Targeting Stem Cells in the Realm of Drug-resistant Breast Cancer

BC therapy: conventional approaches

In the genome era, rapid advances in molecular understanding have subdivided BC into ten interclasts.² However, based on the current clinical practices, BC is known to have four primary subtypes. Luminal BCs are positive for steroid hormone receptors (estrogen receptor [ER] and progesterone receptor [PR]), which is further classified into two groups (A and B). The luminal A (ER+/PR+/HER2−) type tumors are less aggressive than other subtypes and take much longer to grow as well. These cancer cells also respond better to hormonal interventions and have a better prognosis.⁴ Luminal B subgroup (ER+/PR+/HER2+) typically shows high Ki67 proliferative index marker as well as HER2 expression. BC cells belonging to luminal B subgroup usually show poorer prognosis than luminal A, but respond better to standard chemotherapy. Since patients of this subgroup also show high HER2 expression, targeted therapy for HER2 might also be employed in some cases.⁴ In HER2+, BCs, which have amplification or overexpression of the HER2/ERBB2 oncogene, are generally treated with anti-HER2 therapies including the antibody drug trastuzumab and small molecule inhibitor lapatinib. Basal-like BC lacks the hormonal receptors as well as HER2 receptor and therefore is often known as triple negative breast cancer (TNBC). Standard chemotherapeutic regimens involving platinum-based drugs are majorly administered for treating TNBCs.

Majority of BC patients (~77%) have hormonal receptor-positive diseases, which comprise 23.7% from ER+/PR+/HER2− (luminal A) and ~53% from ER+/PR+/HER2+ (luminal B). Approximately, 23%–30% of BC patients show HER2 amplification. TNBC represents about 10%–12% of the total BC population.⁴ Endocrine therapy is currently the gold standard treatment regimen to treat the hormone receptor+ BCs. This therapy works either by making the hormone effect ineffective or by lowering the hormone level itself. Therapeutic drugs prescribed to the patients include 1) tamoxifen, which acts by blocking the estrogen uptake by ER; 2) exemestane, anastrozole, and letrozole that belong to aromatase inhibitor class of drugs, which inhibits the conversion of androgens to estrogens thereby depleting estrogen in the body; 3) leuprolide and goserelin (luteinizing hormone-releasing hormone analogs), these drugs suppress the synthesis of hormone from the ovary; and 4) fulvestrant (a specific ER inhibitor), which makes it suitable for refractory BC patients. Administration of the above drugs for treating hormone receptor+ BC is recommended until there is clinical resistance or metastasis, where chemotherapy is employed.⁵ As different endocrine drugs work by distinct mechanism, a combinatorial approach can show improved efficacy. However, the effectiveness of this combination treatment has not been proved well in the patient scenario.⁵ Therefore, the current consensus is that both endocrine therapy-naïve advanced BC and high endocrine-sensitive patients can benefit from the combination endocrine therapy.⁶

The patient group having HER2 gene amplification or protein overexpression is generally administered molecular targeted therapy; a range of targeted drugs have been approved as single agent or in combination with standard chemo regimen. The receptor-targeted therapeutic agents include 1) trastuzumab (specific anti-HER2 monoclonal antibody [mAb]); 2) ado-trastuzumab emtansine, which is trastuzumab conjugated with emtansine (microtubule inhibitor); 3) pertuzumab (specific anti-HER2 mAb with distinct binding site on HER2 extracellular region compared to trastuzumab); 4) lapatinib, a small molecule inhibitor (TKI) capable of inhibiting both HER2 and epidermal growth factor receptor (EGFR) signaling. The standard regimen for early stage HER2+ cases includes neoadjuvant therapy with a combination of HER2 targeted therapy and chemotherapy.⁷ Subsequently, this treatment is followed by surgery, radiotherapy, and 1 year of HER2-targeted therapy. Endocrine adjuvant can be added based on the specific receptor status in patient. The successful advent of molecular targeted therapy against HER2+ BC can be seen by the substantial increase in overall survival (OS) of patients from ~1.5–5 years.⁷

