Faculty Bibliography

STUDY OBJECTIVES: To evaluate volatile organic compounds (VOCs) in the breath as tumor markers in lung cancer. Alkanes and monomethylated alkanes are oxidative stress products that are excreted in the breath, the catabolism of which may be accelerated by polymorphic cytochrome p450-mixed oxidase enzymes that are induced in patients with lung cancer. DESIGN: Combined case-control and cross-sectional study. SETTING: Five academic pulmonary medicine services in the United States and the United Kingdom. Patients and participants: One hundred seventy-eight bronchoscopy patients and 41 healthy volunteers. INTERVENTION: Breath samples were analyzed by gas chromatography and mass spectroscopy to determine alveolar gradients (ie, the abundance in breath minus the abundance in room air) of C4-C20 alkanes and monomethylated alkanes. MEASUREMENTS: Patients with primary lung cancer (PLC) were compared to healthy volunteers, and a predictive model was constructed using forward stepwise discriminant analysis of the alveolar gradients. This model was cross-validated with a leave-one-out jackknife technique and was tested in two additional groups of patients who had not been used to develop the model (ie, bronchoscopy patients in whom cancer was not detected, and patients with metastatic lung cancer [MLC]). RESULTS: Eighty-seven of 178 patients had lung cancer (PLC, 67 patients; MLC, 15 patients; undetermined, 5 patients). A predictive model employing nine VOCs identified PLC with a sensitivity of 89.6% (60 of 67 patients) and a specificity of 82.9% (34 of 41 patients). On cross-validation, the sensitivity was 85.1% (57 of 67 patients) and the specificity was 80.5% (33 of 41 patients). The stratification of patients by tobacco smoking status, histologic type of cancer, and TNM stage of cancer revealed no marked effects. In the two additional tests, the model predicted MLC with a sensitivity of 66.7% (10 of 15 patients), and it classified the cancer-negative bronchoscopy patients with a specificity of 37.4% (34 of 91 patients). CONCLUSIONS: Compared to healthy volunteers, patients with PLC had abnormal breath test findings that were consistent with the accelerated catabolism of alkanes and monomethylated alkanes. A predictive model employing nine of these VOCs exhibited sufficient sensitivity and specificity to be considered as a screen for lung cancer in a high-risk population such as adult smokers

Chromium(VI) [Cr(VI)] is a ubiquitous environmental and industrial contaminant. Cr(VI) exposure is strongly associated with a higher incidence of human lung cancer, but the mechanism of Cr(VI) carcinogenicity remains unclear. Cigarette smoking has been known as the prominent cause of lung cancer, and polycyclic aromatic hydrocarbons (PAHs), the major carcinogens in cigarette smoke, have been suggested as being responsible for the initiation and development of lung cancer. It has been reported that lung cancer from workers exposed to Cr(VI) has a high percentage of G to T transversion mutations in the non-transcribed strand of the p53 gene, a hallmark of PAH-induced mutation. Cr(VI) is a weak mutagen although it can induce a high percentage of G to T transversion mutations. These results raise the possibility that Cr(VI) may enhance PAH binding at the p53 gene in lung tissue. To test this possibility, we have determined the effect of Cr(VI) exposure on benzo[a]pyrene diol epoxides (BPDE)-DNA binding at total genomic DNA level and at the p53 gene in normal human lung fibroblast cells. We found that in lung cells Cr(VI) pre-exposure does not affect the BPDE-DNA binding at the total genomic DNA level or at exons 5, 6 and 9 of the p53 gene; however, it greatly enhances BPDE-DNA binding at exons 7 and 8 of the p53 gene, especially at mutational hotspots of lung cancer: codons 248, 273 and 282 of the p53 gene. No enhancement of BPDE-DNA binding in the p53 was observed when naked genomic DNA isolated from Cr(VI)-exposed cells was modified with BPDE in vitro. These results suggest that Cr(VI) exposure may enhance chromatin structure-dependent carcinogen-DNA binding. This effect may contribute to the synergism of Cr(VI) and BPDE on mutagenesis and cell transformation, and may also contribute to the higher incidence of lung cancer in Cr(VI)-exposed populations

Tuberculosis is the seventh leading cause of morbidity and mortality in the world, with eight million cases per year. Animal and human studies demonstrate an enrichment of CD4 cells at sites of disease, with a more favorable clinical course when there is a Th1 response with the presence of gamma interferon (IFN-gamma). We previously treated patients who had multidrug-resistant tuberculosis with recombinant IFN-gamma (rIFN-gamma) in aerosol form and were able to convert smear-positive cases to smear negative with 12 treatments over 1 month. We hypothesized that rIFN-gamma would induce signal transducer and activator of transcription (STAT) and interferon regulatory factor (IRF) binding activity in alveolar macrophages (AM). AM treated in vitro showed clear upregulation of STAT-1 and IRF-1 by rIFN-gamma. STAT-1 was not activated and IRF-1 was only weakly induced after 1 day of infection by Mycobacterium tuberculosis TN913. In bronchoalveolar lavage (BAL) cells obtained from 10 of 10 tuberculosis patients 10 +/- 2 days post-antituberculosis treatment, there was no detectable STAT-1 or IRF-1 DNA-binding activity. After 4 weeks of treatment with rIFN-gamma aerosol in addition to the antituberculosis drugs, 10 of 10 patients had increased STAT-1, IRF-1, and/or IRF-9 DNA-binding activity in BAL cells from lung segments shown radiographically to be involved and in those shown to be uninvolved. Symptoms and chest radiographs improved, and amounts of macrophage inflammatory cytokines and human immunodeficiency virus type 1 (HIV-1) viral loads (in five of five HIV-1-coinfected patients) declined in the second BAL specimens. rIFN-gamma aerosol induces signal transduction and gene expression in BAL cells and should be evaluated for efficacy in a randomized, controlled clinical trial

