Ursula Koldovsky

Regulation of CD95 (APO‐1/ FAS) ligand and receptor expression in squamous‐cell carcinoma by interferon‐γ and cisplatin

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.

Investigations by cell-mediated immunologic tests and therapeutic trials with thymopentin in vaginal mycoses

Objective: According to unsatisfactory therapeutic results in patients with chronically recurrent vaginal candidosis, we investigated if immunologic patient factors could be found and treated. Methods: In 42 women with chronically recurrent and 20 women with acute Candida albicans vulvovaginitis, as well as 14 women with C. glabrata vaginitis, the following investigations were carried out: identification of yeast species; quantification of T lymphocytes and their subpopulations in sera; proliferation tests of T lymphocytes in vitro; treatment of 18 patients with chronically recurrent vaginal candidosis with the synthetic T-lymphocyte- stimulator thymopentin; and, finally, control of the above-mentioned parameters in the clinical course. Results: Women with C. albicans vulvovaginitis showed fewer T lymphocytes and subpopulations in the peripheral blood than healthy women. Only the number of non-specific killer (NK) cells, however, was significantly lower in cases of acute C. albicans vulvovaginitis. In women with C. glabrata vaginitis, the number of T lymphocytes in the blood was within the normal range. In vitro proliferation tests using mitogens, bacterial antigens, and commercially available candida antigens with and without addition of thymopentin were carried out on the T lymphocytes of women with chronically recurrent C. albicans vulvovaginitis. These tests revealed no significant differences compared with the other patients with C. albicans infections. The patients were treated with thymopentin. Those women who revealed an increase of initially low numbers of T-helper cells recovered from vaginal candidosis after thymopentin treatment. Conclusions: The peripheral T lymphocytes may be diminished in patients with chronically recurrent C. albicans vaginitis, and immunologic treatment can reduce the relapse rate.

Immunological investigations in vaginal mycoses

Summary. In 42 women with chronically recurrent and 20 women with acute Candida albicans vulvovaginitis, as well as 14 women with Candida glabrata vaginitis, the following investigations were carried out: determination of protein content and secretory immunoglobulin A (sIgA) in the cervicovaginal secretion by a self‐modified ELISA technique; determination of immunocells and cellbound IgA in the cervicovaginal secretion by immunofiuorescence and nephelometric analysis of IgA in the serum. The results were compared with those of 77 pre‐menopausal non‐pregnant women with or without intake of anti‐ovulants, 17 healthy pregnant women and four hysterectomised pre‐menopausal women.

Immunological Findings in Patients with Chronically Recurrent Vaginal Candidosis and New Therapeutic Approaches

Summary: Eighteen patients with chronically recurrent vaginal candidosis showed low T‐lymphocyte counts twice as frequent as a control group of 55 women. The patients were treated with moles locally and lymphocyte stimulating pentapeptide thymopentin. The prolongation of disease‐free intervals and a cure was mainly seen in the patients with low T‐cell values before therapy. In vitro‐proliferation assays upon stimulation with Candida albicans bore no correlation with the course of the disease. We suspect a failure in the co‐operation of the immune cells, caused by differing strong responses to the Candida albicans stimulation.

Is there a role for the Fas-/Fas-Ligand pathway in chemoresistance of human squamous cell carcinomas of the head and neck (SCCHN)?

The aim of the present investigation was to determine the expression of the Fas-receptor/ligand system in established cell lines of squamous cell carcinomas of the head and neck (SCCHN), and to study its functional impact on chemotherapy-induced apoptosis in these SCCHN cell lines. We observed constitutive expression of Fas and FasL in 13 SCCHN cell lines by RT-PCR, Southern-blotting and immunocytochemistry, respectively. Administration of the agonistic Fas-antibody CH-11 led to a significant reduction of viable cells in the colorimetric MTT-assay in 5 out of 13 (38%) cell lines tested and preincubation with Interferon-gamma (IFN-gamma) rendered 3 (23%) primarily resistant cell lines sensitive. Cisplatin (cDDP) and bleomycin (BLM) caused dose-dependent cytotoxicity in all cell lines as determined by the 50% inhibitory concentration (IC(50)) and induction of apoptosis. Furthermore, both antineoplastic agents led to an enhanced surface expression of Fas and FasL in all cell lines, and this effect was independent of the respective p53-status. This upregulation of Fas/FasL surface expression increased preexisting Fas-sensitivity only, but failed to make primarily resistant cell lines undergo Fas-mediated growth reduction or apoptosis. Vice versa, blockade of Fas-receptor-ligand-interactions by monoclonal antibodies directed against FasL was able to attenuate the cytotoxic effect of cDDP and BLM in 2 out of 5 (40%) cell lines tested only. In conclusion, in contrast to many other solid tumors, the Fas/FasL-system does not seem to play an exclusive role in anticancer drug mediated apoptosis in SCCHN.

Experiences with the human tumor colony-forming assay in gynecologic malignancies

SummaryTumor samples from 74 patients with gynecologic malignancies including breast cancer were processed in a soft agar colony-forming assay. None of the samples resulted in a pure single cell suspension. Of the 10 samples meeting our criteria of evaluability for chemosensitivity testing, only 5 samples showed in vitro sensitivity to any drug. Of the 3 evaluable correlations between in vitro and in vivo results, 2 were correct. Due to the low rate of evaluable samples the assay has only limited value in the assignment of chemotherapeutic drugs for patients treated at our institution.

Evaluation of a soft-agar colony forming assay in gynecologic, tumors and breast cancer

Some human tumor cell lines are able to prevent chloroethylnitrosourea-ind~ced DNA-interStrand crosslinks by removing O -chloroethyl DNA monoadducts before cross,inks can form. The same cell lines can ~epair 0 -methyl-guanine adducts by guanine-O -~ethyl-transferase an~ were designated as Mer (methylation repair ). Methylating agents like MNNG or MNU are able to inactivate this repair system followed by an increase in chloroethylnitrosourea-induced DNA intersSrand crosslinks and in vitro cytotoxicity in Mer tumor cell lines. With regard to the potential benefit of such combinations for the treatment of human tumors knowledge of whole animal and organ toxicity is indispensible. We determined the combination toxicity index of MNU plus BCNU according to a method proposed by Skipper. The sums of the decimal fractions of the LDInS of both agents were opposed to the observed 9 ortality. The LDl0(sum) was calculated by probit analysis. Altogether 14 different combinations of both agents were investigated. The results revealed a combination toxicity index of 0.32, the lowest value reported so far for the combination of 2 cytostatic drugs. The main targets of toxicity were the intestinal ~ract and the bone marrow. The results suggest that a clinical application of such combinations should be performed with the respective precautions. Institut fGr Toxikologie und Chemotherapie, Deutsches Erebsforschungszentrum, Im Neuenheimer Feld 280, D-6900 Heidelberg S 23