Ingrid Marschitz

Expression of Apo‐1/Fas (CD95), Bcl‐2, Bax and Bcl‐x in myeloma cell lines: relationship between responsiveness to anti‐Fas mab and p53 functional status

The down‐regulation of apoptosis may be an essential mechanism for tumour cell expansion in slowly proliferating tumours such as multiple myeloma. We studied eight myeloma cell lines for the presence of Bcl‐2, which inhibits apoptosis, of Bax, which counteracts Bcl‐2, of Bcl‐xL and Bcl‐xS, which act in an anti‐ and pro‐apoptotic fashion, respectively, and of Apo‐1/Fas, which induces programmed cell death, when activated by the Apo‐1/Fas ligand or the relevant monoclonal antibody (mab). All cell lines constitutively expressed homogenous amounts of Bcl‐2, but displayed different amounts of Bax and Bcl‐x proteins. The Apo‐1/Fas antigen could be detected in seven out of eight myeloma lines, but expression levels varied considerably. The relative expression levels of Apo‐1/Fas correlated with that of Bax, but not with that of Bcl‐2 or Bcl‐x subtypes. Furthermore, the effectiveness of the Apo‐1/Fas mab was associated with the relative expression levels of the Apo‐1/Fas and with that of the Bax antigen, but not with that of the Bcl‐2 and Bcl‐x antigens. We further showed that wild‐type p53 function is not required for Apo‐1/Fas‐induced apoptosis, nor is it necessary for the expression of Bax or Apo‐1/Fas antigens in myeloma.  In conclusion, our results suggest a p53‐independent co‐regulation of Apo‐1/Fas and Bax, as well as a role for Bax in Apo‐1/Fas‐induced apoptosis in myeloma.

The role of soluble CD23 in distinguishing stable and progressive forms of B-chronic lymphocytic leukemia.

Soluble CD23 (sCD23) has been recognized as an important prognostic parameter in patients with chronic lymphocytic leukemia (B-CLL) at early clinical stages. There is, however, no clear information on its prognostic significance in advanced stages and on its role as an indicator for aggressive or indolent courses of disease. Therefore, sCD23 was measured in the serum of 145 patients at diagnosis and serial determinations were carried out for 8 years in 38 patients. The results indicate that in patients with identical clinical stages at first presentation the disease could take different courses depending on initial sCD23 concentrations below or above specific threshold levels (860 and 5900   U/ml). sCD23 higher than these thresholds was associated with faster progression into upper clinical stages. Furthermore, sCD23-doubling time (sCD23-DT) indicated that patients with long DT progressed slowly, while those with short DT had more aggressive disease. Particularly in patients with advanced disease stages, long sCD23-DT indicated development of smoldering disease. Since sCD23 levels reflect total tumor mass, determination of sCD23-DT has probably a better predictive value than lymphocyte doubling time. It appears that B-CLL patients can be divided into different risk categories according to initial determinations of sCD23 and that sCD23-DT is an additional important parameter in predicting disease progression.

Analysis of Bcl-2 Protein Expression in Chronic Lymphocytic Leukemia A Comparison of Three Semiquantitation Techniques

A number of studies revealed that high expression of the proto-oncogene bcl-2 correlated with poor prognosis or resistance to chemotherapy in some tumors but predicted a favorable clinical course in other neoplasias. In these studies, however, different immunologic techniques for Bcl-2 detection were used, raising the question of whether the methods applied were comparable. Using chronic lymphocytic leukemia (CLL) cells, the aims of our study were as follows: (1) to determine the reproducibility of Bcl-2 semiquantitation by immunocytochemistry, flow cytometry, or immunoblotting; (2) to study the agreement between results obtained by these methods; and (3) to examine the association between Bcl-2 expression in tumor cells of 99 patients with CLL and clinical parameters. We found that determination of Bcl-2 expression by immunocytochemistry was reproducible and the results were comparable with those of flow cytometry and immunoblotting. In the patient collective examined, Bcl-2 expression did not reflect the extent of tumor mass, but higher levels were found more often in patients with progressive disease.