Loss of Heterozygosity on Chromosomes 9q and 16p in Atypical Adenomatous Hyperplasia Concomitant with Adenocarcinoma of the Lung
2001; Elsevier BV; Volume: 159; Issue: 5 Linguagem: Inglês
10.1016/s0002-9440(10)63041-6
ISSN1525-2191
AutoresKazuya Takamochi, Tsutomu Ogura, Kenji Suzuki, Hidenori Kawasaki, Yukiko Kurashima, Tomoyuki Yokose, Atsushi Ochiai, Kanji Nagai, Y Nishiwaki, Hiroyasu Esumi,
Tópico(s)Medical Imaging and Pathology Studies
ResumoAtypical adenomatous hyperplasia (AAH) has recently been implicated as a precursor to lung adenocarcinoma. We previously reported loss of heterozygosity (LOH) in tuberous sclerosis (TSC) gene-associated regions to frequently be observed in lung adenocarcinoma with multiple AAHs. In this study, we analyzed LOH in four microsatellite loci on 9q, including the TSC1 gene-associated region, and four loci on 16p, including theTSC2 gene-associated region, in both 18 AAHs and 17 concomitant lung adenocarcinomas from 11 patients. Seven of 18 (39%) AAHs and 9 of 17 (53%) adenocarcinomas displayed LOH on 9q. Five (28%) AAHs and seven (41%) adenocarcinomas harbored LOH at loci adjacent to the TSC1 gene. Four of 18 (22%) AAHs and 6 of 17 (35%) adenocarcinomas displayed LOH on 16p. One (6%) AAH and five (29%) adenocarcinomas harbored LOH at loci adjacent to theTSC2 gene. These findings may indicate a causal relationship of LOH on 9q and 16p in a fraction of AAH lesions and adenocarcinomas of the lung. Especially, the frequencies of LOH on 9q and at the TSC1 gene-associated region were high. The TSC1 gene or another neighboring tumor suppressor gene on 9q might be involved in an early stage of the pathogenesis of lung adenocarcinoma. Atypical adenomatous hyperplasia (AAH) has recently been implicated as a precursor to lung adenocarcinoma. We previously reported loss of heterozygosity (LOH) in tuberous sclerosis (TSC) gene-associated regions to frequently be observed in lung adenocarcinoma with multiple AAHs. In this study, we analyzed LOH in four microsatellite loci on 9q, including the TSC1 gene-associated region, and four loci on 16p, including theTSC2 gene-associated region, in both 18 AAHs and 17 concomitant lung adenocarcinomas from 11 patients. Seven of 18 (39%) AAHs and 9 of 17 (53%) adenocarcinomas displayed LOH on 9q. Five (28%) AAHs and seven (41%) adenocarcinomas harbored LOH at loci adjacent to the TSC1 gene. Four of 18 (22%) AAHs and 6 of 17 (35%) adenocarcinomas displayed LOH on 16p. One (6%) AAH and five (29%) adenocarcinomas harbored LOH at loci adjacent to theTSC2 gene. These findings may indicate a causal relationship of LOH on 9q and 16p in a fraction of AAH lesions and adenocarcinomas of the lung. Especially, the frequencies of LOH on 9q and at the TSC1 gene-associated region were high. The TSC1 gene or another neighboring tumor suppressor gene on 9q might be involved in an early stage of the pathogenesis of lung adenocarcinoma. Carcinogenesis is a multistep process that results from an accumulation of genetic alterations in oncogenes and tumor suppressor genes. It is reasonable to regard each preneoplastic lesion as possibly having a characteristic genetic change and it is essential to investigate the biological features of preneoplastic lesions to elucidate the pathogenesis of carcinomas. In lung cancers, squamous dysplasia has long been recognized as a preneoplastic lesion of squamous cell carcinoma.1Sozzi G Miozzo M Donghi R Pilotti S Cariani CT Pastorino U Della Porta G Pierotti MA Deletions of 17p and p53 mutations in preneoplastic lesions of the lung.Cancer Res. 1992; 52: 6079-6082PubMed Google Scholar, 2Bennett WP Colby TV Travis WD Borkowski A Jones RT Lane DP Metcalf RA Samet JM Takeshima Y Gu JR Vähäkangas KH Soini Y Pääkkö P Welsh JA Trump BF Harris CC p53 protein accumulates frequently