Study protocol for a randomized, controlled trial using a novel, family-centered diet treatment to prevent nonalcoholic fatty liver disease in Hispanic children.

BACKGROUND: Non-alcoholic fatty liver disease (NAFLD) is the leading liver disorder among U.S. children and is most prevalent among Hispanic children with obesity. Previous research has shown that reducing the consumption of free sugars (added sugars + naturally occurring sugars in fruit juice) can reverse liver steatosis in adolescents with NAFLD. This study aims to determine if a low-free sugar diet (LFSD) can prevent liver fat accumulation and NAFLD in high-risk children. METHODS: In this randomized controlled trial, we will enroll 140 Hispanic children aged 6 to 9 years who are ≥50th percentile BMI and without a previous diagnosis of NAFLD. Participants will be randomly assigned to either an experimental (LFSD) or a control (usual diet + educational materials) group. The one-year intervention includes removal of foods high in free sugars from the home at baseline, provision of LFSD household groceries for the entire family (weeks 1-4, 12, 24, and 36), dietitian-guided family grocery shopping sessions (weeks 12, 24, and 36), and ongoing education and motivational interviewing to promote LFSD. Both groups complete assessment measures at baseline, 6, 12, 18, and 24 months. Primary study outcomes are percent hepatic fat at 12 months and incidence of clinically significant hepatic steatosis (>5%) + elevated liver enzymes at 24 months. Secondary outcomes include metabolic markers potentially mediating or moderating NAFLD pathogenesis. DISCUSSION: This protocol describes the rationale, eligibility criteria, recruitment strategies, analysis plan as well as a novel dietary intervention design. Study results will inform future dietary guidelines for pediatric NAFLD prevention. TRIAL REGISTRATION: ClinicalTrials.gov, NCT05292352.

Sugar intake by type (added vs. naturally occurring) and physical form (liquid vs. solid) and its varying association with children's body weight, NHANES 2009-2014.

BACKGROUND: Sugars and their primary dietary sources (milk, fruits, sweetened foods and beverages) are associated, in different ways, with a range of health outcomes, including obesity. The contribution made to total sugar intake and how the different types and forms of sugar associate with body weight is unclear. OBJECTIVE: To describe sugar consumption and examine its association with weight status among U.S. children by sugar type [added {AS} vs. naturally occurring {NOS}] and form (solid vs. liquid). DESIGN: Cross-sectional dietary data (2 24-h recalls) from children 2-19 years in the National Health and Nutrition Examination Survey, 2009-2014 (n = 8136) were used to estimate the amount of each type and form of sugar by age and weight status. Linear regression models tested trends and the multivariate adjusted association between the different sugars and weight status. RESULTS: Mean total sugar, AS, and NOS was 118.1 g [25.3% total energy {TE}], 71.5 g (14.8% TE), 46.7 g (10.5% TE), respectively. AS in sugar-sweetened (non-dairy) beverages and NOS in juices contributed 6.9% and 2.4% of TE, respectively. Only %TE from AS (controlled for potential demographic, lifestyle confounders) was associated with change in body mass index z-score (BMIz) [AS in beverages: BMIz ß + 0.01 {95% CI: 0.002, 0.03}; AS in foods: BMIz ß - 0.03 {95% CI: -0.04, -0.02}]. CONCLUSION: Dietary sugars, most of which are AS, are a major contributor of calories in the diets of U.S. children. Only AS in non-dairy sources were associated with weight although the direction differed by the form consumed. AS in beverages were associated positively and those in foods were associated inversely with children's weight status.

Pathways and mechanisms linking dietary components to cardiometabolic disease: thinking beyond calories.

