid citrate buffer) were tested. For the demonstration of most antigens and for reduction of background staining, heating in acid citrate buffer was preferred. In the case of elastase and von Willebrand Factor (factor VIIIrAg) pepsin pre-treatment was optimal, whereas Ulex europaeus agglutinin (UEA-1) and alpha-smooth muscle actin (alpha-SMA) required no pretreatment. Staining patterns obtained after 48 h EDTA decalcification and short electrolytic decalcification were identical. Both methods allowed recognition of HLA-A and HLA-B antigens, isolated CD34+ cells, mono-histiocytic cells (CD68+), myeloid cells (elastase and myeloperoxidase), erythroid cells (glycophorin C) and of megakaryocytic cells (Factor VIIIrAg). A relative simple lymphocyte-subset analysis was possible in decalcified paraffin sections allowing recognition of B-cells (CD20+) and T-cells (CD3+ and CD45RO+) in frequencies comparable to frozen sections. Suitable stromal cell staining was achieved by vimentin and desmin antibodies, whereas the bone marrow capillary network was visualized by CD34, factor VIIIrAg and UEA-1. CONCLUSIONS: This immunohistochemical study indicates that all cellular components of the haematopoietic microenvironment of the bone marrow can be identified in decalcified paraffin sections using antigen retrieval methods and that the time of decalcification can be reduced to 1-1.5 h.