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Evolving Paradigms in Triple-Negative Breast Cancer> >

Breast cancer is a leading cause of cancer death in women, with 12% to 20% of cases classified as triple-negative breast cancer (TNBC). These tumors are characterized by a lack of estrogen receptors (ER), progesterone receptors (PR), and human epidermal growth factor receptor (HER2), which limits the use of trastuzumab and hormonebased treatments. Research into treatment options for TNBC is of significance, because it is often diagnosed in younger women and African American women, and because of its aggressive nature, its poor prognosis, and the lack of specific targeted therapies.

Furthermore, when compared with other subtypes, TNBC has a median survival of only12 months and is associated with visceral metastases and recurrence within 3 years of diagnosis.

The 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.

Breast cancer is divided into 18 different 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. Further studies into DNA microarrays allowed division based on gene expressions, including hormone receptors. The breast cancer subtypes luminal A, luminal B, basallike, normal-like, and HER2 positive have individualized treatments and prognoses.

Triple negative breast cancer is a heterogeneous group of tumors with an absence of HER2, ER, and hormone receptor (HR) and can be further divided into apocrine, adenoid cystic, metaplastic, and medullary histopathologic subtypes. Frequently encountered mutations in TNBC include those in

Figure 1. Intrinsic breast cancer and TNBC subtypes relationships.

Adjacent to the basement membrane are basal cells, and there is a subgroup of breast cancer that expresses basal-like genes. Basal-like breast cancer occurs more often in younger patients, is more aggressive, and regularly has

mutations. Basal-like breast cancers mostly fall into to the TNBC classification; however,15% to 54% are positive for PR, ER, and HER2.3 An estimated 75% of patients with TNBC have basal-like markers.

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.

Interestingly, studies have indicated a role for loss of

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.

With the goal of improving survival in TNBC, research is focused on identifying potential molecular targets and markers to guide treatment options for this heterogeneous group of breast cancer.

As stated above, growth factor receptors such as EGFR are expressed in a large portion of basal-like TNBC; other potential targets in TNBC include vascular endothelial growth factor receptor (VEGFR) and fibroblast growth factor receptor (FGFR). Amplification of FGFR1 is found in approximately 9% and FGFR2 is found in 2% to 4% of TNBC.

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 immunomodulary and luminal androgen receptor subtype that involves androgen signaling. Each subtype has distinct pharmacologic targets and can help guide treatment (

Figure 2. Some potential targeted treatments in TNBC.

Triple-negative breast cancer has consistent epidemiologic characteristics that further highlight the need for research into targeted therapies for this prognostically poor disease.

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 of age, to be black or Hispanic, and 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, non-Hispanic black women had a higher incidence of TNBC.

Differences in ethnicity and rates of TNBC represent areas of research to identify mutations or genetic factors that influence younger African-American women to develop TNBC. Also, there is a higher likelihood of high-grade and TNBC with

Studies have also indicated an association between TNBC and obesity. For example, one study found that 50% of patients with TNBC were obese, younger in age, 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 progesterone-receptor and estrogen-receptor negative.

The Women&rsquo;s Health Initiative found an increased risk of ER-positive TNBC and 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 reported a shorter period of breastfeeding. Some studies have also indicated an increased risk of TNBC with oral contraceptive use.

Data from the Women&rsquo;s Health Initiative indicated that TNBC was not associated with tobacco use. Interestingly, when compared with never drinkers, those who drank alcohol had a lower risk of TNBC.

Finally, lower socioeconomic status is associated with a higher risk for developing TNBC.

Including this diverse and vulnerable population at higher risk for TNBC into clinical trials for therapeutic targets is a challenge in breast cancer research.

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.

Typically, mammographic characteristics of TNBC are a hyperdense mass with calcifications, oval- or lobular-shaped mass, and margins that are indistinct or are circumscribed. However, as much as 18% of the 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 but 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.

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 ER or PR 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 (FISH) to insure the 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.

TNBC is typically classified by histology as high-grade invasive ductal carcinoma. Likewise, TNBC may have pushing borders, central necrosis, cellular fibrous proliferation, and thick-walled vessels; TNBC is associated with histologic and intertumor 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, CK14, CK17, CK5/6, laminin, p16, and fatty acid-binding protein.

mutations may be found in more than 75% of TNBC and basal-like breast cancer.

