R. Kimmig

Immunotherapy of malignant ascites with trifunctional antibodies

A new class of intact bispecific antibodies shows unmet effector qualities by activation of not only T cells but also simultaneous activation of Fcγ receptor type I/III+ cells (macrophages, NK‐cells and DC). These trifunctional antibodies (trAb) lead to efficient specific killing of targeted tumor cells without any pre‐ or co‐stimulation. This concept was investigated in vivo in patients with malignant ascites in a clinical situation that allowed monitoring of tumor cell elimination and correlation with clinical effects. In a prospective study, 8 patients with malignant ascites due to peritoneal carcinomatosis were treated with intraperitoneal application of trAb, which bound either the EpCAM‐ or Her2/neu‐antigen on tumor cells. Treatment consisted of 4–6 applications within 9–23 days with a total amount of 145–940 μg. Seven of eight patients required no further paracentesis during follow‐up or until death with a mean paracentesis‐free interval of 38 weeks (median = 21.5, range = 4–136). Tumor cell monitoring showed a complete elimination of tumor cells in ascites already at total doses as low as 40–140 μg. Clinical response with disappearance of ascites accumulation was seen in all patients, which was correlated with elimination of tumor cells (p = 0.0014). Severe adverse events were not observed. Clinically relevant side effects were fever, moderate abdominal pain and skin reactions. Intraperitoneal immunotherapy with trAb showed convincing efficacy in patients with malignant ascites. This treatment offers new therapeutic options for patients with peritoneal carcinomatosis.

Screening for cervical neoplasia by self-assessment for human papillomavirus DNA.

We evaluated self-collected vaginal human papillomavirus tests for cervical cancer screening and compared it with the specimens taken directly from the cervix in 247 patients at high-risk for cervical disease. The sample taken by the patient showed human papillomavirus DNA in a higher percentage than the sample taken by a doctor. Sensitivity of high-risk human papillomavirus types for high-grade cervical precursors (CIN 2/3) and invasive cervical cancers were 93% for both methods.

Photodynamic therapy in women with cervical intraepithelial neoplasia using topically applied 5-aminolevulinic acid †

Photodynamic therapy (PDT) is a novel treatment modality that produces local tissue necrosis with laser light after prior administration of a photosensitizing agent. We performed a study of topically applied 5‐aminolevulinic acid (5‐ALA) in the photodynamic treatment of women with high‐grade cervical intraepithelial neoplasia (CIN) using fixed 5‐ALA doses and application protocols derived from previous in vitro and in vivo results. Three to 5 hr prior to PDT, 10 ml of a 20% solution of 5‐ALA was topically applied using a cervical cap. PDT was performed with irradiation of 100 J/cm2 at an irradiance of 100–150 mW/cm2 with an argon‐ion‐pumped dye laser at 635 nm. For the endocervix, a specifically designed cylindrical applicator was used. Ten treatment cycles of PDT using 5‐ALA were performed in 7 patients with high‐grade CIN. Non‐thermal laser treatment with 100–150 mW/cm2 was well tolerated. Local toxicity was minor as several patients reported burning sensations and vaginal discharge, but no necrosis, sloughing or scarring occurred. After 3 months, a significant reduction in the size of the ectocervical CIN lesions was noted in only 3 patients, who underwent a second PDT cycle. However, no significant improvement in CIN lesions was noted since cold knife conization revealed persistent CIN in all 7 cases. Therefore, PDT after topical application of 5‐ALA using an irradiation of 100 J/cm2 produces only minimal side effects. However, it does not appear to be effective in treating CIN. Int. J. Cancer 81:34–38, 1999.

Cellular apoptosis susceptibility gene expression in endometrial carcinoma: correlation with Bcl-2, Bax, and caspase-3 expression and outcome.

