Evolving Paradigms in Triple-Negative Breast Cancer: Introduction

Breast cancer is a leading cause of cancer deaths in women, with 12% to 20% of cases being classified as triple-negative breast cancer (TNBC). These tumors are characterized by a lack of estrogen receptors (ERs), progesterone receptors (PRs), and human epidermal growth factor receptors (HER2), which limits the use of trastuzumab and hormonal-based treatments.

Research into treatment options for TNBC is crucial because it is often diagnosed in younger women and African American women and because of its aggressive nature, poor prognosis, and lack of specific targeted therapy.¹Furthermore, compared with other subtypes, TNBC has a median survival of only 12 months and is associated with visceral metastases and recurrence within 3 years of diagnosis.²The heterogeneous nature and limited biomarkers of TNBC have restricted the development of drugs specific to TNBC; however, new research has opened the door for targeted therapies based on distinct morphologic features.^2,3

Pathophysiology of TNBC

Breast cancer is divided into 18 subtypes based on histology and morphologic characteristics. However, this does not take into account disease-specific treatment options and prognosis. Furthermore, accuracy of classification is pathologist-dependent. Additional studies into DNA microarrays allowed division based on gene expression, including expression of hormone receptors. The breast cancer subtypes luminal A, luminal B, basal-like, normal-like, and HER2-positive have individualized treatments and prognoses.⁴

TNBC is a heterogeneous group of tumors with an absence of HER2, ER, and PR that can be further divided into apocrine, adenoid cystic, metaplastic, and medullary histopathologic subtypes. Frequently encountered mutations in TNBC include those in TP53 and PIK3CA (FIGURE 1).³

Figure 1. Intrinsic breast cancer and TNBC subtypes relationships.

Basal-like breast cancer and TNBC

Adjacent to the basement membrane are basal cells. There is a subtype of breast cancer that expresses basal-like genes. Basal-like breast cancer occurs more often in younger patients, is more aggressive, and regularly has TP53 mutations. Basal-like breast cancers mostly fall into the TNBC classification; however, 15% to 54% are positive for PRs, ERs, and HER2.³An estimated 75% of patients with TNBC have basal-like markers.⁵

When taking into account gene expression profiling related to treatment response and prognosis, basal-like breast cancers form a homogenous group, while TNBC does not. Therefore, the actual poor prognosis of TNBC may be due to the basal-like subtype that makes up a large portion of TNBC.⁴Interestingly, studies have indicated a role for loss of BRCA1 function in the development of some basal-like TNBC along with DNA methylation. Likewise, studies have indicated a relationship to basal-like TNBC with epidermal growth factor receptor (EGFR) expression, which is associated with cellular growth and poor outcomes.⁴

Molecular targets and markers of TNBC

With the goal of improving survival in TNBC, research is focusing on identifying potential molecular targets and markers to guide treatment options for this heterogeneous group of breast cancers.⁶As stated above, growth factor receptors such as EGFR are expressed in a large portion of basal-like TNBC; other potential targets in TNBC include the vascular endothelial growth factor receptor (VEGFR) and broblast growth factor receptor (FGFR). Amplification of FGFR1 is found in approximately 9% of TNBC and FGFR2 in 2% to 4% of TNBC.³

Studies have indicated that TNBC can be divided into distinct subtypes that have unique responses to treatment. For example, basal-like subtype 1 is associated with DNA and cell cycle damage. Basal-like subtype 2 has myoepithelial markers and growth factors that signal amplification. Gene expression in growth factors and differentiation characterize the mesenchymal subtypes. Finally, there is an immunomodulatory and luminal androgen receptor subtype that involves androgen signaling. Each subtype has distinct pharmacologic targets and can help guide treatment (FIGURE 2).⁶

Figure 2. Some potential targeted treatments in TNBC.

Epidemiology of TNBC

TNBC has consistent epidemiologic characteristics that further highlight the need for research into targeted therapies for this prognostically poor disease.

