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The failure to treat and control the growth of metastases is the main cause of death in breast cancer (BC) patients. Compared to the traditional method of analyzing circulating tumor DNA (ctDNA), capturing intact circulating tumor cells (CTCs) allows us to more accurately characterize mutations and identify suitable targeted therapies. We used CellCollector to collect peripheral CTCs. Thirty metastatic breast cancer (MBC) patients were enrolled, and 17 were analyzed with next-generation sequencing (NGS) methods. Clinical characteristics were analyzed along with the CTCs enumeration and detection rates. Whole-genome amplification (WGA) was used to amplify the CTC genomic DNA of 127 genes. Patients younger than 45 years old, with brain metastasis, with three or more metastatic sites, or with HER2-positive had the highest number of CTCs collected. The CTCs detection rate was also correlated to the number of metastasis sites. Different metastasis sites such as the brain, viscus, bone, and soft tissue contained specific high-frequency gene mutations. AKT3, MYC, and NT5C2 mutations were only found in brain metastases. APC, BCL2L11, ESRP1, FLT3 mutations were only in the visceral metastases. KEAP1, KIT, MET were the specific mutation genes in patients with bone and soft tissue metastases. These findings provide evidence that we can detect gene mutation information for obtaining the biological characteristics by CTCs using CellCollector. Different metastasis sites contain specific high-frequency mutation genes, which provide guidance to the accurate gene therapy.

Ferroptosis is a newly form of regulated cell death, which has attracted great attention for tumor therapy. Herein, we prepared nanoscale coordination polymer Fe-NQA particles to deliver iron and NQA (vitamin K3 derivative) into tumor cells, which synergistically promoted ferroptotic therapy for inhibiting tumor growth, preventing metastasis, and overcoming radioresistance.

Abraxane® (Abx), an FDA approved albumin-bound paclitaxel nano-formulation, is one of the most common chemical drugs for the treatment of metastatic triple-negative breast cancer (mTNBC). However, acquired resistance and metastasis are critical factors that limit the treatment of mTNBC by Abx. In particular, both the tumor hypoxic microenvironment and the increase in hydrogen peroxide (H2O2) levels via paclitaxel stimulation primarily mediate the resistance to chemotherapy, where multiple drug resistance proteins such as P-gp and tumor invasion-related cytokines such as VEGF are continuously activated to pump out chemical drugs and aggravate tumor metastasis, respectively. Therefore, it is of great importance to combine tumor oxygenation with commercial chemical drugs for overcoming the acquired resistance and metastasis. In this study, a facile method was developed to deposit manganese dioxide (MnO2) onto the surface of Abraxane® (Abx) to form MnO2-modified Abx (M-Abx). The modification process did not change the critical characteristics of the parent Abx, which might have great potential for application in clinics for the treatment of mTNBC. Tumor oxygenation mediated by M-Abx specifically occurs within the H2O2-overexpressed tumor microenvironment, and significantly downregulates the content of tumor progression-related proteins, such as HIF-1α, P-gp, and VEGF. Ultimately, M-Abx treatment results in about a 2-fold increase in inhibition efficiency of tumor growth in both primary and metastatic tumors compared with traditional Abx therapy. Therefore, oxygen-rich chemotherapy was realized to efficiently sensitize paclitaxel, relieve acquired resistance and inhibit tumor metastasis.

Hypoxia, and hypoxia inducible factor-1 (HIF-1), can induce tumor resistance to radiation therapy. To overcome hypoxia-induced radiation resistance, recent studies have described nanosystems to improve tumor oxygenation for immobilizing DNA damage and simultaneously initiate oxygen-dependent HIF-1α degradation. However, HIF-1α degradation is incomplete during tumor oxygenation treatment alone. Therefore, tumor oxygenation combined with residual HIF-1 functional inhibition is crucial to optimizing therapeutic outcomes of radiotherapy. Here, a reactive oxygen species (ROS) responsive nanoplatform is reported to successfully add up tumor oxygenation and HIF-1 functional inhibition. This ROS responsive nanoplatform, based on manganese dioxide (MnO2) nanoparticles, delivers the HIF-1 inhibitor acriflavine and other hydrophilic cationic drugs to tumor tissues. After reacting with overexpressed hydrogen peroxide (H2O2) within tumor tissues, Mn2+ and oxygen molecules are released for magnetic resonance imaging and tumor oxygenation, respectively. Cooperating with the HIF-1 functional inhibition, the expression of tumor invasion-related signaling molecules (VEGF, MMP-9) is obviously decreased to reduce the risk of metastasis. Furthermore, the nanoplatform could relieve T-cell exhaustion via downregulation of PD-L1, whose effects are similar to the checkpoint inhibitor PD-L1 antibody, and subsequently activates tumor-specific immune responses against abscopal tumors. These therapeutic benefits including increased X-ray-induced damage, downregulated resistance, and T-cell exhaustion related proteins expression achieved synergistically the optimal inhibition of tumor growth. Overall, this designed ROS responsive nanoplatform is of great potential in the sensitization of radiation for combating primary and metastatic tumors.

Introduction: Metastasis is thought to result from tumour cell entry into the circulation and migration to distant organs, where the mutation landscape of metastatic breast cancer (MBC) may shift and vary. The genotypic features of circulating tumour cells (CTCs) typically differ from those of primary breast cancer (BC) cells. Gene mutation analysis of CTCs in MBC may benefit patients by identifying those amenable to specific therapies. Currently, CTCs are primarily isolated in vitro from small volumes of blood. The aim of this study was to isolate CTCs in vivo using CellCollector and screen for specific gene mutations in cells from different metastasis organ sites and molecular subtypes in MBC patients. Methods: In this study, we used a novel technology, CellCollector, to collect peripheral CTCs. Thirty MBC patients were enrolled, and 17 were analysed with next-generation sequencing (NGS) methods. Clinical characteristics were analysed along with CTC enumeration and detection rates. Whole-genome amplification (WGA) was used to amplify the CTC genomic DNA of 127 genes. Results: We isolated CTCs in vivo from 20 of 30 MBC patients (66.7%), with a median and mean (range) of 2 (0-15) CTCs. In non-cancer patients, no CTCs were detected. We analysed CTC enumeration and the detection rate in different clinical characteristic subgroups. We found that in their corresponding subgroups, patients younger than 45 years old, with brain metastasis, with three or more metastasis organ sites, or with HER2-positive subtypes had the highest CTC medians and means.As far as clinical characteristics were concerned, the number of CTCs seemed correlated with more advanced clinical characteristics. In the one metastasis organ, two metastasis organs and three or more metastasis organs subgroups, the CTC detection rates were 38.5% (5/13), 77.8% (7/9), and 100.0% (8/8), respectively. The CTC detection rate correlated with the number of metastasis organs; patients with more metastasis organ sites had higher CTC detection rates. We also found that different metastasis organs and molecular subtypes contain high-frequency mutation genes, and also contain unique gene mutations. Conclusions: In MBC, CellCollector can be used to collect intact CTCs, from which we can obtain gene mutation information. Different metastasis organs and molecular subtypes may have corresponding unique mutations, which may provide a basis for future gene therapy. CTCs enumeration data and correlations to clinicopathologic characteristicsPatients' characteristics CTCs enumeration RangemedianmeanTotal0-1522Age (years) Citation Format: Geng C, Li S, Yang S, Shi J, Ding Y, Gao W, Cheng M, Sun Y, Xie Y, Sang M. In vivo isolation of circulating tumour cells using CellCollector and detection of gene mutations in different metastasis organ sites in breast cancer [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P3-01-18.