SCC (squamous cell carcinoma) antigen is a subfraction of tumour antigen TA-4 isolated from cervical squamous cell carcinomas. Serum concentrations of SCC antigen were measured by radioimmunoassay in 382 control subjects, 70 women with cervical intraepithelial neoplasia (CIN), 517 with cervical carcinoma and 203 with other gynaecological carcinomas. Elevated SCC antigen levels (greater than 2.5 ng/ml) were found in 4% of normal controls, in 7% of women with CIN I-III, in 2%-23% with various forms of genital adenocarcinomas, in 55% with primary and in 76% of those with recurrent cervical squamous cell carcinoma. The positivity rate of the antigen was correlated with tumour stage (FIGO) and lymph node involvement of primary cervical squamous cell carcinomas. During long-term follow-up serum levels of SCC antigen were found to be concordant with tumour activity in 74% of cases. Patients with still elevated marker levels after therapy had twice the recurrence rate of women with normal serum values. Routine determination of SCC antigen during follow-up of cervical cancer is recommended.

We show by mKNA hybridization analysis and immunostaining using a mouse monoclonal antibody (moAb) to recombinant human interleukin 9 (IL-9) that both primary and cultured Hodgkin and Reed-Sternberg (II-RS) cells produce IL-9 transcripts and protein and express surface binding sites for IL-9. In addition, the growth of H-RS cells obtained from the HDLM-2 line (abundantly producing IL-9 transcripts) was significantly inhibited when anti-IL-9 moAb or an IL-9 antisense oligodeoxyribonucleotide was added to cultures. Excess addition of recombinant human IL-9 relieved the effects of anti-IL-9 moAb on HDLM-2 growth. Growth of H-RS cells of the KM-H2 line, which displays only low amounts of IL-9 detectable upon hybridization of polyadenylic acidselected RNA only, was not affected by anti-IL-9 nmAh. The proliferatile capacity of KM-H2 cells in soft agar and liquid suspension cultures was, however, augmented at least 3-fold when cells were exposed to recombi nent human IL-9. In conclusion, our results show that IL-9 is expressed by H-RS cells and point to a possible role of this molecule as a growth factor for these cells.

Speakers: S. Russell, Cambridge; M. Strauss, Berlin; P.A. Furth, Baltimore; N.S. Yang, Wisconsin; R. Ktihn, Cologne; F. Herrmann, Berlin; C. Bordignon, Mailand; C. Schatz, Strasbourg; W. Storkus, Pittsburgh; T. Pezutto, Berlin; R. Reisfeld, La Jolla; B. Groner, Freiburg; J.M. Heard, Paris; M. Blaese, Bethesda; G. Forni, Turin; S. Brocke, Stanford; F. De Plaen, Brussels; T. Blankenstein, Berlin; M. Lotze, Pittsburgh; G. Kraus, La Jolla; M. Kopf, Basel; R. Boucher, Chapel Hill; J.C. Glorioso, Pittsburgh

Transforming growth factor-beta (TGF-beta) is a potent inhibitor of growth factor-stimulated hematopoiesis in normal and leukemic conditions. Using the factor-dependent myelogenous leukemia cell lines GF-D8 and Mo7, we show that TGF-beta interferes with stem cell factor (SCF)-induced proliferation by downmodulating c-jun gene expression. The ability of SCF to induce accumulation of c-jun transcripts was abolished when TGF-beta was present in culture. Transcriptional nuclear run-on assays indicated that TGF-beta relieved the capacity of SCF to enhance the transcriptional rate of the c-jun gene. Deletion analysis of the c-jun promoter furthermore showed that SCF was activating the c-jun promoter via the NF-jun transcription factor. Gel mobility shift assays showed that SCF increased the binding activity of NF-jun to its recognition site within 5 to 15 minutes. Binding activity peaked at 1 hour after exposure to SCF and declined to starting levels within 4 hours. The ability of SCF to enhance NF-jun binding activity was also dose-dependent in the range of 5 to 100 ng/mL. Exposure of GF-D8 and Mo7 cells to TGF-beta before the addition of SCF antagonized SCF-induced NF-jun binding. Moreover, whereas SCF was capable of functionally activating a heterologous promoter containing the NF-jun binding site, pretreatment of GF-D8 cells with TGF-beta abolished transcriptional activation of this heterologous promoter. These findings indicate that SCF-mediated activation of c-jun via NF-jun is crucial for the SCF-inducible proliferative response and is inhibited by TGF-beta. In additional experiments, the antisense technique was used. Treatment of GF-D8 and Mo7 cells with an antisense oligodeoxyribonucleotide directed against the translation initiation site of c-jun abolished the capacity of SCF to induce a proliferative response, whereas sense and nonsense oligomers had no effect. Taken together, our data indicate that the counteracting modulation of the binding activity of NF-jun by SCF and TGF-beta regulates the expression of the c-jun gene and thereby the proliferative state of the GF-D8 and Mo7 target.

