Inherent growth advantage of (pre)malignant hepatocytes associated with nuclear translocation of pro-transforming growth factor alpha.

The pro-peptide of transforming growth factor alpha (proTGFalpha) was recently found in hepatocyte nuclei preparing for DNA replication, which suggests a role of nuclear proTGFalpha for mitogenic signalling. This study investigates whether the nuclear occurrence of the pro-peptide is involved in the altered growth regulation of (pre)malignant hepatocytes. In human hepatocarcinogenesis, the incidence of proTGFalpha-positive and replicating nuclei gradually increased from normal liver, to dysplastic nodules, to hepatocellular carcinoma. ProTGFalpha-positive nuclei almost always were in DNA synthesis. Also, in rat hepatocarcinogenesis, proTGFalpha-positive nuclei occurred in (pre)malignant hepatocytes at significantly higher incidences than in unaltered hepatocytes. For functional studies unaltered (GSTp(-)) and premalignant (GSTp(+)) rat hepatocytes were isolated by collagenase perfusion and cultivated. Again, DNA synthesis occurred almost exclusively in proTGFalpha-positive nuclei. GSTp(+) hepatocytes showed an approximately 3-fold higher frequency of proTGFalpha-positive nuclei and DNA replication than GSTp(-) cells. Treatment of cultures with the mitogen cyproterone acetate (CPA) elevated the incidence of proTGFalpha-positive nuclei and DNA synthesis in parallel. Conversely, transforming growth factor beta1 (TGFbeta1) lowered both. These effects of CPA and TGFbeta1 were significantly more pronounced in GSTp(+) than in GSTp(-) hepatocytes. In conclusion, nuclear translocation of proTGFalpha increases in the course of hepatocarcinogenesis and appears to be involved in the inherent growth advantage of (pre)malignant hepatocytes.

A magnetic probe to retrieve broken metallic parts of instruments during laparoscopic procedures.

To facilitate retrieval of metallic fragments out of the abdominal cavity during laparoscopic procedures, a semiflexible magnetic probe was devised. Efficacy and safety was demonstrated in two clinical cases without need for additional trocar placement or conversion to laparotomy.

Contralateral breast cancer: molecular differentiation between metastasis and second primary cancer.

Previous cancer in one breast is a strong known risk factor for cancer in the contralateral breast. Differences in tumor histology and nuclear grading are applied to distinguish between a metastatic spread and a second primary cancer, although cancers of the breast often share the same histological features. Comparison of genetic alterations in paired tumors may provide the most reliable approach for discerning clonal relationships, hence uncovering the presence or absence of multiple primary cancers. We compared tumors from 33 patients with cancer in both breasts for mutations in the p53 gene. With this molecular approach, we were able to define the relationship within paired tumors in 13 patients. The paired tumors of two patients shared the same mutation, revealing the second lesion in one case as a contralateral metachronous lymph node metastasis appearing 29 months after first surgery, and in the other as a spread to the opposite breast. In 11 patients, mutations were either discordant or solely present in one of the lesions, confirming the diagnosis of bilateral breast cancer. Histopathological evaluation had failed to provide firm diagnosis in nine out of 11 instances on account of concordances in pathological parameters such as histological type and grading. In our study, we could show that bilateral breast malignancies most frequently represent two primary breast cancers. We could also demonstrate that contralateral breast cancer spread does occur. Standard pathological assessment allowed a firm diagnosis only in the presence of different histological types.

Evaluation of the United States Food and Drug Administration-approved scoring and test system of HER-2 protein expression in breast cancer.

PURPOSE: The purpose of this study was to investigate the prognostic significance of assessment of human epidermal growth factor receptor (HER)-2 oncogene protein overexpression of breast cancer tissue by the United States Food and Drug Administration (FDA)-approved HercepTest and grading system (negative, 0 or 1+; weakly positive, 2+; strongly positive, 3+). Furthermore, results of the HercepTest were correlated with immunohistochemical results obtained using different antibodies and protocols and with HER-2 oncogene gene amplification assessed by fluorescence in situ hybridization (FISH). EXPERIMENTAL DESIGN: HER-2 status in 303 patients with lymph node-positive breast cancer was investigated by using a rabbit polyclonal antibody (DAKO) by conventional immunohistochemistry and by applying the HercepTest. Furthermore, the monoclonal antibody CB-11 was used in conventional immunohistochemistry and with the NexES automatic stainer, which is also under consideration for FDA approval for determination of eligibility for Herceptin therapy. Results were compared with FISH analysis performed in all 2+ and 3+ specimens (103 cases) and 104 HER-2-negative specimens. RESULTS: 3+ positive carcinomas were found in 8.9-15.7% of specimens. FISH revealed that almost exclusively 3+ positive cases were amplified, with the HercepTest and the NexES automatic stainer giving the best results. In univariate analysis, staining with the HercepTest revealed a significantly worse prognosis in 3+ cases. Also, 3+ cases were significantly associated with lower estrogen receptor levels and histological grade III tumors. CONCLUSIONS: This study shows that the results of the FDA-approved HER-2 grading and test system correlated strongly with findings in FISH. Furthermore, HercepTest proved to be of prognostic relevance. Strict adherence to the given protocols is critical.

TP53 mutation and p53 overexpression for prediction of response to neoadjuvant treatment in breast cancer patients.