Calories from any food have the potential to increase risk for obesity and cardiometabolic disease because all calories can directly contribute to positive energy balance and fat gain. However, various dietary components or patterns may promote obesity and cardiometabolic disease by additional mechanisms that are not mediated solely by caloric content. Researchers explored this topic at the 2017 CrossFit Foundation Academic Conference 'Diet and Cardiometabolic Health - Beyond Calories', and this paper summarizes the presentations and follow-up discussions. Regarding the health effects of dietary fat, sugar and non-nutritive sweeteners, it is concluded that food-specific saturated fatty acids and sugar-sweetened beverages promote cardiometabolic diseases by mechanisms that are additional to their contribution of calories to positive energy balance and that aspartame does not promote weight gain. The challenges involved in conducting and interpreting clinical nutritional research, which preclude more extensive conclusions, are detailed. Emerging research is presented exploring the possibility that responses to certain dietary components/patterns are influenced by the metabolic status, developmental period or genotype of the individual; by the responsiveness of brain regions associated with reward to food cues; or by the microbiome. More research regarding these potential 'beyond calories' mechanisms may lead to new strategies for attenuating the obesity crisis.

Is there a role for PCR-SSCP among the methods for missense mutation detection of TP53 gene?

Mutation analysis methods have increased in variety during the past years. High-throughput microarray methods have especially increased in popularity. However, new methods require reference points, and not all of the methods are equal in sensitivity and specificity. Furthermore, the detection of unknown missense mutations, such as unknown TP53 mutations in human tumors, for clinical purposes requires great accuracy, which may be difficult to acquire with the current high-throughput methods. For these reasons, the classical methods, such as PCR-manual sequencing and PCR-SSCP, are still valuable and necessary.

Environmental tobacco smoke, genetic susceptibility, and risk of lung cancer in never-smoking women.

BACKGROUND: Exposure to environmental tobacco smoke (ETS) is considered to be a major lung cancer risk factor for never smokers. We investigated the hypothesis that never-smoking women who are exposed to ETS and develop lung cancer are a genetically susceptible population. METHODS: Archival tumor tissues were analyzed from 106 never-smoking women enrolled in a case-control study of ETS (and other personal and environmental factors) and lung cancer risk. We analyzed germline polymorphisms in genes that have been associated with cancer susceptibility and whose products activate (cytochrome P450 1A1 [CYP1A1]) and detoxify (glutathione S-transferases M1 [GSTM1] and T1 [GSTT1]) chemical carcinogens found in tobacco smoke. RESULTS: When compared with never smokers who had no ETS exposure and developed lung cancer (n = 55), never smokers with exposure to ETS who developed lung cancer (n = 51) were more likely to be deficient in GSTM1 activity (i.e., were GSTM1 null) because of a genetic polymorphism in the GSTM1 gene (odds ratio = 2.6; 95% confidence interval = 1.1-6.1). A statistically significant rising trend in risk occurred with increasing ETS exposure (two-sided P =. 02), reaching a more than sixfold excess risk in those exposed to 55 pack-years of ETS (ETS pack-year = ETS produced by an active smoker, within a confined space such as a room, who smokes one pack of cigarettes a day for a year). No evidence was found of associations between GSTT1 deficiency or the CYP1A1 valine variant and lung cancer risk due to ETS exposure. CONCLUSIONS: A common genetic polymorphism divides the population of never smokers into two groups of approximately equal size, one (homozygous carriers of the GSTM1 null allele) that has a statistically significant greater risk of lung cancer from ETS than the other (heterozygous or homozygous carriers of the wild-type GSTM1 allele).

p53 mutations, ras mutations, and p53-heat shock 70 protein complexes in human lung carcinoma cell lines.

The p53 tumor suppressor gene is frequently mutated and the K-ras oncogene is occasionally mutated in primary specimens of human lung carcinomas. These mutated genes also cooperate in the immortalization and neoplastic transformation of rodent cells. To determine whether these mutations are necessary for maintenance of the immortalized and/or neoplastically transformed states of human bronchial epithelial cells, the p53 gene and regions of the ras (K-, H-, and N-) genes were sequenced in nine human lung carcinoma cell lines. Detection of p53 mutations by polymerase chain amplification and direct DNA sequencing was corroborated by p53 immunocytochemistry and coimmunoprecipitation of p53 with heat shock protein 70. p53 and ras genes were frequently, but not always, mutated in the carcinoma cell lines. These data are consistent with the hypothesis that multiple genetic changes involving both protooncogenes and tumor suppressor genes occur during lung carcinogenesis.