in early bronchial neoplasia.Cancer Res. 1993; 53: 4817-4822PubMed Google Scholar However, the etiology of adenocarcinoma, one of the major histological types of lung cancer, is not well understood. Several genetic alterations in atypical adenomatous hyperplasia (AAH), such as K-ras orp53 mutations or loss of heterozygosity (LOH) on chromosomes 3p, 9p, or 17p, have been reported.3Hung J Kishimoto Y Sugio K Virmani A McIntire DD Minna JD Gazdar AF Allele-specific chromosome 3p deletions occur at an early stage in the pathogenesis of lung carcinoma.JAMA. 1995; 273: 558-563Crossref PubMed Scopus (306) Google Scholar, 4Kishimoto Y Sugio K Hung JY Virmani AK McIntire DD Minna JD Gazdar AF Allele-specific loss in chromosome 9p loci in preneoplastic lesions accompanying non-small-cell lung cancers.J Natl Cancer Inst. 1995; 87: 1224-1229Crossref PubMed Scopus (203) Google Scholar, 5Kitaguchi S Takeshima Y Nishisaka T Inai K Proliferative activity, p53 expression and loss of heterozygosity on 3p, 9p and 17p in atypical adenomatous hyperplasia of the lung.Hiroshima J Med Sci. 1998; 47: 17-25PubMed Google Scholar, 6Kohno H Hiroshima K Toyozaki T Fujisawa T Ohwada H p53 mutation and allelic loss of chromosome 3p, 9p of preneoplastic lesions in patients with nonsmall cell lung carcinoma.Cancer. 1999; 85: 341-347Crossref PubMed Scopus (69) Google Scholar, 7Cooper CA Carby FA Bubb VJ Lamb D Kerr KM Wyllie AH The pattern of K-ras mutation in pulmonary adenocarcinoma defines a new pathway of tumour development in the human lung.J Pathol. 1997; 181: 401-404Crossref PubMed Scopus (76) Google Scholar, 8Westra WH Baas IO Hruban RH Askin FB Wilson K Offerhaus GJ Slebos RJ K-ras oncogene activation in atypical alveolar hyperplasias of the human lung.Cancer Res. 1996; 56: 2224-2228PubMed Google Scholar, 9Sagawa M Saito Y Fujimura S Linnoila RI K-ras point mutation occurs in the early stage of carcinogenesis in lung cancer.Br J Cancer. 1998; 77: 720-723Crossref PubMed Scopus (46) Google Scholar, 10Ohshima S Shimizu Y Takahama M Detection of c-Ki-ras gene mutation in paraffin sections of adenocarcinoma and atypical bronchioloalveolar cell hyperplasia of human lung.Virchows Arch. 1994; 424: 129-134PubMed Google Scholar These genetic abnormalities and other immunohistochemical or morphometric abnormalities in AAH overlap with those of adenocarcinomas.11Kitamura H Kameda Y Ito T Hayashi H Nakamura N Nakatani Y Inayama Y Kanisawa M Cytodifferentiation of atypical adenomatous hyperplasia and bronchioloalveolar lung carcinoma: immunohistochemical and ultrastructural studies.Virchows Arch. 1997; 431: 415-424Crossref PubMed Scopus (41) Google Scholar, 12Kitamura H Kameda Y Nakamura N Inayama Y Nakatani Y Shibagaki T Ito T Hayashi H Kimura H Kanisawa M Atypical adenomatous hyperplasia and bronchoalveolar lung carcinoma. Analysis by morphometry and the expressions of p53 and carcinoembryonic antigen.Am J Surg Pathol. 1996; 20: 553-562Crossref PubMed Scopus (132) Google Scholar, 13Kodama T Biyajima S Watanabe S Shimosato Y Morphometric study of adenocarcinomas and hyperplastic epithelial lesions in the peripheral lung.Am J Clin Pathol. 1986; 85: 146-151Crossref PubMed Scopus (93) Google Scholar Thus, AAH has been implicated as a preneoplastic lesion of lung adenocarcinoma, and listed as a precursor lesion in the World Health Organization 1999 classification of lung tumors.14Travis WD Corrin B Shimosato Y Brambilla E World Health Organization: Histological Typing of Lung and Pleural Tumours. ed 3. Springer-Verlag, Berlin1999Crossref Google Scholar Tuberous sclerosis (TSC) is a relatively common autosomal-dominant disease that causes mental retardation, seizures, and multiple hamartomas in many organs including the brain, eyes, kidney, skin, and heart. Mutations of either theTSC1 or the TSC2 gene are responsible for this disease.15van Slegtenhorst M de Hoogt R Hermans C Nellist M Janssen B Verhoef S Lindhout D van den Ouweland A Halley D Young J Burley M Jeremiah S Woodward K Nahmias J Fox M Ekong R Osborne J Wolfe J Povey S Snell RG Cheadle JP Jones AC Tachataki M Ravine D Sampson JR Reeve MP Richardson P Wilmer F Munro C Hawkins TL Sepp T Ali JBM Ward S Green AJ Yates JRW Kwiatkowska J Henske EP Short MP Haines JH Jozwiak S Kwiatkowski DJ Identification of the tuberous sclerosis gene TSC1 on chromosome 9q34.Science. 