Britt-Marie Ljung

The gene expression signatures of melanoma progression

Because of the paucity of available tissue, little information has previously been available regarding the gene expression profiles of primary melanomas. To understand the molecular basis of melanoma progression, we compared the gene expression profiles of a series of nevi, primary melanomas, and melanoma metastases. We found that metastatic melanomas exhibit two dichotomous patterns of gene expression, which unexpectedly reflect gene expression differences already apparent in comparing laser-capture microdissected radial and vertical phases of a large primary melanoma. Unsupervised hierarchical clustering accurately separated nevi and primary melanomas. Multiclass significance analysis of microarrays comparing normal skin, nevi, primary melanomas, and the two types of metastatic melanoma identified 2,602 transcripts that significantly correlated with sample class. These results suggest that melanoma pathogenesis can be understood as a series of distinct molecular events. The gene expression signatures identified here provide the basis for developing new diagnostics and targeting therapies for patients with malignant melanoma.

Detection of breast cancer cells in ductal lavage fluid by methylation-specific PCR

If detected early, breast cancer is curable. We tested cells collected from the breast ducts by methylation-specific PCR (MSP). Methylated alleles of Cyclin D2, RAR-beta, and Twist genes were frequently detected in fluid from mammary ducts containing endoscopically visualised carcinomas (17 cases of 20), and ductal carcinoma in situ (two of seven), but rarely in ductal lavage fluid from healthy ducts (five of 45). Two of the women with healthy mammograms whose ductal lavage fluid contained methylated markers and cytologically abnormal cells were subsequently diagnosed with breast cancer. Carrying out MSP in these fluid samples may provide a sensitive and powerful addition to mammographic screening for early detection of breast cancer.

In situ analyses of genome instability in breast cancer

Transition through telomere crisis is thought to be a crucial event in the development of most breast carcinomas. Our goal in this study was to determine where this occurs in the context of histologically defined breast cancer progression. To this end, we assessed genome instability (using fluorescence in situ hybridization) and other features associated with telomere crisis in normal ductal epithelium, usual ductal hyperplasia, ductal carcinoma in situ and invasive cancer. We modeled this process in vitro by measuring these same features in human mammary epithelial cell cultures during ZNF217-mediated transition through telomere crisis and immortalization. Taken together, the data suggest that transition through telomere crisis and immortalization in breast cancer occurs during progression from usual ductal hyperplasia to ductal carcinoma in situ.

Breast tumor copy number aberration phenotypes and genomic instability

BackgroundGenomic DNA copy number aberrations are frequent in solid tumors, although the underlying causes of chromosomal instability in tumors remain obscure. Genes likely to have genomic instability phenotypes when mutated (e.g. those involved in mitosis, replication, repair, and telomeres) are rarely mutated in chromosomally unstable sporadic tumors, even though such mutations are associated with some heritable cancer prone syndromes.MethodsWe applied array comparative genomic hybridization (CGH) to the analysis of breast tumors. The variation in the levels of genomic instability amongst tumors prompted us to investigate whether alterations in processes/genes involved in maintenance and/or manipulation of the genome were associated with particular types of genomic instability.ResultsWe discriminated three breast tumor subtypes based on genomic DNA copy number alterations. The subtypes varied with respect to level of genomic instability. We find that shorter telomeres and altered telomere related gene expression are associated with amplification, implicating telomere attrition as a promoter of this type of aberration in breast cancer. On the other hand, the numbers of chromosomal alterations, particularly low level changes, are associated with altered expression of genes in other functional classes (mitosis, cell cycle, DNA replication and repair). Further, although loss of function instability phenotypes have been demonstrated for many of the genes in model systems, we observed enhanced expression of most genes in tumors, indicating that over expression, rather than deficiency underlies instability.ConclusionMany of the genes associated with higher frequency of copy number aberrations are direct targets of E2F, supporting the hypothesis that deregulation of the Rb pathway is a major contributor to chromosomal instability in breast tumors. These observations are consistent with failure to find mutations in sporadic tumors in genes that have roles in maintenance or manipulation of the genome.