Genetic analysis of human esophageal tumors from two high incidence geographic areas: frequent p53 base substitutions and absence of ras mutations.

Esophageal squamous cell carcinoma (ESC) samples from patients residing in Uruguay and in Normandy, France, where alcoholic beverages and tobacco smoke are major risk factors, were analyzed for point mutations in the p53 tumor suppressor gene. Among 34 tumors (15 from Normandy and 19 from Uruguay) 15 point mutations in the p53 gene that result in amino acid substitutions or chain termination were identified by polymerase chain reaction amplification of exons 5-8 and direct DNA sequencing. Base substitutions in ESC from these high-incidence areas are dispersed over the midregion of the p53 gene. There are differences between ESC and other types of gastrointestinal cancer in the nature of frequent base substitutions. CpG to TpG transitions were far less prevalent in these ESC than in colorectal tumors, whereas G to T transversions, rarely found in colon cancers, were found in one-fourth of the ESC samples. Base substitutions at A:T pairs constitute an important fraction of ESC p53 mutations, in contrast to mutation patterns in most other types of solid tumors. In contrast to the frequent mutation of the p53 gene in these samples, no mutations in the H-, K-, or N-ras genes were found in 16 tumors from Uruguay by direct sequencing of exons in which transforming mutations are known to occur. A previous study on ras mutations in ESC from France was also negative (M. C. Hollstein et al., Cancer Res., 48: 5119-5123, 1988). The role of distinct etiological factors in generating these differences and the potential for linking patient exposure histories with patterns of p53 mutations in high risk populations are considered.

p53 and Kirsten-ras mutations in human mesothelioma cell lines.

Twenty cell lines from 17 individuals with malignant mesothelioma have been examined for p53 alterations by direct sequencing of genomic DNA, by evaluation of mRNA expression levels, and by immunocytochemical analysis of p53 protein expression in comparison with normal human pleural mesothelial cells. The results of this study show p53 abnormalities in cell lines from 3 individuals. These include 2 point mutations and one null cell line. Interestingly, while both cell lines with point mutations exhibit high levels of p53 protein, normal mesothelial cells as well as 12 of the mesotheliomas evaluated express low but significant levels. In addition, sequencing of K-ras at codons 12, 13, and 61 reveals wild-type sequence in all 20 mesothelioma cell lines. The capacity to induce tumors in athymic nude mice did not correlate with the presence of a p53 mutation or elevated p53 protein levels. These data suggest that neither p53 alteration nor K-ras activation constitutes a critical step in the development of human mesothelioma.

Integrity of p53 in hepatitis B x antigen-positive and -negative hepatocellular carcinomas.

Inactivation of the tumor suppressor p53 seems to be important to the pathogenesis of hepatocellular carcinoma (HCC) associated with chronic hepatitis B virus infection. Although this inactivation may be due to mutations in the p53 gene, recent evidence suggests that the hepatitis B virus-encoded X antigen (HBxAg) binds to and inactivates wild-type p53. Hence, experiments were designed to test the hypothesis that there is a low frequency of p53 mutations in HBxAg-positive HCC. HBxAg and p53 were assayed by immunohistochemistry (IHC) in HCC and nontumor liver from 16 Chinese patients, half of whom were hepatitis B surface antigen carriers. HBxAg was detectable in tumor and/or nontumor cells from all patients by IHC; six of these samples also had detectable p53. To determine whether p53 detection by IHC, and hence stabilization, is associated with mutation, sequencing of p53 exons 5-8 was performed with each patient sample. Wild-type sequences were found in 13 of 16 HBxAg-positive cases (81%). Hence, HBxAg is a common marker of HCC that correlates with the persistence of wild-type p53 among both carriers and noncarriers. The low frequency of p53 mutations in HCC in these patients implies that p53 inactivation may occur predominantly by complex formation with HBxAg.

p53 and K-ras mutations in lung cancers from former and never-smoking women.