One of the most common structural chromosomal aberrations in acute myeloblastic leukemia (AML) is the pericentric inversion of chromosome 16, inv(16) (p13q22) which results in the rearrangement of the CBFβ- and the MYH11-gene and the transcription of a chimeric CBFβ/MYH11 fusion mRNA. In a retrospective analysis, 329 patients (pts.) with de novo AML were evaluated for CBFβ/MYH11 fusion transcripts using reverse transcriptase polymerase chain reaction (RT-PCR). In 14/329 pts. (4%) CBFβ/MYH11 fusion transcripts could be detected. All pts. displayed the same breakpoint at bp 1921 of the MYH11-gene. In 4 pts. the inv(16) could not be detected by conventional cytogenetical analysis: 2 pts. showed a normal chromosome 16, in another 2 pts. a deletion del(16q22) was detected. Combined cytogenetical analysis and fluorescence in situ hybridization (FISH) with a-satellite probes for the chromosomes 7, 8, X and Y revealed additional numerical chromosomal aberrations in 4/14 pts. (+8, n=2; +8 and +21, 11=1; +9 and +21, n=1). Five pts. were analysed for CBFβ/MYH11 fusion transcripts in complete remission (CR) after polychemotherapy. 4/5 pts. became PCRnegative after induction and/or consolidation therapy; 1 of these pts. relapsed after 10 months (mo) and was again CBFβ/MYH11 positive. After additional therapy this pt. achieved a 2nd CR but remained PCR-positive. The other 3 pts. are in ongoing CR after 10 to 94 mo. 1/5 pts. remained PCR-positive even after high-dose AraC consolidation and relapsed after 8 mo. Taken together, RTPCR is a rapid and sensitive tool for diagnosis of inv(16) positive AML which might be more sensitive than conventional cytogenetical analysis. The clinical relevance of MRD detection in clinical CR has to be determined in prospective studies.

The reciprocal translocations t(8;21) and t(15;17) are among the most common chromosomal aberrations in patients (pts.) with acute myeloblastic leukemia (AML). Both translocations result in chimeric fusion mRNAs [AML1/ETO for t(8;21) and PML/RARa for t(15;17)] which can be detected by specific amplification with reverse transcriptase polymerase chain reaction (RT-PCR). In addition to primary diagnosis, RT-PCR can be used in these pts. for the sensitive monitoring of minimal residual disease (MRD) in complete remission (CR) after intensive polychemotherapy. In this study, we analysed 9 pts. with t(8;21) and 16 pts. with t(15;17) in CR for MRD using nested RT-PCR for AML1/ETO and PML/RARa. In the t(8;21) group 4/9 pts. became PCR-negative after induction and/or consolidation therapy; 3 of these pts. are in ongoing CR after 12 to 72 months (mo). 1 pt. relapsed after 24 mo. 5/9 pts. remained PCRpositive after chemotherapy; 3 of these pts. are in ongoing CR after 2 to 48 mo, 1 pt. relapsed after 12 mo and 1 pt. died in CR. In the t(15;17) group 15/16 pts. became PCR-negative after 1 or 2 cycles of induction chemotherapy. 13 of these pts. are in ongoing CR after 3 to 78 mo. 2 pts relapsed after 21 and 22 mo. and in both pts. clinical relapse was preceeded by the recurrence of PML/RARα fusion transcripts in morphological CR. 1/16 pts. remained PCR-positive after induction and consolidation therapy but became negative after allogeneic bone marrow transplantation and is in CR after 15 mo. Taken together, in most of the t(15;17) positive AML pts. the leukemic clone can be reduced below the detection level by aggressive chemotherapy. Because the recurrence of PML/RARa transcripts in CR is correlated with later clinical relapse, serial PCR-analyses should be performed in these pts. In contrast, a considerable proportion of AML-pts. with t(8;21) remain PCR-positive after chemotherapy apparently without correlation to clinical outcome. Further prospective studies are necessary to determine whether PCR-analyses are of clinical relevance in these pts.

Cytosine arabinoside (ara-C) has been used extensively in the treatment of acute myelogenous leukemia (AML) for many years (see [4] for review). Besides the conventional low-dose regimen worked out by Frei et al. [16], high dose regimens [29] as well as combinations of ara-C with anthracycline derivatives [30] have further expanded and improved today’s therapeutic repertoire for remission induction of AML. Despite considerable success in improving complete remission (CR) rates using new ara-C-based treatment modalities, we still face appaIlingly high relapse rates. Further improvement of AML induction therapy is thus mandatory.

