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.

Furthermore, 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.

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 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.

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).

Figure 1. Intrinsic breast cancer and TNBC subtypes relationships.

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.

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 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.

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.

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 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.

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).

Figure 2. Some potential targeted treatments in TNBC.

TNBC 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, 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.

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.

Furthermore, these patients had significantly decreased disease-free survival (DFS) compared with patients of normal weight.

The Women&rsquo;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.

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, 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 in 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 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.

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.

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.

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.

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.

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.

A recent study revealed a novel prognostic model using AR expression and BRCA1, stratified by HR status, that may be more predictive of DFS.

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.

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.

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.

Positive 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.

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.

Evolving Paradigms in Triple-Negative Breast Cancer: Introduction

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 anthracycline-based, 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 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.

Evolving Paradigms in Triple-Negative Breast Cancer: Conclusion

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Evolving Paradigms in Triple-Negative Breast Cancer: References

With a lack of ERs, PRs, and HER2 as potential treatment targets, TNBC cannot be treated with conventional breast cancer hormonal-based or trastuzumab-based treatments. 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% women with the BRCA1 mutation have the TNBC subtype that influences the type of surgery selected. Although 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 overall survival (OS) and DFS. 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.

Radiation therapy (RT) after BCS has shown to be beneficial in the long term for patients with breast cancer. Therefore, RT is indicated in TNBC after mastectomy or after BCS despite some controversy.

For example, one study suggested possible radio resistance due to ER-negative status in TNBC. This is thought to be related to more time available for DNA damage repair from radiation in ER-negative cells.

Conversely, as a result of the aggressive and rapid-growing nature of TNBC, RT after BCS may not be as effective as mastectomy. Because many patients with TNBC have a mutated BRCA1 gene, which lacks double-stranded DNA repair, theoretically they should be very radiosensitive. Some researchers suggest that RT of the surrounding breast tissue after BCS could eliminate remaining mutated BRCA1 foci and lessen the risk of local or regional recurrence.

Therefore, in light of the conflicting data, either BCS or modified radical mastectomy is considered an acceptable choice in early stages of TNBC. Research has indicated an improvement in 5-year relapse-free survival (RFS) and OS with the addition of RT to modified radical mastectomy. Treatment may also include neo-adjuvant chemotherapy with or without RT. Retrospective data indicate that patients treated with chemotherapy and reclassified as stage I or II may not benefit from RT compared with patients with persistent nodal disease. However, with a lack of controlled, prospective studies, the standard of care continues to involve RT after neoadjuvant chemotherapy.

Although TNBC is considered to be quite responsive to chemotherapy, the combination of aggressive behavior and metastatic course, with a short disease-free period, can make the prognosis of TNBC poor. Chemotherapeutic targets include cell proliferation with anthracyclines, p53 with taxanes, and DNA repair complexes with platinum and taxane regimens.

Platinum therapies, such as cisplatin, lead to double-strand breaks and cross linking of double-stranded DNA. In cases of the BRCA1 mutation, platinum therapy leads to cell death by inability to repair the damaged DNA. With research indicating that TNBC may be more sensitive to platinum-based regimens, there is a new push for understanding how to incorporate these regimens into breast cancer treatments. For example, 21% of patients with TNBC treated with neoadjuvant cisplatin in a phase II study had complete response (CR) with 50% showing a good response.

Ezzat and colleagues combined cisplatin with paclitaxel and found a 63% partial response rate and a 28% CR rate in patients with HER2-negative, ER-negative breast cancer. Likewise, patients with TNBC treated with a combination of neoadjuvant cisplatin, paclitaxel, and epirubicin for 8 weeks had a 65% pathologic CR.

Taxanes target genetic instability and have been shown to be beneficial in both TNBC and basal-like breast cancer. Meta-analyses show benefit with adjuvant treatment with taxanes, and some research indicates that taxanes may be more effective in receptor-negative cancers. Furthermore, taxanes were associated with a CR of 45% in basal-like breast cancer compared with 6% in luminal subtypes. In addition, patients with TNBC who were treated with 4 cycles of 5-uorouracil, epirubicin, and cyclophosphamide (FEC), followed by 8 weeks of paclitaxel, demonstrated more benefit than those treated with 6 cycles of FEC.

