Breast Cancer Xenograft Mouse Model Research Articles

Drug screening on digital microfluidics for cancer precision medicine

Drug screening based on in-vitro primary tumor cell culture has demonstrated potential in personalized cancer diagnosis. However, the limited number of tumor cells, especially from patients with early stage cancer, has hindered the widespread application of this technique. Hence, we developed a digital microfluidic system for drug screening using primary tumor cells and established a working protocol for precision medicine. Smart control logic was developed to increase the throughput of the system and decrease its footprint to parallelly screen three drugs on a 4 × 4 cm2 chip in a device measuring 23 × 16 × 3.5 cm3. We validated this method in an MDA-MB-231 breast cancer xenograft mouse model and liver cancer specimens from patients, demonstrating tumor suppression in mice/patients treated with drugs that were screened to be effective on individual primary tumor cells. Mice treated with drugs screened on-chip as ineffective exhibited similar results to those in the control groups. The effective drug identified through on-chip screening demonstrated consistency with the absence of mutations in their related genes determined via exome sequencing of individual tumors, further validating this protocol. Therefore, this technique and system may promote advances in precision medicine for cancer treatment and, eventually, for any disease.

Rapamycin prevents cyclophosphamide-induced ovarian follicular loss and potentially inhibits tumour proliferation in a breast cancer xenograft mouse model.

Rapamycin prevents cyclophosphamide-induced ovarian follicular loss and potentially inhibits tumour proliferation in a breast cancer xenograft mouse model.

Abstract 4149: Real-time intravital trafficking of tumor infiltrating lymphocytes in breast cancer models

Abstract Cancer immunotherapy utilizing the immunoreactive efficiency of immune cells via adoptive cell therapy (ACT) has been a novel method of cancer treatment. In tumor microenvironment, immune cell reactivity of tumor-infiltrating lymphocytes (TILs) can facilitate cancer cell attack, which finally derives efficient suppression of tumor growth. However, the conventional evaluation for cancer immunotherapy has been generally performed by ex vivo methods, providing restricted information on lymphocyte motility and immune cell dynamics within the tumor microenvironment. In vivo data with the dynamic behavior of TILs in breast cancer can provide new insight to investigate novel targets of immune checkpoint inhibitors. In this research, longitudinal intravital imaging of motility trafficking of TILs in breast cancer animal models has been performed by utilizing intravital microscopy to unveil the immunoreactive dynamics of T cell activity in early stage. In vivo analysis of TIL motility within different breast cell lines, including 4T1 (mouse breast cancer cell), MDA-MB-231 (human triple-negative breast cancer cell), and MCF7 (human breast cancer cell), respectively, has been investigated for immune responses to cancer. We’ve monitored spatiotemporal dynamics of TIL over 11 days in breast cancer xenograft mouse model generated by tumor implantation into the dorsal skinfold imaging chamber and adoptive T cell transfer by blood circulation. In the results, the adoptive immunoreactivity of TIL has been differentiated by the timepoint of adoptive T cell transfer, distance proximity of T cells in tumors, and breast cancer types. Furthermore, anti-cancer effect coordinated with dendritic cells behaviors showed induced immune responses with accumulated dendritic cells with increased TIL count over time in breast cancer model treated with various cancer therapy such as immunotherapy, radiation therapy and anti-cancer drug (anti-PD-L1), which implied a relationship between programmed death-ligand 1 expression levels and TIL/dendritic cell motility. The result demonstrates that intravital TIL motility trafficking analysis in cancer model can be an invaluable assessment method to explain the detailed cellular/molecular mechanism of immune response to cancer involved with T lymphocytes, dendritic cells, and immune checkpoint inhibitors to accelerate the efficiency of cancer immunotherapy. Citation Format: Soyeon Ahn, Kubra Akyildiz, Hyunseok Kim, Pilhan Kim. Real-time intravital trafficking of tumor infiltrating lymphocytes in breast cancer models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4149.

