Almost 2% of Spanish breast cancer families are associated to germline pathogenic mutations in the ATM gene.

PURPOSE: There is still a considerable percentage of hereditary breast and ovarian cancer (HBOC) cases not explained by BRCA1 and BRCA2 genes. In this report, next-generation sequencing (NGS) techniques were applied to identify novel variants and/or genes involved in HBOC susceptibility. METHODS: Using whole exome sequencing, we identified a novel germline mutation in the moderate-risk gene ATM (c.5441delT; p.Leu1814Trpfs*14) in a family negative for mutations in BRCA1/2 (BRCAX). A case-control association study was performed to establish its prevalence in Spanish population, in a series of 1477 BRCAX families and 589 controls further screened, and NGS panels were used for ATM mutational screening in a cohort of 392 HBOC Spanish BRCAX families and 350 patients affected with diseases not related to breast cancer. RESULTS: Although the interrogated mutation was not prevalent in case-control association study, a comprehensive mutational analysis of the ATM gene revealed 1.78% prevalence of mutations in the ATM gene in HBOC and 1.94% in breast cancer-only BRCAX families in Spanish population, where data about ATM mutations were very limited. CONCLUSION: ATM mutation prevalence in Spanish population highlights the importance of considering ATM pathogenic variants linked to breast cancer susceptibility.

BRCA2 gene: a candidate for clinical testing in familial colorectal cancer type X.

Familial colorectal cancer type X (FCCX) encompasses a group of families with dominant inheritance pattern of colorectal cancer (CRC) but no alteration in any known CRC susceptibility gene. Therefore, the explanation of their susceptibility is a priority to offer an accurate genetic counseling. We screened the 27 coding exons and exon-intron boundaries of BRCA2 in 48 FCCX probands. We identified 29 variants including a frameshift mutation. Deleterious variant c.3847_3848delGT p.(Val1283Lysfs*2) showed cosegregation with disease as well as loss of heterozygosity (LOH) in CRC tumor DNA. This is the first evidence of germline BRCA2 pathogenic mutation associated with CRC risk. Furthermore, missense variants c.502C>A p.(Pro168Thr), c.5744C>T p.(Thr1915Met) and c.7759C>T p.(Leu2587Phe) were proposed as candidate risk alleles based on cosegregation, LOH tumor analysis and in silico testing.

Detailed characterization of MLH1 p.D41H and p.N710D variants coexisting in a Lynch syndrome family with conserved MLH1 expression tumors.

Lynch syndrome (LS) is an autosomal dominant cancer-susceptibility disease caused by inactivating germline mutations in mismatch repair (MMR) genes. Variants of unknown significance (VUS) are often detected in mutational analysis of MMR genes. Here we describe a large family fulfilling Amsterdam I criteria carrying two rare VUS in the MLH1 gene: c.121G > C (p.D41H) and c.2128A > G (p.N710D). Collection of clinico-pathological data, multifactorial analysis, in silico predictions, and functional analyses were used to elucidate the clinical significance of the identified MLH1 VUS. Only the c.121G > C variant cosegregated with LS-associated tumors in the family. Diagnosed colorectal tumors were microsatellite unstable although immunohistochemical staining revealed no loss of MMR proteins expression. Multifactorial likelihood analysis classified c.2128A > G as a non-pathogenic variant and c.121G > C as pathogenic. In vitro functional tests revealed impaired MMR activity and diminished expression of c.121G > C. Accordingly, the N710 residue is located in the unconserved MLH1 C-terminal domain, whereas D41 is highly conserved and located in the ATPase domain. The obtained results will enable adequate genetic counseling of c.121G > C and c.2128A > G variant carriers and their families. Furthermore, they exemplify how cumulative data and comprehensive analyses are mandatory to refine the classification of MMR variants.

Identification of a founder EPCAM deletion in Spanish Lynch syndrome families.