Somatic p53 mutations are common in lung cancer. Active cigarette smoking is positively correlated with the total frequency of p53 mutations and G:C to T:A transversions on the nontranscribed (DNA coding) strand. Mutational hotspots within the p53 gene, e.g., codon 157, have been identified for tobacco-related lung cancer, whereas these same mutations are found rarely in other cancers. Such data implicate specific p53 mutations as molecular markers of smoking. Because limited data exist concerning the p53 mutation frequency and spectra in ex-smokers and nonsmokers, we have analyzed p53 and K-ras mutations in 126 lung cancers from a population-based case-control study of nonsmoking (n = 117) or ex-smoking (n = 9) women from Missouri with quantitative assessments of exposure to environmental tobacco smoke. Mutations in the p53 gene were found in lung cancers from lifetime nonsmokers (19%) and ex-smokers (67%; odds ratio, 9.08; 95% confidence interval, 2.06-39.98). All deletions were found in tumors from patients who were either ex-smokers or nonsmokers exposed to passive smoking. The G:C to A:T transitions (11 of 28; 39%) were the most frequent p53 mutations found and clustered in tumors from lifetime nonsmokers without passive smoke exposure. The incidence of K-ras codon 12 or 13 mutations was 11% (14 of 115 analyzed) with no difference between long-term ex-smokers and nonsmokers. These and other results indicate that p53 mutations occur more commonly in smokers and ex-smokers than in never-smokers. Such comparisons provide additional evidence of genetic damage caused by tobacco smoke during lung carcinogenesis.

p53 mutation and protein accumulation during multistage human esophageal carcinogenesis.

Preinvasive lesions of squamous cell carcinoma are well defined morphologically and provide a model for multistage carcinogenesis. Since alterations in the p53 tumor suppressor gene occur frequently in invasive esophageal squamous cell carcinoma, we examined a set of preinvasive lesions to investigate the timing of p53 mutation. Surgically resected tissues from nine patients with esophageal squamous cell carcinoma contained precursor lesions which had not yet invaded normal tissues. Immunohistochemistry showed high levels of p53 protein in both preinvasive lesions and invasive carcinomas in six cases; sequence analysis of all invasive tumors identified p53 missense mutations in two cases. Preinvasive lesions from both tumors with mutations plus one wild-type tumor were microdissected and sequenced. In one patient there were different mutations in the invasive carcinoma (codon 282, CGGarg > TGGtrp) and a preinvasive lesion (codon 272, GTGval > T/GTGleu/val). In a second case, an invasive carcinoma had a mutation in codon 175 (CGCarg > CAChis), and adjacent preinvasive lesions contained a wild-type sequence. A carcinoma and preinvasive lesion from the third case contained high levels of protein and a wild-type DNA sequence. Therefore, p53 mutation may precede invasion in esophageal carcinogenesis, and multifocal esophageal neoplasms may arise from independent clones of transformed cells. The timing of p53 protein accumulation is favorable for an intermediate biomarker in multistage esophageal carcinogenesis.

Alterations of p14ARF, p53, and p73 genes involved in the E2F-1-mediated apoptotic pathways in non-small cell lung carcinoma.

Overexpression of E2F-1 induces apoptosis by both a p14ARF-p53- and a p73-mediated pathway. p14ARF is the alternate tumor suppressor product of the INK4a/ARF locus that is inactivated frequently in lung carcinogenesis. Because p14ARF stabilizes p53, it has been proposed that the loss of p14ARF is functionally equivalent to a p53 mutation. We have tested this hypothesis by examining the genomic status of the unique exon 1beta of p14ARF in 53 human cell lines and 86 primary non-small cell lung carcinomas and correlated this with previously characterized alterations of p53. Homozygous deletions of p14ARF were detected in 12 of 53 (23%) cell lines and 16 of 86 (19%) primary tumors. A single cell line, but no primary tumors, harbored an intragenic mutation. The deletion of p14ARF was inversely correlated with the loss of p53 in the majority of cell lines (P = 0.02), but this relationship was not maintained among primary tumors (P = 0.5). E2F-1 can also induce p73 via a p53-independent apoptotic pathway. Although we did not observe inactivation of p73 by either mutation or DNA methylation, haploinsufficiency of p73 correlated positively with either p14ARF or p53 mutation or both (P = 0.01) in primary non-small cell lung carcinomas. These data are consistent with the current model of p14ARF and p53 interaction as a complex network rather than a simple linear pathway and indicate a possible role for an E2F-1-mediated failsafe, p53-independent, apoptotic pathway involving p73 in human lung carcinogenesis.