Ethnicity

The Carolina Breast Cancer Study found a higher rate of basal-like breast cancer in premenopausal African American women compared with postmenopausal African American women and non—African American women. Furthermore, the basal-like subtype is associated with a shorter survival, which may help explain the poorer prognosis for breast cancer in young African American women. Likewise, data from the California Cancer Registry showed that women with TNBC were more likely to be younger than 40 years, to be African American or Hispanic, and to have a poor prognosis, with a 5-year survival of 14% in black women.

When looking at invasive breast cancers in women of all ages, African Americans had a higher incidence of TNBC, with a higher likelihood of high-grade disease with BRCA1 mutations. Differences in ethnicity and rates of TNBC represent areas of research to identify mutations or genetic factors that in uence younger African American women to develop TNBC.

Weight

Studies have also indicated an association between TNBC and obesity. For example, one study found that 50% of patients with TNBC were obese, younger, and had larger tumors. Likewise, obesity was associated with lymph node metastasis in breast cancer irrespective of receptor classification. When not accounting for HER2 status, being overweight, having recently given birth, black race, and younger at first pregnancy were associated with breast cancer that was PR- and ER-negative. A retrospective study found that being overweight or obese was significantly correlated with TNBC progression.⁷Furthermore, these patients had significantly decreased disease-free survival (DFS) compared with patients of normal weight.

Hormonal and pregnancy factors

The Women’s Health Initiative found an increased risk of ER-positive TNBC and a decreased risk of TNBC with nulliparous women. Likewise, a higher number of births was positively associated with TNBC risk. When comparing TNBC to non-TNBC patients, women with TNBC also reported a shorter period of breastfeeding; a recent study presented evidence on the protective effects of longer lactation against pregnancy-associated TNBC.⁸Some studies have also indicated an increased risk of TNBC with oral contraceptive use.

Other risk factors

Data from the Women’s Health Initiative indicated that TNBC was not associated with tobacco use. Interestingly, compared with never-drinkers, those who drank alcohol had a lower risk of TNBC.⁵Finally, lower socioeconomic status is associated with a higher risk for developing TNBC. Including this diverse and vulnerable population at higher risk for TNBC in clinical trials for therapeutic targets is a challenge in breast cancer research.^9​

Diagnosis of TNBC

Although mammography and ultrasound may be able to pick up on the smooth borders associated with TNBC tumors, in practice, diagnosis of TNBC typically combines imaging and immunohistochemical (IHC) techniques.¹⁰

Typically, mammographic characteristics of TNBC are a hyperdense mass with calcications, oval- or lobular-shaped mass, and margins that are indistinct or are circumscribed. However, as many as 18% of cases of TNBC are not clear on mammogram. Sensitivity of detecting TNBC with ultrasound is high (92%-100%) and can reveal a distinct and circumscribed mass and, possibly with posterior acoustic enhancement, can indicate fluid or necrosis. However, these features are common in abscesses, cysts, and benign tumors, as well.

Similarly, magnetic resonance imaging (MRI) has a high sensitivity for detecting TNBC. The most common and predictive feature of TNBC found on MRI is rim enhancement (76%) with smooth margins, lobular shape, mass enhancement, and elevated T2 signaling within the tumor.

Ultimately, combining MRI, ultrasound, and mammography has the highest sensitivity for diagnosing breast cancer—even though it may be associated with an elevated false-positive rate. Furthermore, most TNBCs are not detected with imaging, but after they are palpable masses or have led to nipple discharge or pain.¹¹

The College of American Pathologists (CAP) and the American Society of Clinical Oncology (ASCO) have published guidelines using IHC methods to determine the HER2, PR, and ER status of breast cancer. Several important considerations have been emphasized. First, a tumor cell should be considered positive for ERs or PRs if at least 1% is immunoreactive versus previous thresholds of less than 10%. Also, it is important to use normal breast tissue as an internal control to avoid false-negative results. When HER2 status is equivocal, ASCO/CAP guidelines advocate using fluorescent in situ hybridization to ensure proper classification and allow for appropriate treatment choices. Finally, metastatic or relapsed breast cancer may have different characteristics from primary malignancy and, therefore, a confirmatory biopsy may be needed.¹⁰