ZC3H12A inhibits tumor growth and metastasis of breast cancer under hypoxic condition via the inactivation of IL-17 signaling pathway

ABSTRACT Hypoxia is a major contributor to tumor microenvironment (TME) and metastasis in most solid tumors. We seek to screen hypoxia-related genes affecting metastasis in breast cancer and to reveal relative potential regulatory pathway. Based on gene expression profiling of GSE17188 dataset, differential expressed genes (DEGs) were identified between highly metastatic breast cancer cells under hypoxia and samples under normoxia. The protein–protein interaction (PPI) network was utilized to determine hub genes. The gene expression profiling interactive analysis database (GEPIA2) and quantitative reverse-transcription polymerase chain reaction (qRT-PCR) were employed to quantify hub genes. Moreover, overexpression of zinc finger CCCH-type containing 12A (ZC3H12A) was performed both in breast cancer cells and xenograft mouse model to determine the role of ZC3H12A. We identified 134 DEGs between hypoxic and normoxic samples. Based on PPI analysis, 5 hub genes interleukin (IL)-6, GALN (GAL), CD22 molecule (CD22), ZC3H12A and TNF receptor associated factor 1 (TRAF1) were determined; the expression levels of TRAF1, IL-6, ZC3H12A and GAL were remarkably downregulated while CD22 was upregulated in breast cancer cells. Besides, patients with higher expression of ZC3H12A had favorable prognosis. Overexpression of ZC3H12A could inhibit metastasis and tumor growth of breast cancer; overexpression of ZC3H12A downregulated the expression of IL-17 signaling pathway-related proteins such as IL-17 receptor A (IL-17RA), IL-17A and nuclear factor κB activator 1 (Act1). This study reveals ZC3H12A and IL-17 signaling pathway as potential therapeutic targets for hypoxic breast cancer.

The HDAC inhibitor HFY-4A improves TUSC2 transcription to induce immunogenic cell death in breast cancer

We managed to explore the function of HFY-4A, a novel histone deacetylases (HDACs) inhibitor, on breast cancer as well as its potential mechanisms. MCF7 and T47D cells were treated with 0.8, 1.6 or 3.2 μM HFY-4A for 0–72 h, following of which CCK-8, colony formation, EdU staining, flow cytometry, Transwell, and wound healing assays were carried out. Western blot, immunohistochemistry, and ELISA were conducted for assaying the expression of immunogenic cell death (ICD)-related proteins. The interaction between HFY-4A, HDAC1, and tumor suppressor candidate 2 (TUSC2) was evaluated by chromatin immunoprecipitation assay. Further, the function of HFY-4A in breast cancer progression in vivo was evaluated using xenograft mouse models. HFY-4A inhibited the proliferation, migration, and invasion, and induced apoptosis of breast cancer cells in a dose-dependent manner. HFY-4A dose-dependently caused the ICD of breast cancer cells, as evidenced by the significant high levels of high-mobility group box 1 (HMGB1), calreticulin (CRT), heat shock protein 70 (HSP70), and HSP90. Interestingly, HFY-4A could facilitate TUSC2 transcription by promoting acetylation of histones on the TUSC2 promoter. The results of rescue assays revealed that HFY-4A repressed proliferation and mobility, but enhanced apoptosis and ICD through facilitating TUSC2 transcription in breast cancer. In breast cancer xenograft mouse models, HFY-4A was verified to inhibit tumor growth via upregulating TUSC2. HFY-4A could inhibit breast cancer cell proliferation and mobility, and enhanced apoptosis and ICD through facilitating TUSC2 transcription.

Potential risk of tamoxifen: gut microbiota and inflammation in mice with breast cancer.

Tamoxifen is an effective anti-tumor medicine, but evidence has been provided on tamoxifen-related inflammation as well as its impact on gut microbiota. In this study, we aimed to investigate tamoxifen-induced gut microbiota and inflammation alteration. We established a BC xenograft mouse model using the MCF-7 cell line. 16S rRNA gene sequencing was used to investigate gut microbiota. qRT-PCR, western blotting, and cytometric bead array were used to investigate inflammation-related biomarkers. Various bioinformatic approaches were used to analyze the data. Significant differences in gut microbial composition, characteristic taxa, and microbiome phenotype prediction were observed between control, model, and tamoxifen-treated mice. Furthermore, protein expression of IL-6 and TLR5 was up-regulated in tamoxifen-treated mice, while the mRNA of Tlr5 and Il-6, as well as protein expression of IL-6 and TLR5 in the model group, were down-regulated in the colon. The concentration of IFN-γ, IL-6, and IL12P70 in serum was up-regulated in tamoxifen-treated mice. Moreover, correlation-based clustering analysis demonstrated that inflammation-negatively correlated taxa, including Lachnospiraceae-UCG-006 and Anaerotruncus, were enriched in the model group, while inflammation-positively correlated taxa, including Prevotellaceae_UCG_001 and Akkermansia, were enriched in the tamoxifen-treated group. Finally, colon histologic damage was observed in tamoxifen-treated mice. Tamoxifen treatment significantly altered gut microbiota and increased inflammation in the breast cancer xenograft mice model. This may be related to tamoxifen-induced intestinal epithelial barrier damage and TLR5 up-regulation.