Archival analysis of p53 genetic and protein alterations in Chinese esophageal cancer.

A strategy and methods for archival analysis of genetic and protein alterations in the p53 tumor-suppressor gene are presented. The tumor series includes 43 paraffin-embedded esophageal carcinomas from two high-incidence regions in the People's Republic of China. More than half contained elevated p53 protein levels which were detected by a high-titer polyclonal antiserum and a sensitive immunohistochemical method. To estimate the frequency of underlying mutations, DNA was isolated from conventional paraffin sections, amplified by the polymerase chain reaction, and examined by dideoxy termination sequencing. Analysis of exons 5-8 in a subset of 10 tumors revealed mis-sense point mutations in 4 out of 5 immunostain-positive tumors and a mutation encoding a stop codon in 1 of 5 immunostain-negative tumors. In this report of archival material, we conclude that detectable levels of p53 protein correlate closely with the occurrence of mis-sense mutations. Furthermore, these methods render large repositories of paraffin-embedded tumor and non-tumor tissues accessible to analysis. Immunohistochemical screening for elevated protein levels followed by sequence analysis represents an efficient strategy for the evaluation of the p53 mutational spectrum.

Nitric oxide synthase, cyclooxygenase 2, and vascular endothelial growth factor in the angiogenesis of non-small cell lung carcinoma.

We have investigated the hypothesis that nitric oxide synthase (NOS2), cyclooxygenase-2 (COX2), and vascular endothelial growth factor (VEGF) protein levels individually demonstrate a direct correlation with microvessel density (MVD) and clinical outcome in human non-small cell lung cancer (NSCLC). Furthermore, we hypothesized that MVD may explain the propensity of certain histological lung cancer subtypes for early metastasis via a hematological route. Immunohistochemically, we studied the protein expression levels of NOS2, COX2, and VEGF and MVD by counting CD31-reactive blood vessels (BVs) in 106 surgically resected NSCLC specimens. NOS2, COX2, and VEGF immunoreactivity were observed in 48, 48, and 58%, respectively, of the study subjects, and their levels correlated with MVD at the tumor-stromal interphase (P < or = 0.001). More adenocarcindmas and large cell carcinomas displayed overexpression of NOS2 when compared with squamous cell carcinoma (SCC; r = 0.44; P < 0.001). NOS2 and COX2 levels were found to correlate positively with VEGF status (r = 0.44; P < 0.001, 0.01, and 0.03, respectively). These results attest to the significant interaction of these factors in the angiogenesis of NSCLC. Although neither angiogenic factors nor MVD correlated with patient survival, the latter correlated with tumor clinical stage in both squamous (SCC; 73 BVs/mm2) and non-SCC (78 BVs/mm2) tumors. These results indicate that angiogenesis is a complex process that involves multiple factors including NOS2, COX2, and VEGF. Furthermore, the role of angiogenesis in the biology of various histological lung cancer types may be different. The complexity of angiogenesis may explain the modest results observed in antiangiogenesis therapy that target a single protein.

p21waf1/cip1 and transforming growth factor beta 1 protein expression correlate with survival in non-small cell lung cancer.

