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Defining the IBC Phenotype

Defining the IBC Phenotype

Inflammatory breast cancer (IBC) is an aggressive and lethal form of breast cancer. It is also an entity for which no consensus exists regarding its clinical definition. The current nomenclature is considered a misnomer since its clinical presentation is not caused by inflammatory components but mainly by lymphatic obstruction. The pathologic hallmark—dermal lymphatic invasion by tumor emboli—may arise in any histopathologic breast carcinoma type and is not sufficient to make the diagnosis.[1] These points should be considered as we examine the molecular determinants behind IBC, and recognize that an improved molecular definition could identify a specific molecular profile for IBC.

What We Know Now
In view of recent advances in molecular profiling and targeted therapies for cancer, there has been much interest in IBC phenotype investigation. It is important to emphasize that besides the classical descriptive markers of breast cancer (hormonal receptors [HR], epidermal growth factor receptor [EGFR], human epidermal growth factor receptor 2 [HER2], and p53) and the specific determinants of IBC (RhoC GTPase and WISP3), so well described by Houchens and Merajver, other molecules or pathways have been well described.

Anderson et al used the definition, “inflammatory carcinoma, including diffuse (beyond that directly overlying the tumor) dermal lymphatic permeation or infiltration” to select IBC cases from the Surveillance, Epidemiology, and End Results (SEER) database and compared them to locally advanced breast cancer (LABC). They concluded that younger age at diagnosis, poorer tumor grade, and negative estrogen receptors were more predictive of IBC than of LABC. These different prognostic factor profiles distinguished two different biologic entities—IBC and LABC.[1] These investigators also observed that the incidence rates for IBC increased until the age of 50 years, reaching a plateau after that, whereas in LABC the rates increased for all ages. For IBC patients, this age effect was not altered by estrogen receptor–positive or –negative status. LABC patients, however, had increased rates of estrogen receptor–positive tumors with advancing age.

Charafe-Jauffret et al defined IBC as “T4d tumors with no mandatory presence of dermal lymphatic emboli” in their analysis of 80 database IBC patients in a comparison with 552 consecutive non-IBC cases.[2] Expression of E-cadherin, EGFR, estrogen receptor (ER), MIB1/Ki67, HER2, MUC1, progesterone receptor (PR), and p53 was studied by immunohistochemistry and tissue microarray. Five markers were significantly associated with IBC in a multivariate analysis: E-cadherin, ER-negative status, MIB1, MUC1 cytoplasmic staining, and HER2 positivity (2+ or 3+). Most of these are not specific markers of IBC. In their analysis, however, the researchers found that a breast tumor with this full phenotype at diagnosis carried a 90.5% probability of an IBC diagnosis.

One of these studied molecules, E-cadherin, has been repeatedly linked to IBC. Interestingly, E-cadherin has a controversial role, is generally thought to act as a tumor suppressor, and is expected to be absent in tumors associated with increased invasiveness and high metastatic potential.[3] Conversely, E-cadherin facilitates intercellular adhesion and enables the formation of cohesive tumor emboli as seen in IBC.[4] E-cadherin–positive staining was shown to be significantly different between cases of IBC (35.4%) and non-IBC (10.3%).[5] Although this protein was expressed in both IBC and non-IBC, in IBC tumors it was extensively present.

In addition to E-cadherin, alterations in LIBC/WISP3 and ARHC/RhoC have been specifically linked to the IBC phenotype.[6] Many of these proteins are altered by hypoxia, environmental stress, and proinflammatory cytokines.[7,8] Avid embolus formation and rapid tumor growth—both prominent features of IBC—could generate hypoxia. Hypoxia is a known factor in driving angiogenesis and contributes to drug and radiation resistance.[9] Additional studies are necessary to identify genetic or epigenetic mechanisms determining this pattern of altered gene expression and to determine the cell types responsible for the altered expression of each gene.

Genetic Mechanisms
Aiming to identify genes specifically linked to a molecular profile of IBC, Biche et al[10] used real-time reverse transcription–polymerase chain reaction (RT-PCR) to quantify the mRNA expression of 538 genes in pooled IBC samples compared to pooled non-IBC samples. They found that 27 genes were upregulated and none were downregulated in IBC. The upregulated genes mainly encoded transcription factors (JUN, FOS, FOSB, MYCN, and SNAIL1), growth factors (vascular endothelial growth factor [VEGF], DTR/HB-EGF, IGFBP7, IL6, ANGPT2, EREG, CCL3, MIP1A, and CCL5/RANTES), and growth factor receptors (TBXA2R, TNFRSF10A/TRAILR1, and ROBO2).