The CD95 (APO-1/Fas) receptor, which is able to induce apoptotic cell death upon oligomerization by its natural ligand or by anti-CD95 antibodies, is broadly distributed in normal and malignant hematopoietic cells. However, its functionality could be demonstrated only in a limited number of cell types, and molecular events responsible for the resistance of cells to CD95-mediated apoptosis remain to be elucidated. The bcl-2 protein which has been associated with increased resistance to various apoptotic stimuli has been shown to block partially the CD95-mediated apoptosis in murine cell lines, but not in human lymphocytes. In AML, a limited number of cases has yet been tested as to their susceptibility to CD95-induced apoptosis. Using flow cytometry, we investigated both expression and functionality of CD95 and bcl-2 in untreated leukemic cells collected from 46 de novo AML patients, and correlated the results with the expression of the progenitor cell associated CD34 antigen and with growth characteristics of leukemic cell samples in vitro. Our study revealed that almost all (>90%) AML samples expressed both bcl-2 and CD95. The level of bcl-2 expression, quantified as mean fluorescence intensity, was significantly higher in CD34+ than in CD34− AML (p<0.001) while that of CD95 was significantly lower in CD34+ leukemic cell samples (p<0.001). The apoptotic effect of crosslinking of CD95 by anti-CD95 antibody was observed in 88% of CD34− AML but only in 29% of CD34+ AML samples (p<0.01). No correlation between the extent of CD95-induced apoptosis and the level of CD95 and bcl-2 expression was found. Autonomous growth was observed in both CD34+ and CD34− samples (67 and 84% of cases, respectively). In CD34+ AML, all cell samples susceptible to CD95-mediated apoptosis exhibited autonomous growth. By contrast, CD34− samples were highly sensitive to CD95−triggering independent of their growth characteristics. In conclusion, our data reveal a high susceptibility to CD95− triggering of AML with a CD34− phenotype, and suggest that this subgroup of AML may represent an appropriate target for therapeutic strategies based on tumor-specific T-cell cytotoxicity involving CD95-CD95L interactions.

In a previous study for patients with advanced MDS, AML after MDS, and secondary AML, a remission rate about 45% was obtained after intensified chemotherapy. In order to increase the remission rate, a prospective trial with three cycles of idarubicin 10 mg/m2 x 3d (2d in cycle 2 + 3), continuous infusion of ara-C 100 mg/m2 x 7d (5d in cycle 2 + 3), etoposide 100 mg/m2 x 5d (cycle 1–3), followed by amsacrine 60 mg/m2 x 5d and ara-C 100 mg/m2 x 5d in cycle 4, with G-CSF priming in the first two cycles was initiated. From February 1994 until October 1996, a total of 67 patients (34 male, 33 female) with RAEB-t (n = 11), secondary AML (n = 8), or AML following MDS (n = 48) were entered. 44 patients (age 33–75 years, median 58 years; 9 RAEB-t, 32 AML following MDS, 3 secondary AML) have finished induction therapy. After the first course of chemotherapy, 27 patients had a complete remission (CR 61.4%), 7 patients showed a partial remission (PR 15.9%), and 10 patients were non-responders (NR 22.7%). After cycle 2, 4 additional patients entered CR resulting in an overall CR rate of 70.5%. Overall, 4 patients died from early death, 4 from progression or relapse. Seven patients underwent stem cell transplantation. In course I, neu-tropenia (< 500/μ1) lasted 15 (range 1–40) days, while the median number of days with febrile neutropenia was 13 (range 1–34). Thrombocytopenia (< 20000/μl) lasted for a median of 9 (range 8–58) days. In conclusion, the intensive chemotherapy with idarubicin/ara-C/VP-16 combined with G-CSF priming results in a promising rate of remissions and is well tolerated.

The lipopolysaccharide binding protein (BLP) is of major importance for endotoxin recognition, presentation and subsequent cytokine induction in immune cells. As a member of a growing family of structurally and functionally related proteins, LBP is synthesized in hepatocytes and constitutively secreted into the bloodstream. During the acute-phase response, however, LBP levels rise substantially. In this article the mechanisms of induction of LBP protein synthesis are highlighted. Induction of LBP in hepatocytes is the result of transcriptional and posttranscriptional mechanisms, as shown by nuclear run-on and RNA half-life experiments. Cloning of the 5' flanking region of the LBP gene gave results consistent with the LBP promoter as a typical acute-phase protein promoter. Reporter-gene assays employing the Luciferase gene and mutation variants of the LBP promoter revealed that integrity of a common acute-phase promoter motif, binding STAT-3, is essential for activation of the LBP promoter. Elucidating the transcriptional activation mechanism could show the way how to therapeutically lower LBP levels in high-risk patients in order to reduce their susceptibility to Gram-negative septic shock.

We describe a patient presenting with systemic lupus erythematosus (SLE) and concomitant low-grade (lg) non-Hodgkin’s lymphoma of the B cell type (B-NHL). Although the association of autoimmune disorder and lymphoma is well conceived, there is only scarce information available as to the simultaneous occurrence of both disease conditions in one patient. As in this patient diagnosis of lg B-NHL was also based on the detection of a monoclonal population of CD5+ B lymphocytes, and given that the polyclonal expansion of CD5+ B cells has been previously reported in rheumatoid arthritis (RA), Sjögren’s syndrome (SS) and single cases of SLE, the observations we made in this patient led us to discuss the role of the CD5+ population in the development of rheumatic disorders and concomitant lymphoid malignancy. Moreover, since impaired production rates of interleukin 3 (IL-3) and interleukin 4 (IL-4) have been associated with an abnormal expansion of CD5+ lymphoma cells and seeing that soluble interleukin 2 receptor (sIL-2R) serum levels were found to be positively correlated with disease activity both in SLE and lg B-NHL, these parameters were investigated and related to the patient’s disease state throughout the entire clinical observation period.