Germline deletions at the 3'-end of EPCAM have been involved in the etiology of Lynch syndrome (LS). The aim of this study was to characterize at the molecular level Spanish families harboring EPCAM deletions. Non-commercial multiplex ligation-dependent probe amplification (MLPA) probes and long-range polymerase chain reaction (PCR) amplification were used to characterize each deletion. Haplotyping was performed by analyzing eight microsatellite markers and five MSH2single nucleotide polymorphisms (SNPs). Methylation of MSH2 was analyzed by methylation specific-MLPA. Tumors diagnosed in seven Spanish families harboring EPCAM deletions were almost exclusively colorectal. Mosaicism in MSH2 methylation was observed in EPCAM deletion carrier samples, being average methylation levels higher in normal colon and colorectal tumors (27.6% and 31.1%), than in lymphocytes and oral mucosa (1.1% and 0.7%). Three families shared the deletion c.858 + 2568_*4596del, with a common haplotype comprising 9.9 Mb. In two families the novel EPCAM deletion c.858 + 2488_*7469del was identified. This study provides knowledge on the clinical and molecular characteristics of mosaic MSH2 epimutations. The identification of an EPCAM founder mutation has useful implications for the molecular diagnosis of LS in Spain.

Correlation between response to neoadjuvant chemotherapy and survival in locally advanced breast cancer patients.

BACKGROUND: Measurement of residual disease following neoadjuvant chemotherapy that accurately predicts long-term survival in locally advanced breast cancer (LABC) is an essential requirement for clinical trials development. Several methods to assess tumor response have been described. However, the agreement between methods and correlation with survival in independent cohorts has not been reported. PATIENTS AND METHODS: We report survival and tumor response according to the measurement of residual breast cancer burden (RCB), the Miller and Payne classification and the Response Evaluation Criteria in Solid Tumors (RECIST) criteria, in 151 LABC patients. Kappa Cohen's coefficient (К) was used to test the agreement between methods. We assessed the correlation between the treatment outcome and overall survival (OS) and relapse-free survival (RFS) by calculating Harrell's C-statistic (c). RESULTS: The agreement between Miller and Payne classification and RCB classes was very high (К = 0.82). In contrast, we found a moderate-to-fair agreement between the Miller and Payne classification and RECIST criteria (К = 0.52) and RCB classes and RECIST criteria (К = 0.38). The adjusted C-statistic to predict OS for RCB index (0.77) and RCB classes (0.75) was superior to that of RECIST criteria (0.69) (P = 0.007 and P = 0.035, respectively). Also, RCB index (c = 0.71), RCB classes (c = 0.71) and Miller and Payne classification (c = 0.67) predicted better RFS than RECIST criteria (c = 0.61) (P = 0.005, P = 0.006 and P = 0.028, respectively). CONCLUSIONS: The pathological assessment of tumor response might provide stronger prognostic information in LABC patients.

Increased frequency of 20q gain and copy-neutral loss of heterozygosity in mismatch repair proficient familial colorectal carcinomas.

Many hereditary nonpolyposis colorectal cancers (CRCs) cannot be explained by Lynch syndrome. Other high penetrance genetic risk factors are likely to play a role in these mismatch repair (MMR)-proficient CRC families. Because genomic profiles of CRC tend to vary with CRC susceptibility syndromes, our aim is to analyze the genomic profile of MMR-proficient familial CRC to obtain insight into the biological basis of MMR-proficient familial CRC. We studied 30 MMR-proficient familial colorectal carcinomas, from 15 families, for genomic aberrations, including gains, physical losses, and copy-neutral loss of heterozygosity LOH (cnLOH) using SNP array comparative genomic hybridization. In addition, we performed somatic mutation analysis for KRAS, BRAF, PIK3CA and GNAS. The frequency of 20q gain (77%) is remarkably increased when compared with sporadic CRC, suggesting that 20q gain is involved in tumor progression of familial CRC. There is also a significant increase in the frequency of cnLOH and, as a consequence, a reduced frequency of physical loss compared with sporadic CRC. The most frequent aberrations observed included gains of 7p, 7q, 8q, 13q, 20p and 20q as well as physical losses of 17p, 18p and 18q. Most of these changes are also observed in sporadic CRC. Mutations in KRAS were identified in 37% of the MMR-proficient CRCs, and mutations in BRAF were identified in 16%. No mutations were identified in PIK3CA or chromosome 20 candidate gene GNAS. We show that the patterns of chromosomal instability of MMR-proficient familial CRC are clearly distinct from those from sporadic CRC. Both the increased gain on chromosome 20 and the increased levels of cnLOH suggest the presence of yet undiscovered germline defects that can, in part, underlie the cancer risk in these families.

Glutathione S-transferase P1 c.313A > G polymorphism could be useful in the prediction of doxorubicin response in breast cancer patients.

