Obesity and survival among women with ovarian cancer: results from the Ovarian Cancer Association Consortium.

BACKGROUND: Observational studies have reported a modest association between obesity and risk of ovarian cancer; however, whether it is also associated with survival and whether this association varies for the different histologic subtypes are not clear. We undertook an international collaborative analysis to assess the association between body mass index (BMI), assessed shortly before diagnosis, progression-free survival (PFS), ovarian cancer-specific survival and overall survival (OS) among women with invasive ovarian cancer. METHODS: We used original data from 21 studies, which included 12 390 women with ovarian carcinoma. We combined study-specific adjusted hazard ratios (HRs) using random-effects models to estimate pooled HRs (pHR). We further explored associations by histologic subtype. RESULTS: Overall, 6715 (54%) deaths occurred during follow-up. A significant OS disadvantage was observed for women who were obese (BMI: 30-34.9, pHR: 1.10 (95% confidence intervals (CIs): 0.99-1.23); BMI: ⩾35, pHR: 1.12 (95% CI: 1.01-1.25)). Results were similar for PFS and ovarian cancer-specific survival. In analyses stratified by histologic subtype, associations were strongest for women with low-grade serous (pHR: 1.12 per 5 kg m(-2)) and endometrioid subtypes (pHR: 1.08 per 5 kg m(-2)), and more modest for the high-grade serous (pHR: 1.04 per 5 kg m(-2)) subtype, but only the association with high-grade serous cancers was significant. CONCLUSIONS: Higher BMI is associated with adverse survival among the majority of women with ovarian cancer.

Mass spectrometric techniques for characterizing low-molecular-weight resins used as paint varnishes.

The molecular structure of three low-molecular-weight resins used as paint varnishes has been characterized by use of an approach based on three different mass spectrometric techniques. We investigated the ketone resin MS2A, the aldehyde resin Laropal A81, and the hydrocarbon resin Regalrez 1094, now commonly used in restoration. To date, the molecular structures of these resins have not been completely elucidated. To improve current knowledge of the chemical composition of these materials, information obtained by gas chromatography-mass spectrometry (GC/MS), pyrolysis-gas chromatography-mass spectrometry (Py/GC/MS), and electrospray ionization mass spectrometry (ESI-Q-ToF) was combined. Analysis, in solution, of the whole polymeric fraction of the resins by flow-injection ESI-Q-ToF, and of the non-polymeric fraction by GC-MS, enabled us to identify previously unreported features of the polymer structures. In addition, the Py-GC/MS profiles that we obtained will help to enhance the databases currently available in the literature. The proposed approach can be extended to other low-molecular-weight resins used as varnishes in conservation.

A round robin exercise in archaeometry: analysis of a blind sample reproducing a seventeenth century pharmaceutical ointment.

Chemical analysis of ancient residues of pharmaceutical or cosmetic preparations such as balms or ointments is made problematic by the high complexity of these mixtures, composed of organic and inorganic materials. Consequently, a multi-analytical approach and special caution in the interpretation of the results are necessary. In order to contribute to the improvement of analytical strategies for the characterization of complex residues and to reconstruct ancient medical practices, a replica of a pharmaceutical formulation of the seventeenth century was prepared in the laboratory according to a historically documented recipe. In a round robin exercise, a portion of the preparation was analysed as a blind sample by 11 laboratories using various analytical techniques. These included spectroscopic, chromatographic and mass spectrometric methods. None of the laboratories was able to completely reconstruct the complex formulation, but each of them gave partial positive results. The round robin exercise has demonstrated that the application of a multi-analytical approach can permit a complete and reliable reconstruction of the composition. Finally, on the basis of the results, an analytical protocol for the study of residues of ancient medical and pharmaceutical preparations has been outlined.

Study on the impregnation of archaeological waterlogged wood with consolidation treatments using synchrotron radiation microtomography.