Combination of novel oncolytic herpesvirus with paclitaxel as an efficient strategy for breast cancer therapy.

New strategies are needed to improve the treatment of patients with breast cancer (BC). Oncolytic virotherapy is a promising new tool for cancer treatment but still has a limited overall durable antitumor response. A novel replicable recombinant oncolytic herpes simplex virus type 1 called VG161 has been developed and has demonstrated antitumor effects in several cancers. Here, we explored the efficacy and the antitumor immune response of VG161 cotreatment with paclitaxel (PTX) which as a novel oncolytic viral immunotherapy for BC. The antitumor effect of VG161 and PTX was confirmed in a BC xenograft mouse model. The immunostimulatory pathways were tested by RNA-seq and the remodeling of tumor microenvironment was detected by Flow cytometry analysis or Immunohistochemistry. Pulmonary lesions were analyzed by the EMT6-Luc BC model. In this report, we demonstrate that VG161 can significantly represses BC growth and elicit a robust antitumor immune response in a mouse model. The effect is amplified when combined with PTX treatment. The antitumor effect is associated with the infiltration of lymphoid cells, including CD4+ T cells, CD8+ T cells, and NK cells (expressing TNF and IFN-γ), and myeloid cells, including macrophages, myeloid-derived suppressor cells, and dendritic cell cells. Additionally, VG161 cotreatment with PTX showed a significant reduction in BC lung metastasis, which may result from the enhanced CD4+ and CD8+ T cell-mediated responses. The combination of PTX and VG161 is effective for repressing BC growth by inducing proinflammatory changes in the tumor microenvironment and reducing BC pulmonary metastasis. These data will provide a new strategy and valuable insight for oncolytic virus therapy applications in primary solid or metastatic BC tumors.

Abstract 6126: Antitumor effect of Iso-mukaadial acetate in MCF-7 human cancer xenograft models

Abstract Antitumor drugs used today have shown significant efficacy and are derived from natural products such as plants. Iso-mukaadial acetate has previously been shown to possess anticancer properties by inducing apoptosis. The purpose of this study was to investigate the therapeutic effect of Iso-mukaadial acetate in the breast cancer xenograft mice model. Female athymic mice were used and were inoculated with breast cancer cells subcutaneously after culturing. Group one served as a negative control group (no treatments) and group two positive control group (cisplatin) which was administered intravenously. Iso-mukaadial acetate treatment was administered orally to group three(100 mg/kg) and group four (500 mg/kg). Blood was collected (70 µL) from the tail vein on day zero, day seven and day 14. Tumor regression was measured, and a pharmacokinetic study was conducted. Estimation of Serum Parameters for Hepatic and Renal Indices were examined using an Automated Chemistry Analyzer. Histopathological analysis was conducted to evaluate morphological changes before and after treatment under a light microscope. Tumor shrinkage was observed after treatment. In conclusion, Iso-mukaadial acetate inhibited tumor progression in breast cancer athymic mice without major side effects. Citation Format: Portia Raphela-Choma, Lesetja R. Motadi, Mpho S. Choene. Antitumor effect of Iso-mukaadial acetate in MCF-7 human cancer xenograft models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6126.

Preclinical Activity of 4-Demethyl-4-cholesteryloxycarbonylpenclomedine in Melanoma.