BACKGROUND: Identification of predicting factors for anthracyclines-based chemotherapy remains a clinical challenge. Glutathione S-transferase (GSTs) enzymes detoxify chemotherapy drugs and their metabolites. Several polymorphisms in GST genes result in reduced or no activity of the enzymes. Specifically, GSTM1 and GSTT1 genes are polymorphically deleted, the polymorphism GSTP1 c.313A>G (rs1695) determines the amino acid substitution Ile105Val, where the Val-containing enzyme has reduced activity. Also, GSTA1*B allele has reduced levels of GSTA1 enzyme. Several polymorphisms in GSTs have been associated with differences in survival for cancer patients treated with chemotherapy. PATIENTS AND METHODS: We genotyped a total of five polymorphisms in GSTM1, GSTT1, GSTP1 and GSTA1 genes in 159 patients with locally advanced breast cancer, treated with single-agent doxorubicin or docetaxel (Taxotere). Gene expression microarrays were performed in 67 breast tumor samples. We correlate this data with treatment outcome. RESULTS: In multivariate analysis, patients homozygous GG for GSTP1 c.313A>G SNP had a lower risk of chemoresistance when treated with doxorubicin (odds ratio 0.106; confidence interval 0.012-0.898; P=0.040). No association was found in the docetaxel arm. Also, we found that GSTP1 expression varied significantly among breast cancer molecular subtypes. CONCLUSIONS: GSTP1 may constitute another tool contributing to individualized anthracycline-based therapy.

Genomic predictors of response to doxorubicin versus docetaxel in primary breast cancer.

UNLABELLED: Taxanes and anthracyclines improve the outcome of early breast cancer, although the benefit is limited to a small proportion of patients and are toxic. We prospectively looked for predictors of response to these drugs. EXPERIMENTAL DESIGN: Four cycles of doxorubicin (75 mg/m²) or docetaxel (100 mg/m²) were compared as presurgical chemotherapy for breast cancer. Biomarkers were determined by immunohistochemistry and fluorescent in situ hybridization using prechemotherapy core biopsies. Tumors were also classified into one of the molecular intrinsic subtypes using an immunohistochemical panel of five biomarkers and genomic profiles. Single genes and intrinsic subtypes were correlated with response to doxorubicin versus docetaxel. Among the 204 evaluable patients, significant predictors of sensitivity in multivariate analysis were low topo2a expression and ER-negative status for doxorubicin and small tumor size and ER-negative status for docetaxel. Predictors of resistance in multivariate analysis were triple-negative status (ER/PgR/HER2 negative by IHC/FISH) for doxorubicin, and high TNM stage for docetaxel. Triple-negative tumors were associated with topo2a overexpression more than the other subtypes. In 94 patients with gene expression profiles, docetaxel was superior to doxorubicin in the basal-like subtype (good pathological response rate - PCR + class I of 56 vs. 0%; P = 0.034); no significant differences were observed in the other subtypes when comparing these two drugs. Low topo2a expression and ER-negative status were predictors of response to doxorubicin, while small tumor size and ER-negative status predicted response to docetaxel. Docetaxel was superior to doxorubicin in triple-negative/basal-like tumors, while no significant differences were seen in the remaining intrinsic subtypes.

Irinotecan-cetuximab-bevacizumab as a salvage treatment in heavily pretreated metastatic colorectal cancer patients: a retrospective observational study.

BACKGROUND: The objective was to evaluate the efficacy of irinotecan-cetuximab-bevacizumab in combination as a salvage treatment for heavily pretreated metastatic colorectal cancer patients. METHODS: A total of 39 patients resistant to both oxaliplatin and irinotecan were included in this retrospective study. Treatment consisted of irinotecan 180/m(2) every 14 days, weekly cetuximab standard dose and bevacizumab 5 mg/kg every 14 days. RESULTS: Partial response was observed in 8 patients (20%), stable disease in 24 (61%) and progressive disease in 7 (18%). Overall response rate in KRAS wild type was 6/22 (27%) and in mutated KRAS it was 2/15 (13%). Median time to progression was 8 months (6.4-9.4) and median overall survival 12 months (10.1-13.8). Overall, grade 3-4 adverse events were observed in 24 patients (62%). CONCLUSIONS: This regimen is active and moderately well tolerated in heavily pretreated advanced colorectal patients. However, caution is advisable when interpreting these results, because they run against the findings of two large phase III trials.

Gene expression profiling integrated into network modelling reveals heterogeneity in the mechanisms of BRCA1 tumorigenesis.