Oncolytic replicating adenoviruses are a promising new modality for the treatment of cancer. Despite the assumed biologic advantage of continued viral replication and spread from infected to uninfected cancer cells, early clinical trials demonstrate that the efficacy of current vectors is limited. In xenograft tumor models using immune-incompetent mice, wild-type adenovirus is also rarely able to eradicate established tumors. This suggests that innate immune mechanisms may clear the virus or that barriers within the tumor prevent viral spread. The aim of this study was to evaluate the kinetics of viral distribution and spread after intratumoral injection of virus in a human tumor xenograft model. After intratumoral injection of wild-type virus, high levels of titratable virus persisted within the xenograft tumors for at least 8 weeks. Virus distribution within the tumors as determined by immunohistochemistry was patchy, and virus-infected cells appeared to be flanked by tumor necrosis and connective tissue. The close proximity of virus-infected cells to the tumor-supporting structure, which is of murine origin, was clearly demonstrated using a DNA probe that specifically hybridizes to the B1 murine DNA repeat. Importantly, although virus was cleared from the circulation 6 hr after intratumoral injection, after 4 weeks systemic spread of virus was detected. In addition, vessels of infected tumors were surrounded by necrosis and an advancing rim of virus-infected tumor cells, suggesting reinfection of the xenograft tumor through the vasculature. These data suggest that human adenoviral spread within tumor xenografts is impaired by murine tumor-supporting structures. In addition, there is evidence for continued viral replication within the tumor, with subsequent systemic dissemination and reinfection of tumors via the tumor vasculature. Despite the limitations of immune-incompetent models, an understanding of the interactions between the virus and the tumor-bearing host is important in the design of effective therapies

BACKGROUND: Mutations in ras genes are commonly found in human cancers and in animal models. Although mutations at codons 12, 13, and 61 of H-, N- and K-ras genes can activate their oncogenic function, mutations at codon 12 of K-ras are the most common mutations found among the three ras genes in human cancers. To investigate whether codon 12 of human K-ras is especially susceptible to carcinogens and/or whether carcinogen-DNA adducts at this codon are repaired less efficiently, we examined tobacco smoke carcinogen-induced DNA damage in normal human bronchial epithelial and fibroblast cells. METHODS: We used the UvrABC nuclease incision method in combination with ligation-mediated polymerase chain reaction to map the distribution of DNA adducts induced by benzo[a]pyrene diol epoxide (BPDE) and other bulky carcinogens within exons 1 and 2 in H-ras, N-ras, and K-ras. We also analyzed BPDE-DNA adduct repair efficiency in these three genes using the same method. RESULTS: Codons 12 and 14 of the K-ras gene were hotspots for carcinogen-DNA adduct formation, with little and no adduct formation at codons 13 and 61, respectively. The BPDE-DNA adducts formed at codon 14 were repaired almost twice as quickly as those formed at codon 12. There was some BPDE-DNA adduct formation at codons 12 of H-ras and N-ras, but this codon was not a hotspot. Furthermore, no substantial difference in repair rates between codon 12 and the other codons analyzed (codons 3 and 18) was observed in either the H-ras or N-ras genes. CONCLUSION: These findings link the human cancer mutational hotspot at codon 12 of K-ras to preferential DNA damage and poor repair

Gene transfer of p53 induces cell death in most cancer cells, and replication-defective adenoviral vectors expressing p53 are being evaluated in clinical trials. However, low transduction efficiency limits the efficacy of replication-defective vector systems for cancer therapy. The use of replication-competent vectors for gene delivery may have several advantages, holding the potential to multiply and spread the therapeutic agent after infection of only a few cells. However, expression of a transgene may adversely affect viral replication. We have constructed a replicating adenoviral vector (Adp53rc) that expresses high levels of p53 at a late time point in the viral life cycle and also contains a deletion of the adenoviral death protein (ADP). Adp53rc-infected cancer cells demonstrated high levels of p53 expression in parallel with the late expression pattern of the adenoviral fiber protein. p53 expression late in the viral life cycle did not impair effective virus propagation. Survival of several lung cancer cell lines was significantly diminished after infection with Adp53rc, compared with an identical p53-negative control virus. p53 expression also improved virus release and spread. Interestingly, p53 was more cytotoxic than the ADP in cancer cells but less cytotoxic than the ADP in normal cells. In conclusion, late expression of p53 from a replicating virus improves tumor cell killing and viral spread without impairing viral replication. In addition, in combination with a deletion of the ADP, specificity of tumor cell killing is improved.