Mohammed A. Khan

Molecular epidemiology of human cancer risk: gene-environment interactions and p53 mutation spectrum in human lung cancer.

The p53 tumour suppressor gene is at the crossroads of a network of cellular pathways including cell cycle checkpoints, DNA repair, chromosomal segregation, and apoptosis. These pathways have evolved to maintain the stability of the genome during cellular stress from DNA damage, hypoxia, and activated oncogenes. The high frequency of p53 mutations in human cancer is a reflection of the importance of p53 involvement in this network of pathways during human carcinogenesis. An electronic database containing p53 mutations from more than 9000 cancers (http://www.iarc.fr/p53/homepage.html) can be used to generate hypotheses for further clinical, epidemiological, and laboratory investigations. For example, one can hypothesize that (a)p53 mutations vary in their pathobiological significance; (b) cellular content influences the selection of p53 mutations in clonally derived cancers; (c) the location and type of mutation within the p53 gene provide clues to functional domains in the gene product; and (d) the p53 mutation spectrum can be a molecular link between aetiological agents and human cancer. This review will focus on the role of p53 and cancer susceptibility genes in the molecular pathogenesis and epidemiology of human lung cancer. Copyright

Common Genetic Variation in TP53 Is Associated with Lung Cancer Risk and Prognosis in African Americans and Somatic Mutations in Lung Tumors

Lung cancer is primarily caused by tobacco smoking, but susceptibility is likely modified by common genetic variation. In response to many forms of cellular stress, including DNA damage, the p53 protein functions to induce cell cycle arrest, DNA repair, senescence, or apoptosis. We hypothesized that common TP53 haplotypes modulate pathways of lung carcinogenesis and lung cancer susceptibility or prognosis. To investigate our hypothesis, 14 polymorphisms in TP53, including haplotype tagging and coding single nucleotide polymorphisms, were genotyped in two studies from the greater Baltimore, Maryland area. One study is a case-control study and the second is a case-only study for which TP53 mutational spectra data are available. African Americans with Pro-T-A-G-G haplotypes of the combined TP53 polymorphisms TP53_01 (rs1042522), TP53_65 (rs9895829), TP53_66 (rs2909430), TP53_16 (rs1625895), and TP53_11 (rs12951053) had both an increased risk for lung cancer (odds ratio, 2.32; 95% confidence interval, 1.18-4.57) and a worsened lung cancer prognosis (hazards ratio, 2.38; 95% confidence interval, 1.38-4.10) compared with those with Arg-T-A-G-T haplotypes. No associations of TP53 polymorphisms with lung cancer were observed in Caucasians. In the case-only study, several polymorphisms in TP53 and TP53 haplotypes, overlapping regions of TP53 associated with risk and prognosis in African Americans, were associated with increased odds of somatic TP53 mutation in lung tumors in Caucasians. In conclusion, common genetic variation in TP53 could modulate lung cancer pathways, as suggested by the association with lung cancer in African Americans and somatic TP53 mutation frequency in lung tumors. (Cancer Epidemiol Biomarkers Prev 2007;16(2):214–22)

Chronic Inflammation Promotes Retinoblastoma Protein Hyperphosphorylation and E2F1 Activation

Chronic inflammation contributes to tumorigenesis. The retinoblastoma protein (pRb), in its hyperphosphorylated form, releases E2 promoter binding factor-1 (E2F1), which drives cell proliferation. Here, we show that pRb is hyperphosphorylated in both mouse and human colitis. In turn, pRb hyperphosphorylation is associated with release of E2F1 from pRb, resulting in the activation of E2F1 target molecules involved in proliferation and apoptosis. These observations provide insight into the in vivo mechanisms associated with chronic colon inflammation and increased colon cancer risk.

Macrophages, nitric oxide and microRNAs are associated with DNA damage response pathway and senescence in inflammatory bowel disease.