In favourable conditions of low temperature and low oxygen concentration, archaeological waterlogged wooden artefacts, such as shipwrecks, can survive with a good state of preservation. Nevertheless, anaerobic bacteria can considerably degrade waterlogged wooden objects with a significant loss in polysaccharidic components. Due to these decay processes, wood porosity and water content increase under ageing. In such conditions, the conservation treatments of archaeological wooden artefacts often involve the replacement of water with substances which fill the cavities and help to prevent collapse and stress during drying. The treatments are very often expensive and technically difficult, and their effectiveness very much depends on the chemical and physical characteristics of the substances used for impregnation. Also important are the degree of cavity-filling, penetration depth and distribution in the structure of the wood. In this study, the distribution in wood cavities of some mixtures based on polyethylene glycols and colophony, used for the conservation of waterlogged archaeological wood, was investigated using synchrotron radiation X-ray computed microtomography (SR-microCT). This non-destructive imaging technique was useful for the study of the degraded waterlogged wood and enabled us to visualise the morphology of the wood and the distribution of the materials used in the wood treatments. The study has shown how deposition is strictly related to the dimension of the wooden cavities. The work is currently proceeding with the comparison of synchrotron observations with the data of the solutions viscosity and with those of the properties imparted to the wood by the treatments.

The epidemiology and risk factors of head and neck cancer: a focus on human papillomavirus.

Head and neck cancer was the eighth leading cause of cancer death worldwide in 2000. Although the incidence of head and neck squamous cell carcinoma (HNSCC) in the United States is relatively low, survival is poor and has not improved for several decades. While tobacco and alcohol are the primary risk factors for HNSCC development, epidemiological studies report a strong association with human papillomavirus (HPV) in a subset of HNSCC. More than 95% of cervical squamous cell carcinomas are linked to persistent HPV infection; evidence demonstrates that HPV is a necessary carcinogen. Not all HPV-positive HNSCC express the viral oncogenes (E6 and E7), which suggests that HPV may function as a carcinogen in a smaller proportion of HNSCC. This review presents our current understanding of the relationship between HPV and HNSCC, and describes future research directions that may lead to a better understanding of the involvement of HPV in head and neck cancer.

hMENA(11a) contributes to HER3-mediated resistance to PI3K inhibitors in HER2-overexpressing breast cancer cells.

Human Mena (hMENA), an actin regulatory protein of the ENA/VASP family, cooperates with ErbB receptor family signaling in breast cancer. It is overexpressed in high-risk preneoplastic lesions and in primary breast tumors where it correlates with HER2 overexpression and an activated status of AKT and MAPK. The concomitant overexpression of hMENA and HER2 in breast cancer patients is indicative of a worse prognosis. hMENA is expressed along with alternatively expressed isoforms, hMENA(11a) and hMENAΔv6 with opposite functions. A novel role for the epithelial-associated hMENA(11a) isoform in sustaining HER3 activation and pro-survival pathways in HER2-overexpressing breast cancer cells has been identified by reverse phase protein array and validated in vivo in a series of breast cancer tissues. As HER3 activation is crucial in mechanisms of cell resistance to PI3K inhibitors, we explored whether hMENA(11a) is involved in these resistance mechanisms. The specific hMENA(11a) depletion switched off the HER3-related pathway activated by PI3K inhibitors and impaired the nuclear accumulation of HER3 transcription factor FOXO3a induced by PI3K inhibitors, whereas PI3K inhibitors activated hMENA(11a) phosphorylation and affected its localization. At the functional level, we found that hMENA(11a) sustains cell proliferation and survival in response to PI3K inhibitor treatment, whereas hMENA(11a) silencing increases molecules involved in cancer cell apoptosis. As shown in three-dimensional cultures, hMENA(11a) contributes to resistance to PI3K inhibition because its depletion drastically reduced cell viability upon treatment with PI3K inhibitor BEZ235. Altogether, these results indicate that hMENA(11a) in HER2-overexpressing breast cancer cells sustains HER3/AKT axis activation and contributes to HER3-mediated resistance mechanisms to PI3K inhibitors. Thus, hMENA(11a) expression can be proposed as a marker of HER3 activation and resistance to PI3K inhibition therapies, to select patients who may benefit from these combined targeted treatments. hMENA(11a) activity could represent a new target for antiproliferative therapies in breast cancer.

Selective 'in synthesis' labelling of peptides by fluorochromes.

