Chemoresistance is a major obstacle in triple negative breast cancer (TNBC), the most aggressive breast cancer subtype. Here we identify hypoxia-induced ECM re-modeler, lysyl oxidase (LOX) as a key inducer of chemoresistance by developing chemoresistant TNBC tumors in vivo and characterizing their transcriptomes by RNA-sequencing. Inhibiting LOX reduces collagen cross-linking and fibronectin assembly, increases drug penetration, and downregulates ITGA5/FN1 expression, resulting in inhibition of FAK/Src signaling, induction of apoptosis and re-sensitization to chemotherapy. Similarly, inhibiting FAK/Src results in chemosensitization. These effects are observed in 3D-cultured cell lines, tumor organoids, chemoresistant xenografts, syngeneic tumors and PDX models. Re-expressing the hypoxiarepressed miR-142-3p, which targets HIF1A, LOX and ITGA5, causes further suppression of the HIF-1α/LOX/ITGA5/FN1 axis. Notably, higher LOX, ITGA5, or FN1, or lower miR-142-3p levels are associated with shorter survival in chemotherapy-treated TNBC patients. These results provide strong pre-clinical rationale for developing and testing LOX inhibitors to overcome chemoresistance in TNBC patients.

Trastuzumab-refractory, HER2 (human epidermal growth factor receptor 2)-positive breast cancer is commonly treated with trastuzumab emtansine (T-DM1), an antibody-drug conjugate of trastuzumab and the microtubule-targeting agent, DM1. However, drug response reduces greatly over time due to acquisition of resistance whose molecular mechanisms are mostly unknown. Here, we uncovered a novel mechanism of resistance against T-DM1 by combining whole transcriptome sequencing (RNA-Seq), proteomics and a targeted small interfering RNA (siRNA) sensitization screen for molecular level analysis of acquired and de novo T-DM1-resistant models of HER2-overexpressing breast cancer. We identified Polo-like kinase 1 (PLK1), a mitotic kinase, as a resistance mediator whose genomic as well as pharmacological inhibition restored drug sensitivity. Both acquired and de novo resistant models exhibited synergistic growth inhibition upon combination of T-DM1 with a selective PLK1 inhibitor, volasertib, at a wide concentration range of the two drugs. Mechanistically, T-DM1 sensitization upon PLK1 inhibition with volasertib was initiated by a spindle assembly checkpoint (SAC)-dependent mitotic arrest, leading to caspase activation, followed by DNA damage through CDK1-dependent phosphorylation and inactivation of Bcl-2/xL. Furthermore, we showed that Ser70 phosphorylation of Bcl-2 directly regulates apoptosis by disrupting the binding to and sequestration of the pro-apoptotic protein Bim. Importantly, T-DM1 resistance signature or PLK1 expression correlated with cell cycle progression and DNA repair, and predicted a lower sensitivity to taxane/trastuzumab combination in HER2-positive breast cancer patients. Finally, volasertib in combination with T-DM1 greatly synergized in models of T-DM1 resistance in terms of growth inhibition both in three dimensional (3D) cell culture and in vivo. Altogether, our results provide promising pre-clinical evidence for potential testing of T-DM1/volasertib combination in T-DM1 refractory HER2-positive breast cancer patients for whom there is currently no treatment available.

An important goal of self-assembly research is to develop a general methodology applicable to almost any material, from the smallest to the largest scales, whereby qualitatively identical results are obtained independently of initial conditions, size, shape and function of the constituents. Here, we introduce a dissipative self-assembly methodology demonstrated on a diverse spectrum of materials, from simple, passive, identical quantum dots (a few hundred atoms) that experience extreme Brownian motion, to complex, active, non-identical human cells (~10 17 atoms) with sophisticated internal dynamics. Autocatalytic growth curves of the self-assembled aggregates are shown to scale identically, and interface fluctuations of growing aggregates obey the universal Tracy-Widom law. Example applications for nanoscience and biotechnology are further provided.

◥TACC3, a transforming acidic coiled-coil (TACC) family member, is frequently upregulated in a broad spectrum of cancers, including breast cancer. It plays critical roles in protecting microtubule stability and centrosome integrity that is often dysregulated in cancers; therefore, making TACC3 a highly attractive therapeutic target. Here, we identified a new TACC3-targeting chemotype, BO-264, through the screening of in-house compound collection. Direct interaction between BO-264 and TACC3 was validated by using several biochemical methods, including drug affinity responsive target stability, cellular thermal shift assay, and isothermal titration calorimetry. BO-264 demonstrated superior antiproliferative activity to the two currently reported TACC3 inhibitors, especially in aggressive breast cancer subtypes, basal and HER2þ, via spindle assembly checkpoint-dependent mitotic arrest, DNA damage, and apoptosis, while the cytotoxicity against normal breast cells was negligible. Furthermore, BO-264 significantly decreased centrosomal TACC3 during both mitosis and interphase. BO-264 displayed potent antiproliferative activity ($90% have less than 1 mmol/L GI 50 value) in the NCI-60 cell line panel compromising of nine different cancer types. Noteworthy, BO-264 significantly inhibited the growth of cells harboring FGFR3-TACC3 fusion, an oncogenic driver in diverse malignancies. Importantly, its oral administration significantly impaired tumor growth in immunocompromised and immunocompetent breast and colon cancer mouse models, and increased survival without any major toxicity. Finally, TACC3 expression has been identified as strong independent prognostic factor in breast cancer and strongly prognostic in several different cancers. Overall, we identified a novel and highly potent TACC3 inhibitor as a novel potential anticancer agent, inducing spindle abnormalities and mitotic cell death.

Centrosome amplification (CA) is a hallmark of cancer that is strongly associated with highly aggressive disease and worse clinical outcome. Clustering extra centrosomes is a major coping mechanism required for faithful mitosis of cancer cells with CA that would otherwise undergo mitotic catastrophe and cell death. However, its underlying molecular mechanisms have not been fully described. Furthermore, little is known about the processes and players triggering aggressiveness of cells with CA beyond mitosis. Here, we identified Transforming Acidic Coiled-Coil Containing Protein 3 (TACC3) to be overexpressed in tumors with CA, and its high expression is associated with dramatically worse clinical outcome. We demonstrated, for the first time, that TACC3 forms distinct functional interactomes regulating different processes in mitosis and interphase to ensure proliferation and survival of cancer cells with CA. Mitotic TACC3 interacts with the Kinesin Family Member C1 (KIFC1) to cluster extra centrosomes for mitotic progression, and inhibition of this interaction leads to mitotic cell death via multipolar spindle formation. Interphase TACC3 interacts with the nucleosome remodeling and deacetylase (NuRD) complex (HDAC2 and MBD2) in nucleus to inhibit the expression of key tumor suppressors (e.g., p21, p16 and APAF1) driving G1/S progression, and its inhibition blocks these interactions and causes p53-independent G1 arrest and apoptosis. Notably, inducing CA by p53 loss/mutation increases the expression of TACC3 and KIFC1 via FOXM1 and renders cancer cells highly sensitive to TACC3 inhibition. Targeting TACC3 by guide RNAs or small molecule inhibitors strongly inhibits growth of organoids and breast cancer cell line- and patient-derived xenografts with CA by induction of multipolar spindles, mitotic and G1 arrest. Altogether, our results show that TACC3 is a multifunctional driver of highly aggressive breast tumors with CA and that targeting TACC3 is a promising approach to tackle this disease.