Evolving Paradigms in Triple Negative Breast Cancer: Introduction

This article is part I of a series. View parts II and III: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.¹Furthermore, 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.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.^2,3Pathophysiology of TNBCBreast 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.⁴Triple 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 inTP53andPIK3CA(FIGURE 1).³Figure 1. Intrinsic breast cancer and TNBC subtypes relationships.Basal-like breast cancer and TNBCAdjacent 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 hasTP53mutations. 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.⁵When 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 ofBRCA1function 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.⁴Molecular targets and markers of TNBCWith 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.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 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.³Studies 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).⁶Figure 2. Some potential targeted treatments in TNBC.Epidemiology of TNBCTriple-negative breast cancer has consistent epidemiologic characteristics that further highlight the need for research into targeted therapies for this prognostically poor disease.EthnicityThe 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 withBRCA1mutations.WeightStudies 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.Hormonal and pregnancy factorsThe 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.Other risk factorsData 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.⁵Finally, 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.⁷Diagnosis of TNBCAlthough 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.⁸Typically, 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.⁹The 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.⁸TNBC 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.BRCA1mutations may be found in more than 75% of TNBC and basal-like breast cancer.⁹Metaplastic, 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.⁹Emerging alternative biomarkers for TNBCThere 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.⁸Androgen 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.⁹Another 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.¹⁰Involved 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.⁹Another 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.⁹Treatment of TNBCSurgical treatment of TNBCWith 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.¹In 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 theBRCA1mutation 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.¹¹Research 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 in TNBCRadiation therapy (RT) after BCS has shown to be beneficial for patients with breast cancer long term. Therefore, it is indicated in TNBC after mastectomy or after BCS despite some controversy.¹For example, one study suggested possible radioresistance 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.¹¹Conversely, because 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 aBRCA1gene mutation, which lacks double-stranding DNA repair, theoretically they should be very radiosensitive. Some researchers suggest that RT of the surrounding breast tissue after BCS could eliminate remaining mutatedBRCA1foci and lessen the risk of local or regional recurrence.1Therefore, 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 and OS with the addition of RT to modified radical mastectomy. Treatment may also include neoadjuvant chemotherapy with or without RT. Retrospective data indicate that patients treated with chemotherapy 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.¹²Chemotherapy in TNBCAlthough TNBC is considered to be quite responsive to chemotherapy, a combination of aggressive behavior and metastatic course with a short disease-free period makes the prognosis of TNBC poor. Chemotherapeutic targets include cell proliferation with anthracyclines, P53 with taxanes, and DNA repair complexes with platinum and taxane regimens.¹Platinum-based regimensPlatinum therapies, such as cisplatin, lead to double-strand breaks and cross linking of double-stranded DNA. In cases ofBRCA1mutation, the agent 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 platinum 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.¹³Ezzat et al combined cisplatin with paclitaxel and found a 63% partial response rate and 28% CR rate in patients with HER2-negative and 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.¹Taxane-based regimensTaxanes target genetic instability and have been shown to be beneficial in TNBC and in basal-like breast cancer. Meta-analyses show benefit with adjuvant treatment with taxanes and some research indicates that taxanes may be more effective in receptornegative 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-fluorouracil, epirubicin, and cyclophosphamide (FEC), and then 8 weeks of paclitaxel weekly, demonstrated more benefit than those treated with 6 cycles of FEC.