p21waf1/cip1 encodes a cyclin-dependent kinase inhibitor that is transcriptionally activated by the p53 tumor suppressor gene, transforming growth factor beta 1 (TGF-beta 1), AP2, and other pathways. Because p21waf1/cip1, p53, and TGF-beta 1 all regulate apoptosis and the cell cycle, we tested the hypothesis that their relative protein levels would correlate with biological features including the survival of non-small cell lung cancer (NSCLC) patients. We conducted an immunohistochemical analysis of p21waf1/cip1 and TGF-beta 1 and identified four patient groups with distinct survival outcomes. Concordant p21waf1/cip1 and TGF-beta 1 expression (i.e., either high p21waf1/cip1 and high TGF-beta 1 expression or low p21waf1/cip1 and low TGF-beta 1 expression) predicted 70% disease-free survival at 2000 days of follow-up. Discordant p21waf1/cip1 and TGF-beta 1 expression (i.e., either high p21waf1/cip1 and low TGF-beta 1 expression or low p21waf1/cip1 and high TGF-beta 1 expression) predicted 35% disease-free survival (P = 0.0003; log-rank test). These survival relationships were not attributable to differences in grade, stage, or p53 status. Although current models do not fully explain these complex interactions, most of these data fit a paradigm whereby TGF-beta 1 regulation determines NSCLC survival. In addition to the survival correlation, we found that high p21waf1/cip1 protein expression correlated with high tumor grade (P = 0.014). There is little evidence that p21waf1/cip1 protein levels accurately predict p53 mutation status in NSCLC; specifically, 20 of 48 (42%) tumors with p53 mutations contained high levels of p21waf1/cip1 protein. These findings indicate that p21waf1/cip1 immunohistochemical analysis may provide useful information concerning the biological properties of NSCLC.

Sugars and adiposity: the long-term effects of consuming added and naturally occurring sugars in foods and in beverages.

OBJECTIVE: The aim of this study was to determine if the association with adiposity varies by the type (added vs. naturally occurring) and form (liquid vs. solid) of dietary sugars consumed. METHODS: Data from the 10-year National Heart, Lung, and Blood Institute (NHLBI) Growth and Health Study (n = 2,021 girls aged 9-10 years at baseline; n = 5,156 paired observations) were used. Using mixed linear models, 1-year changes in sugar intake, body mass index z-score (BMIz) and waist circumference (WC) were assessed. RESULTS: The results showed mean daily added sugar (AS) intake: 10.3 tsp (41 g) liquid; 11.6 tsp (46 g) solid and naturally occurring sugar intake: 2.6 tsp (10 g) liquid; 2.2 tsp (9 g) solid. Before total energy adjustment, each additional teaspoon of liquid AS was associated with a 0.222-mm increase in WC (p = 0.0003) and a 0.002 increase in BMIz (p = 0.003). Each teaspoon of solid AS was associated with a 0.126-mm increase in WC (p = 0.03) and a 0.001 increase in BMIz (p = 0.03). Adjusting for total energy, this association was maintained only between liquid AS and WC among all and between solid AS and WC among those overweight/obese only. There was no significant association with naturally occurring sugar. CONCLUSIONS: These findings demonstrate to suggest a positive association between AS intake (liquid and solid) and BMI that is mediated by total energy intake and an association with WC that is independent of it.

p53 is not mutated in hepatocellular carcinomas from Alaska Natives.

Hepatocellular carcinoma is common among Alaska Natives. The known risk factor in this population is hepatitis B viral infection; fungal toxins, including aflatoxin B1, have not been detected in foodstuffs. In this series of 14 patients (including 4 siblings and 2 second cousins), 3 patients were less than 12 years old at diagnosis of hepatocellular carcinoma, 8 patients were 13-24 years old, and 3 patients were more than 60 years old. Since p53 mutations occur in 29% of hepatocellular carcinomas worldwide, we tested the tumors for p53 mutations and serum samples for anti-p53 antibodies. Serum samples from these 14 patients did not contain detectable levels of anti-p53 antibodies. Loss of heterozygosity within the p53 locus was not detected in any of 9 informative cases. Immunohistochemical analysis for p53 protein accumulation was negative in all of 11 tumors. DNA sequence analysis of 12 tumor samples showed no evidence of p53 mutation in the highly conserved regions included in exons 5-8. These data, combined with one case from a previous report, indicate a mutation frequency of 0 of 13, which differs significantly from the worldwide frequency of 29% (chi 2 3.9; P = 0.048). These results indicate that liver carcinogenesis among Alaska Natives occurs independently of a traditional p53 pathway. The familial clustering and early onset in this population strongly suggest an inherited genetic predisposition to develop liver cancer. Germline mutations in a tumor suppressor or a cancer susceptibility gene are likely. Future studies of these samples should include investigations of candidate suppressor or susceptibility genes which map to chromosomal regions commonly deleted in liver cancers.

p53 mutations in lung cancer following radiation therapy for Hodgkin's disease.