Biomarker Expression and Risk of Subsequent Tumors After Initial Ductal Carcinoma In Situ Diagnosis

BACKGROUND Studies have failed to identify characteristics of women who have been diagnosed with ductal carcinoma in situ (DCIS) and have a high or low risk of subsequent invasive cancer. METHODS We conducted a nested case-control study in a population-based cohort of 1162 women who were diagnosed with DCIS and treated by lumpectomy alone from 1983 to 1994. We collected clinical characteristics and information on subsequent tumors, defined as invasive breast cancer or DCIS diagnosed in the ipsilateral breast containing the initial DCIS lesion or at a regional or distant site greater than 6 months after initial treatment of DCIS (N = 324). We also conducted standardized pathology reviews and immunohistochemical staining for the estrogen receptor (ER), progesterone receptor, Ki67 antigen, p53, p16, epidermal growth factor receptor-2 (ERBB2, HER2/neu oncoprotein), and cyclooxygenase-2 (COX-2) on the initial paraffin-embedded DCIS tissue. Competing risk models were used to determine factors associated with risk of subsequent invasive cancer vs DCIS, and cumulative incidence survival functions were used to estimate 8-year risk. RESULTS Factors associated with subsequent invasive cancer differed from those associated with subsequent DCIS. Eight-year risk of subsequent invasive cancer was statistically significantly (P = .018) higher for women with initial DCIS lesions that were detected by palpation or that were p16, COX-2, and Ki67 triple positive (p16(+)COX-2(+)Ki67(+)) (19.6%, 95% confidence interval [CI] = 18.0% to 21.3%) than for women with initial lesions that were detected by mammography and were p16, COX-2, and Ki67 triple negative (p16(-)COX-2(-)Ki67(-)) (4.1%, 95% CI = 3.4% to 5.0%). In a multivariable model, DCIS lesions that were p16(+)COX-2(+)Ki67(+) or those detected by palpation were statistically significantly associated with subsequent invasive cancer, but nuclear grade was not. Eight-year risk of subsequent DCIS was highest for women with DCIS lesions that had disease-free margins of 1 mm or greater combined with either ER(-)ERBB2(+)Ki67(+) or p16(+)COX-2(-)Ki67(+) status (23.6%, 95% CI = 18.1% to 34.0%). CONCLUSION Biomarkers can identify which women who were initially diagnosed with DCIS are at high or low risk of subsequent invasive cancer, whereas histopathology information cannot.

PRIMARY INTRAOCULAR LYMPHOMA (OCULAR RETICULUM CELL SARCOMA) DIAGNOSIS AND MANAGEMENT

The authors retrospectively reviewed the diagnosis and management of 20 intraocular lymphoma patients who initially presented with either ocular or central nervous system (CNS) disease. As the ophthalmic community has become more aware of this entity, the interval between symptoms and diagnosis has significantly shortened. Diagnosis can usually be made on cytopathologic examination of vitreous cells. However, in three cases more than one vitreous biopsy was necessary. Results of cytologic examination appeared to be more accurate than those of conventional lymphocyte surface marker studies in the diagnosis of an intraocular lymphoma. Long-term survival occurred in some patients treated with a combination of intrathecal chemotherapy and ocular/CNS irradiation.

Genetic alterations in primary breast cancers and their metastases: direct comparison using modified comparative genomic hybridization.