¹³It is unclear if anthracyclines alone are beneficial to patients with TNBC but evidence supports the combination of anthracyclines with taxanes. Despite evidence of TNBC benefiting from regimens that include taxanes, in vitro studies indicate a possible resistance to taxanes in BRCA1- mutated tumors.¹Anthracycline-based regimensAnthracycline-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.¹³Likewise, 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 andBRCA1carriers are as sensitive to anthracyclinecontaining treatments. For example, when looking at patients with TNBC treated with FEC, those who were carriers ofBRCA1had a CR rate of 17% versus 42% in non-BRCA1carrier 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.¹Adjuvant chemotherapy for TNBCBecause 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 algorithmbased NCCN guidelines for node-negative TNBC recommends no adjuvant chemotherapy for breast tumors 0.5 cm or less and chemotherapy in tumors greater than 1 cm. However, 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).¹²Figure 3. General treatment algorithm for adjuvant chemotherapy for early TNBC.Adapted from www.NCCN.org.Neoadjuvant chemotherapy in TNBCWhether 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.¹²Neoadjuvant 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.¹TNBC and basal-like breast cancers have higher CR rates to neoadjuvant breast cancer treatments than luminal breast cancers, despite overall poorer outcomes.¹²There 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 TNBCTNBC 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.¹Several 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.¹²However, several trials have indicated no specific benefit of using taxanes in TNBC over other breast cancer types.¹Platinum agents have shown efficacy inBRCA1cancers 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.¹²Investigational Targets for TNBCBecause there are no approved targeted treatments for TNBC, chemotherapy continues to be the primary treatment of TNBC.¹⁰Identifying 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.¹⁰Patients 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.¹⁴The 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.¹⁵mTOR/AKT/PI3K Pathways and InhibitorsMutations inP53,PTEN, andPI3Kare 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.⁹Loss 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).¹⁶Figure 4. The distribution of TNBC subtypes.Adapted from The Cancer Genome Atlas. Bar graphs illustrate the percentage of subtype in TNBC.In a phase II trial of everolimus and carboplatin for patients with metastatic TNBC, Singh et al found the clinical benefit rate was 36% (95% CI, 21.1-57.4) with an OS of 16.6 months (95% CI, 7.3 months to not reached) and median PFS of 3 months (95% CI, 1.6- 4.6 months). The reported AEs were thrombocytopenia, neutropenia, and anemia.¹⁷A study by Yunokawa et al found that everolimus had favorable activity against basal-like TNBC cell lines. Furthermore, there was an association between CK5/6 and EGFR markers and a positive response to everolimus in TNBCs.¹⁸There are reports that activation of mTOR might be associated with resistance to cisplatin, which in theory could be avoided with the use of everolimus.¹A study looking at temsirolimus with neratinib in patients with metastatic TNBC or HER2-amplified breast cancer is ongoing (NCT001111825). The aim of the open-label study is to identify the benefit and safety of this combination in TNBC or HER2-amplified breast cancer that progressed despite treatment.¹⁹Finally, therapies that target both PI3K and mTOR, such as BEZ235, demonstrate targeted inhibition and tolerability. Furthermore, TNBC with LAR or mesenchymal-like expression,PIK-3CAmutation, and loss of PTEN may be more sensitive to these dual inhibitors.The most common AEs reported with these inhibitors are rash, nausea, diarrhea, mucositis, and hyperglycemia.²⁰Vascular Endothelial Growth Factor PathwayThe highly proliferative nature of TNBC requires that it has persistent angiogenesis and therefore it is logical that these tumors would have higher expression of VEGF. Bevacizumab is an antiVEGF monoclonal antibody.²O&rsquo;Shaughnessy et al conducted a meta-analysis of patients with TNBC with first-line treatments and bevacizumab. Patients demonstrated a 35% decrease in risk of death or disease progression and response rate improved by 19%.²¹Furthermore, when bevacizumab and docetaxel were compared with placebo and docetaxel in EGFR2-negative metastatic breast cancer, patients demonstrated an increase in PFS.