A new method is described for producing fluorescently-tagged peptides containing specific internal derivatives of lysyl residues. The technique employs the base-labile Boc-Lys(Fmoc)-COOH derivative with base-catalyzed removal of the Fmoc protecting group during peptide synthesis and subsequent fluorescent derivatization of the deprotected epsilon-amino group of lysine. By this technique, other lysine residues and the alpha-amino group of the fragment remain unmodified, which could have some value in studies where it might be required to tag a single individual lysine residue within the peptide, but not the amino terminus. In spite of the fact that poly-substituted peptides are badly soluble and might seldom find a practical application, this technique also allows the introduction of different fluorochromes at different lysyl residues within the peptide, thus obtaining double fluorescence. The method, fast and easy, requires a limited number of manual operations during the automatic synthesis of peptides. Although peptide synthesizers provided with an oscillating glass reactor are more suitable for the manual interventions described, this technique might be also adapted to the newer instruments utilizing continuous-flow columns.

Allelic variants of aromatase and the androgen and estrogen receptors: toward a multigenic model of prostate cancer risk.

PURPOSE: The purpose of this study was to determine whether polymorphisms in the CAG repeat in exon 1 of the androgen receptor (AR), two intronic restriction sites in the estrogen receptor (ESR1 XbaI and ESR1 PvuII), and an Arg264Cy5 substitution in the aromatase gene (CYP19) contribute to prostate cancer risk. EXPERIMENTAL DESIGN: A case-control study was performed with 88 Caucasian prostate cancer patients and 241 Caucasian male controls. Logistic regression models were used to assess individual and joint contributions of genotypes to prostate cancer risk. RESULTS: For single polymorphisms, only the AR repeat number was significantly related to increased prostate cancer risk [age- and body mass index (BMI)-adjusted odds ratio (OR), 1.14; 95% confidence interval (CI), 1.04-1.25], suggesting a 14% increase in risk for each missing CAG repeat. When subjects were classified as either long (> or =23 AR CAG repeats) or short (<23 repeats) carriers, a significant increase in risk was also observed (age- and BMI-adjusted OR, 1.75; 95% CI, 1.05-2.95; P = 0.04). The aromatase C/T was associated with an increase in risk of borderline significance (age- and BMI-adjusted OR, 2.50; 95% CI, 0.99-6.28). When examining the effects of two polymorphisms on prostate cancer risk, homozygosity for the ESR1 XbaI restriction site together with a longer AR was more frequent among controls (32%) than cases (18%; age- and BMI-adjusted OR, 0.39; 95% CI, 0.19-0.78). The aromatase C/C genotype together with a longer AR was also more frequent among controls (55%) than cases (41%; age- and BMI-adjusted OR, 0.51; 95% CI, 0.30-0.89). CONCLUSIONS: Estrogen and aromatase may play a role in prostate cancer. A multigenic model of prostate cancer susceptibility is also supported.

Reproductive risk factors for epithelial ovarian cancer according to histologic type and invasiveness.

PURPOSE: Differences in histology among the subtypes of epithelial ovarian tumors suggest possible differences in their etiologies. We examined reproductive risk factors for epithelial ovarian cancer according to histologic subtype and tumor invasiveness. METHODS: We conducted a population-based, case-control study of associations between reproductive risk factors and epithelial ovarian cancer in the Delaware Valley from 1994 to 1998. Cases age 20 to 69 years with a recent diagnosis of epithelial ovarian cancer (n = 767) were compared to community controls (n = 1367) frequency matched by age. RESULTS: With few exceptions, we found significant risk reduction for each histologic subtype of epithelial ovarian cancer by using oral contraceptive, bearing children, and having a tubal ligation; for each subtype, there was significant increased risk associated with a family history of the disease. There were no significant differences among histologic subtypes in the magnitude of the odds ratios for OC use, parity, breastfeeding, tubal ligation, hysterectomy, family history of breast or ovarian cancer, use of noncontraceptive estrogens, age at menarche, and age at menopause. There were also few differences between invasive and borderline tumors, except that women with borderline tumors were significantly younger than women with invasive disease (44.7 years vs. 52.0 years, p < 0.001). Among serous tumors only, women with borderline tumors were more likely to use oral contraceptives than women with invasive tumors (OR = 2.28 95% CI 1.20-4.35). CONCLUSION: The results of this study suggest that reproductive risk factors do not differ among histologic subtypes of epithelial ovarian cancers.