Breast tumor development and progression are thought to be driven by an accumulation of genetic alterations, but little is known about the specific changes that occur during the metastatic process. We analyzed pairs of primary breast cancers and their matched lymph node metastases from 11 patients, pairs of primaries and distant metastases from three patients, and pairs of primaries, and local recurrences from two patients by using comparative genomic hybridization (CGH). Simultaneous hybridization analysis of primary versus matched lesion DNAs from 11 patients was also performed (modified CGH). This modified approach was useful not only for confirming CGH results but also for demonstrating quantitative differences between aberrations present at both sites. Frequent chromosomal changes present at both sites (>35% of 16 cases) were 1q, 8q, and 17q gains and 6q, 8p, 9q, 13q, 16q, 17p, and Xp losses. The total number of aberrations detected exclusively in the lymph nodes or distant metastases was higher than that in the primary tumors (2.5 vs. 0.7, P< 0.05). We found high‐level amplifications in four metastases (two lymph nodes and two distant metastases), but none in any primary tumor. These findings suggest that progression from primary breast cancer to metastasis may be associated with the acquisition of further genetic changes. Although further investigations are required, it was of interest that 3 of 11 patients (27%) showed 18q loss solely in their lymph node metastases. Genes Chromosom. Cancer 19:267–272, 1997.

HER2/Neu and the Ets transcription activator PEA3 are coordinately upregulated in human breast cancer.

HER2/Neu is overexpressed in 25 – 30% of all human breast cancers as a result of both gene amplification and enhanced transcription. Transcriptional upregulation of HER2/neu leads to a 6 – 8-fold increased abundance of its mRNA per gene copy and likely results from the elevated activity of transcription factors acting on the HER2/neu promoter. Here we report that transcripts of PEA3, an ETS transcription factor implicated in oncogenesis, were increased in 93% of HER2/Neu-overexpressing human breast tumor samples. Analyses to uncover the molecular basis for elevated PEA3 transcripts in HER2/Neu-positive breast tumors revealed that the HER2/Neu receptor tyrosine kinase initiated an intracellular signaling cascade resulting in increased PEA3 transcriptional activity; transcriptionally-activated PEA3 stimulated HER2/neu and PEA3 gene transcription by binding to sites in the promoters of these genes. PEA3 also activates transcription of genes encoding matrix-degrading proteinases, enzymes required for tumor cell migration and invasion. These findings implicate PEA3 in the initiation and progression of HER2/Neu positive breast cancer, and suggest that PEA3 and signaling proteins affecting its regulation are appropriate therapeutic targets.

Environmental factors and the risk of salivary gland cancer.

Cancer of the major salivary glands is rare, and little is known about its etiology. We conducted a population‐based case‐control study to elucidate the risk factors for these tumors. Of 199 cases diagnosed with salivary gland tumors between 1989 and 1993, 150 (75%) were interviewed. We subsequently excluded nine cases based on review of pathology specimens. We identified 271 controls through random‐digit dialing and the Health Care Finance Administration files; 191 (70%) were interviewed. Therapeutic medical radiation treatment to the head or neck [odds ratio (OR) = 2.6; 95% confidence interval (CI) = 0.84–8.1], full mouth dental x‐rays (OR = 1.6; 95% CI = 1.0–2.7), and ultraviolet light treatment to the head or neck (OR = 1.9; 95% CI = 0.89–4.3) were associated with increased risk. These elevations in risk were largely limited to those exposed before 1955, when the exposure dose was substantially higher. Occupational exposure to radiation/radioactive materials (OR = 2.4; 95% CI = 1.0–5.4) and nickel compounds/alloys (OR = 6.0; 95% CI = 1.6–22.0), as well as employment in the rubber industry (OR = 7.0; 95% CI = 0.80–60.3), increased risk. In men, current smoking (OR = 2.1; 95% CI = 0.98–4.7) and heavy alcohol consumption (OR = 2.5; 95% CI = 1.1–5.7) were associated with risk, but these factors were not strongly related to salivary gland cancer in women.