1997; 277: 805-808Crossref PubMed Scopus (1369) Google Scholar, 16Identification and characterization of the tuberous sclerosis gene on chromosome 16: The European Chromosome 16 Tuberous Sclerosis Consortium.Cell. 1993; 75: 1305-1315Abstract Full Text PDF PubMed Scopus (1487) Google Scholar The lesion most commonly described in the lung is lymphangioleiomyomatosis, which occurs in 1% of patients with TSC and affects only females.17Capron F Ameille J Leclerc P Mornet P Barbagellata M Reynes M Rochemaure J Pulmonary lymphangioleiomyomatosis and Bourneville's tuberous sclerosis with pulmonary involvement: the same disease?.Cancer. 1983; 52: 851-855Crossref PubMed Scopus (75) Google Scholar Multifocal micronodular pneumocyte hyperplasia (MMPH) has also been described as a rare pulmonary manifestation of TSC.18Muir TE Leslie KO Popper H Kitaichi M Gagne E Emelin JK Vinters HV Colby TV Micronodular pneumocyte hyperplasia.Am J Surg Pathol. 1998; 22: 465-472Crossref PubMed Scopus (98) Google Scholar MMPH is so similar to AAH morphologically that histological distinction between MMPH and AAH is difficult.19Guinee D Singh R Azumi N Singh G Przygodzki RM Travis W Koss M Multifocal micronodular pneumocyte hyperplasia: a distinctive pulmonary manifestation of tuberous sclerosis.Mod Pathol. 1995; 8: 902-906PubMed Google Scholar We previously reported that LOH in theTSC gene-associated regions was frequently observed in lung adenocarcinoma with multiple AAHs.20Suzuki K Ogura T Yokose T Nagai K Mukai K Kodama T Nishiwaki Y Esumi H Loss of heterozygosity in the tuberous sclerosis gene associated regions in adenocarcinoma of the lung accompanied by multiple atypical adenomatous hyperplasia.Int J Cancer. 1998; 79: 384-389Crossref PubMed Scopus (38) Google Scholar In this study, we analyzed microsatellite alterations at several microsatellite loci including the TSC gene-associated regions in both AAH lesions and concomitant adenocarcinomas to clarify the stage of lung adenocarcinoma pathogenesis in which these genetic alterations are involved. From November 1997 through June 1998, 126 patients underwent surgical resection of lung cancer at our hospital. AAH was found in 22 patients with adenocarcinoma, 2 patients with adenosquamous carcinoma, 1 with squamous cell carcinoma, and 1 with large cell carcinoma. AAH is much more frequently found in patients with adenocarcinoma than in those with other histological subtypes. We analyzed LOH on chromosomes 9q and 16p in 18 AAHs and 17 adenocarcinomas from 11 patients. The patients' characteristics are summarized in Table 1. There were four males and seven females, and their ages ranged from 47 to 74 years. Seven of the 11 (64%) patients were non-smokers. Two patients had a past history of malignancy. Three patients had a family history of malignancy in first-degree relatives; two were lung cancers, one a gastric cancer. There were three patients with multiple adenocarcinomas, and nine with multiple AAHs. All patients with multiple adenocarcinomas had concomitant multiple AAHs.Table 1Clinicopathological Characteristics of PatientsPatientAgeGenderSmoking (pack-years)Past history of malignancyFamily history of malignancyNumber of adenocarcinomasNumber of AAH174M41−−42272F0−+ (lung, sister)†Affected organ and family member with history of malignancy.11360F0+ (thyroid)*Organs affected by a past malignancy.