Deregulation of proliferation and apoptosis is known to contribute to neoplastic transformation and growth. Using specific antibodies for the cellular apoptosis susceptibility (CAS) gene, caspase-3, Bcl-2, and Bax, we examined the protein expression in 89 endometrial carcinomas and in 56 samples of nonneoplastic adjacent endometrium for comparison. Immunostaining results were scored with regard to approximate percentage of positive tumor cells (<10%, 10% to 50%, >50%) and relative immunostaining intensity (1+, 2+, 3+). In nonneoplastic endometrium, CAS protein was expressed in 70.6%, Bax in 64%, caspase-3 in 52%, and Bcl-2 in 87%. In neoplastic tissue, CAS was present in 93% of the tumors, Bax in 88%, caspase-3 in 77%, and Bcl-2 in 51%. Bcl-2:Bax ratio was >1 in 9 cases (10%). In cases of atrophy (n=24) and simple (n=10) and complex (n=22) hyperplasia in the adjacent endometrium, lower levels of expression compared with carcinoma were observed for CAS (p=0.003), caspase-3 (p=0.034), and Bax (p=0.04) and higher levels for Bcl-2, although for this protein the results were not statistically significant (p=0.32). There was no association between immunoscores and FIGO stage. High caspase-3 levels were seen in endometrioid tumor type (p=0.017). CAS expression was higher in grade 3 tumors (p=0.002) and older patients (p=0.013). All tumors of younger patients (<50 years) were Bcl-2 negative (p=0.037). Caspase-3 correlated positively with CAS (p=0.008), Bax (p=0.04), and low Bcl-2:Bax ratio (p=0.043), and inversely (as a trend) with Bcl-2 (p=0.056). Survival analysis (Kaplan-Meier and Cox regression) established a strong association between prognosis and stage, grade, and histologic type (all p≤0.0036). In addition, shorter survival was observed for patients whose tumors contained >50% of positive cells for caspase-3 (p=0.024) or for CAS (p=0.04). Age, Bcl-2, Bax, and Bcl-2:Bax ratio did not provide prognostic information. Our results suggest a role of CAS, Bcl-2, Bax, and caspase-3, which are apparently involved in the progressive deregulation of proliferation and apoptosis leading from simple and complex hyperplasia to carcinoma. In addition, CAS and caspase-3 protein levels may be useful markers in predicting the outcome of the patients.

Second-line carboplatin and gemcitabine in platinum sensitive ovarian cancer—a dose-finding study by the Arbeitsgemeinschaft Gynäkologische Onkologie (AGO) Ovarian Cancer Study Group

BACKGROUNDnDespite the progress that has been achieved in the last years, recurrence rates in ovarian cancer patients are still considerably high and the majority of patients ultimately become candidates for second-line treatment. Carboplatin reinduction is a broadly adopted regimen in patients with recurrences occurring six months or later after first-line treatment. Gemcitabine is among the candidates as combination partner in second-line regimens.nnnPATIENTS AND METHODSnWe performed a study with escalating doses of gemcitabine combined with carboplatin in 26 platinum-pretreated patients with recurrent ovarian cancer and a treatment-free interval of 6+ months. Dose-limiting toxicity (DLT) and a maximum tolerable dose (MTD) recommendable for further trials was evaluated.nnnRESULTSnThe DLT was myelosuppression, mainly thrombocytopenia. No dose limiting non-hematological toxicities were observed. The MTD of gemcitabine was 1,000 mg/m2 given on days 1 + 8 of a three-week schedule combined with carboplatin AUC 4 given on day 1. The majority of evaluable patients showed an objective response (62.5%), and median progression-free and overall survival were 10 and 18+ months, respectively.nnnCONCLUSIONnGemcitabine-carboplatin given according to the MTD is well tolerated and active against recurrent platinum-sensitive disease. A randomized trial comparing carboplatin with or without gemcitabine in platinum-sensitive ovarian cancer has already been initiated.