Metaplastic, medullary, and salivary gland breast cancers are typically triple negative on immunophenotypical testing. Metaplastic breast cancers tend to express EGFR, are heterogeneous, and are associated with poor prognosis. Medullary breast cancers are categorized in the basal-like subgroup with well-circumscribed margins, high mitotic rate, diffuse lymphoid infiltrate, syncytial growth pattern, and lack of glandular structures. However, medullary cancers tend to have a better prognosis despite being high grade. Salivary glandtype 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.

There are emerging alternatives to ER, PR, and HER2 as IHC biomarkers for TNBC. These include testing for cytokeratin (CK) 5/6 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.

Androgen receptor stimulates tumorigenesis in ER-negative tumors and is estimated to be expressed in 30% of TNBC. AR is also involved in the activation of HER2 and Wnt signaling pathways by inducing transcription of HER3 and Wnt7B.

Another potential molecular biomarker and therapeutic target for TNBC is the glycoprotein nonmetastatic melanoma 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.

Involved in both basal-like and TNBC, EGFR is a tyrosine kinase that is related to HER2. Gene amplification of EGFR has been reported in TNBC to be zero in one study, to more than 50% in other studies. Likewise, it is reported that EGFR is in 65% to 72% of basallike carcinomas.

Another area of research for molecular markers in TNBC is FGFR, which is involved in cell proliferation, migration, and survival. TNBC of the mesenchymal type has amplified expression of the FGF.

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.

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 coactivation of this pathway in some TNBCs.

With a lack of ER, PR, and HER2 receptors as potential treatment targets, TNBC cannot be treated with conventional breast cancer hormone-based or trastuzumab-based treatments. Therefore, patients are typically treated with a combination of surgery, chemotherapy, and radiation.

In fact, an emphasis is placed on local and regional treatment such as radiation and surgery for early-stage disease.

With the exception of advanced and inoperable metastatic breast cancer, breast-conserving surgery (BCS), simple mastectomy, or radical mastectomy have remained initial treatments for breast cancer. However, 75% to 85% of women with the

mutation have the TNBC subtype that influences the type of surgery selected. While mastectomy to reduce risk of recurrence is preferred over BCS, it is not clear if surgical recommendations should be altered for patients with TNBC.

For example, a retrospective study of patients with TNBC treated with mastectomy compared with BCS assessed OS and disease-free survival. The investigators concluded that BCS in TNBC should be considered in selected patients.

Research also indicates that BCS is an appropriate option in TNBC because recurrence after BCS is no higher than that for other types of breast cancer. Furthermore, when choosing a surgical approach, the preferences of the patient as well as clinical and pathologic variables should be considered.

Articles

Clinical Articles

Breast cancer is a leading cause of cancer death in women, with 12% to 20% of cases
classified as triple-negative breast cancer (TNBC).

Evolving Paradigms in Triple Negative Breast Cancer: Introduction

While TNBC is a heterogeneous group of breast cancers there is increasing effort to subdivide it into more homogenous groups that may help in more accurate prognosis and response to therapy.

Le Du et al propose that five molecular subtypes of TNBC have a high potential to drive clinical trials. These include the basal-like, mesenchymal-like, the immune-associated, androgen receptor overexpression in the luminal/apocrine group, and HER2-enriched.

Because of genetic instability of the tumors, the cancer can have molecular changes throughout the course of the disease. Therefore, it is not known if molecular classification at the time of diagnosis will continue to be predictive of response after treatment as the tumor evolves. Less invasive methods, such as analysis of circulating tumor DNA or circulating tumor cells, may be an accurate method of monitoring clinical response to treatment and personalizing the treatment as the disease progresses. As more research continues into the molecular subtypes of TNBC, there is the potential to uncover additional biomarkers (

Figure 5. Triple-Negative Breast Cancer classifications.

An additional target under research in the treatment of TNBC is the notch signaling pathway, which is involved in self-renewal, angiogenesis, proliferation, and apoptosis. TNBC xenograft models with NOTCH1 rearrangements were found to be sensitive to gamma-secretase inhibitors.

PF-03084014 is a selective gamma-secretase inhibitor that is thought to have antitumor activity by blocking activation of notch receptors.