−43451F0−+ (lung, mother)13547M4−−12662F0−−36775F0−−11865M45−−12965M70+ (stomach)−151061F0−−121169F0−+ (stomach, father)12* Organs affected by a past malignancy.† Affected organ and family member with history of malignancy. Open table in a new tab All resected specimens prefixed with 100% methanol were sliced at 5-mm intervals and examined macroscopically. Appropriate tissue sections were fixed with 100% methanol and embedded in paraffin. It is difficult to extract enough amount of DNA for molecular analysis from a tiny lesion, such as AAH. DNA extracted from formalin-fixed materials is often fragmented artificially. Using methanol fixation, even relatively higher molecular weight DNA was preserved well.21Noguchi M Furuya S Takeuchi T Hirohashi S Modified formalin and methanol fixation methods for molecular biological and morphological analyses.Pathol Int. 1997; 47: 685-691Crossref PubMed Scopus (74) Google Scholar Primary lung adenocarcinomas and AAHs were evaluated microscopically by conventional hematoxylin and eosin (H&E) staining. The pathological characteristics of the 17 adenocarcinomas and 18 AAHs were assessed by two pathologists (AO and TY). Histological typing of adenocarcinomas was performed according to the World Health Organization classification of lung tumors.14Travis WD Corrin B Shimosato Y Brambilla E World Health Organization: Histological Typing of Lung and Pleural Tumours. ed 3. Springer-Verlag, Berlin1999Crossref Google Scholar Nuclear atypia was categorized into three grades: 1, nuclei that were uniform in size and slightly larger than those of reactive type II alveolar epithelial cells; 2, nuclei that were in uniform size and up to twice the size of reactive type II alveolar epithelial cells; 3, nuclei of various sizes and more than twice the size of reactive type II alveolar epithelial cells. The mitotic indices were divided into three grades: 1, ≤5 mitotic cells/10 high-power fields; 2, 6 to 15/10 high-power fields; 3, ≥16/10 high-power fields. The scar grades were classified into four grades based on fibrotic foci in the tumors: 1, no or minimal desmoplasia; 2, fibroblastic tissue with a small amount of collagen; 3, fibroblastic tissue with moderate to abundant collagen; 4, hyalinized tissue.22Shimosato Y Suzuki A Hashimoto T Nishiwaki Y Kodama T Yoneyama T Kameya T Prognostic implications of fibrotic focus (scar) in small peripheral lung cancers.Am J Surg Pathol. 1980; 4: 365-373Crossref PubMed Scopus (277) Google Scholar We analyzed solitary AAHs in this study, AAHs continuous with or directly adjacent to a primary adenocarcinoma were not included. The histological diagnosis of AAH was based on the following criteria, as previously described.23Suzuki K Nagai K Yoshida J Yokose T Kodama T Takahashi K Nishimura M Kawasaki H Yokozaki M Nishiwaki Y The prognosis of resected lung carcinoma associated with atypical adenomatous hyperplasia: a comparison of the prognosis of well-differentiated adenocarcinoma associated with atypical adenomatous hyperplasia and intrapulmonary metastasis.Cancer. 1997; 79: 1521-1526Crossref PubMed Scopus (67) Google Scholar 1) The lesion had well-defined boundaries and consisted of proliferation of single-layered atypical epithelial cells without central scar formation or collapse (Figure 1A). 2) The cytoplasm was eosinophilic, and the cells often had a rounded or domed appearance resembling either type II pneumocytes or Clara cells. 3) The atypical cells in AAH had usually hyperchromatic nuclei and inconspicuous nucleoli, but the atypia was less marked than that of adenocarcinoma cells. AAHs were classified into two grades:24Yokose T Yuji I Atushi O High prevalence of atypical adenomatous hyperplasia of the lung in autopsy specimens from elderly patients with malignant neoplasms.Lung Cancer. 