BACKGROUND: Gene expression profiling has distinguished sporadic breast tumour classes with genetic and clinical differences. Less is known about the molecular classification of familial breast tumours, which are generally considered to be less heterogeneous. Here, we describe molecular signatures that define BRCA1 subclasses depending on the expression of the gene encoding for oestrogen receptor, ESR1. METHODS: For this purpose, we have used the Oncochip v2, a cancer-related cDNA microarray to analyze 14 BRCA1-associated breast tumours. RESULTS: Signatures were found to be molecularly associated with different biological processes and transcriptional regulatory programs. The signature of ESR1-positive tumours was mainly linked to cell proliferation and regulated by ER, whereas the signature of ESR1-negative tumours was mainly linked to the immune response and possibly regulated by transcription factors of the REL/NFkappaB family. These signatures were then verified in an independent series of familial and sporadic breast tumours, which revealed a possible prognostic value for each subclass. Over-expression of immune response genes seems to be a common feature of ER-negative sporadic and familial breast cancer and may be associated with good prognosis. Interestingly, the ESR1-negative tumours were substratified into two groups presenting slight differences in the magnitude of the expression of immune response transcripts and REL/NFkappaB transcription factors, which could be dependent on the type of BRCA1 germline mutation. CONCLUSION: This study reveals the molecular complexity of BRCA1 breast tumours, which are found to display similarities to sporadic tumours, and suggests possible prognostic implications.

Lynch syndrome in a 15-year-old boy.

Hereditary nonpolyposis colorectal cancer (HNPCC), or Lynch syndrome, dominantly inherited, is characterized by the development of a variety of cancers due to germline mutations in DNA mismatch repair genes (MMR). This syndrome was diagnosed in a 15-year-old boy because his father and grandmother were also found to have the same kind of cancer. Microsatellite instability prompted a search for germline mutations in the MLH1, MSH2, MSH6, and PMS2 genes. Use of immunohistochemical staining for MMR proteins, genomic sequencing, and deletion studies, evidenced MSH2 axonal deletion. Neoplastic lesions of colon are most often encountered in the adult population but can, on rare occasions, be found in younger patients. We would like to emphasize the importance of suspecting Lynch syndrome and performing genetic studies, even in young patients, when there is a family history of colorectal cancer.

Dual role of LOH at MMR loci in hereditary non-polyposis colorectal cancer?

hMLH1 and hMSH2 can be considered tumor suppressor genes, as both alleles must be inactivated in order to lose the mismatch repair (MMR) function. In this regard, it has been proposed that LOH at MMR loci is a common Knudson's second-hit mechanism in HNPCC patients. However, experimental evidence supporting this view is scarcely found in the literature. We have performed a comprehensive analysis of LOH in 45 HNPCC tumors carrying a germline alteration in MMR loci. Overall, we have detected LOH at MMR loci in 56% of the cases. However, up to 40% of the LOH events targeted the mutant allele, arguing against a second-hit role in these tumors. Interestingly, the age at diagnosis was significantly older in these patients. To explain this and previous data, we propose a dual role for LOH at MMR loci in HNPCC.

A haplotype containing the p53 polymorphisms Ins16bp and Arg72Pro modifies cancer risk in BRCA2 mutation carriers.

Germline mutations in the BRCA1 and BRCA2 genes confer a high lifetime risk of developing breast and other cancers; however, remarkable differences exist regarding disease manifestation in mutation carriers. It has been suggested that other genetic and/or environmental factors modify not only the appearance but also the age of onset and type of tumor in BRCA1/2-associated cases. The aim of the present study was to investigate the role of two p53 polymorphisms (c.97-147ins16bp and c.215c>g, p.Arg72Pro) as potential modifiers. For this purpose we investigated the possible association between the two polymorphisms and disease status in 447 BRCA1/2 mutation carriers belonging to 170 Spanish breast and/or ovarian cancer families. Genotype and haplotype analyses revealed that the presence of a specific haplotype carrying the allele without the 16-bp insertion and the variant allele for the Arg72Pro (No Ins-72Pro haplotype) was associated with an earlier age of onset in BRCA2 mutation carriers. We found an increased risk of developing a first primary tumor (breast or ovarian) before 35 years of age for individuals who carried at least one No Ins-72Pro haplotype (OR: 2.69; 95% CI: 1.15-6.29; P=0.022). We confirmed these data by a functional study in which we compared different p53 genotypes in relation to their apoptotic response after cell treatment with a cytotoxic drug (AraC). Our results revealed a decrease in p53 apoptotic rate associated with the No Ins-72Pro haplotype.