Specificities of rabbit antisera to multiple antigen (MAP) peptides.

Two multiple antigen peptides consisting of 6 and 7 aminoa cid residues, respectively, plus a 12-residue fragment, used as a control, all linked to a polylysine core, were used as immunogens in rabbits in order to obtain an immune response. Rabbit antisera against such polymers were then tested in ELISA against a panel of antigens in order to analyze the specificities of the resulting antibodies. The responses were different for all three immunogens, being partially or totally directed, for two of the three compounds, including the 12-residue control MAP peptide, against the polylysyl core, which is considered as non immunogenic. The third MAP polymer was practically unable to elicit an immune response.

Flexibility of amino acid residues at position four of nonapeptides enhances their binding to human leucocyte antigen (HLA) molecules.

The binding affinity of synthetic nonapeptides to human leucocyte antigens (HLA) molecules of the A0201 allotype, the most common in Caucasian, is enhanced or reduced by suitable amino acid substitutions at position 4, as a result of increased or decreased chain flexibility. A higher flexibility of the bond at this position correlates with an easier accommodation of the fragment into the HLA groove, while rigidity of the peptide chain appears to interfere. These data are based on two lines of evidence: a) most natural high affinity ligands for HLA-A0201 possess, at position 4, flexible residues b) substitutions of such residues by rigid amino acids results in a decrease of binding affinity.

Isolation, characterization and comparison of antipeptide and antiprotein rabbit antibodies to the pi-isoform of glutathione S-transferase.

The main linear epitopes of pi-glutathione transferase (pi-GST, EC 2.5.1.18), an enzyme related to cancer progression in a restricted number of tumours, were identified by testing in ELISA the reactivities of polyclonal anti-pi-GST rabbit sera against a panel of 51 overlapping decapeptides, covering the whole 216-residue sequence of the protein. Several major reactivity peaks were detected, each covering two or three adjacent peptides. The most active fragments were then reconstructed by conventional solid-phase synthesis, linked to Sepharose, and used as affinity ligands for isolating specific anti-pi-GST antibody subsets. A second group of antisera was then prepared in rabbits by using as immunogens some of the above described synthetic fragments, linked to a carrier protein, and antipeptide antibodies purified by affinity chromatography. An ELISA test was then performed, using as antigens a panel of peptides and different isoforms of GST, in order to establish whether antibodies isolated from total anti-pi-GST sera would display higher reactivity and specificity, as compared to traditional antipeptide antibodies. Binding data clearly confirm that the formers might be indeed better reagents for the detection and possibly quantitation of pi-GST.

Definition of essential amino acid residues in the recognition of a peptide by a mouse monoclonal antibody.

A mouse monoclonal antibody reacting in ELISA with a synthetic peptide representing a linear amino acid stretch of the protein antigen was tested on all overlapping 5-mer to 9-mer fragments of the peptide, as prepared by multi-pin synthesis. Analysis of the binding data suggests that several residues in the peptide might be relatively unrelevant for recognition, while few others seem to play a critical role as key residues. On the basis of such observations, we attempted to reconstruct an alternative essential epitope by introducing multiple amino acid substitutions in the 9-mer peptide exhibiting the best binding activity, and then tested its ability to be recognized by the monoclonal antibody.

Copper resinate: preparation, characterisation and study of degradation.

