Maurice Chazal

Prognostic Value of Tumoral Thymidylate Synthase and p53 in Metastatic Colorectal Cancer Patients Receiving Fluorouracil-Based Chemotherapy: Phenotypic and Genotypic Analyses

PURPOSE The aim of this multicenter prospective study was to evaluate the role of intratumoral parameters related to fluorouracil (FU) sensitivity in 103 metastatic colorectal cancer patients receiving FU-folinic acid. PATIENTS AND METHODS Liver metastatic biopsy specimens were obtained for all patients and primary tumor biopsy specimens for 54 patients. Thymidylate synthase (TS), folylpolyglutamate synthetase, and dihydropyrimidine dehydrogenase were measured by radioenzymatic assays; TS promoter polymorphism (2R/2R v 2R/3R v 3R/3R) was determined by polymerase chain reaction; and p53 protein and mutations were analyzed by immunoluminometric assay and denaturing gradient gel electrophoresis, respectively. RESULTS p53 mutations were observed in 56.7% of metastases. TS activity was significantly higher in 2R/3R tumors as compared with 2R/2R or 3R/3R. TS activity in metastasis was the only parameter linked to clinical responsiveness (responders exhibited the lower TS, P =.047). Univariate Cox analyses demonstrated that TS activity in primary tumor (the greater the TS, the poorer the survival; P =.040), TS promoter polymorphism in primary tumor (risk of death of 2R/3R v 2R/2R, 2.68; P =.035), and p53 stop mutation in metastasis (risk of death of stop mutations v wild type, 3.14; P =.018) were the only significant biologic predictors of specific survival. Stepwise analysis did not discriminate between TS activity and TS polymorphism. CONCLUSION Present results confirm the value of tumoral TS activity for predicting FU responsiveness, point out the importance of detailed p53 mutation analysis for predicting survival, and suggest that TS genotype in primary tumor carries a prognostic value similar to that of TS activity.

K-Ras Mutations and Treatment Outcome in Colorectal Cancer Patients Receiving Exclusive Fluoropyrimidine Therapy

Purpose: K-Ras mutations predict resistance to anti–epidermal growth factor receptor (EGFR) monoclonal antibodies. Because combinations of anti-EGFR with 5-fluorouracil (5-FU)-based chemotherapy are promising treatments, we analyzed the effect of K-Ras mutations in patients having received exclusive 5-FU therapy. Experimental Design: This study was conducted on 93 stage IV colorectal cancer patients with unresectable measurable liver metastasis receiving 5-FU-leucovorin (56 men and 37 women; 77 cancer deaths). Liver metastases (n = 93) along with primary tumors (n = 48) were analyzed for K-Ras mutations (codons 12 and 13), p53 mutations (exons 4-9), p53 polymorphism (codon 72), thymidylate synthase (TS) polymorphism (28-bp repeats including G>C mutation), methylenetetrahydrofolate reductase polymorphism (677C>T, 1298A>C), thymidylate synthase (TS) activity, dihydropyrimidine dehydrogenase activity, folylpolyglutamate synthase activity, and p53 protein expression. Results: Thirty-six of 93 (38.7%) metastases were K-Ras mutated (30 at codon 12 and 6 at codon 13). Mutated primary tumors (16 of 48) matched perfectly with mutated metastases. The additional analyzed tumor markers were not different between K-Ras mutated and wild-type tumors. The objective response rate was 37%: 44.4% in K-Ras mutated versus 32.1% in wild-type K-Ras metastasis (P = 0.27). Low TS activity in metastasis was the only significant predictor of tumor response (P = 0.047). K-Ras status did not influence specific survival. Conclusions: The present data indicate a perfect concordance of K-Ras mutations between primary and liver metastasis and suggest that any predictive and/or prognostic value of K-Ras mutations in treatments combining anti-EGFR monoclonal antibodies with 5-FU should be exclusively linked to the anti-EGFR agent.