Classification of BRCA1 missense variants of unknown clinical significance.

BACKGROUND: BRCA1 is a tumour suppressor with pleiotropic actions. Germline mutations in BRCA1 are responsible for a large proportion of breast-ovarian cancer families. Several missense variants have been identified throughout the gene but because of lack of information about their impact on the function of BRCA1, predictive testing is not always informative. Classification of missense variants into deleterious/high risk or neutral/low clinical significance is essential to identify individuals at risk. OBJECTIVE: To investigate a panel of missense variants. METHODS AND RESULTS: The panel was investigated in a comprehensive framework that included (1) a functional assay based on transcription activation; (2) segregation analysis and a method of using incomplete pedigree data to calculate the odds of causality; (3) a method based on interspecific sequence variation. It was shown that the transcriptional activation assay could be used as a test to characterise mutations in the carboxy-terminus region of BRCA1 encompassing residues 1396-1863. Thirteen missense variants (H1402Y, L1407P, H1421Y, S1512I, M1628T, M1628V, T1685I, G1706A, T1720A, A1752P, G1788V, V1809F, and W1837R) were specifically investigated. CONCLUSIONS: While individual classification schemes for BRCA1 alleles still present limitations, a combination of several methods provides a more powerful way of identifying variants that are causally linked to a high risk of breast and ovarian cancer. The framework presented here brings these variants nearer to clinical applicability.

Regulation of glycogenolysis in the liver of the newborn rat in vivo. Inhibitory effect of glucose.

Newborn rats were injected immediately after delivery with glucose or glucose plus mannoheptulose, and the time-courses of liver glycogen, plasma glucose insulin and glucagon concentration were studied. The administration of glucose prevented both liver glycogenolysis and the increase in plasma glucagon concentration, which normally occurs immediately after delivery. In addition, the administration of glucose prevented the decrease of plasma glucose and insulin concentration which normally occurs during the first hour of extrauterine life. Supplementation of glucose with mannoheptulose prevented the increase of plasma insulin concentrations caused by the administration of glucose; liver glycogenolysis, however, was not stimulated in these circumstances. The increase in the rate of glycogenolysis caused by the administration of glucagon was prevented in newborn rats previously treated with glucose. These results suggest that glucose exerts an inhibitory effect on the stimulation of neonatal liver glycogenolysis by glucagon.

Pre-test prediction models of BRCA1 or BRCA2 mutation in breast/ovarian families attending familial cancer clinics.

OBJECTIVE: To test whether statistical models developed to calculate pre-test probability of being a BRCA1/2 carrier can differentiate better between the breast/ovarian families to be referred to the DNA test laboratory. STUDY DESIGN: A retrospective analysis was performed in 109 Spanish breast/ovarian families previously screened for germline mutations in both the BRCA1 and BRCA2 genes. Four easy to use logistic regression models originally developed in Spanish (HCSC model), Dutch (LUMC model), Finnish (HUCH model), and North American (U Penn model) families and one model based on empirical data of Frank 2002 were tested. A risk counsellor was asked to assign a subjective pre-test probability for each family. Sensitivity, specificity, negative and positive predictive values, and areas under receiver operator characteristics (ROC) curves were calculated in each case. Correlation between predicted probability and mutation prevalence was tested. All statistical tests were two sided. RESULTS: Overall, the models performed well, improving the performances of a genetic counsellor. The median ROC curve area was 0.80 (range 0.77-0.82). At 100% sensitivity, the median specificity was 30% (range 25-33%). At 92% sensitivity, the median specificity was 42% (range 33.3-54.2%) and the median negative predictive value was 93% (range 89.7-98%). BRCA1 families tended to score higher risk than BRCA2 families in all models tested. CONCLUSIONS: All models increased the discrimination power of an experienced risk counsellor, suggesting that their use is valuable in the context of clinical counselling and genetic testing to optimise selection of patients for screening and allowing for more focused management. Models developed in different ethnic populations performed similarly well in a Spanish series of families, suggesting that models targeted to specific populations may not be necessary in all cases. Carrier probability as predicted by the models is consistent with actual prevalence, although in general models tend to underestimate it. Our study suggests that these models may perform differently in populations with a high prevalence of BRCA2 mutations.

