Papillary carcinoma of the breast represents approximately 0.5% of all newly diagnosed cases of breast cancer. The prevalence of both invasive and in situ papillary carcinoma seems to be greater older postmenopausal women, and -in relative terms-in males. Histologic features of the tumor include cellular proliferations surrounding fibrovascular cores, with or without invasion. In this review, characteristics of both in situ and invasive disease are outlined. Immunohistochemical analyses of papillary carcinoma suggest the utility of markers such as smooth muscle myosin heavy chain, calponin, p63 and high molecular weight keratins, which can characterize the myoepithelial cell layer. With respect to radiographic evaluation of papillary carcinoma, ultrasonography is the most extensively studied imaging modality, though magnetic resonance mammography has potential utility. Available data suggest improved outcome for papillary carcinoma as compared to invasive ductal carcinoma. Treatment-related information for patients with papillary carcinoma is limited, and patterns noted in available series suggest a variable approach to this disease. The scarcity of information underscores the need for further treatment- and outcome-related studies in papillary carcinoma of the breast.

Purpose

Trastuzumab is a monoclonal antibody targeted to the Her2 receptor and approved for treatment of Her2-positive breast cancer. Among patients who initially respond to trastuzumab therapy, resistance typically arises within one year. BT/HerR cells are trastuzumab-resistant variants of Her2-positive BT474 breast cancer cells. The salient feature of BT/HerR cells is failure to down-regulate PI3K/Akt signaling upon trastuzumab binding. The current work addresses the mechanism of sustained signaling in BT/HerR cells, focusing on the protein kinase A (PKA) pathway.

Experimental Design

We performed microarray analysis on BT/HerR and BT474 cell lines to identify genes that were up- or down-regulated in trastuzumab resistant cells. Specific genes in the PKA pathway were quantified using RT-PCR and Western hybridization. SiRNA transfection was used to determine the effects of gene knockdown on cellular response to trastuzumab. Electrophoretic mobility shift assays were used to measure cAMP-responsive element binding activity under defined conditions. Immunohistochemistry was used to analyze protein expression in clinical samples.

Results

BT/HerR cells had elevated PKA signaling activity and several genes in the PKA regulatory network had altered expression in these cells. Down-regulation of one such gene, the PKA-RIIα regulatory subunit, conferred partial trastuzumab resistance in Her2-positive BT474 and SK-Br-3 cell lines. Forskolin activation of PKA also produced significant protection against trastuzumab-mediated Akt dephosphorylation. In patient samples, PKA signaling appeared to be enhanced in residual disease remaining after trastuzumab-containing neoadjuvant therapy.

Conclusions

Activation of PKA signaling may be one mechanism contributing to trastuzumab resistance in Her2-positive breast cancer. We propose a molecular model by which PKA confers its effects.

Metabolism of oxygen, while central to life, also produces reactive oxygen species (ROS) that have been implicated in processes as diverse as cancer, cardiovascular disease, and aging. It has recently been shown that central nervous system stem cells1, 2 and hematopoietic stem cells and early progenitors3-6 contain lower levels of ROS than their more mature progeny and that these differences appear to be critical for maintaining stem cell function. We hypothesized that epithelial tissue stem cells and their cancer stem cell (CSC) counterparts may also share this property. Here we show that normal mammary epithelial stem cells contain lower concentrations of ROS than their more mature progeny cells. Congruently, subsets of CSCs in some human and murine breast tumors contain lower ROS levels than corresponding non-tumorigenic cells (NTCs). Consistent with ROS being critical mediators of ionizing radiation-induced cell killing7, 8, CSCs in these tumors develop less DNA damage and are preferentially spared after irradiation compared to NTCs. Lower ROS levels in CSCs are associated with increased expression of free radical scavenging systems. Pharmacologic depletion of ROS scavengers in CSCs significantly decreases their clonogenicity and results in radiosensitization. These results indicate that, similar to normal tissue stem cells, subsets of CSCs in some tumors contain lower ROS levels and enhanced ROS defenses compared to their non-tumorigenic progeny, which may contribute to tumor radioresistance.