Characterization of centromere alterations in liposarcomas

Supernumerary ring and large marker chromosomes are a characteristic of atypical lipomas and well‐differentiated liposarcomas (ALP‐WDLPS) and are composed of amplified 12q14–15 sequences in association with variable segments from other chromosomes. Although stably transmitted, these chromosomes contain centromeric alterations, showing no detectable alpha‐satellite sequences. We performed C‐banding, fluorescence in situ hybridization, and immunostaining with anti‐centromere antibodies in 8 cases of liposarcomas with supernumerary rings and large markers, including 5 ALP‐WDLPS and 3 dedifferentiated‐LPS and high‐grade LPS. Our results with alpha‐satellite probes and anti‐CENPB antibodies confirm the lack of detectable alpha‐satellite sequences in the five ALP‐WDLPS supernumerary chromosomes, whereas centromeric activity was proved by the detection of kinetochores by using anti‐CENPC antibodies. In contrast, the high grade and dedifferentiated liposarcomas showed a different pattern. In 2 cases, amplified chromosome 12 sequences, including amplification of alpha‐satellite 12 sequences in 1 case, were present on chromosomes with typical centromeres. In another case, the rings were similar to WDLPS‐ALP rings, but a large marker contained a chromosome 5 centromere and amplified alpha‐satellite sequences from chromosome 8. ALP‐WDLPS is the first example of a tumor class for which the presence of stable analphoid chromosomes is a constant and specific abnormality. Formation of newly derived centromeres, so‐called neocentromeres, could be an original and effective way to maintain a selective advantage in neoplastic cells by conferring stability to the supernumerary chromosomes of ALP‐WDLPS. The activation of normally non‐centromeric sequences might be obtained by an epigenetic mechanism due to the peculiar chromatin conformation of these highly complex chromosomes.

Determination of microsatellite instability, p53 and K-RAS mutations in hepatic metastases from patients with colorectal cancer: relationship with response to 5-fluorouracil and survival.

In vitro and clinical studies have suggested that high‐frequency microsatellite instability (MSI‐H) phenotype, p53 and K‐ras mutations might influence the response to chemotherapy in a variety of tumors, including primary colorectal cancers (CRC). Unresectable hepatic metastases from CRC are commonly treated with 5‐fluorouracil (5FU) and folinic acid. Since several new active drugs are now used for treating CRC, molecular determinants predictive to response to 5FU would thus be crucial for optimizing indications of chemotherapy to those patients. MSI‐H phenotype, p53 and K‐ras status were characterized in a prospective study of 56 patients with CRC metastatic to the liver and treated with 5FU‐based chemotherapy. The objective response rate after a 3‐month treatment was 32.1%. The prevalence of p53 mutations, K‐ras mutations and MSI‐H phenotype was 62.5%, 30.3% and 1.8%, respectively. No significant association was found between response to chemotherapy and p53 mutations (78% mutated tumors in responders vs. 55% in nonresponders; p = 0.10) and K‐ras mutations (39% mutated tumors in responders vs. 26% in nonresponders; p = 0.34). Survival was longer for patients with p53‐mutated metastases than for patients with unresected wild‐type p53 metastases (median survival 15 months vs. 17 months; p = 0.06). The determination of the MSI‐H phenotype, p53 and K‐ras status in hepatic metastases from CRC does not discriminate a group of patients that should preferentially benefit from 5FU‐based chemotherapy. The prognosis of patients with treated liver metastases is better when p53 is mutated.

Treatment of hepatic metastases of colorectal cancer by electrochemotherapy: An experimental study in the rat ☆

BACKGROUND Electrochemotherapy, which consist of local or systemic administration of a cytotoxic agent followed by application of electric pulses to a tumor, has proved effective for various types of tumors in animals and for cutaneous and head and neck cancers in human beings. This is the first study to investigate the efficacy of electrochemotherapy for treatment of hepatic metastases of colorectal cancer in the rat. METHODS After induction of a solitary hepatic metastasis in 36 male BDIX rats, the animals were randomized to one of four groups: B-E-(no treatment), B+E-(intratumoral bleomycin), B-E+ (application of electric pulses to the tumor), and B+E+ (electrochemotherapy: intratumoral bleomycin followed by application of electric pulses). RESULTS Groups B-E and B-E+ had no tumor response. Group B+E had one partial response. Group B+E+ had seven partial responses and two complete responses. The difference in terms of response between group B+E+ and the other three groups was statistically significant (P < .05). Comparison of the mean posttherapy tumor volumes (B-E-, 50.6 mm3; B+E-, 58.7 mm3; B-E+, 46 mm3; and B+E+, 5.65 mm3) revealed a significantly smaller residual tumor in group B+E+ than in the other three groups (P < .05). CONCLUSIONS Electrochemotherapy is an effective means to reduce the volume of hepatic metastases of colorectal cancer in the rat. Additional research is required to determine the optimum treatment duration, dose effects, volume of tumor that can be treated by electrochemotherapy, and impact on survival. Such experimental studies are indispensable prerequisites for clinical trials.