It is unclear if anthracyclines alone are beneficial to patients with TNBC, but evidence supports the combination of anthracyclines with taxanes. Despite evidence of TNBC benefitting from regimens that include taxanes, in vitro studies indicate a possible resistance to taxanes in BRCA1-mutated tumors.

Anthracycline-based chemotherapy targets and destabilizes DNA and thus is considered important in the treatment of breast cancer and TNBC. Liedtke et al compared patients with TNBC with patients with breast cancer but not with TNBC and found that treatment with an anthracycline-containing regimen yielded a 3-year disease-free survival that was similar in both groups (CR 94% versus 98%). However, patients with TNBC who did not meet a pathologic CR had worse outcomes for 3-year disease-free survival than did patients who did not have TNBC (68% vs 88%, P = .0001). This was thought to be secondary to an earlier relapse rate, which was much higher in the TNBC group that had incomplete response.

Likewise, younger patients with TNBC who received high-dose, rapidly cycled treatments containing epirubicin and cyclophosphamide versus a conventional adjuvant regimen demonstrated a 71% 5-year disease-free survival versus 26% in the conventional group.

When taking into the account the heterogeneity of TNBC, it is not clear if TNBC and

carriers are as sensitive to anthracycline-containing treatments. For example, when looking at patients with TNBC treated with FEC, those who were carriers of

carrier TNBC. Finally, pooled data indicate no benefit with anthracycline-containing regimens in HER2-negative breast cancers. Subgroup analyses of studies using anthracyclines in TNBC show mixed results in basal-like breast cancers and TNBC.

Because there are no approved targeted treatments for TNBC, adjuvant treatment with an anthracycline-based and taxane-based regimen for early-stage TNBC is common, based on recommendations of the National Cancer Comprehensive Network (NCCN). The 5-year risk of recurrence in small, node-negative TNBC is low and therefore has a lower treatment benefit. The algorithm-based NCCN guidelines for node-negative TNBC do not recommend adjuvant chemotherapy for breast tumors 0.5 cm or less. For tumors 0.6 cm to 1.0 cm, chemotherapy can be considered after a balanced discussion with the patient. For patients with lymph node-positive TNBC with any size tumor, adjuvant chemotherapy with an anthracycline/taxane-based regimen is recommended in the guidelines (FIGURE 3).

Figure 3. General treatment algorithm for adjuvant chemotherapy for early TNBC.

Whether or not to consider neoadjuvant chemotherapy in TNBC takes into account several factors. The first involves the resectability of the tumor and lymph nodes, with the goal of having clear margins. The second factor is the likelihood that the tumor could be reduced enough to allow for BCS versus mastectomy.

Neoadjuvant chemotherapy allows for treatment efficacy more rapidly than conventional adjuvant therapies. Interestingly, studies comparing neoadjuvant treatment in non-TNBC and TNBC have demonstrated higher response rates and better predicted long-term outcomes in the TNBC groups. Factors that predict a favorable response rate to neoadjuvant treatment are Ki-67 expression and ER negativity.

TNBC and basal-like breast cancers have higher CR rates to neoadjuvant breast cancer treatments than luminal breast cancers, despite overall poorer outcomes.

There is an association between survival outcomes and the rate of CR; therefore, CR is becoming a valuable endpoint when assessing the efficacy of neoadjuvant chemotherapy. Retrospective analysis of neoadjuvant anthracycline/ taxane-based treatment in patients with TNBC demonstrated higher CR rates (38% vs 12%) than in patients with non-TNBC and longer disease-free survival in patients with evidence of CR. Finally, encouraging results and new strategies to manage side effects have led to interest in platinum-based treatments for neoadjuvant treatment of TNBC.

Chemotherapy in metastatic and advanced TNBC

TNBC is characterized as metastatically aggressive with metastases commonly to the brain, liver, and lung. Because there are no standard treatments for metastatic TNBC, treatment planning should consider the patient&rsquo;s performance status and preference, comorbidities, prior treatments, and disease characteristics.

