S. Bader

BACKGROUND: The recently identified RASSF1 locus is located within a 120-kilobase region of chromosome 3p21.3 that frequently undergoes allele loss in lung and breast cancers. We explored the hypothesis that RASSF1 encodes a tumor suppressor gene for lung and breast cancers. METHODS: We assessed expression of two RASSF1 gene products, RASSF1A and RASSF1C, and the methylation status of their respective promoters in 27 non-small-cell lung cancer (NSCLC) cell lines, in 107 resected NSCLCs, in 47 small-cell lung cancer (SCLC) cell lines, in 22 breast cancer cell lines, in 39 resected breast cancers, in 104 nonmalignant lung samples, and in three breast and lung epithelial cultures. We also transfected a lung cancer cell line that lacks RASSF1A expression with vectors containing RASSF1A complementary DNA to determine whether exogenous expression of RASSF1A would affect in vitro growth and in vivo tumorigenicity of this cell line. All statistical tests were two-sided. RESULTS: RASSF1A messenger RNA was expressed in nonmalignant epithelial cultures but not in 100% of the SCLC, in 65% of the NSCLC, or in 60% of the breast cancer lines. By contrast, RASSF1C was expressed in all nonmalignant cell cultures and in nearly all cancer cell lines. RASSF1A promoter hypermethylation was detected in 100% of SCLC, in 63% of NSCLC, in 64% of breast cancer lines, in 30% of primary NSCLCs, and in 49% of primary breast tumors but in none of the nonmalignant lung tissues. RASSF1A promoter hypermethylation in resected NSCLCs was associated with impaired patient survival (P =.046). Exogenous expression of RASSF1A in a cell line lacking expression decreased in vitro colony formation and in vivo tumorigenicity. CONCLUSION: RASSF1A is a potential tumor suppressor gene that undergoes epigenetic inactivation in lung and breast cancers through hypermethylation of its promoter region.

BACKGROUND: Analysis of exhaled breath, especially of volatile organic compounds (VOCs), is of increasing interest in the diagnosis of lung cancer. Compared with other methods of breath analysis, ion mobility spectrometry (IMS) offers a tenfold higher detection rate of VOCs. By coupling the ion mobility spectrometer with a multicapillary column as a pre-separation unit, IMS offers the advantage of an immediate twofold separation of VOCs with visualisation in a three-dimensional chromatogram. The total analysis time is about 500 s compared with gas chromatography/mass spectrometry (GC/MS) of about 1 h. It therefore seemed reasonable to test IMS in breath analysis. METHODS: In a pilot study, 32 patients with lung cancer were subjected to a breath analysis by IMS. Their IMS chromatograms were compared with those of 54 healthy controls. An IMS that was built for special clinical application was used to identify characteristic peaks of VOCs which might be relevant for the diagnosis of lung cancer in exhaled air of 10 ml volume. RESULTS: By a combination of 23 peak regions within the IMS chromatogram, patients with lung cancer, including a patient with carcinoma in situ, were classified and differentiated from healthy persons with an error rate of zero. CONCLUSION: Breath analysis by IMS can detect a discriminating combination of VOCs in patients with lung cancer. By pattern recognition without the need for chemical analysis of the underlying VOCs, IMS has the potential to facilitate lung cancer diagnosis.

Semaphorins and collapsins make up a family of conserved genes that encode nerve growth cone guidance signals. We have identified two additional members of the human semaphorin family [human semaphorin A(V) and human semaphorin IV] in chromosome region 3p21.3, where several small cell lung cancer (SCLC) cell lines exhibit homozygous deletions indicative of a tumor suppressor gene. Human semaphorin A(V) has 86% amino acid homology with murine semaphorin A, whereas semaphorin IV is most closely related to murine semaphorin E, with 50% homology. These semaphorin genes are approximately 70 kb apart flanking two GTP-binding protein genes, GNAI-2 and GNAT-1. In contrast, other human semaphorin gene sequences (human semaphorin III and homologues of murine semaphorins B and C) are not located on chromosome 3. Human semaphorin A(V) is translated in vitro into a 90-kDa protein, which accumulates at the endoplasmic reticulum. The human semaphorin A(V) (3.4-kb mRNA) and IV (3.9- and 2.9-kb mRNAs) genes are expressed abundantly but differentially in a variety of human neural and nonneural tissues. Human semaphorin A(V) was expressed in only 1 out of 23 SCLCs and 7 out of 16 non-SCLCs, whereas semaphorin IV was expressed in 19 out of 23 SCLCs and 13 out of 16 non-SCLCs. Mutational analysis in semaphorin A(V) revealed mutations (germ line in one case) in 3 of 40 lung cancers. Our data suggest the need to determine the function of human semaphorins A(V) and IV in nonneural tissues and their role in the pathogenesis of lung cancer.

A feasibility study using an ion mobility spectrometer coupled with a multi-capillary column (MCC) was started to identify characteristic peaks of volatile compounds in exhaled human breath samples of 10 ml volume. The breath of 20 patients with sarcoidosis and suspicion of sarcoidosis because of mediastinal lymph node enlargement was investigated. Using a set of procedures for data processing and scoring a sector of interest was determined within the IMS-chromatogram. It could be shown that a procedure related to a single peak in the IMS-chromatogram delivers differentiation into the two groups of patients with confirmed sarcoidosis and those suffering no sarcoidosis. The potential biomarker is characterized by the following parameters: inverse mobility (1/K(0)) 0.53 ± 0.01 V s cm(-2)-retention time 22 ± 5 s. These results are a first step in breath analysis by MCC/IMS in patients with sarcoidosis by an automated procedure applied to IMS-chromatograms directly.

The MBD4 gene is involved in the repair of mutation at methyl-CpG dinucleotides. In microsatellite unstable tumours MBD4 can itself be mutated at an exonic polynucleotide tract. By analysing DNA from microdissected tumour samples we have found that both frequency and pattern of mutation are more significant than originally reported. © 2000 Cancer Research Campaign http://www.bjcancer.com