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In an attempt to characterize early B cell development including the commitment of progenitor cells to the B cell lineage, we generated and compared genomewide gene expression profiles of human hematopoietic stem cells (HSCs) and pre-B cells (PBCs) by using serial analysis of gene expression. From more than 100,000 serial analysis of gene expression tags collected from human CD34 + HSCs and CD10 + CD19 + PBCs, 42,399 unique transcripts were identified in HSCs but only 16,786 in PBCs, suggesting that more than 60% of transcripts expressed in HSCs were silenced during or after commitment to the B cell lineage. On the other hand, mRNAs of pre-B cell receptor (pre-BCR)-associated genes are virtually missing in HSCs but account for more than 10% of the transcriptome of PBCs, which also show increased expression of apoptosis-related genes. Both concentration of the transcriptional repertoire on pre-BCR-related genes together with marked up-regulation of apoptosis mediators in PBC might reflect selection for the expression of a functional pre-BCR within the bone marrow. Besides known regulator genes of early B cell development such as PAX5 , E2A , and EBF , the most abundantly expressed genes in PBCs include ATM , PDGFRA , SIAH1 , PIM2 , C/EBPB , WNT16 , and TCL1 , the role of which has not been established yet in early B cell development.

Interaction of CD95 (Apo-1/Fas) and its ligand (CD95L) plays an important role in the regulation of the immune response, since CD95+ lymphocytes may be killed after engagement of the CD95 receptor. Studying the CD95/CD95L system in 40 cases of breast cancer, the malignant cells expressed CD95L, but lost CD95 expression, when compared with non-malignant mammary tissue. Jurkat T cells incubated on breast cancer sections underwent CD95L-specific apoptosis. The rate of apoptosis correlated with the CD95L mRNA levels of the tissue samples. In four breast cancer cell lines, CD95L expression was increased by interferon-gamma (IFN-gamma), which resulted in higher levels of CD95L-specific apoptosis in co-cultured Jurkat T cells. Since IFN-gamma is mainly secreted by activated T cells, up-regulation of CD95L in breast cancer cells in response to IFN-gamma may thus counterselect activated tumour-infiltrating T cells and favour the immune escape of breast cancer. As demonstrated by inhibition of matrix metalloproteinases, CD95L expressed on breast cancer cells can also be shed from the cell membrane into the culture supernatant. Supernatants derived from cultured breast cancer cells induced apoptosis in Jurkat T cells via CD95L. In breast cancer patients, depletion of CD4+ and CD8+ peripheral blood lymphocytes was significantly correlated with CD95L expression in the tumours. This might be suggestive for a relationship between CD95L expression by breast cancer and systemic immunosuppression.

CD95 ligand expression has been observed in various malignancies. Studying the CD95 ligand (CD95L) and receptor (CD95) system in eight non-malignant mammary tissues and 40 breast cancer tissues, mRNA and protein expression was determined by quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) and immunofluorescence. mRNA levels of CD95L correlated positively ( r=0·90; p< 0·01) and transmembrane CD95 inversely ( r=−0·88; p< 0·01) with histopathological grading of the breast tumours: CD95L mRNA levels were low in adenomas, but increased by 20-fold in grade I, 120-fold in grade II, and 310-fold in grade III breast cancer. In contrast, CD95 mRNA levels were low in high-grade carcinomas, but high in benign mammary tissues. Since CD95L acts as an efficient inducer of apoptosis in CD95+ cells, apoptotic cells were identified on the tissue sections. Tumour-infiltrating lymphocytes and stromal cells in close proximity to CD95L-expressing breast cancer underwent apoptosis. As a functional test, CD95+ target cells were cultured on breast cancer tissue sections. The target cells underwent apoptosis when cultured on breast cancer sections, but could be rescued when CD95L was specifically blocked by a CD95–Fc fusion molecule. The data suggest an inverse regulation of CD95 ligand and receptor expression during dedifferentiation of breast cancer. Killing of bystander cells by the CD95L-expressing breast tumour could be involved in tissue invasion. Copyright © 1999 John Wiley & Sons, Ltd.

CD95 (Apo-1/Fas) ligand (CD95L) expression has been observed in various malignancies. In human primary cell lines from a squamous cell carcinoma (SCC) of the vulva, the effect of cisplatin (CDDP) and IFNγ on the expression of CD95L and its 2 receptor isoforms, CD95 transmembrane (CD95tm) and CD95 soluble receptor, was studied at the mRNA and protein levels. Addition of CDDP and IFNγ increased CD95L mRNA levels in the primary cell line 6-fold and 1.7-fold, respectively. In comparison, CD95tm mRNA levels were diminished by CDDP but increased 8-fold upon IFNγ challenge. CD95L expressed by SCC cells was functionally relevant since these cells were able to induce CD95-specific apoptosis in autologous lymphocytes from the SCC-bearing patient. Thus, CD95L expression in SCC may contribute to tumor-associated immunosuppression, which may be modulated by CDDP and IFNγ. In tumor samples of the primary SCC, CD95L expression was enhanced in the area of the border between invasive tumor tissue and surrounding stroma cells. The locally restricted over-expression of CD95L was congruent with the arrangement of apoptotic stroma cells in the direct vicinity of invading tumor tongues, suggesting a role as invasion factor for CD95L. Int. J. Cancer 80:564–572, 1999. © 1999 Wiley-Liss, Inc.

Resistance to CD95 (Apo-1/Fas)-mediated apoptosis is a typical feature of breast cancer cells. Recent studies identified deleterious mutations of the CD95 gene not only in a variety of B cell lymphomas but also in a number of solid tumour entities. Therefore, we amplified and sequenced selected regions (including regulatory promoter regions and the last exon coding for the death domain) of the CD95 gene from 48 breast cancer cases and 10 breast cancer cell lines but no mutation was found. In the presence of both polymorphic allele, loss of heterozygosity was excluded in 27 informative cases. We conclude that relevant somatic mutations of the CD95 gene occur, if at all, at a very low frequency and are not the primary cause for resistance to CD95-mediated apoptosis in breast cancer.

Resistance to CD95 (Apo-1/Fas)-mediated apoptosis is a typical feature of breast cancer cells. Recent studies identified deleterious mutations of the CD95 gene not only in a variety of B cell lymphomas but also in a number of solid tumor entities. Therefore, we amplified and sequenced selected regions of the CD95 gene from 48 breast cancer cases and 10 cell lines but no mutation was found. In the presence of both polymorphic alleles, loss of heterozygosity was excluded in 27 informative cases. We conclude, that relevant somatic mutations of the CD95 gene occur, if at all, at a low frequency and are not the primary cause for resistance to CD95-mediated apoptosis in breast cancer. © 2001 Wiley-Liss, Inc.