The value of p53 to predict the cytotoxic effect of two commonly used chemotherapy regimens was assessed in patients with advanced breast cancer. Response to a DNA-damaging combination therapy [fluorouracil, epirubicin, cyclophosphamide (FEC] considered to induce p53-dependent apoptosis was compared with a microtubule stabilizing therapy (paclitaxel) expected to be independent of p53 function. The p53 status of the patients' breast tumors was assessed using both immunohistochemistry (IHC) and direct sequencing of the entire p53 gene. p53 findings were correlated with treatment response, and linkage between p53 function and cellular response was assessed by terminal deoxynucleotidyl transferase-mediated nick end labeling assay. In a series of 67 breast tumors, 19% had TP53 gene mutations, 40% had a positive p53 IHC, and 12% had both. In the FEC group, treatment failure was related to both the presence of TP53 gene mutations (P = 0.0029) and a positive IHC (P < 0.0001). Apoptosis was almost exclusively found in tumors having normal p53 in both parameters (P < 0.0001). In the paclitaxel group, treatment response was neither related to apoptosis nor to normal p53. Combination of sequencing and IHC results revealed a significant association between abnormal p53 and response to paclitaxel (P = 0.011). We found TP53 mutations, as well as p53 protein overexpression, to be associated with response to chemotherapy. Whereas clinical response to FEC was found to be dependent on normal p53, the cytotoxicity of paclitaxel was related to defective p53. The efficiency of paclitaxel during mitosis might be supported by lack of G1 arrest due to p53 deficiency. Therefore, patients with p53-deficient tumors may benefit from paclitaxel.

Inhibition of proliferation and induction of apoptosis in soft tissue sarcoma cells by interferon-alpha and retinoids.

Uncontrolled proliferation and a defect of apoptosis constitute crucial elements in the development and progression of tumours. Among many other biological response modifiers known to influence these mechanisms, the efficacy of retinoids and interferons in the treatment of various malignant entities is currently matter of discussion. In the present study, we have investigated the effects of 9-cis-retinoic acid (9cRA), 13-cis-retinoic acid (13cRA), all-trans-retinoic acid (tRA) and interferon-alpha on proliferation and apoptosis of human soft tissue sarcoma (STS) cell lines HTB-82 (rhabdomyosarcoma), HTB-91 (fibrosarcoma), HTB-92 (liposarcoma), HTB-93 (synovial sarcoma) and HTB-94 (chondrosarcoma) in relation to p53 genotype as well as p53 expression. HTB-91, HTB-92 and HTB-94 STS cells exhibited mutant p53, whereas wild-type p53 was found in HTB-93 STS cells, and a normal p53 status in HTB-82 STS cells, carrying a silent point mutation only. Interferon-alpha, irrespective of p53 status, inhibited the proliferation of all five cell lines dose- and time-dependently. Similarly, 9cRA, 13cRA and tRA decreased the proliferation of HTB-82 and HTB-93 STS cells, whereas the proliferation of p53-mutated HTB-91, HTB-92 and HTB-94 STS cells remained unchanged. Furthermore, only 9cRA and tRA were capable of inducing apoptosis in HTB-82 and HTB-93 STS cells, whereas HTB-91, HTB-92 and HTB-94 STS cells did not undergo apoptosis under the influence of 9cRA or tRA. Retinoic acid receptor (RAR)-alpha and RAR-beta mRNA were not detectable by Northern blot analysis in the five STS cell lines, whereas mRNA for the universal retinoic acid receptor, RAR-gamma, was expressed in all STS cell lines indicating that retinoid resistance was not associated with a lack of RAR expression. Apoptosis was not induced by interferon-alpha or 13cRA in any of the five STS cell lines tested. Our results indicate that within the panel of tested STS cell lines, inhibition of proliferation and induction of apoptosis result from different mechanisms which differ in their dependence upon the presence of intact p53.

The TP53 genotype but not immunohistochemical result is predictive of response to cisplatin-based neoadjuvant therapy in stage III non-small cell lung cancer.

BACKGROUND: The cytotoxic effects of cisplatin and anthracyclins have been attributed to apoptosis induction, which has been recognized as a major function of the TP53 gene. The TP53 gene appears to be mutated in about 50% of cases of non-small cell lung cancer. A possible dependence of chemotherapy response on TP53 genotype was evaluated retrospectively in a group of patients with advanced non-small cell lung cancer and induction treatment. METHODS: Patients with complete or partial remission were compared with those with stable or progressive disease with respect to TP53 genotype and overall survival. Mutations in the TP53 gene were detected by complete direct sequencing (exons 2-11). RESULTS: A normal TP53 genotype proved to be significantly associated with major response to chemotherapy (P <.001). Overall, no association was found between p53 protein expression and TP53 genotype. A normal TP53 genotype was found to be highly sensitive in predicting response to treatment, whereas a mutant genotype was revealed to be specific in predicting lack of response. The difference in overall length of survival was significant between patients exhibiting a normal TP53 genotype (corresponding to those whose disease responded to chemotherapy) and patients showing mutant TP53 genotype (corresponding to those who had disease resistant to chemotherapy, P =.027). CONCLUSIONS: In a small cohort of patients with advanced non-small cell lung cancer we found a direct link between normal TP53 genotype and response to cisplatin-based induction treatment and also between mutant genotype and resistance to treatment, whereas p53 immunohistochemical result was predictive of neither.