TNBC is typically classified by histology as high-grade invasive ductal carcinoma. Likewise, TNBC may have pushing borders, central necrosis, cellular brous proliferation, and thick-walled vessels; TNBC is associated with histologic and intratumor heterogeneity. TNBC and basal-like breast cancers share many histologic features, such as high-grade tumors, stromal lymphocytic response, and pushing borders. However, basal-like tumors may express other biomarkers, such as p53, EGFR, cyto-keratin (CK)14, CK17, CK5/6, laminin, p16, and fatty acid-binding protein. BRCA1 mutations may be found in more than 75% of TNBC and basal-like breast cancer.¹¹

Metaplastic, medullary, and salivary gland breast cancers are typically triple negative on immunophenotypic testing. Metaplastic breast cancers tend to express EGFR, are heterogeneous, and are associated with poor prognosis. Medullary breast cancers are categorized into the basal-like subgroup with well-circumscribed margins, a high mitotic rate, diffuse lymphoid in ltrate, syncytial growth pattern, and lack of glandular structures. However, medullary cancers tend to have a better prognosis despite being high-grade. Salivary gland-type breast cancers are uncommon and, although typically triple negative, are considered low-grade. The adenoid cystic carcinoma is also uncommon and known to express TP63 and c-KIT.¹¹

Emerging alternative biomarkers for TNBC

There are emerging alternatives to ERs, PRs, and HER2 as IHC biomarkers for TNBC. These include testing for CK5/6 and EGFR for basal-like breast cancer. Likewise, testing for other markers, such as androgen receptor (AR)-positive gene expression and DNA repair alterations like BRCA1, open the door for potential targets for TNBC and the basal-like subtype.¹⁰

The AR stimulates tumorigenesis in ER-negative tumors and is estimated to be expressed in 30% of TNBCs. ARs are also involved in the activation of HER2 and Wnt-signaling pathways by inducing transcription of HER3 and Wnt7B.¹¹A recent study revealed a novel prognostic model using AR expression and BRCA1, stratified by HR status, that may be more predictive of DFS.¹²

Another potential molecular biomarker and therapeutic target for TNBC is the glycoprotein non-metastatic gene B (gpNMB), which is expressed in many cancers, especially in basal-like and TNBC. gpNMB is often expressed in aggressive, metastatic breast cancers and is associated with an invasive phenotype. Evidence suggests that gpNMB allows for breast cancer cells to metastasize and grow secondary to vasculature recruitment.¹³

Involved in both basal-like and TNBC, EGFR is a tyrosine kinase and related to HER2. However, the range of gene amplication of EGFR in TNBC was reported to be zero in one study compared with more than 50% in other studies. Likewise, it is reported that EGFR is present in 65% to 72% of basal-like carcinomas.¹¹

Another area of research for molecular markers in TNBC is FGFR, which is involved in cell proliferation, migration, and survival. Mesenchymal-type TNBC has amplifed expression of FGFR. Tumor progression depends on angiogenesis and the VEGF pathway. High levels of VEGF are typically found in TNBC and may be associated with a poorer prognosis.

A recent study evaluated fascin-1, an actin-bundling protein, via immunohistochemical analysis as a novel diagnostic marker of TNBC.¹⁴Positive and strong expression rates of fascin-1 were significantly higher in TNBC cases compared with HER2-enriched, luminal A, or luminal B breast cancer subtypes. Sensitivity and specificity approached 90% in this study, suggesting the use of strong positive fascin-1 expression as a diagnostic marker.

Finally, mammalian target of rapamycin (mTOR), a serine-threonine protein kinase, is involved in the PI3K/AKT pathway, which affects cellular transformation and cell migration. Studies have indicated co-activation of this pathway in some TNBCs.¹¹