Several systemic treatment options are available for advanced and metastatic HER2-negative breast cancers. Treatment of advanced TNBC follows the same algorithm as for other advanced subtypes, with single chemotherapy agents for asymptomatic disease and combination regimens for rapid progression and symptomatic forms.

Based on a randomized phase III study, the first-line treatment is paclitaxel.

However, several trials have indicated no specific benefit of using taxanes in TNBC over other breast cancer types.

cancers and also have some response in sporadic TNBC. For instance, in first-line and second-line treatment for advanced TNBC both cisplatin and carboplatin demonstrated a 30% response rate.

Because there are no approved targeted treatments for TNBC, chemotherapy continues to be the primary treatment of TNBC.

Identifying potential treatment targets for TNBC may open the door to more personalized treatment.

Glycoprotein nonmetastatic melanoma b (gpNMB)

Glycoprotein nonmetastatic melanoma B overexpression is associated with TNBC, along with other cancers, and is associated with a decrease in OS and disease-free survival and poor prognosis. In breast tissue, gpNMB expression is thought to regulate the ability of cells to allow tumor growth, metastasis, and recruit vasculature.

Glembatumumab vedotin (CDX-011) is an antibody-drug conjugate that binds to gpNMB and releases auristatin E, which leads to cell cycle arrest and subsequent apoptosis. Research has indicated that the efficacy of glembatumumab depends on the amount of gpNMB expressed on the surface of the targeted cell.

Initial studies involved locally advanced or metastatic breast cancer and unresectable melanoma. Patients with breast cancer who received the maximum tolerated dose every 3 weeks demonstrated 62% tumor shrinkage. Furthermore, the progression-free survival (PFS) for patients with gpNMB-positive breast cancer was 17.3 weeks versus 9.1 weeks.

The EMERGE trial aimed to understand the safety and efficacy of glembatumumab in gpNMB-positive metastatic breast cancer with previous intensive treatment.

Patients included in the study had at least 5% of epithelial or stromal cells expressing gpNMB. They were randomly assigned to either investigators&rsquo; choice of chemotherapy (n = 41) or to glembatumumab (n = 83). The investigators concluded that glembatumumab vedotin was well tolerated. However, the primary endpoint was not met for all the study subjects, with overall objective response rate of 6% in the glembatumumab group versus 7%. For tumors with overexpression of gpNMB (greater than 25%) the overall objective response was 30%. The glembatumumab group had less hematologic toxicity but more rash, neuropathy, alopecia, and pruritus.

The METRIC trial is recruiting study participants to demonstrate the effectiveness and safety of glembatumumab vedotin in advanced TNBC (NCT01997333). This randomized, multicenter trial is looking at PFS as the primary outcome measure. Secondary outcomes include objective response rate, OS, adverse events (AEs), and duration of response. Patients will be randomized to either glembatumumab vedotin or to capecitabine.

are associated with TNBC and inhibition studies indicate that targeting mTOR, a serine-threonine protein kinase, may have more antitumor effect than therapies that target tyrosine kinase receptors. The PI3K/AKT pathway regulates the mTOR complexes, and its activation is associated with cellular transformation. Likewise, overexpression of this pathway is related to poor cancer prognosis. Other effects of this pathway include cell migration, transcription, cell cycle progression, and survival.

Loss of PTEN is associated with mTOR activation, which is common to TNBC tumors. Two mTOR inhibitors, everolimus and temsirolimus, have been studied in HER2-negative and TNBC (FIGURE 4).

Figure 4. The distribution of TNBC subtypes.

Adapted from The Cancer Genome Atlas. Bar graphs illustrate the percentage of subtype in TNBC.

Triple Negative Breast Cancer

Evolving Paradigms in Triple-Negative Breast Cancer: Introduction

Evolving Paradigms in Triple-Negative Breast Cancer: Conclusion

Evolving Paradigms in Triple-Negative Breast Cancer: References

Evolving Paradigms in Triple-Negative Breast Cancer: Treatment