This paper describes a method for the synthesis of Copper Resinate, which disappeared from artists' palettes in the eighteenth century. This was carried out by interpreting ancient recipes following a scientific approach. Its characterisation using Fourier Transform-Infrared Spectrometry and Gas Chromatography-Mass Spectrometry demonstrated that it is a mixture containing copper and oxidised abietic acids, mainly dehydroabietic and 7-oxo-dehydroabietic acids, formed during the preparation of the pigment and the curing of the paint layer. The composition of copper resinate paint layers, artificially aged by U.V. irradiation at 365 nm (UV), heating (T), and exposed to atmospheric pollutants (NOX) in a climatic chamber, was investigated. The combination of irradiation and temperature produced a change in colour along with a significant increase in the recovered amount of 7-oxo-dehydroabietic acid. The identification of copper resinate in a sample from an old painting should be related to the presence of the following resin compounds which are stable in the ageing process: dehydroabietic and 7-oxo-dehydroabietic acid pimaradienic acids. Photo-oxidation of the resin acids co-ordinated with copper seem to be the most probable decay mechanism responsible for the colour change in the pigment.

Progesterone receptor variation and risk of ovarian cancer is limited to the invasive endometrioid subtype: results from the Ovarian Cancer Association Consortium pooled analysis.

There is evidence that progesterone plays a role in the aetiology of invasive epithelial ovarian cancer. Therefore, genes involved in pathways that regulate progesterone may be candidates for susceptibility to this disease. Previous studies have suggested that genetic variants in the progesterone receptor gene (PGR) may be associated with ovarian cancer risk, although results have been inconsistent. We have established an international consortium to pool resources and data from many ovarian cancer case-control studies in an effort to identify variants that influence risk. In this study, three PGR single nucleotide polymorphisms (SNPs), for which previous data have suggested they affect ovarian cancer risk, were examined. These were +331 C/T (rs10895068), PROGINS (rs1042838), and a 3' variant (rs608995). A total of 4788 ovarian cancer cases and 7614 controls from 12 case-control studies were included in this analysis. Unconditional logistic regression was used to model the association between each SNP and ovarian cancer risk and two-sided P-values are reported. Overall, risk of ovarian cancer was not associated with any of the three variants studied. However, in histopathological subtype analyses, we found a statistically significant association between risk of endometrioid ovarian cancer and the PROGINS allele (n=651, OR=1.17, 95% CI=1.01-1.36, P=0.036). We also observed borderline evidence of an association between risk of endometrioid ovarian cancer and the +331C/T variant (n=725 cases; OR=0.80, 95% CI 0.62-1.04, P=0.100). These data suggest that while these three variants in the PGR are not associated with ovarian cancer overall, the PROGINS variant may play a modest role in risk of endometrioid ovarian cancer.

11q13 amplification status and human papillomavirus in relation to p16 expression defines two distinct etiologies of head and neck tumours.

Two distinct etiologies of head and neck squamous cell carcinoma (HNSCC) have been proposed, DNA damage owing to tobacco and alcohol exposure and human papillomavirus (HPV) oncogene-mediated transformation. Common genetic alterations in HNSCC include TP53 mutations, 11q13 amplification (amp) and CDKN2A/p16 mutations or promoter methlyation. However, in HPV+ HNSCC it is frequent to observe wild-type TP53 and expression of p16. The relationship of this unusual pattern with 11q13 amp has not been tested. In a retrospective study on 125 HNSCC patients, only 17% (five out of 30) of HPV+ vs 44% (39 out of 89) of HPV - tumours expressed 11q13 amp (adjusted odds ratio (OR)=0.2, 95% confidence interval (CI)=0.1-0.6). A subpopulation of tumours (n=69) were classified according to the three molecular markers, TP53, p16 and 11q13 amp. In addition to wild-type TP53, and p16 expression, HPV+ tumours were more likely not to be amplified at 11q13 (OR=6.5, 95% CI=1.8-23.9). As HPV+ HNSCC lack the genetic alterations which are common in other tumours, we hypothesise that HPV infection may represent an early event in the HNSCC carcinogenic process, thus suggesting a distinct molecular pathway.

Aims and limitations in the use of antipeptide antibodies in molecular biology.

Antibodies to peptides obtained by synthesis and, to a much lesser extent, prepared by enzymatic digestion of proteins, have been widely used in the last ten years in a variety of immunochemical and biological investigations. There are however several limitations in the correct utilization of such reagents. In fact, in spite of their 'predetermined specificity', antipeptide antibodies often fail to discriminate related molecules, and their reactivity with native proteins may be scarce or even absent, even if the peptide has been selected from surface regions of the protein. Our critical point of view, concerning two main aspects of antipeptide antibody features, i.e. specificity and reactivity, will be presented here, as confronted with information from the available literature. We have selected a restricted number of references among hundreds of publications dealing with antipeptide antibodies: for sure we neglected outstanding papers on the subject, and we apologize in advance.

