STAT family of transcription factors in breast cancer: Pathogenesis and therapeutic opportunities and challenges.

Breast cancer is the most commonly diagnosed cancer and second-leading cause of cancer deaths in women. Breast cancer stem cells (BCSCs) promote metastasis and therapeutic resistance contributing to tumor relapse. Through activating genes important for BCSCs, transcription factors contribute to breast cancer metastasis and therapeutic resistance, including the signal transducer and activator of transcription (STAT) family of transcription factors. The STAT family consists of six major isoforms, STAT1, STAT2, STAT3, STAT4, STAT5, and STAT6. Canonical STAT signaling is activated by the binding of an extracellular ligand to a cell-surface receptor followed by STAT phosphorylation, leading to STAT nuclear translocation and transactivation of target genes. It is important to note that STAT transcription factors exhibit diverse effects in breast cancer; some are either pro- or anti-tumorigenic while others maintain dual, context-dependent roles. Among the STAT transcription factors, STAT3 is the most widely studied STAT protein in breast cancer for its critical roles in promoting BCSCs, breast cancer cell proliferation, invasion, angiogenesis, metastasis, and immune evasion. Consequently, there have been substantial efforts in developing cancer therapeutics to target breast cancer with dysregulated STAT3 signaling. In this comprehensive review, we will summarize the diverse roles that each STAT family member plays in breast cancer pathobiology, as well as, the opportunities and challenges in pharmacologically targeting STAT proteins and their upstream activators in the context of breast cancer treatment.

Design, synthesis and analysis of novel sphingosine kinase-1 inhibitors to improve oral bioavailability.

The sphingomyelin pathway is important in cell regulation and determining cellular fate. Inhibition of sphingosine kinase isoform 1 (SK1) within this pathway, leads to a buildup of sphingosine and ceramide, two molecules directly linked to cell apoptosis, while decreasing the intracellular concentration of sphingosine-1-phosphate (S1P), a molecule linked to cellular proliferation. Recently, an inhibitor capable of inhibiting SK1 in vitro was identified, but also shown to be ineffective in vivo. A set of compounds designed to assess the impact of synthetic modifications to the hydroxynaphthalene ring region of the template inhibitor with SK1 to obtain a compound with increased efficacy in vivo. Of these fifteen compounds, 4A was shown to have an IC50 = 6.55 µM with improved solubility and in vivo potential.

Correction: Regua et al. TrkA Interacts with and Phosphorylates STAT3 to Enhance Gene Transcription and Promote Breast Cancer Stem Cells in Triple-Negative and HER2-Enriched Breast Cancers. Cancers 2021, 13, 2340.

In the original publication [...].

Antibody-Drug Conjugates for the Treatment of HER2-Positive Breast Cancer.

Human epidermal growth factor receptor 2 (HER2) receptor tyrosine kinase is overexpressed in 20-30% of breast cancers and is associated with poor prognosis and worse overall patient survival. Most women with HER2-positive breast cancer receive neoadjuvant chemotherapy plus HER2-targeted therapies. The development of HER2-directed therapeutics is an important advancement in targeting invasive breast cancer. Despite the efficacy of anti-HER2 monoclonal antibodies, they are still being combined with adjuvant chemotherapy to improve overall patient outcomes. Recently, significant progress has been made towards the development of a class of therapeutics known as antibody-drug conjugates (ADCs), which leverage the high specificity of HER2-targeted monoclonal antibodies with the potent cytotoxic effects of various small molecules, such as tubulin inhibitors and topoisomerase inhibitors. To date, two HER2-targeting ADCs have been approved by the FDA for the treatment of HER2-positive breast cancer: Ado-trastuzumab emtansine (T-DM1; Kadcyla®) and fam-trastuzumab deruxtecan-nxki (T-Dxd; Enhertu®). Kadcyla and Enhertu are approved for use as a second-line treatment after trastuzumab-taxane-based therapy in patients with HER2-positive breast cancer. The success of ADCs in the treatment of HER2-positive breast cancer provides novel therapeutic advancements in the management of the disease. In this review, we discuss the basic biology of HER2, its downstream signaling pathways, currently available anti-HER2 therapeutic modalities and their mechanisms of action, and the latest clinical and safety characteristics of ADCs used for the treatment of HER2-positive breast cancer.

