5.3.

Breast Cancer Risk Indicators and Precursor Lesions Proliferative lesions of the breast reported to be associated with increased risk of breast cancer development are termed breast cancer risk indicators (Dupont and Page 1985; Dupont et. Al. 1993; Page and Dupon 1990, 1991, 1992; Tavassoli and Devilee 2003). To be considered breast cancer precursors, these lesions should be clonal, neoplastic proliferations and have histological, immunohistochemical and molecular features similar to and consistent with those of matched invasive breast cancers, wither synchronous or metachronous. Progression of one of these lesions to invasive cancer is by no means a centainty and only a proportion of them even evolve to an invasive breast cancer, and this they are in fact, non-obligate precursors. On the other hand, benign lesions, that often involve both luminal and myoppthelial celss (e.g. sclerosing adnonsis), have neither been associated with significantly increased risk of breast cancer development (Dupont and Page 1985; Dupont et al. 1993; Pge and Dupont 1990, 1991, 1992, 1998; Pge et al. 1985) nor display genetic aberrations in common with those of true precursors and invasive breast cancer (Washington et al. 2000). The term low grade breast neoplasia family has recently been coined to describe a group of in situ lesions that includes Columnar Cell Lesions (CCL), Flat Epithelial Atypia (FEA), ADH, ALH, LCIS and low grade DCIS (Fig. 5.2), and invasive lesions reported to be found in association with these nob-obligate precursors (i.e. tubular carcinoma, invasive cribriform cacionma, classic ILC and low grad IDC) (Abdel-Fatah et al. 2007, 2008). These lesions co-exist at frequencies that could not be justified by chance alone. They are characterized by low histological grade, expression of hormone receptors, lack of HER2 expression and HER2 gene amplification, and genetic aberrations usually found in low grade breast cancers (i.e. deletions of 16q and gains of lq). Until recently, only high grade DCIS was recognized as a precursor of high grade breast cancer (Reis-Fillho et al. 2005a; Simson et al. 2005a). Recent studies on Microglandular Adenosis (MGA), that was previously considered a hyperplastic lesion, demonstrated a frequent association with high grade breast cancers and that MGA harbors genomic aberrations identical to those found in matched invasive cancers (Geyer et al. 2009a; Shein et al. 2009). These observations suggest that MGA may constitute a non-obligate precursor of a subgroup of high grade breast cancers, in particular those of triple negative (i.e. ER-, PR-, HER2-) phenotype (Geyer et al. 2009; Shin et al. 2009). It is beyond the scope of this chapter discuss in exhaustive detail the pathology of breast cancer risk indicators and precursors and the readers are referred to excellent reviews and textbooks. In the following sections, the genetic features of normal breast, breast cancer precursors and risk indicators are discussed. Based on the concepts of the low grade breast neoplasia family and on the differences between low and high grade breast cancer outlined above, we have subdivided the known nonobligatory breast cancer precursors into low and high historical grade. 5.3.1. Normal Breast Determining the significant of molecular genetic changes in normal breast cells remains a matter of debate. Several studies have identified loss of heterozygosity (LOH) in normal cells (Deng et al. 1996; Larson et al. 2002; Lakhni et al. 1999) (e.g. in 13% of normal TDLUs and in 44% patients (Larson et al. 1998)). However, the loci affected in normal TDLUs and adjacent neoplastic cells are identical in only a

minority of cases. In addition, in neoplastic cells LOH affects all informative markers mapping to a given locus whereas in normal cells single markers are often involved (Lakhai et al. 1999; Larson et al. 1998). Furthermore, the prevalence of LOH in normal TDLUs seems not to increase according to the proximity with the invasive cancer (Larson et al. 2002). In considering this, the reader should note that LOH does not equate with deletion of a given locus; in fact, there are several lines of evidence to demonstrate that loss of single microsatellite markers often stems from endoreduplication and gene conversion/mitotic recombination (Lambros et al. 2007; Cleton-Jansen et al. 2004). The genetic changes found in morphologically normal cells seem to be subte and show little overlap with the most frequent genetic changes described in DCIS and invasive carcinoma; their mere presence is not sufficient to suggest that these cells may be non-obligate precursors. LOH has also been identified in stromal cells around pre-invasive and invasive lesions. The genetic changes in normal stormal and epithelial cells appear to be independent from distance to the tumor (Moinfar et al. 2000a). finally, the reader is reminded that the majority of studies documenting the presence of LOH in normal epithelial, myoepithelial and, specially, stromal cells of the breast were performed with DNA extracted from microdissected, formalin-fixed paraffin-embedded samples. Formal fixation and paraffin embedding have been shown to cause artifacts of PCR amplification (Campbell et al. 2009: Holliday et al, 2009; Farrand et al. 2002) and may constitute one of the reasons for the surprisingly high prevalence of LOH in normal TDLUs and stromal cells. Notwithstanding the potential artefactual nature of some of the genetic aberrations documented in morphologically normal cells, the identification of LOH in cell (Lakhani et al. 1999(. Studies reporting on analysis of X chromosome inactivation patterns (Tsai et al. 1996( and expression levels of glucose-6-phosphate in patients harboring the Mediterranean mutation of the G6PD gene (Novelli et al. 2003) have supported this hypothesis, and suggest that clonality patches can be larger than sing TDLUs. 5.3.2. Benign Proliferative and Non-Proliferative Breast Lesions Numerous epdeminological studies (Bordian et al. 1993: Kamal et al. 1992) have demonstrated the low relative risk of development of cancer conferred by a diagnosis of benign entities, such as sclerosing adenosis (Fig. 5.1a), fibrosis, and cysts. Data on the genetic features of non-proliferative breast lesions are scant. 5.3.2.1 Radial Scars Radial scars (RS) are benign lesions characterized by obliterated ducts and infiltrating tubules, line by epithelial and myoepithelial cells, surrounded by contracted ducts and lobules exhibiting variable epithelial hyperplasia, duct ectasia, adenosis and papillomatosis (Rosen 2001). The role of RS in breast cancer development is difficult to ascertain based on the available data (Andersen and Gram 1984; Andersen and Battersy 1985; Fisher et al. 1979). On the one hand some studies report an increased risk of breast cancer development (proportional to RS size) (Dupont and Page 1985; Jcobs et al, 1999; Sloane and Mayers 1993; Manfrin et al, 2008), and allelic imbalances often affected in breast cancer (e.g. 16q and 8p) identified through the analysis of formalin-fixed, paraffin-embedded RS samples are consistent with a potential role of RS in breast cancer progression (Iqbal et al, 2002).

Moreover, one must consider reports of a high incidence (32%) of carcinomas in asymptomatic patients with RS (Manfrin et al. 2008) and the fact that invasive cancers found in association with RS were of low histological grade, consistent with previous reports of 16q LOH in RS. Despite the lines of evidence suggesting that at least a subgroup of RSs may be non-obligate precursors, it is clear that additional data obtained from the analysis of fresh/frozen samples with higher resolution methods are required to establish whether RS are risk indicators of breast cancer development or if they constitute true non-obligate precursors of DCIS and invasive breast cancer.