Background Cellular senescence can be a functional barrier to carcinogenesis. We hypothesized that inflammation modulates carcinogenesis through senescence and DNA damage response (DDR). We examined the association between senescence and DDR with macrophage levels in inflammatory bowel disease (IBD). In vitro experiments tested the ability of macrophages to induce senescence in primary cells. Inflammation modulating microRNAs were identified in senescence colon tissue for further investigation. Methodology/Principal Findings Quantitative immunohistochemistry identified protein expression by colon cell type. Increased cellular senescence (HP1γ; P = 0.01) or DDR (γH2A.X; P = 0.031, phospho-Chk2, P = 0.014) was associated with high macrophage infiltration in UC. Co-culture with macrophages (ANA-1) induced senescence in >80% of primary cells (fibroblasts MRC5, WI38), illustrating that macrophages induce senescence. Interestingly, macrophage-induced senescence was partly dependent on nitric oxide synthase, and clinically relevant NO• levels alone induced senescence. NO• induced DDR in vitro, as detected by immunofluorescence. In contrast to UC, we noted in Crohn’s disease (CD) that senescence (HP1γ; P<0.001) and DDR (γH2A.X; P<0.05, phospho-Chk2; P<0.001) were higher, and macrophages were not associated with senescence. We hypothesize that nitric oxide may modulate senescence in CD; epithelial cells of CD had higher levels of NOS2 expression than in UC (P = 0.001). Microarrays and quantitative-PCR identified miR-21 expression associated with macrophage infiltration and NOS2 expression. Conclusions Senescence was observed in IBD with senescence-associated β-galactosidase and HP1γ. Macrophages were associated with senescence and DDR in UC, and in vitro experiments with primary human cells showed that macrophages induce senescence, partly through NO•, and that NO• can induce DDR associated with senescence. Future experiments will investigate the role of NO• and miR-21 in senescence. This is the first study to implicate macrophages and nitrosative stress in a direct effect on senescence and DDR, which is relevant to many diseases of inflammation, cancer, and aging.

p53 mutation spectrum in relation to GSTM1, CYP1A1 and CYP2E1 in surgically treated patients with non-small cell lung cancer

p53 mutation status was analysed in relation to DNA polymorphisms of GSTM1, CYP1A1 and CYP2E1 from 105 surgically resected non-small cell lung cancer cases. Demographic factors, smoking, occupation, family history, tumour histology, grade and stage were taken into account. p53 mutations, detected either directly by DNA sequencing (P = 0.04, adjusted for smoking) or indirectly by immunostaining (P = 0.06), were overrepresented among CYP1A1 variants. Mutations in exon 8 and transitions at CpG sites in the p53 gene were favoured in this subset. There was no relation between the individual gene polymorphisms or p53 mutations and disease-free survival by Kaplan-Meier analysis. The finding of excess CYP1A1 heterozygotes in individuals with p53 mutations after adjustment for smoking suggests that CYP1A1 activation contributes to lung cancer via p53 inactivation.

Polymerization of soy protein digests by microbial transglutaminase for improvement of the functional properties

Abstract The protease- and acid-treated soy proteins were polymerized by microbial transglutaminase (TGase) in order to improve their functional properties. Although the protease digests and acid hydrolysates were considerably insoluble, the soy protein digests or hydrolysates polymerized by TGase were soluble, despite being composed of higher molecular weight fractions ((11.8–99.4) × 10 6 ) compared to that of the native soy protein (0.48 × 10 6 ). The surface hydrophobicity of the polymerized proteins was greatly decreased, compared to that of the protease digests and acid hydrolysates, suggesting that the exposed hydrophobic residues of the polymerized peptides were buried inside the polymerized molecules. The emulsifying properties of the polymerized soy proteins were greatly improved compared to those of the untreated, protease- or acid-treated proteins. The foaming properties of the polymerized soy proteins were also improved. The bitterness of the protease digests and acid hydrolysates of soy proteins was diminished by the polymerization with TGase treatment.

Influences of Chronic Mild Stress Exposure on Motor, Non-Motor Impairments and Neurochemical Variables in Specific Brain Areas of MPTP/Probenecid Induced Neurotoxicity in Mice

Parkinsons disease (PD) is regarded as a movement disorder mainly affecting the elderly population and occurs due to progressive loss of dopaminergic (DAergic) neurons in nigrostriatal pathway. Patients suffer from non-motor symptoms (NMS) such as depression, anxiety, fatigue and sleep disorders, which are not well focussed in PD research. Depression in PD is a predominant /complex symptom and its pathology lies exterior to the nigrostriatal system. The main aim of this study is to explore the causative or progressive effect of chronic mild stress (CMS), a paradigm developed as an animal model of depression in1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (25 mg/kg. body wt.) with probenecid (250 mg/kg, s.c.) (MPTP/p) induced mice model of PD. After ten i.p. injections (once in 3.5 days for 5 weeks) of MPTP/p or exposure to CMS for 4 weeks, the behavioural (motor and non-motor) impairments, levels and expressions of dopamine (DA), serotonin (5-HT), DAergic markers such as tyrosine hydroxylase (TH), dopamine transporter (DAT), vesicular monoamine transporters—2 (VMAT 2) and α-synuclein in nigrostriatal (striatum (ST) and substantia nigra (SN)) and extra-nigrostriatal (hippocampus, cortex and cerebellum) tissues were analysed. Significantly decreased DA and 5-HT levels, TH, DAT and VMAT 2 expressions and increased motor deficits, anhedonia-like behaviour and α-synuclein expression were found in MPTP/p treated mice. Pre and/or post exposure of CMS to MPTP/p mice further enhanced the MPTP/p induced DA and 5-HT depletion, behaviour abnormalities and protein expressions. Our results could strongly confirm that the exposure of stress after MPTP/p injections worsens the symptoms and neurochemicals status of PD.