2000; 29: 125-130Abstract Full Text Full Text PDF PubMed Scopus (37) Google Scholar 1) Low-grade AAH; the cell density was low to moderate, and the cells were arranged in a single layer, intermittently or focally and continuously, on the alveolar septa (Figure 1B). Their nuclei were mostly small, but occasionally large, and exhibited lesser degrees of variation in size, shape, and hyperchromasia than high-grade AAH. 2) High-grade AAH; the cells were continuously and densely arranged in a single layer, but did not exhibit the piled-up structure often observed in adenocarcinoma (Figure 1C). Their nuclei showed significant atypia, but lacked the margin irregularity and eosinophilic nucleoli observed in frank adenocarcinoma. Serial 5-μm slices were made from each block for microdissection. The slides were dewaxed with xylene and rehydrated in a graded alcohol series, then stained with H&E to define the location of AAHs or adenocarcinomas. Microdissection was performed under a microscope (BX50W1; Olympus, Tokyo, Japan) by using a microcapillary tube drawn to a thin tip with a micropipette puller (PC-10; Narishige, Tokyo, Japan) and a joystick-operated hydraulic micromanipulator (ONO-125; Olympus-Narishige, Tokyo, Japan). The microdissected cells were allowed to adhere to Parafilm (American Can, Greenwich, CT) and placed in 500-μl microcentrifuge tubes. Normal lymph node tissue or normal lung tissue was scraped with a 27-gauge needle to provide a normal control. DNA was extracted with a DNA extractor WB kit (Wako Pure Chemicals, Osaka, Japan). The DNA concentration was adjusted to ∼50 cell equivalents per μl. We analyzed LOH on chromosome 9q (including the TSC1 gene-associated region) and 16p (including the TSC2 gene-associated region), using the following microsatellite markers. Four markers on 9q from centromere to telomere, D9S146 (9q13), D9S149 (9q34), D9S150 (9q34), andDBH (9q34);25Henske EP Ozelius L Gusella JF Haines JL Kwiatkowski DJ A high-resolution linkage map of human 9q34.1.Genomics. 1993; 17: 587-591Crossref PubMed Scopus (24) Google Scholar and four markers on 16p from centromere to telomere, D16S300 (16p11.1-11.2),D16S292 (16p13.12-13.13), D16S291 (16p13.1), andD16S525 (16p13.3).26Thompson AD Shen Y Holman K Sutherland GR Callen DF Richards RI Isolation and characterisation of (AC)n microsatellite genetic markers from human chromosome 16.Genomics. 1992; 13: 402-408Crossref PubMed Scopus (94) Google Scholar, 27Shen Y Holman K Doggett NA Callen DF Sutherland GR Richards RI Dinucleotide repeat polymorphisms at the D16S525, D16S359, D16S531 and D16S522 loci.Hum Mol Genet. 1994; 3: 210Crossref PubMed Scopus (7) Google Scholar PCR was performed in a 20-μl volume of a mixture containing 10 mmol/L Tris (pH 8.3), 50 mmol/L KCl, 1.0 to 1.5 mmol/L MgCl2, 200 μmol/L of each Cy 5′-end labeled primer (Pharmacia Biotech, Tokyo, Japan), 0.25 U of Taq polymerase (TaKaRa Biomedicals, Shiga, Japan) and then cycled 36 to 38 times in a GeneAmp PCR System 9600 (Perkin Elmer, Foster City, CA); each cycle consisted of 1 minute at 94°C for denaturation, 2 minutes at 55 to 62°C for annealing, 1 minute at 72°C for strand elongation, and 7 minutes at 72°C for final elongation. The PCR products were diluted with a loading buffer consisting of 95% formamide, 20 mmol/L ethylenediaminetetraacetic acid (pH 8.0), and Dextran blue, and denatured for 5 minutes at 98°C. The samples were electrophoresed on 5% polyacrylamide gels containing 8.3 mol/L urea for 3 hours at 34 W using an ALFred DNA sequencer (Pharmacia Biotech). To confirm the reproducibility of the experiment, all cases were examined at least twice by independent PCR and electrophoresis. LOH