TNBC is aggressive by nature and defiant to treat as well when compared to hormone-positive and HER2+ BC. TNBC can be further subdivided into six subtypes based on transcriptomic heterogeneity and response to chemotherapy. These subtypes are mesenchymal (M), a mesenchymal stem-like (MSL), basal-like (BL1 and BL2), a luminal androgen receptor (LAR), and an immunomodulatory (IM) type.⁸ Both M and MSL subtypes have enhanced expression of factors regulating epithelial–mesenchymal transition (EMT), but intriguingly only the MSL subtype has diminished expression of genes involved in proliferation. The BL1 subtype is categorized by augmented expression of cell cycle and DNA damage repair genes, while the BL2 subtype shows higher expression of growth factor receptors and myoepithelial markers. The LAR subtype is regulated by the androgen receptor (AR) and characterized by luminal gene expression. The IM subtype comprises of BC cells encoding immune checkpoint regulatory genes such as programmed cell death protein 1 (PD-1) and programmed death-ligand 1 (PD-L1), antigens, and immunomodulatory cytokines. Detailed analysis shows activation of immune signal transduction pathways in this subtype, which is likely from both the tumor cells and infiltrating lymphocytes.⁸

Until now, standard chemotherapy remains the mainstay of treatment in TNBCs. The absence of the receptors precludes the application of targeted therapies against advanced stage disease. The only US Food and Drug Administration (FDA)-approved therapy is chemotherapy drugs such as anthracycline, taxane, and platinum drugs with or without bevacizumab.⁹ The median OS of patients with metastatic disease ranges between 9 months and 1 year.⁹ Given the suboptimal treatment outcome with standard therapeutic agents, identification of novel targets and therapy is the need of the hour.

Even with the development of so many different agents, the BC patient scenario is still very disappointing. This can be attributed to the innate biology of the cancer cells to outsmart the current therapies. Over the past decade, it has been identified that cancer cells employ various strategies to overcome the cytotoxic effects such as activation of other signaling pathways, altered metabolism, change in the cell cycle machinery, and epigenetic changes to name a few. This knowledge has led to the development of agents with potential to overcome the resistance of BC cells (Figure 2). In the following sections, we discuss some of the promising therapeutic strategies that are being investigated to treat drug-resistant BC.

BC therapy: developments in the challenging dogma

Hormonal therapy-resistant BC

Development of resistance in hormone receptor-positive BC against their targeted treatment agents is now a well-established phenomenon. Resistant cancer is often metastatic in nature and the underpinning genomic alterations occur majorly in ER cascade. However, other signaling pathways might also get activated and are involved. Mammalian target of phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt)/rapamycin (mTOR) (PI3K–Akt–mTOR) signaling circuit is considered as one of the prime contributing factors to the resistance in a variety of cancers including BC with hormonal drug resistance.¹⁰ This signaling cascade is reported to be overactivated in almost 70% of BC, with PIK3CA (PI3K catalytic subunit p110α) being the frequently mutated and/or amplified genes.¹¹ In addition, activation of escape pathways like HER2 signaling as well as altered cell cycle kinetics has been observed to mediate resistance against ER therapies Thus, to tackle ER+ metastatic BC, there is a need to develop novel therapeutic approach with a potential to either minimize or reverse drug resistance. To address the conundrum of resistance, spectrum of different chemotherapeutic agents has been developed.

PI3K inhibitors

Combinatorial therapies targeting both hormonal receptors and PI3K/AKT/mTOR pathways have been appraised to reverse the resistance to hormonal therapies. Combinatorial treatment with PI3K inhibitors and aromatase inhibitors has been employed as a second-line of treatment for advanced luminal A cases. Buparlisib (a pan-class I PI3K inhibitor) was reported to considerably improve progression-free survival (PFS) in patients, specifically in those having PIK3CA mutation. However, PI3K inhibitors such as pilaralisib,¹² voxtalisib,¹² and buparlisib¹³ cannot be employed in treating patients due to their high toxicity. Recently, taselisib and alpelisib are also under Phase III trials (NCT02340221 and NCT02437318, respectively) and are reported to be efficacious primarily due to their high selectivity and lesser toxicity. These α-specific PI3K inhibitors showed promising results in patients harboring PIK3CA mutations.¹⁴ Irrespective of PIK3CA status, both taselisib¹⁵ and pictillisib¹⁶in combination with letrozole or anastrozole were found to augment antitumor effects in early luminal A patients when employed as neoadjuvant treatment. Buparlisib and alpelisib are currently under Phase II efficacy investigation (NCT01923168).

mTOR inhibitors

Everolimus (derived from sirolimus) has been approved by the US FDA for treating ER/PR + advanced BC in combination with exemestane. Everolimus has also been employed in combination with letrozole, but its clinical efficacy was a failure as it could not reverse the resistant BC.¹⁷ Another derivative of sirolimus, that is, temsirolimus, was a complete defeat as it could hardly show any clinical benefits either as first-line therapy in combination with letrozole or as a single agent in second-line therapy in advanced ER/PR + BCs.¹⁸