Eight novel germline MLH1 and MSH2 mutations in hereditary non-polyposis colorectal cancer families from Spain.

Germline mutations in the MLH1 and MSH2 genes, account for the majority of HNPCC families. We have screened such families from Spain by using DGGE analysis and subsequent direct sequencing techniques. In eight families we identified six novel MLH1 and two novel MSH2 mutations comprising one frame shift mutation (c.1420 del C), two missense mutations (L622H and R687W), two splice site mutations (c.1990-1 G>A and c.453+2 T>C and one nonsense mutation (K329X) in the MLH1 gene as well as two frame shift mutations (c.1979-1980 del AT and c.1704-1705 del AG) in the MSH2 gene. Our analysis contributes to the further characterization of the mutational spectrum of MLH1 and MSH2 genes in HNPCC families.

Insulin-like growth factor-I serum levels show a midembryogenesis peak in chicken that is absent in growth-retarded embryos cultured ex ovo.

Insulin-like growth factor-I (IGF-I) is the primary mediator of GH action after birth, but its role as a regulator of prenatal growth is unclear. In a previous study we showed that IGF-I mRNA was expressed in chicken embryos beginning at the blastoderm stage (day 0, newly laid egg). Here we present the ontogeny of serum IGF-I in normal and growth-retarded chicken embryos. Serum samples were pooled from multiple embryos starting on day 4 of development in ovo until hatching (day 21). Extracts of day 2 and 3 whole embryos were also studied. IGF-binding proteins were removed by filtration on Sep-Pak C-18 cartridges. IGF-I was quantitated by a heterologous RIA validated for chicken species. Embryonic IGF-I showed a HPLC profile similar to that of adult chicken serum IGF-I. Serum IGF-I was measurable on day 6 of development (approximately 0.04 ng/ml), reached a peak on day 15 (18 ng/ml), and decreased to a low concentration (0.2 ng/ml) the day before hatching. Embryos cultured ex ovo showed progressive growth retardation after day 10 of development, and by day 20 their weight was 50% of normal. The serum IGF-I concentration of ex ovo cultured embryos was normal on day 10, but remained low until day 21, without the midembryogenesis rise observed in normal embryos. These results support the concept that IGF-I may have a role in general embryonic growth in addition to any paracrine/autocrine action in individual tissues.

Differential prognosis of replication error phenotype and loss of heterozygosity in sporadic colorectal cancer.

Several distinct genetic alterations have been associated with colorectal tumorigenesis. This study investigated the frequency of microsatellite instability, also known as replication error (RER), and loss of heterozygosity (LOH) at six chromosome regions in sporadic colorectal cancer (CRC). Eighty-six tumour and paired normal mucosa samples were included in the study. A polymerase chain reaction (PCR)-based technique was performed to analyse six (CA)n dinucleotide repeats located near or within regions containing important genes implicated in the complex process of colorectal tumorigenesis (chromosomes 2p, 3p, 5q, 11p, 17p and 18q). Overall, LOH frequency was higher in RER-tumours (25/46, 54.3%) compared with RER+ tumours (9/40, 22.5) (P = 0.04). To investigate prognostic implications, survival analysis was performed for 66 patients. Compared with RER- tumours, patients with RER+ tumours at 2p, 3p, 5q, 11p or 18q were found to have an improved prognosis (overall survival, P = 0.02 and disease-free survival (DFS) P = 0.005) this variable being an independent prognostic factor by multivariate analysis (P = 0.001). Overall survival of patients whose tumours were LOH+ was significantly shorter compared with those without LOH (overall survival, P = 0.008 and DFS, P = 0.01). Thus, tumours displaying RER+ and LOH+ phenotype, as established by microsatellite analysis, show a differential prognosis. These data indicate that this may be a useful tool for the identification of patients at different risks affected by CRC.

p53 exon 7 mutations as a predictor of poor prognosis in patients with colorectal cancer.

We have studied 61 resected colorectal adenocarcinomas in order to investigate p53 mutations as a prognostic factor for this pathology. Mutations in exons 5-9 of the p53 gene were analyzed by the polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) technique followed by sequencing. Our data indicate that p53 exon 7 mutations were prevalent in the latest stages of colorectal carcinogenesis and patients bearing this alteration had the worst prognosis. Therefore, according to our results, mutations affecting exon 7 of the p53 gene could be considered as a useful marker of biological aggressiveness for colorectal cancer.