was considered to be present if the reduction rate of the height of the allele in the tumor was >40%, as previously defined.28Shiseki M Kohno T Nishikawa R Sameshima Y Mizoguchi H Yokota J Frequent allelic losses on chromosomes 2q, 18q, and 22q in advanced non-small cell lung carcinoma.Cancer Res. 1994; 54: 5643-5648PubMed Google Scholar The two-sided Fisher's exact test was used for statistical analysis (Stat View-J 5.0, Macintosh). A P value 2 cm85 (0.64)*Numbers in parentheses are P values (Fisher's exact test).3 (1.0)4 (0.64)3 (0.62)Histological subtype BAC84231 Other than BAC95 (1.0)4 (0.62)4 (1.0)4 (0.29)Vascular invasion Negative105443 Positive74 (1.0)2 (1.0)3 (1.0)2 (1.0)Lymphatic permeation Negative115342 Positive64 (0.62)3 (0.60)3 (0.64)3 (0.28)Nuclear atypia 1 or 262110 3117 (0.34)5 (0.33)6 (0.30)5 (0.10)Mitotic index 1125544 2 or 354 (0.29)1 (0.60)3 (0.59)1 (1.0)Scar grade 1 or 294332 3 or 485 (0.64)3 (1.0)4 (0.64)3 (0.62)BAC, bronchioloalveolar carcinoma.* Numbers in parentheses are P values (Fisher's exact test). Open table in a new tab Table 3Relationships between Histological Characteristics of AAH and LOH on Chromosomes 9q and 16pnLOH on 9qLOH on 16pTSC1LOHTSC2LOHSize <5 mm106140 ≥5 mm81 (0.07)*Numbers in parentheses are P values (Fisher's exact test).3 (0.27)1 (0.31)1 (0.44)Grade Low104240 High83 (1.0)2 (1.0)1 (0.31)1 (0.44)Low, low-grade AAH; high, high-grade AAH.* Numbers in parentheses are P values (Fisher's exact test). Open table in a new tab BAC, bronchioloalveolar carcinoma. Low, low-grade AAH; high, high-grade AAH. Little is known about the etiology of lung adenocarcinoma, as compared with squamous cell carcinoma. AAH has recently been implicated as a preneoplastic lesion of lung adenocarcinoma. Miller29Miller RR Bronchioloalveolar cell adenomas.Am J Surg Pathol. 1990; 14: 904-912Crossref PubMed Scopus (143) Google Scholar initially proposed a pulmonary adenoma-carcinoma sequence analogous to that of the colon. AAH is much more frequently associated with adenocarcinoma than other histological subtypes of lung cancer.30Colby TV Wistuba II Gazdar A Precursors to pulmonary neoplasia.Adv Anat Pathol. 1998; 5: 205-215Crossref PubMed Scopus (74) Google Scholar Multiple AAHs are occasionally detected in patients with multiple lung adenocarcinomas.29Miller RR Bronchioloalveolar cell adenomas.Am J Surg Pathol. 1990; 14: 904-912Crossref PubMed Scopus (143) Google Scholar In our series, the histological subtype was adenocarcinoma in 22 of 26 (85%) lung cancer patients with AAH, and all patients with multiple adenocarcinomas had concomitant multiple AAHs. These findings suggest that AAH and adenocarcinoma might be induced by common etiological factors. AAH has been reported to have several genetic alterations in common with lung adenocarcinoma. Mutations of K-ras codon 12 have variously been detected in 15 to 50% of AAHs and 22 to 42% of adenocarcinomas.7Cooper CA Carby FA Bubb VJ Lamb D Kerr KM Wyllie AH The pattern of K-ras mutation in pulmonary adenocarcinoma defines a new pathway of tumour development in the human lung.J Pathol. 1997; 181: 401-404Crossref PubMed Scopus (76) Google Scholar, 8Westra WH Baas IO Hruban RH Askin FB Wilson K Offerhaus GJ Slebos RJ K-ras oncogene activation in atypical alveolar hyperplasias of the human lung.Cancer Res. 1996; 56: 2224-2228PubMed Google Scholar, 9Sagawa M Saito Y Fujimura S Linnoila RI K-ras point mutation occurs in the early stage of carcinogenesis in lung cancer.Br J Cancer. 