Reproducibility in the assessment of postmenopausal ovaries with transvaginal ultrasound.

OBJECTIVE: The goal of this study was to evaluate the interexaminer variation in the assessment of postmenopausal ovaries using transvaginal ultrasound (TVU). METHODS: One hundred eighty-eight cancer screening trial participants undergoing TVU were reassessed by a second TVU examination. RESULTS: Although first examiners tended to describe significantly larger left (P < 0.001) but smaller right (P = 0.036) ovaries, as well as fewer ovarian abnormalities, examiners agreed on the test interpretation 93% of the time (kappa = 0.846). In only two cases (1%) were the differences in interpretation such that the two examiners recommended different follow-up procedures. CONCLUSIONS: Because of the high fatality rate of ovarian cancer, early detection remains the best way to combat this devastating disease. TVU is one screening technique we are currently evaluating in a cancer screening trial. To help ensure screening test reproducibility, we have followed explcit protocols for training and certifying all TVU examiners, as well as for conducting TVU examinations. This study demonstrates that by adhering to specific training, certification, and examination protocols, TVU reproducibility is excellent. Such protocols may well serve as a standard for TVU training and examination.

Interferon-gamma (IFN-gamma) can counteract the in vitro inhibitory effect of an HIV p24 immunosuppressive heptapeptide.

Previous work from our laboratory demonstrated that a synthetic heptapeptide (Ch7), corresponding to a conserved sequence of HIV core protein p24 (aa 232-238), was able to specifically abrogate antigen-induced responses in cultures of normal human peripheral blood lymphocytes (PBL). In the present study we show that Ch7 did not inhibit the induction of IFN-gamma-secreting cells nor the accumulation of IFN-gamma mRNA in antigen-stimulated cultures. However, delayed addition of recombinant human IFN-gamma to Ch7-suppressed cultures was able to restore fully the capacity to mount an antigen-specific antibody response. Thus, although the Ch7 immunosuppressive effect may not be directly related to a decreased production of IFN-gamma, an increased level of this cytokine is certainly able to counteract the negative effect of the peptide.

MHC-peptide binding: dimers of cysteine-containing nonapeptides bind with high affinity to HLA-A2.1 class I molecules.

Small peptides, 8-10 amino acids long, derived from degradation of cytoplasmic proteins by a proteasome-proteinase complex, are usually presented and recognized by CD8+ cytolytic T lymphocytes (CTLs) associated with major histocompatibility complex (MHC) class I molecules. Recently synthetic peptides were used for the in vitro induction of tumor-specific CTLs, offering another strategy in the study of the immune-response repertoire and providing a new tool in cancer vaccination and immunotherapy. Peptides derived from otherwise normal proteins, overexpressed in many tumors as products of the protooncogene, may represent a target for an immune response. This is the case of HER-2/neu gene (also known as ErbB-2), encoding a cysteine-rich glycoprotein transmembrane receptor with tyrosine kinase activity (gp185neu). Recent data, demonstrating that HLA-A2.1-related peptides are able to stimulate in vitro CD8+ lymphocytes, Prompted us to study the binding to HLA-A2.1 molecules of several gp185 synthetic peptides containing a cystein residue and to define the relevance of this amino acid residue in the reduced or oxidated form of the sulfhydryl group. We found that monomers and their homodimers, linked by a disulfide bridge, bind to HLA-A2.1 molecules with overlapping affinity. These results suggest that additional amino acids of the nonapeptide do not prevent the binding and the HLA refolding through chemical or sterical interactions. This might be of particular relevance for the in vivo processing of cysteine-rich proteins. Because ErbB-2 molecules, as tumor-differentiation antigens in melanoma, are cysteine-rich molecules, it may be relevant to evaluate the possible role of the cystine residues interacting with the T-cell receptor. The recognition of these heterodimers by CD8+ lymphocytes will require functional in vivo studies.