Some genes involved in angiogenesis were upregulated in Biche’s study, whereas others had similar expression levels in IBC and non-IBC. Confirming its angiogenic phenotype, IBC has been associated with increased microvessel density and endothelial cell proliferation in different analyses.[11,12] Van der Auwera et al[13] used real-time RT-PCR and found a significantly higher expression of VEGF-C and VEGF-D in IBC. Although VEGF-A expression was not altered, VEGFR2 was increased in IBC compared to non-IBC. A pilot study was conducted to evaluate the use of bevacizumab (Avastin), an anti-VEGF monoclonal antibody, in IBC. Bevacizumab was used alone for one cycle and then in combination with chemotherapy in previously untreated IBC patients (n = 20) or LABC patients (n = 1), with an objective response rate of 67%.[14] Tumor samples from these patients were collected to study gene expression profiles and gene ontology.[15] The gene classes for VEGFR activity (including genes for platelet-derived growth factor receptors [PDGFR-α and PDGFR-β]) and the gene class for cell motility (including genes for cell adhesion molecule CD31) were significantly associated with responders to bevacizumab and chemotherapy. The proteins CD31 and PDGFR-β were also identified by immunohistochemistry as significantly associated with responders, confirming the gene expression.

Bertucci et al[16] compared gene expression in IBC and non-IBC tumors through DNA microarray. Genes overexpressed in IBC included ARHQ (of RhoC GTPase family), RAB1A (a small GTPase), tyrosine kinase SYK, and FNTA (which encodes farnesyltransferase α subunit). Overexpressed genes in this study also included some genes from the “basal cluster” with underexpression of the “luminal cluster,” findings also described by Van Laere et al in another study.[17] Other overexpressed genes in Bertucci’s analysis are involved in local inflammatory processes (CSCL2, BMP4, SCGB1A1, FPRL1, VCAM1), cell cycle (CCNG2, CDC37, CCT2), apoptosis (DAD1, ALS2CR2), transport (CRABP1, SLC18A2, SLC22A4, SLC2A12), and transcription (ARNT, DTR, NPAS2, SIX3). Genes found underexpressed in IBC (BRE, GPC4, THBS4, PTPRA) encode proteins involved in negative regulation of cell motility, invasion, or angiogenesis.

In a subsequent analysis of the same patients’ samples, Bertucci et al[18] demonstrated that the five molecular subtypes described initially by Perou[19] exist in both non-IBC and IBC tumors, and were associated with similar histologic and clinical features. Their findings were again confirmed by Van Laere et al,[20] who also described a predominance of HER2-overexpressing and basal-like subtypes in IBC, as opposed to a predominance of combined luminal A, luminal B, and normal-like cluster in non-IBC. The five molecular subtypes do not provide the explanation for the differences between IBC and non-IBC in these studies, and their differences are probably linked to other molecular profiles.

The microarray cDNA analysis performed by Van Laere et al[17] to identify IBC’s molecular signature revealed the overexpression of a large number of nuclear factor (NF)-κB target genes. NF-κB is a transcription factor that controls inflammatory gene expression.[21] NF-κB also stimulates tumoral invasiveness, angiogenesis, and lymphangiogenesis.[22] Such activities could explain the aggressive behavior of IBC.

Also studied in IBC is the c-MET proto-oncogene, which encodes for the high-affinity tyrosine kinase receptor for hepatocyte growth factor (HGF) or scatter factor. c-Met and HFG play a role in cell migration, morphogenic differentiation, organization of three-dimensional tubular structures, cell growth, and angiogenesis.[23,24] Deregulation of c-Met and HGF has been shown to correlate with poor outcome in breast carcinoma.[25,26] c-Met protein immunohistochemical expression in 41 IBC tumors was compared with 480 non-IBC infiltrating ductal carcinomas.[5] c-Met was expressed in 100% of IBC cases but in only 64% of non-IBC cases. This study also found that phosphoinositide 3-kinase (PI3K) protein was concomitantly overexpressed with c-Met, supporting the hypothesis that c-Met activation in IBC induces downstream signaling through the PI3K pathway. c-Met is a potential therapeutic target for small molecules or biologic inhibitors of downstream signal transduction.

Future or Next Steps
Many genes and proteins appear to be possible candidates for an IBC genetic or molecular profile. However, none have proven to be specifically and exclusively linked to IBC, and most of the studies have been conducted with small groups of patients, largely because of the relative rareness of IBC. Future studies with larger samples, such as one being initiated in North Africa[27] are absolutely necessary because, as described here, if a specific molecular profile is responsible for IBC vs non-IBC differences, it has not yet been fully determined. It is still unclear as to whether IBC is a distinct entity or one end of the spectrum of breast cancer.

 

References

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