IL-6/JAK/STAT3 Signaling in Breast Cancer Metastasis: Biology and Treatment.

Breast cancer is the most commonly diagnosed cancer in women. Metastasis is the primary cause of mortality for breast cancer patients. Multiple mechanisms underlie breast cancer metastatic dissemination, including the interleukin-6 (IL-6)-mediated signaling pathway. IL-6 is a pleiotropic cytokine that plays an important role in multiple physiological processes including cell proliferation, immune surveillance, acute inflammation, metabolism, and bone remodeling. IL-6 binds to the IL-6 receptor (IL-6Rα) which subsequently binds to the glycoprotein 130 (gp130) receptor creating a signal transducing hexameric receptor complex. Janus kinases (JAKs) are recruited and activated; activated JAKs, in turn, phosphorylate signal transducer and activator of transcription 3 (STAT3) for activation, leading to gene regulation. Constitutively active IL-6/JAK/STAT3 signaling drives cancer cell proliferation and invasiveness while suppressing apoptosis, and STAT3 enhances IL-6 signaling to promote a vicious inflammatory loop. Aberrant expression of IL-6 occurs in multiple cancer types and is associated with poor clinical prognosis and metastasis. In breast cancer, the IL-6 pathway is frequently activated, which can promote breast cancer metastasis while simultaneously suppressing the anti-tumor immune response. Given these important roles in human cancers, multiple components of the IL-6 pathway are promising targets for cancer therapeutics and are currently being evaluated preclinically and clinically for breast cancer. This review covers the current biological understanding of the IL-6 signaling pathway and its impact on breast cancer metastasis, as well as, therapeutic interventions that target components of the IL-6 pathway including: IL-6, IL-6Rα, gp130 receptor, JAKs, and STAT3.

Breast cancer extracellular vesicles-derived miR-1290 activates astrocytes in the brain metastatic microenvironment via the FOXA2→CNTF axis to promote progression of brain metastases.

Mechanisms underlying breast cancer brain metastasis (BCBM) are still unclear. In this study, we observed that extracellular vesicles (EVs) secreted from breast cancer cells with increased expression of tGLI1, a BCBM-promoting transcription factor, strongly activated astrocytes. EV-derived microRNA/miRNA microarray revealed tGLI1-positive breast cancer cells highly secreted miR-1290 and miR-1246 encapsulated in EVs. Genetic knockin/knockout studies established a direct link between tGLI1 and both miRNAs. Datamining and analysis of patient samples revealed that BCBM patients had more circulating EV-miRs-1290/1246 than those without metastasis. Ectopic expression of miR-1290 or miR-1246 strongly activated astrocytes whereas their inhibitors abrogated the effect. Conditioned media from miR-1290- or miR-1246-overexpressing astrocytes promoted mammospheres. Furthermore, miRs-1290/1246 suppressed expression of FOXA2 transcription repressor, leading to CNTF cytokine secretion and subsequent activation of astrocytes. Finally, we conducted a mouse study to demonstrate that astrocytes overexpressing miR-1290, but not miR-1246, enhanced intracranial colonization and growth of breast cancer cells. Collectively, our findings demonstrate, for the first time, that breast cancer EV-derived miR-1290 and miR-1246 activate astrocytes in the brain metastatic microenvironment and that EV-derived miR-1290 promotes progression of brain metastases through the novel EV-miR-1290→FOXA2→CNTF signaling axis.

TrkA Interacts with and Phosphorylates STAT3 to Enhance Gene Transcription and Promote Breast Cancer Stem Cells in Triple-Negative and HER2-Enriched Breast Cancers.