1998; 77: 720-723Crossref PubMed Scopus (46) Google Scholar, 10Ohshima S Shimizu Y Takahama M Detection of c-Ki-ras gene mutation in paraffin sections of adenocarcinoma and atypical bronchioloalveolar cell hyperplasia of human lung.Virchows Arch. 1994; 424: 129-134PubMed Google Scholar LOH on chromosome 3p was detected in 10 to 18% of AAHs and 12 to 67% of adenocarcinomas.4Kishimoto Y Sugio K Hung JY Virmani AK McIntire DD Minna JD Gazdar AF Allele-specific loss in chromosome 9p loci in preneoplastic lesions accompanying non-small-cell lung cancers.J Natl Cancer Inst. 1995; 87: 1224-1229Crossref PubMed Scopus (203) Google Scholar, 5Kitaguchi S Takeshima Y Nishisaka T Inai K Proliferative activity, p53 expression and loss of heterozygosity on 3p, 9p and 17p in atypical adenomatous hyperplasia of the lung.Hiroshima J Med Sci. 1998; 47: 17-25PubMed Google Scholar, 6Kohno H Hiroshima K Toyozaki T Fujisawa T Ohwada H p53 mutation and allelic loss of chromosome 3p, 9p of preneoplastic lesions in patients with nonsmall cell lung carcinoma.Cancer. 1999; 85: 341-347Crossref PubMed Scopus (69) Google Scholar LOH on chromosome 9p was detected in 5 to 13% of AAHs and 19 to 50% of adenocarcinomas.4Kishimoto Y Sugio K Hung JY Virmani AK McIntire DD Minna JD Gazdar AF Allele-specific loss in chromosome 9p loci in preneoplastic lesions accompanying non-small-cell lung cancers.J Natl Cancer Inst. 1995; 87: 1224-1229Crossref PubMed Scopus (203) Google Scholar, 5Kitaguchi S Takeshima Y Nishisaka T Inai K Proliferative activity, p53 expression and loss of heterozygosity on 3p, 9p and 17p in atypical adenomatous hyperplasia of the lung.Hiroshima J Med Sci. 1998; 47: 17-25PubMed Google Scholar, 6Kohno H Hiroshima K Toyozaki T Fujisawa T Ohwada H p53 mutation and allelic loss of chromosome 3p, 9p of preneoplastic lesions in patients with nonsmall cell lung carcinoma.Cancer. 1999; 85: 341-347Crossref PubMed Scopus (69) Google Scholar We demonstrated AAHs to harbor LOH on chromosomes 9q and 16p, LOHs that are also found in lung adenocarcinomas. The frequency of LOH on 9q, especially in theTSC1 gene associated regions, was high. Petersen and colleagues31Petersen I Bujard M Petersen S Wolf G Goeze A Schwendel A Langreck H Gellert K Reichel M Just K du Manoir S Cremer T Dietel M Ried T Patterns of chromosomal imbalances in adenocarcinoma and squamous cell carcinoma of the lung.Cancer Res. 1997; 57: 2331-2335PubMed Google Scholar reported LOH on 9q34 to be significantly associated with lung adenocarcinoma. Our results were also consistent with this observation. The relatively high frequency of LOH on 9q34 in AAH lesions and adenocarcinomas suggests that a novel tumor suppressor gene for lung adenocarcinomas may exist in this region, and theTSC1 gene is one candidate. Hung and colleagues3Hung J Kishimoto Y Sugio K Virmani A McIntire DD Minna JD Gazdar AF Allele-specific chromosome 3p deletions occur at an early stage in the pathogenesis of lung carcinoma.JAMA. 1995; 273: 558-563Crossref PubMed Scopus (306) Google Scholar and Kishimoto and colleagues4Kishimoto Y Sugio K Hung JY Virmani AK McIntire DD Minna JD Gazdar AF Allele-specific loss in chromosome 9p loci in preneoplastic lesions accompanying non-small-cell lung cancers.J Natl Cancer Inst. 1995; 87: 1224-1229Crossref PubMed Scopus (203) Google Scholar detected more frequent and extensive 3p and 9p losses in carcinomas than in corresponding preneoplastic lesions, and the identical allele was lost among them, an allele-specific loss. Their findings were consistent with our results. Six of seven AAHs with LOH on 9q and all AAHs with LOH on 16p showed LOH at only one microsatellite locus. However, the allelic losses on 9q and 16p were more extensive in adenocarcinomas than in AAHs. In particular, the rate of complete LOH on 9q was high (78%) in adenocarcinomas. This progressive loss on 9q during progression might suggest existence of more than one tumor suppressor gene on 9q for lung adenocarcinoma. In addition, the identical allele was lost among multiple AAHs and concomitant adenocarcinomas in the same patients. These observations