Gemcitabine/carboplatin (GC) vs. carboplatin (C) in platinum sensitive recurrent ovarian cancer (OVCA). Results of a Gynecologic Cancer Intergroup randomized phase III trial of the AGO OVAR, the NCIC CTG and the EORTC GCG

5005 Background: Most patients (pts) with OVCA relapse. ICON 4 - AGO OVAR 2.2 results suggest combination C-based therapy may be superior to single agent C. GC is a feasible less neurotoxic regimen and thus of interest to evaluate in this setting.nnnMETHODSnIn this phase III trial pts with platinum sensitive recurrent OVCA (≥ 6 months after the end of primary therapy) were randomized to receive C (AUC 4 d1) plus G (1000 mg/m2 d1 and 8) or C (AUC 5 d1) every 3 weeks. The primary objective compared progression free survival (PFS) in both arms. Secondary objectives were response rate (RR) and duration, overall survival (OS), toxicity and quality of life (QoL).nnnRESULTSnFrom 09/99 and 04/02, 356 pts (178 GC, 178 C) were randomized. Both study arms were well balanced for baseline disease characteristics. Median number of cycles was 6 for GC (0-10) and C (0-9). The weekly dose intensity was 98.2% for C (C arm) and 96.2% for C and 75.6% (92.8% for d 1 and 63.4% for d8) for G (GC arm). Grade 3/4 hematologic toxicities were significant higher in the GC arm (anemia 27.4% of pts. vs. 8.0%, neutropenia 70.3% vs. 12.0%, thrombocytopenia 34.9% vs. 11.4%). G-CSF or GM-CSF was more often used in GC (23.6% of pts. vs. 10.1%) as well as red cell transfusions (37.1% vs. 14.6%). Clinical sequelae were similar between arms: febrile neutropenia (1.1% GC vs. 0% C) and infections (0.6% for both arms). Grade 3/4 non-hematologic toxicities were infrequent in both arms (< 5%), especially for sensory neuropathy (1.1% GC vs. 1.7% C). Overall RR for GC was 47.2% (95% CI: 39.9-54.5%) and 30.9% (95% CI: 24.1-37.7) for C (p= 0.0016). GC pts reported significantly faster palliation of abdominal symptoms as well as significantly improved global QoL. With a median follow up of 13 months, median PFS was 8.6 mo (95%CI: 8.0-9.7) for GC and 5.8 mo (95% CI: 5.2-7.1) for C (HR 0.72 [95% CI: 0.57-0.90], p= 0.0038, e= 311). OS data are immature and this study was not powered to detect differences in OS.nnnCONCLUSIONnThe combination of G plus C improves PFS and QoL in platinum sensitive recurrent OVCA patients with acceptable toxicity. [Table: see text].

Topotecan versus treosulfan, an alkylating agent, in patients with epithelial ovarian cancer and relapse within 12 months following 1st-line platinum/paclitaxel chemotherapy. A prospectively randomized phase III trial by the Arbeitsgemeinschaft Gynaekologische Onkologie Ovarian Cancer Study Group (AGO-OVAR).

OBJECTIVEnEffective therapies with a low rate of side effects are warranted in the 2nd-line setting in ovarian cancer. Both topotecan and the alkylating agent treosulfan have demonstrated efficacy in this patient group and are broadly used in Germany. Therefore, we started a prospectively randomized phase III trial comparing these two drugs in early recurrent ovarian cancer.nnnMETHODSnPatients having relapsed after platinum-taxane therapy were randomized to receive either topotecan or treosulfan. Stratification depended on platinum sensitivity (stratum 1: up to 6 months after primary chemotherapy, stratum 2: 6 to 12 months).nnnRESULTSnA total of 274 patients were treated either with topotecan (136 patients) or treosulfan (138). Hematologic toxicity was significantly more frequent with topotecan but without severe clinical consequences. Non hematologic toxicity was similar in both study arms. Overall survival was significantly longer with topotecan (p=0.0023), with a median of 55.0 weeks versus 41.0 weeks as well as progression-free survival (p=0.0020) with a median of 23.1 weeks versus 12.7 weeks. Similar results were found for stratum 2 subgroup. Overall response rate was 27.5% for topotecan and 16.0% for treosulfan (p=0.0307). In stratum 1 progression-free survival was 18.1 weeks for topotecan and 9.4 weeks for treosulfan (p=0.0476), but there was no difference in overall survival in this prognostic poor subgroup.nnnCONCLUSIONSnThis randomized phase III trial could detect superiority of topotecan versus treosulfan in patients with recurrent disease after platinum-paclitaxel combination therapy. Our experience indicates that optimization of systemic treatment could improve outcome even in this poor prognostic subgroup of patients with relapsed ovarian cancer.