²²Subgroup analysis of patients with TNBC revealed an 8.2-month median PFS in the bevacizumab and docetaxel group. Adding bevacizumab to the docetaxel regimen did not worsen the toxicity profile; however, there were more grade 3 to 4 AEs in the bevacizumab group, in cluding febrile neutropenia, neutropenia, and hypertension.²In contrast, the RIBBON-1 trial comparing chemotherapy (capecitabine, taxane-based, or anthracycline-based) with bevacizumab showed improvements in PFS when used as a first-line treatment for metastatic breast cancer.²³However, these results did not show clear benefit with the addition of bevacizumab in patients with TNBC.The RIBBON-2 trial used chemotherapy plus bevacizumab in patients with metastatic breast cancer who had already received cytotoxic therapy. The TNBC group demonstrated an increase in median PFS of 3.3 months (P= .0006).²The use of bevacizumab with anthracycline-based and taxanebased regimens in the neoadjuvant setting resulted in significantly increased CR rates in the GeparQuinto trial.²⁴Recently developed therapies, such as ramucirumab, bind to VEGFR2 and therefore may have more complete angiogenesis inhibition.²A phase III clinical trial of docetaxel plus ramucirumab is under way for patients with HER2-negative metastatic, unresectable, or locally recurrent breast cancer (NCT00703326).²⁵Two therapies target cell surface-receptor tyrosine kinases such as VEGF receptors (VEGFRs), which are important for tumor angiogenesis. Sunitinib and sorafenib are anti-VEGFR tyrosine kinase inhibitors (TKIs) that have been studied in the TNBC subgroup.²Sunitinib was evaluated in a phase II study for patients with metastatic breast cancer who were previously heavily treated with taxanes and anthracyclines. The overall response rate (ORR) was 11%, but 56% required dose adjustment secondary to AEs such as nausea, diarrhea, fatigue, and mucosal inflammation.²⁶The patients with TNBC demonstrated a relative response rate of 15%.²Another multikinase inhibitor is sorafenib, which has activity against tumor angiogenesis and proliferation.²A trial comparing capecitabine and sorafenib with capecitabine and placebo in patients with advanced or metastatic HER2-negative breast cancer found an improvement in PFS in the sorafenib arm. However, the dose used in the study was associated with rash, diarrhea, neutropenia, hypertension, and mucosal inflammation, AEs that required discontinuation in 20% of patients versus 9% in the placebo arm.²⁷Trials specific to the TNBC population are under way for two other TKIs: apatinib28 (NCT01176669) and cediranib29 (NCT01116648).Fibroblast growth factor and receptorFGFR is being investigated in breast cancer because of its role in cellular fibrous proliferation in TNBC. It involves the RAS-RAFmitogen-activated protein kinase (MAPK/ERK) cascade.⁹Molecular profiling of TNBC demonstrated that a subgroup had amplification of FGFR2.30 Furthermore, these cell lines were very susceptible to FGFR inhibition.⁹Lucitanib (E-3810) is an inhibitor of VEGFR1 and FGFR1 and preclinical models highlight its activity against angiogenesis.³¹Using xenografts of advanced TNBC, Bello et al demonstrated lasting tumor regression with a combination of E-3810 and paclitaxel.³²Epidermal growth factor receptorEGFR is another tyrosine kinase within the HER2 family. More than 50% of TNBC and 65% to 72% of basal-line breast cancers have dysregulation of the EGFR pathway.9Adding cetuximab, a monoclonal antibody that targets EGFR, to cisplatin doubled the ORR in patients with metastatic TNBC.³³Likewise, patients demonstrated a longer median PFS and similar OS to cisplatin alone.²The most commonly reported AEs were fatigue, neutropenia, and an acne-like rash.³³In stage IV TNBC, cetuximab plus carboplatin was associated with a response in only 20% of patients. The investigators suggested that although TNBC involves activation of the EGFR pathway, there may be an alternative mechanism for this pathway activation.³⁴Erlotinib used for neoadjuvant treatment of TNBC with carboplatin and docetaxel had a 40% pathologic CR rate. Furthermore, there was a strong correlation between CR andBRCAmutations, with 100% CR in patients withBRCA-mutated disease versus 27% in patients with non-BRCA-mutated disease (P= .006).²Neoadjuvant treatment of TNBC with ixabepilone with and without cetuximab is being studied (NCT01097642).³⁵Androgen receptors in TNBCEvidence suggests that 10% to 35% of all patients with TNBC have AR-positive gene expression.²In patients who are ER positive, AR expression is thought to inhibit proliferation. However, in patients who are ER negative, AR expression may promote tumorigenesis. The efficacy of bicalutamide on TNBC cell lines showed that the LAR subtype was more sensitive.⁹A clinical trial involving bicalutamide in patients with metastatic ER-negative, PR-negative, or AR-positive breast cancer is ongoing (NCT00468715).