A phase II, open-label biomarker study of PF- 030804014 is under way for nonmetastatic chemoresistant TNBC (NCT02338531).

There is evidence that the JAK2/STAT3 pathway disruption may be a potential target in TNBC. Janus kinases (JAKs) are another group of tyrosine kinases involved in cell survival and growth. In basal-like breast cancer, JAK/STAT3 was found to be active and inhibition led to decreased growth in the xenograft models.

Ruxolitinib, an IL-6/JAK/STAT inhibitor, is being studied in a phase I trial for recurrent breast cancer, with a phase II portion specifically for preoperative inflammatory TNBC (NCT02041429).

Trop-2 is overexpressed in some epithelial cancers and involved in cell-to-cell adhesion. The US Food and Drug Administration (FDA) gave a fast track designation to IMMU-132 (isactuzumab govitecan) for progressive metastatic TNBC.

An open-label, phase II study that is not yet recruiting will look at IMMU-132 and carboplatin in patients with TNBC (NCT02161679).

Innate immunosurveillance has the ability to identify cancerous cells and destroy them. However, cancer cells characteristically have the ability to evade the immune system. New evidence points to cytotoxic T-lymphocyte antigen (CTLA) inhibitors and programed cell death-1 inhibitors (PD-1) as potential targeted immunotherapies. Furthermore, the PD-1 ligand, PD-L1, is expressed in about 20% of TNBC and confers a poorer prognosis.

Nanda et al presented an abstract for pembrolizumab (MK-3475), a selective humanized monoclonal antibody that blocks PD-1, in the treatment of advanced TNBC. They found that pembrolizumab was tolerated and effective in heavily pretreated TNBC and thus opens the door for immune checkpoint inhibition in TNBC.

New research into the association of CD44(+)/CD24(-) and TNBC found this relationship may play a role in the aggressive nature of TNBC. Therefore, CD44(+)/CD24(-) may be another future targeted therapy in the TNBC population.

Finally, new research into insulin-like growth factor may provide even more options for targeted therapy. For example, insulin-like growth factor-2 mRNA binding protein 3 (IMP3) is expressed in TNBC, and its function is not completely understood. Evidence now points to IMP3 being involved in breast tumor initiation and more aggressive diease.

Likewise, insulin-like growth factor binding protein-3 (IGFBP-3) is highly expressed in TNBC and may function as a tumor promotor. The mechanism involves interaction with EGFR and growth-stimulatory signaling. Furthermore, the interaction of IGFBP-3 and EGFR may be asscoiated with TNBC chemoresistance. In-depth understanding of this relationship is another potential target for TNBC treatment.

Evolving Paradigms in Triple-Negative Breast Cancer: References> >

While TNBC is a heterogeneous group of breast cancers there is
increasing effort to subdivide it into more homogenous groups
that may help in more accurate prognosis and response to therapy.

Future of TNBC Research and Treatment: A Movement Toward Precision Medicine

The treatment of TNBC continues to be a challenge for physicians because of its limited treatment options, the dearth of approved targeted therapies, high mortality, and poor prognosis. Patients and physicians are limited to chemotherapy that targets DNA repair complexes,

, and cell proliferation by using anthracyclinebased, platinum-based, and taxane-based treatment regimens.

However, research into targeted therapies for TNBC is robust. Further understanding of gene expression in breast cancer has led to classifications in TNBC that may help guide research into targeted therapies.

The ultimate goal is to develop personalized, targeted treatments for TNBC. However, this requires a deep understanding of the interaction of the immune system, oncogenic pathways, and the microenvironments of the tumor.80 Understanding the etiology and ultimately prevention of TNBC requires further research using population-based studies.

New research and the potential for targeted, personalized treatment in TNBC allow room for some optimism. However, because TNBC represents a subset of breast cancer, this requires collaboration in trials and research in order to address TNBC on a large scale.

The treatment of TNBC continues to be a challenge for physicians
because of its limited treatment options, the dearth of approved
targeted therapies, high mortality, and poor prognosis. 

Triple Negative Breast Cancer

Evolving Paradigms in Triple Negative Breast Cancer: Introduction

Future of TNBC Research and Treatment: A Movement Toward Precision Medicine

Evolving Paradigms in Triple-Negative Breast Cancer: Conclusion