indicate that one allele might be inactivated congenitally because of mutation or epigenetically, and the remaining allelic loss would then be acquired. This suggests a possible role of genetic predisposition in the pathogenesis of lung adenocarcinoma with AAH. In our series, 7 of 11 (64%) patients with lung adenocarcinoma and corresponding AAH were non-smokers and 5 of 11 (45%) had either a past or family history of malignancy. These findings might also indicate genetic predisposition. Muir and colleagues18Muir TE Leslie KO Popper H Kitaichi M Gagne E Emelin JK Vinters HV Colby TV Micronodular pneumocyte hyperplasia.Am J Surg Pathol. 1998; 22: 465-472Crossref PubMed Scopus (98) Google Scholar reported that MMPH is distinguishable from AAH by the following features: AAH cells have a greater degree of nuclear-to-cytoplasmic ratio than MMPH, less well circumscribed with peripheral lepidic spread, less prominent interstitial reticuln, and fewer air space macrophages than MMPH. However, some investigators have reported that MMPH is so similar to AAH morphologically that histological distinction between MMPH and AAH is difficult.19Guinee D Singh R Azumi N Singh G Przygodzki RM Travis W Koss M Multifocal micronodular pneumocyte hyperplasia: a distinctive pulmonary manifestation of tuberous sclerosis.Mod Pathol. 1995; 8: 902-906PubMed Google Scholar Thus, we supposed that these two conditions might share some molecular mechanisms of pathogenesis. Genetic alteration in TSC1 gene- associated regions is a candidate. If we postulate that the TSC1 gene itself is responsible for early development of lung adenocarcinoma, there would be an obvious discrepancy. We would more frequently encounter TSC patients with MMPH. However, MMPH is believed to be a rare pulmonary manifestation of TSC, and only a few such cases have been reported.18Muir TE Leslie KO Popper H Kitaichi M Gagne E Emelin JK Vinters HV Colby TV Micronodular pneumocyte hyperplasia.Am J Surg Pathol. 1998; 22: 465-472Crossref PubMed Scopus (98) Google Scholar, 19Guinee D Singh R Azumi N Singh G Przygodzki RM Travis W Koss M Multifocal micronodular pneumocyte hyperplasia: a distinctive pulmonary manifestation of tuberous sclerosis.Mod Pathol. 1995; 8: 902-906PubMed Google Scholar How can we explain this discrepancy? Because MMPH is a subtle pulmonary lesion, underestimation of the incidence of MMPH might be one explanation. The life expectancy of patients with TSC is relatively short,32Shepherd CW Gomez MR Lie JT Crowson CS Causes of death in patients with tuberous sclerosis.Mayo Clin Proc. 1991; 66: 792-796Abstract Full Text Full Text PDF PubMed Scopus (410) Google Scholar and this is another possible reason. We do not understand how many genetic alterations are required for formations of MMPH and AAH. If several genetic alterations are required, low incidence of MMPH association in TSC patients might be explained. This is the third possible reason. The forth and most probable explanation is that TSC1 itself is not the responsible gene but rather that some novel tumor suppressor gene for lung adenocarcinoma exists very close to the TSC1 gene. In our present study, we analyzed only a limited number of lesions. Further study of a large number of adenocarcinomas and AAH lesions is clearly required. We did not analyzed LOH in AAH adjacent to adenocarcinoma, because it was difficult to convincingly differentiate the border between AAH and adenocarcinoma. If we could analyze several adenocarcinomas obviously associated with AAH adjacently, it would become more convincing. In addition, an extensive mutation analysis or demonstrating the suppressed expression of the TSC1 gene in lung adenocarcinoma might be helpful in answering the above question.
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