Evaluation of Abnormal Mammography Results and Palpable Breast Abnormalities

Professional organizations recommend that women at the age of 40 or 50 years start undergoing screening mammography every 1 to 2 years (1-5). Primary care practitioners order most screening mammography examinations and must decide how to evaluate women who have an abnormal result. Evaluation of women with an abnormal mammography result is a common problem because even good-quality mammography facilities generally interpret 5% to 10% of all screening examinations as abnormal. About 90% of women with abnormal results do not have breast cancer (6-9); therefore, a safe and efficient evaluation is crucial. Breast symptoms are also a common problem; primary care practitioners receive approximately 20 presentations per 1000 person-years for the investigation of a breast symptom (10, 11). A breast lump is the most common symptom associated with breast cancer; between 9% and 11% of breast lumps result in a diagnosis of breast cancer (10, 12, 13). The prevalence of breast cancer among women who present with a breast lump increases with age from 1% for women 40 years of age and younger to 9% for women between 41 and 55 years of age to 37% for women age 55 years and older (12). Given that about 1 in 10 women with a breast lump or abnormal mammography result will have breast cancer, primary care practitioners must make a series of decisions to exclude or establish a diagnosis of breast cancer in these women. We review the literature on the evaluation of an abnormal screening mammography result and palpable breast abnormality and present an evidence-based approach with which to evaluate these two common problems. Methods We searched for published manuscripts determining the accuracy of screening and diagnostic mammography, fine-needle aspiration biopsy [FNAB], and core-needle biopsy to detect breast cancer among women with nonpalpable and palpable breast lesions. We systematically searched MEDLINE from January 1966 to March 2003 using the Medical Subject Heading terms or key words mammography, sensitivity, specificity, screening combined with breast cancer or breast neoplasm; or fine-needle biopsy, sensitivity, specificity, palpable mass combined with breast cancer or breast neoplasm; or core needle biopsy, sensitivity, specificity, palpable mass combined with breast cancer or breast neoplasm. We manually searched bibliographies of original and review articles identified in MEDLINE. Studies were included if 1) mammography, FNAB, or core-needle biopsy was performed before a definitive diagnosis was obtained; 2) the study sample included 100 or more women; and 3) breast cancer status was determined from histopathology review of excisional biopsy specimens, from linkage with a state cancer registry or the Surveillance, Epidemiology, and End Results program, or from clinical follow-up of 95% or more of the study sample. Studies of mammography, FNAB, or core-needle biopsy were considered high quality if they had a study sample that was population-based or consecutively sampled and if they determined cancer status in 95% or more of the study sample 1 or more years after the imaging or biopsy test was performed in order to determine the false-negative rate of the test (6, 9, 13-22). The major limitations of studies not considered high quality were incomplete follow-up, convenience study sample, and results that were not age adjusted. We primarily used results of three high-quality studies to describe optimal management strategies to evaluate nonpalpable and palpable breast lesions (13-15). Only one study (14) that evaluated the accuracy of screening mammography reported likelihood ratios or results to calculate likelihood ratios for the six American College of Radiology Breast Imaging and Reporting Data System (BI-RADS) assessment categories (23) for both first and subsequent screening examinations. A study of the accuracy of diagnostic mammography was the only population-based study to report results for nonpalpable and palpable breast lesions (15). The study of the accuracy of FNAB was the only study that gave results that allowed calculation of likelihood ratios for the four cytology assessment categories (13). Evaluation of Abnormal Screening Mammography Examination Screening mammography is performed in asymptomatic women with the goal of discovering invasive breast cancer at an early, curable stage. Screening mammography typically includes two views of each breast (craniocaudal and mediolateral oblique). The sensitivity of mammography ranges from 74% to 95%, and the specificity ranges from 89.4% to 99.1% (9, 14, 16, 17, 24). Sensitivity and specificity are higher for women more than 50 years of age, whereas sensitivity is lower and specificity higher for subsequent examinations compared with first screening examinations (9, 14, 16, 17, 24). The most common (and most worrisome) mammographic abnormalities that are found on screening examinations and that require further evaluation are