³⁶Poly (adenosine diphosphate-ribose) polymerase inhibitorsPoly (adenosine diphosphate-ribose) polymerase (PARP) enzymes, especially PARP1, are involved in single-strand DNA break repair, and loss of this function leads to accumulation of these breaks. Furthermore, the breaks are also repaired by pathways that are related toBRCA1andBRCA2. Preclinical results of inhibition of PARP1 demonstrated that TNBC cells are more sensitive than non-TNBC cells. Several PARP inhibitors are being evaluated as possible therapeutic options in TNBC.An oral PARP inhibitor, olaparib, has shown activity againstBRCA-mutated tumors and as a single agent for advanced breast cancer. The reported AEs were mild and included nausea, vomiting, fatigue, and anemia.In a phase I trial of olaparib plus paclitaxel as first-line or second-line treatment of metastatic TNBC, Dent et al demonstrated an objective response rate in 3 of 9 patients in the first cohort and in 4 of 10 patients in the second cohort.²However, this combination had significant interactions and high rates of neutropenia. Other AEs reported where diarrhea and nausea. The investigators suggested considering alternative dosing and scheduling.³⁷Olaparib is also being investigated with paclitaxel or carboplatin in patients with TNBC (NCT0070770738and NCT00516724).³⁹Likewise, olaparib plus cisplatin is being investigated for use in advanced solid tumors (NCT00782574),⁴⁰including locally advanced TNBC.²Veliparib is another oral PARP that targets both PARP1 and PARP2. A single-arm, phase II trial of the combination of veliparib and temozolomide in patients with metastatic breast cancer, including 15 patients with TNBC,²concluded that the combination was active in metastatic breast cancer with one CR and two partial responses. Progression of disease was noted in 14 patients. The reported AEs included grade 3 and 4 thrombocytopenia and neutropenia.⁴¹Although its mechanism is not well understood, iniparib was developed initially as a PARP inhibitor. However, iniparib has been shown to result in cell-cycle arrest, increases in the effects of DNA damaging pathways, and possible antiproliferative activity in TNBC cell lines.²Combining gemcitabine and carboplatin with and without iniparib for metastatic TNBC in an open-label, phase II study demonstrated improved rate of clinical benefit (34%-56%;P= .01) and ORR (32%-52%;P= .02). Likewise, the OS increased from 7.7 months to 12.3 months (HR for death, 0.57;P= .01). The most common AEs in both groups were neutropenia, anemia, fatigue, leukopenia, and thrombocytopenia. There was little difference in the rate of AEs between the two groups, and thus toxicity was not significantly increased with the addition of iniparib.⁴²However, a phase III trial of the gemcitabine, carboplatin, and iniparib combination in advanced TNBC did not meet the study&rsquo;s primary endpoints.²Studies of neoadjuvant treatment of TNBC with paclitaxel alone or with iniparib (NCT01204125)43 and treatment of TNBC with the PARP inhibitor rucaparib (NCT0104970)⁴⁴are under way.Src tyrosine kinase inhibitorsOverexpression of the Src tyrosine kinase is associated with more metastatic progression and invasion in breast cancer.45 Dasatinib is an oral inhibitor of the Src kinases and was originally approved in leukemia with the Philadelphia chromosome.²Pichot et al demonstrated synergistic activity of doxorubicin and dasatinib in breast cancer cells and interference with migration and invasion in the triple-negative cell line.⁴⁶Likewise, synergy was demonstrated with dasatinib, cetuximab, and cisplatin in TNBC cell lines.⁴⁷Cell viability was reduced and apoptosis was induced more often in the three-drug combination versus cisplatin and cetuximab.²Dasatinib is being studied as adjuvant treatment in women with ER-negative breast cancer, with the aim of preventing breast cancer in the unaffected breast (NCT01471106).⁴⁸Side effects associated with dasatinib include cytopenias, bleeding, pulmonary arterial hypertension, and swelling.⁴⁹Histone deacetylase inhibitorsHistone deacetylase inhibitors (HDACis) such as panobinostat target pathways involved in apoptosis, inhibition of angiogenesis, tumor-suppressor genes, cell-cycle arrest, and invasion. In cancers other than TNBC, panobinostat has demonstrated clinical response with limited side effects.²Mitogen-activated protein-kinase kinaseIn TNBC mouse models, inhibition of the mitogen-activated kinase kinase (MEK) with PI3K/mTOR has shown activity.¹²Trametinib is an oral MEK inhibitor under investigation for multiple malignancies.⁵²A study to understand how MEK functions in TNBC is being conducted in which GSK1120212 is given for a short time (NCT01467310).⁵³Likewise, a combination of trametinib and GSK2141795, an Akt inhibitor, is being studied in metastatic TNBC in a phase II trial (NCT01964924).