An analysis of genetic factors related to risk of inflammatory bowel disease and colon cancer

BACKGROUND AND AIMS Patients with inflammatory bowel disease (IBD) have a higher risk of developing colorectal cancer than the general population. Genome-wide association studies have identified and replicated several loci associated with risk of IBD; however, it is currently unknown whether these loci are also associated with colon cancer risk. METHODS We selected 15 validated SNPs associated with risk of either Crohns disease, ulcerative colitis, or both in previous GWAS and tested whether these loci were also associated with colon cancer risk in a two-stage study design. RESULTS We found that rs744166 in STAT3 was associated with colon cancer risk in two studies; however, the direction of the observation was reversed in TP53 mutant tumors possibly due to a nullification of the effect by mutant p53. The SNP, which lies within intron 1 of the STAT3 gene, was associated with lower expression of STAT3 mRNA in TP53 wild-type, but not mutant, tumors. CONCLUSIONS These data suggest that the STAT3 locus is associated with both IBD and cancer. Further understanding the function of this variant in relation to TP53 could possibly explain the role of this gene in autoimmunity and cancer. Furthermore, an analysis of this locus, specifically in a population with IBD, could help to resolve the relationship between this SNP and cancer.

Immobilization with Atrophy Induces De Novo Expression of Neuronal Nicotinic α7 Acetylcholine Receptors in Muscle Contributing to Neurotransmission

Background:Mature acetylcholine receptor (AChR) isoform normally mediates muscle contraction. The hypothesis that &agr;7AChRs up-regulate during immobilization and contribute to neurotransmission was tested pharmacologically using specific blockers to mature (waglerin-1), immature (&agr;A-OIVA), and &agr;7AChRs (methyllycaconitine), and nonspecific muscle AChR antagonist, &agr;-bungarotoxin. Methods:Mice were immobilized; contralateral limbs were controls. Fourteen days later, anesthetized mice were mechanically ventilated. Nerve-stimulated tibialis muscle contractions on both sides were recorded, and blockers enumerated above sequentially administered via jugular vein. Data are mean ± standard error. Results:Immobilization (N = 7) induced tibialis muscle atrophy (40.6 ± 2.8 vs. 52.1 ± 2.0 mg; P < 0.01) and decrease of twitch tension (34.8 ± 1.1 vs. 42.9 ± 1.5 g; P < 0.01). Waglerin-1 (0.3 ± 0.05 &mgr;g/g) significantly (P = 0.001; N = 9) depressed twitch tension on contralateral (≥97%) versus immobilized side (approximately 45%). Additional waglerin-1 (total dose 1.06 ± 0.12 &mgr;g/g or approximately 15.0 × ED50 in normals) could not depress twitch of 80% or greater on immobilized side. Immature AChR blocker, &agr;A-OIVA (17.0 ± 0.25 &mgr;g/g) did not change tension bilaterally. Administration of &agr;-bungarotoxin (N = 4) or methyllycaconitine (N = 3) caused 96% or greater suppression of the remaining twitch tension on immobilized side. Methyllycaconitine, administered first (N = 3), caused equipotent inhibition by waglerin-1 on both sides. Protein expression of &agr;7AChRs was significantly (N = 3; P < 0.01) increased on the immobilized side. Conclusions:Ineffectiveness of waglerin-1 suggests that the twitch tension during immobilization is maintained by receptors other than mature AChRs. Because &agr;A-OIVA caused no neuromuscular changes, it can be concluded that immature AChRs contribute minimally to neurotransmission. During immobilization approximately 20% of twitch tension is maintained by up-regulation of &agr;-bungarotoxin- and methyllycaconitine-sensitive &agr;7AChRs.

TP53 Mutation Spectrum in Lung Cancer Is Not Different in Women and Men

Whether women are more susceptible to lung cancer than men has been controversial. Several case-control studies suggested that women have greater risk of lung cancer compared with men at similar levels of cigarette smoking, whereas some large cohort studies failed to observe this association. Other studies indicated that lung cancer may have biological characteristics and mechanisms of carcinogenesis that are gender specific. Therefore, we hypothesized that women are more susceptible to the carcinogenic effects of tobacco smoke exposure, as evidenced by a higher frequency of G:C-to-T:A somatic mutations in tumors from women in comparison with men at similar levels of tobacco smoke exposure. To investigate our hypothesis, we examined the TP53 mutational spectrum in a case-only (102 women and 201 men) series study where complete smoking information was available. A similar frequency and type of somatic TP53 mutations were observed in women and men. In conclusion, our study indicates that the TP53 mutation spectrum is similar in women and men. Our results are consistent with a recent large cohort study and summary of previous cohort studies, suggesting that women likely have equivalent susceptibility to lung cancer as men.