Obstetric risks and vertical transmission of hepatitis C virus infection in pregnancy.

Background. Reports of obstetric complications of mothers infected with hepatitis C virus (HCV) are limited and the risk of mother‐to‐infant transmission varies widely. We assessed the course of pregnancy in HCV‐infected women and the rate of vertical transmission.

mRNA expression of ligands of the epidermal-growth-factor-receptor in the uterus

Six different ligands of the epidermal‐growth‐factor receptor (EGFR) have been identified in the past. In some cervical squamous‐cell carcinomas, an increased amount of proteins binding to the EGFR has been reported. In order to identify the mRNA of EGFR ligands (EGFRL), which might be over‐expressed in cervical and endometrial cancers, we performed semi‐quantitative reverse‐transcription/polymerase chain reactions (RT‐PCR) for all 6 EGFRL in RNA extracts of normal and malignant tissue samples of the human uterus. PCR products from RNA extracts of 83 patients were quantitated relative to the housekeeping gene and internal standard pyruvate dehydrogenase by analyzing the PCR kinetics of product synthesis. In extracts of normal cervix, the level of mRNA expression of the EGFRL was significantly higher than in endometrium. No significant difference was detected between normal cervix and cervical carcinomas. However, both in cervical and in endometrial cancers, mRNA expression was non‐parametrically distributed and in some cervical cancers overexpression of transforming growth factor alpha (TGF‐α), amphiregulin or EGF was observed. In endometrial cancers, mRNA levels of all EGFRL were higher than in normal endometrium. This increase was significant (p < 0.005) for TGF‐α and amphiregulin. Thus, TGF‐α mRNA is over‐expressed in approximately 10% of cervical cancers and in the majority of endometrial cancers. Since TGF‐α anti‐sense therapy might represent a future strategy in such cancers, we also determined the absolute level of TGF‐α mRNA expression by quantitative PCR using a cloned standard. Int. J. Cancer 72:581–586, 1997.© 1997 Wiley‐Liss, Inc.

Quantitative determination of the epidermal growth factor receptor in cervical cancer and normal cervical epithelium by 2-color flow cytometry: Evidence for down-regulation in cervical cancer

Expression of epidermal growth factor receptor (EGFR) was quantified by 2‐color flow cytometry in cervical cancer (n = 73) and normal cervical epithelium (n = 11). EGFR was determined using a murine monoclonal EGFR antibody, and number of bound antibodies was quantified adding calibration beads with defined antigenic binding sites. Tumor cells were identified by simultaneous DNA staining. Epithelium of normal cervical tissue was detected by labeling for cytokeratin. Results were compared with EGFR quantification by autoradiography on cryostat sections using a radioligand binding assay. A high degree of correlation was found between the 2 methods. In cervical carcinomas 14,600 binding sites/cell (median; range, 160–283,000 binding sites/cell) were detected, considerably less compared with normal cervical squamous epithelium, which was 30,700 binding sites/cell (median; range, 19,900–44,000 binding sites/cell). This finding clearly contrasts with other reports of enhanced EGFR expression in cervical cancer. The discrepancy may be explained by contamination of tissue homogenates used for radioligand or enzyme immunosorbent assays by non‐epithelial tissue elements with low or absent EGFR expression. Interference with quantitative EGFR determination in epithelial cells may result in false low estimates of EGFR expression predominantly in normal cervical tissue. This should be avoided by identifying tumor and normal epithelial cells prior to analysis. In our study, 63% of cervical cancers expressed low levels of EGFR compared with normal cervical epithelium, and only 10% showed overexpression. There is evidence that cervical carcinomas overexpressing EGFR represent a small, but biologically distinct group of cervical cancers exhibiting enhanced aggressiveness associated with poor survival of the patients. Int. J. Cancer 74:365–373, 1997.