⁵⁴7-Hydroxystauroporine and checkpoint kinase inhibitorsMutations inTP53are common in TNBC and are associated with checkpoint regulation; 7-hydroxystauroporine (UCN-01) is a checkpoint kinase 1 inhibitor.²A phase II trial of the combination of irinotecan and UCN-01 in metastatic TNBC demonstrated limited activity and found that patients with metastatic TNBC and withTP53mutations had a poorer prognosis.⁵⁵The OS in patients with theTP53mutation was 5.5 months versus 20.3 months without theTP53mutation.Proteasome inhibitorsInitially approved for hematologic malignancies, bortezomib is a proteasome inhibitor with preclinical studies indicating activity against breast cancer.²Studies show that bortezomib induces apoptosis in TNBC cell lines⁵⁶and suggest that bortezomib downregulates CIP2A-dependent p-Akt, which may be associated with more aggressive breast cancers.²A recent study by Song et al demonstrated that delivery of bortezomib with nanoparticles increases the circulation half-life and attenuates tumor growth in the basal-like TNBC mice model.⁵⁷Cycline-dependent kinase (CDK) inhibitorsSmall interfering RNAs that target and disable survival gene expression are a potential new area for targeting TNBC cells. For example, Kren et al completed a preclinical study on the use of cyclin dependent kinase 11 (CDK11) and casein kinase 2 (CK2) for RNA interference in TNBC cells. CDK11 is a survival protein kinase involved in the regulation of RNA, mitosis, splicing, and transcription. CK2 is another survival protein kinase that is involved in preventing cancer cell death. Kren et al found that 100% of TNBC cells stained positive for CDK11. When they attempted to downregulate CDK11 and CK2 in the breast cancer cells, they noted a significant decrease in messenger RNA (mRNA) and protein expression, clone survival, and cell viability. It also led to induction of cell death changes.⁵⁸Dinaciclib (MK-7965) is a small molecule that inhibits CDK1, CDK2, CDK5, and CDK9.²A phase II trial investigating the use of dinaciclib versus capecitabine in patients with advanced breast cancer did not show superiority to capecitabine. However, the investigators noted that it did show some antitumor activity and was generally tolerated. The most common AEs were neutropenia, leukopenia, febrile neutropenia, and elevated aspartate aminotransferase.⁵⁹This trial did not evaluate patients with breast cancer based on subpopulations, histologic markers, or molecular markers.²A phase I study evaluating dinaciclib plus epirubicin was conducted to determine the maximum tolerated dose in patients with metastatic TNBC. The dose-limiting toxicities included febrile neutropenia, syncope, and vomiting. The investigators did not escalate the dose past the second cohort because of the toxicities. No treatment responses were noted and the accrual was stopped. The investigators concluded that the combination had substantial toxicities without evidence of efficacy for TNBC.⁶⁰Using TNBC patient-derived xenografts and three established TNBC cell lines, Rajput et al looked at the efficacy of dinaciclib. They concluded that dinaciclib led to cell cycle arrest at the G2/M phase as well as to apoptosis.⁶¹Finally, inBRCA1wild-type TNBC cell lines, investigators demonstrated that the use of CDK inhibition with dinaciclib led to the cancer cells being more sensitive to PARP inhibition with veliparib. The investigators noted that the combination may be an option for TNBC treatment.⁶²Wnt/frizzled pathway targetsWnt signaling, which is involved in cell proliferation, differentiation, and migration, is implicated in several cancers.²Activation of the &beta;-catenin/Wnt pathways is associated with poor clinical outcomes and TNBC.⁶³Transcriptome analysis of TNBC cell lines resulted in 72 Wnt targets that were expressed more often in TNBC, and these may indicate chronic activation of the Wnt pathway in TNBC.⁶⁴Interestingly, the Wnt5a oncogene is thought to be a tumor suppressor gene and signals via the frizzled receptors. When breast cancer cells were analyzed for transcription of Wnt5a, investigators noted lower levels when compared with normal cells. Likewise, in patients with ER-negative status, the lower levels of Wnt5a was associated with a poor outcome.⁶⁵A study recruiting patients (NCT02020291) is investigating a hexapeptide known as Foxy-5 that mimics the activity of the Wnt5a molecule, which is thought to be involved in cancer cell migration. Invasive breast cancers that have lower levels or lack Wnt5a protein are associated with shorter recurrence-free survival. Prospective study participants include those with metastatic breast, colorectal, or prostate cancer and are screened to insure lower or absent levels of Wnt5a.⁶⁶A membrane-bound O-acyltransferase known as PORCN (Porcupine) is involved in a necessary step in the Wnt pathway.⁶⁷In knockout mice models, loss of PORCN is associated with Wnt signaling inhibition.