High risks of lung cancer occur after successful treatment of Hodgkin's disease. In addition to tobacco smoking, other risk factors include radiotherapy, chemotherapy, and immunosuppression, although the relative contributions of each are unknown. We conducted p53 mutational spectrum analysis in second lung cancers after radiation therapy for Hodgkin's disease in the Netherlands and in Ontario, Canada. Lung cancer tissues from 11 patients were analyzed by p53 immunohistochemistry and DNA sequence analysis. All were male cigarette smokers, all received radiation therapy, and six also received chemotherapy. The lung cancers occurred 9.8 years (mean) after treatment. Radiation doses to lung lobes that developed the tumors averaged 5.7 Gy (range, 3.7-11.7 Gy). Sequence analysis showed four missense and two silent p53 point mutations in five patients. There were four G:C-->A:T transitions; three of four mutated deoxyguanines occurred on the coding strand, and one was a CpG site. There were two transversions: one G:C-->C:G and one A:T-->C:G. Despite moderate or heavy smoking histories in all patients, the mutational spectrum appears to differ from usual smoking-related lung cancers in which G:C-->T:A transversions predominate. The absence of G:C-->T:A mutations and the prominence of G:C-->A:T transitions, which are characteristic of radiation and oxidative damage, suggest that radiotherapy might have caused some of the p53 mutations. These data illustrate the potential of mutation analysis to determine causes of human cancer. If confirmed in a larger series, these results imply that some radiation-induced cancers can be distinguished from those caused by other factors.

Late onset Epstein-Barr virus-associated lymphoproliferative disease after allogeneic bone marrow transplant presenting as breast masses.

We report a patient who developed breast masses 17 months after a T cell-depleted partially mismatched related donor (PMRD) bone marrow transplant (BMT) for chronic myeloid leukemia. The patient had severe chronic graft-versus-host disease (GVHD) and the masses were due to Epstein-Barr virus (EBV) lymphoproliferative disease (LPD). The patient expired from fungal pneumonia after chemotherapy for the EBV-LPD.

Infrequent p53 mutations in arsenic-related skin lesions.

Oral arsenic exposure increases the risk for a variety of benign and malignant skin lesions, but the molecular mechanism of the carcinogenic effect is poorly understood. Arsenic-related squamous cell carcinomas of the skin can develop either de novo or progress from Bowen's disease lesions. Arsenic-related basal cell carcinomas develop usually in non-sun-exposed areas and are multiple. Because p53 tumor suppressor protein is a protective cellular molecule against environmental carcinogens and mutations in the p53 gene are frequent in nonmelanoma skin cancers, we studied p53 in 23 premalignant or malignant skin lesions from seven patients with a history of arsenic medication. The eighth patient studied (with six lesions) had a long standing exposure to UV radiation. Accumulation of the p53 protein was detected (with a monoclonal DO-7 antibody) in 78% of the lesions from cases with arsenic exposure. Two of the six (30%) arsenic-related premalignant lesions and in addition one UV related carcinoma in situ lesion were clearly and repeatedly positive when p53 exons 5 to 8 were screened by a nonradioactive single-strand conformation polymorphism (SSCP) analysis. Only one of the arsenic-related lesions was confirmed by sequencing to have a mutation (a CC to TT double transition). No indications of mutations were found among the 18 basal cell carcinoma or two squamous cell carcinoma lesions studied. Our results suggest that the frequent accumulation of p53 protein in arsenic-related skin lesions is not due to p53 mutations. which may not be a prerequisite in the development of arsenic-induced skin cancers.