JAK2-STAT3 and TrkA signaling pathways have been separately implicated in aggressive breast cancers; however, whether they are co-activated or undergo functional interaction has not been thoroughly investigated. Herein we report, for the first time that STAT3 and TrkA are significantly co-overexpressed and co-activated in triple-negative breast cancer (TNBC) and HER2-enriched breast cancer, as shown by immunohistochemical staining and data mining. Through immunofluorescence staining-confocal microscopy and immunoprecipitation-Western blotting, we found that TrkA and STAT3 co-localize and physically interact in the cytoplasm, and the interaction is dependent on STAT3-Y705 phosphorylation. TrkA-STAT3 interaction leads to STAT3 phosphorylation at Y705 by TrkA in breast cancer cells and cell-free kinase assays, indicating that STAT3 is a novel substrate of TrkA. ß-NGF-mediated TrkA activation induces TrkA-STAT3 interaction, STAT3 nuclear transport and transcriptional activity, and the expression of STAT3 target genes, SOX2 and MYC. The co-activation of both pathways promotes breast cancer stem cells. Finally, we found that TNBC and HER2-enriched breast cancer with JAK2-STAT3 and TrkA co-activation are positively associated with poor overall metastasis-free and organ-specific metastasis-free survival. Collectively, our study uncovered that TrkA is a novel activating kinase of STAT3, and their co-activation enhances gene transcription and promotes breast cancer stem cells in TNBC and HER2-enriched breast cancer.

Hedgehog Signaling and Truncated GLI1 in Cancer.

The hedgehog (HH) signaling pathway regulates normal cell growth and differentiation. As a consequence of improper control, aberrant HH signaling results in tumorigenesis and supports aggressive phenotypes of human cancers, such as neoplastic transformation, tumor progression, metastasis, and drug resistance. Canonical activation of HH signaling occurs through binding of HH ligands to the transmembrane receptor Patched 1 (PTCH1), which derepresses the transmembrane G protein-coupled receptor Smoothened (SMO). Consequently, the glioma-associated oncogene homolog 1 (GLI1) zinc-finger transcription factors, the terminal effectors of the HH pathway, are released from suppressor of fused (SUFU)-mediated cytoplasmic sequestration, permitting nuclear translocation and activation of target genes. Aberrant activation of this pathway has been implicated in several cancer types, including medulloblastoma, rhabdomyosarcoma, basal cell carcinoma, glioblastoma, and cancers of lung, colon, stomach, pancreas, ovarian, and breast. Therefore, several components of the HH pathway are under investigation for targeted cancer therapy, particularly GLI1 and SMO. GLI1 transcripts are reported to undergo alternative splicing to produce truncated variants: loss-of-function GLI1ΔN and gain-of-function truncated GLI1 (tGLI1). This review covers the biochemical steps necessary for propagation of the HH activating signal and the involvement of aberrant HH signaling in human cancers, with a highlight on the tumor-specific gain-of-function tGLI1 isoform.

Combined inhibition of JAK2-STAT3 and SMO-GLI1/tGLI1 pathways suppresses breast cancer stem cells, tumor growth, and metastasis.

Triple-negative breast cancer (TNBC) and HER2-positive breast cancer are particularly aggressive and associated with unfavorable prognosis. TNBC lacks effective treatments. HER2-positive tumors have treatment options but often acquire resistance to HER2-targeted therapy after initial response. To address these challenges, we determined whether novel combinations of JAK2-STAT3 and SMO-GLI1/tGLI1 inhibitors synergistically target TNBC and HER2 breast cancer since these two pathways are concurrently activated in both tumor types and enriched in metastatic tumors. Herein, we show that novel combinations of JAK2 inhibitors (ruxolitinib and pacritinib) with SMO inhibitors (vismodegib and sonidegib) synergistically inhibited in vitro growth of TNBC and HER2-positive trastuzumab-resistant BT474-TtzmR cells. Synergy was also observed against breast cancer stem cells. To determine if the combination is efficacious in inhibiting metastasis, we treated mice with intracardially inoculated TNBC cells and found the combination to inhibit lung and liver metastases, and prolong host survival without toxicity. The combination inhibited orthotopic growth, VEGF-A expression, and tumor vasculature of both TNBC and HER2-positive trastuzumab-refractory breast cancer. Lung metastasis of orthotopic BT474-TtzmR xenografts was suppressed by the combination. Together, our results indicated that dual targeting of JAK2 and SMO resulted in synergistic suppression of breast cancer growth and metastasis, thereby supporting future clinical testing.