²Research into this pathway led to the development of a PORCN inhibitor known as LGK974. In vitro and in vivo use of LGK974 has demonstrated inhibition of Wnt signaling. LGK974 has been shown to be effective in breast cancer tumor models as well.⁶⁷Solzak et al found that RNA sequencing data of seven TNBC cells demonstrated hyperactivation and overexpression of both Wnt pathways and PI3K/AKT/mTOR pathways. Likewise, they tested cellbased models against BKM120 and LGK974 combined and alone. They found that the combination produced synergy in its ability to reduce cell viability.⁶⁸A phase I, open-label, dose-escalation study of LGK974 is recruiting (NCT01351103). Patients included in the trial are those with pancreatic adenocarcinoma,BRAF-mutant colorectal cancer, and other cancers with documented genetic abnormalities in the Wnt pathway.⁶⁹Prognosis, recurrence, and metastasis of TNBCDespite all the current research into TNBC treatments, TNBC has a poorer prognosis, is diagnosed earlier in life, is more aggressive, and has a shorter relapse-free survival. Furthermore, the risk of dying secondary to relapse is higher in patients with TNBC.⁴After an average follow-up of 96 months for a group of patients with TNBC, Tian et al found that TNBC subtype was inversely associated with OS (P<.05) and that this diagnosis carries a poor prognosis.⁷⁰Prognosis may also be affected by how the patient initially responds to treatment. For example, patients with TNBC treated with neoadjuvant chemotherapy had higher rates of CR compared with patients with non-TNBCs. However, their 3-year OS and 3-year PFS was lower. These patients had higher rates of visceral metastases and shorter survival after recurrence. However, patients with TNBC who achieved CR had a similar survival rate to those with non-TNBC. Residual disease in TNBC after neoadjuvant chemotherapy was found to carry a worse prognosis than that in patients with non-TNBCs who have residual disease.⁷¹Likewise, TNBC has been found to correlate with younger age, higher nuclear grade, and biological aggressiveness. Rhee et al found a relapse rate of 14.7% in TNBC versus 6.6% in non-TNBC, with a 4-year relapse-free survival rate of 85.5% versus 94.2%. A shorter relapse-free survival was associated with triple-negative status, younger age, and close resection margin on multivariant analysis.⁷²Patients with ipsilateral breast recurrence of TNBC were found to have 5-year survival of 72.7% and disease metastasis-free survival of 48.6% despite standard chemotherapy.⁷³Disease-specific survival at 5 years has been found to be 88% in TNBC, 98% in HR+/HER2—, and interestingly, 86% in HR-/HER2+. The study found a 5-year relapse-free survival of 84% in patients with TNBC, 95% in patients with HR+/HER2&ndash;, and 76% in patients with HR-/HER2+ disease. When Kaplan and Malmgren adjusted the model for race, age, TNM status, and treatment, they found a recurrence risk of 2.32 for TNBC (95% CI, 1.32-4.08) with patients with HR+/HER2&ndash; as the reference group.⁷⁴Finally, when compared with non-TNBCs, OS after recurrence has been found to be significantly lower in TNBC. Likewise, TNBC is associated with a significantly lower disease-free survival compared with non-TNBC.⁷⁵Retrospective analyses of distant metastatic patterns in TNBC demonstrated that pulmonary metastasis is associated with the longest metastatic OS, 16.6 months. This is followed by bone metastasis with 16.3 months and liver metastasis with an OS of 8.9 months. Pleural and brain metastases are associated with an OS of 7.5 and 4.3 months, respectively. Location of first metastasis significantly correlated with survival in the TNBC subgroup, with the lowest survival in brain metastasis.⁷⁶Metastasis to the brain has a higher incidence within the first 5 years of a TNBC diagnosis. Therefore, Chikarmane et al suggest that this should help guide metastatic surveillance and imaging.⁷⁷In a cohort of women followed for approximately 8 years, in the first 5 years after diagnosis women with TNBC had a higher likelihood of distant recurrence (HR, 2.6;P<.0001) and death (HR, 3.2;P<.001). This risk of recurrence seemed to peak at the third year after diagnosis and actually rapidly declined following the peak. Dent noted that although TNBC is considered an aggressive breast cancer, this may actually be temporary in the course of the disease. In this cohort the survival time after recurrence in the TNBC group was significantly shorter at 9 months versus 20 months (P= .02).⁷⁸Finally, survival in TNBC is impacted by age, with distant diseasefree survival and OS being shorter in younger patients even with more aggressive treatment strategies. In patients more than 60 years of age, the average disease-free survival was 8 years versus 4 years in patients 31 to 40 years of age.⁷⁹Click here for references:Evolving Paradigms in Triple-Negative Breast Cancer: References> >