masses and calcifications. The differential diagnosis for a mammographic mass includes cyst, benign nonproliferative lesions, benign proliferative lesions with or without atypia, fibroadenoma, radial scar, intramammary lymph node, lipoma, galactoceles, ductal carcinoma in situ, and invasive cancer. The differential diagnosis for a mammographic calcification includes benign nonproliferative lesions, benign proliferative lesions with or without atypia, fat necrosis, atherosclerosis, dermal lesion, ductal carcinoma in situ, and invasive cancer. No specific mammographic findings are associated with lobular carcinoma in situ. However, when mammographic calcifications are biopsied, lobular carcinoma in situ has been identified adjacent to histologic calcifications located in normal epithelium. Of note, although the differential diagnosis for a mass or calcification is long, all diagnoses other than ductal carcinoma in situ and invasive cancer are benign and require no further evaluation. Radiologists generally describe both masses and calcifications in terms of location, size, and other characteristics (such as shape, borders, and pattern). In addition to describing findings, radiologists make an assessment and recommendation (25). The American College of Radiology recommends one of six assessments for interpretation of a mammographic screening examination (Tables 1 and 2) (23). Abnormal screening mammography assessments are evaluated with diagnostic mammography, ultrasonography, and biopsy. Table 1. American College of Radiology Breast Imaging Reporting and Data System Table 2. Frequency of Screening Mammographic Results and Risk for Breast Cancer Based on Mammographic Result Diagnostic Mammography and Ultrasonography of Nonpalpable Lesions Diagnostic mammography is a comprehensive radiologic examination of a breast abnormality that may allow the radiologist to more definitively classify a finding; it can sometimes be done during the same visit as the screening examination. Diagnostic mammography consists of multiple specialized views, including magnification views or spot compression views. Results of diagnostic mammography are reported by using one of five assessment categories by the American College of Radiology: negative, benign finding, probably benign finding, suspicious abnormality, or highly suggestive of malignancy (23). About 15% of women with nonpalpable cancer will have a diagnostic mammography examination that shows no evidence of cancer (Table 3) (15). Negative likelihood ratios for a normal mammography result in women with a nonpalpable lesion are approximately 0.2 (Table 3). Ultrasonography is of particular value in distinguishing a cyst from a solid lesion on screening or diagnostic mammography. Ultrasonography is 98% to 100% accurate in diagnosing simple cysts when four rigorous sonographic criteria are used to evaluate the lesion (oval or lobulated shape; anechoic, well-defined posterior border; increased through-transmission; and no alteration of surrounding breast parenchyma) (26, 27). Table 3. Accuracy of Diagnostic Mammography in Women with a Nonpalpable Lesion or Breast Lump Biopsy For mammographic abnormalities that are nonpalpable and that require biopsy, image-guided tissue sampling is necessary. Tissue for diagnosis can be obtained by mammography, ultrasonography-guided FNAB, or core-needle biopsy or by open surgical biopsy with needle localization. The sensitivity of needle-localized excisional biopsy is 99% for nonpalpable lesions (28). The most common reason for missing the carcinoma is erroneous placement of the needle guidewire. Image-guided FNAB or core-needle biopsy is quicker, cheaper, and easier than standard-needle localization open biopsy (29-31). For image-guided FNAB or core-needle biopsy, a mammographic x-ray tube is angled to produce two views of the lesion, and the position of the lesion is calculated from the apparent movement of the lesion relative to a fixed reference grid. With use of this information, a needle is placed in the lesion; needle position is confirmed on repeated stereotactic x-ray views. Alternatively, high-resolution ultrasonography can be used to position the needle in real time within the lesion. A fine needle (22- to 25-gauge), an automated core needle (14- to 18-gauge), or a vacuum-assisted biopsy probe (11-gauge) can be used to obtain a sample of breast tissue. With nonpalpable lesions, core-needle biopsy is usually preferred over FNAB because the core sample provides adequate tissue for histologic diagnosis and is more accurate (32, 33). The accuracy of FNAB for nonpalpable lesions varies more than that of core-needle biopsy (sensitivity, 77% to 97%; specificity, 78% to 98%) (32-34); FNAB is also highly operator dependent and more often produces insufficient diagnostic material (33% for FNAB vs. 1.5% for core-needle biopsy) (18, 19). Image-guided FNAB allows safe sampling of very thin breast tissue and of lesions situated close to the chest wall or the skin, where core biopsy is not