Temozolomide plays a role in treating melanoma refractory to immunomodulatory and mitogen-activated protein kinase-targeted approaches, but its efficacy is limited. 4-Demethyl-4-cholesteryloxycarbonylpenclomedine (DM-CHOC-PEN) is a polychlorinated pyridine cholesteryl carbonate. Its mechanism of action is considered to be via alkylation/adduct formation with N7-guanine. It demonstrated activity in intracranial implanted human glioma and breast cancer xenograft mouse models. The activity of DM-CHOC-PEN in melanoma models was assessed. B-16 melanoma cells were exposed to DM-CHOC-PEN at different concentrations to assess proliferation and survival. B-16 cells were implanted subcutaneously into the flank of adult female C57BL mice which were then were treated with 200 mg/kg DM-CHOC-PEN intraperitoneally daily for 5 days in the setting of palpable subcutaneous tumor. Survival was compared to mice treated with temozolomide or saline. Five mice were treated per group. In vitro, the respective half-maximal inhibitory concentrations of DM-CHOC-PEN and temozolomide were 0.5 and ≥3.0 μg/ml. Floating, heavily melanotic cells formed and these cells were separated, analyzed, and contained 10-90 ng DM-CHOC-PEN per 105 cells. The improvement in survival of mice treated with DM-CHOC-PEN or temozolomide relative to saline controls was 142% and 78%, respectively. Longer survival was seen with DM-CHOC-PEN in a C57BL murine model relative to temozolomide and saline-treated controls, supporting the development of clinical trials assessing the efficacy of DM-CHOC-PEN as treatment for metastatic melanoma.

Antitumor activity of iridium/ruthenium complexes containing Nitro -substituted quinoline ligands in vivo and in vitro

Four iridium/ruthenium half-sandwich metal complexes containing 8-hydroxyquinoline ligands were synthesized as anticancer agents. In vitro toxicity results showed that all complexes had outstanding antitumor activity against a variety of tumor cells, especially the MDA-MB-231 cell. The complex could initiate downstream apoptotic signals through the lysosomal damage pathway and the reactive oxygen species (ROS) produced at the same time can further aggravate the damage to lysosomes. In addition, the complex significantly inhibited the growth of tumors in the breast cancer mouse xenograft model in vivo.

Pirfenidone suppressed triple-negative breast cancer metastasis by inhibiting the activity of the TGF-β/SMAD pathway.

Among breast cancer patients, metastases are the leading cause of death. Despite decades of effort, little progress has been made to improve the treatment of breast cancer metastases, especially triple-negative breast cancer (TNBC). The extracellular matrix plays an important role in tumour growth and metastasis by causing its deposition, remodelling, and signalling. As we know, the process of fibrosis results in excessive amounts of extracellular matrix being deposited within the cells. So, it will be interesting to study if the use of anti-fibrotic drugs in combination with conventional chemotherapy drugs can produce synergistic antitumor effects. In this study, we assessed the efficacy of Pirfenidone (PFD), an FDA-approved medication for the treatment of idiopathic pulmonary fibrosis, on TNBC cells as well as its anti-tumour effects in xenograft tumour model. PFD inhibited in a dose-dependent manner breast cancer cell proliferation, migration, and invasion, while promoted their apoptosis in vitro. PFD also suppressed TGF-β-induced activation of Smad signalling pathway and expression level of EMT-inducing transcription factors (e.g. SNAI2, TWIST1, ZEB1) as well as the mesenchymal genes such as VIMENTIN and N-Cadherin. On the contrary, the expression level of epithelial marker gene E-Cadherin was up-regulated in the presence of PFD. In vivo, PFD alone exerted a milder but significant anti-tumour effect than the chemotherapy drug nanoparticle albumin-bound paclitaxel (nab-PTX) did in the breast cancer xenograft mouse model. Interestingly, PFD synergistically boosted the cancer-killing effect of nab-PTX. Furthermore, Our data suggest that PFD suppressed breast cancer metastasis by inhibiting the activity of the TGFβ/SMAD pathway.

Chimeric antigen receptor T cells engineered to recognize the P329G-mutated Fc part of effector-silenced tumor antigen-targeting human IgG1 antibodies enable modular targeting of solid tumors

BackgroundChimeric antigen receptor (CAR) T cell therapy has proven its clinical utility in hematological malignancies. Optimization is still required for its application in solid tumors. Here, the lack of cancer-specific...

PLAG co-treatment increases the anticancer effect of Adriamycin and cyclophosphamide in a triple-negative breast cancer xenograft mouse model

Chemotherapy induces tumor cell death and inhibits tumor progression, but the accompanying immune responses in the surrounding dying tissue cause significant inflammation. These responses, such as excessive neutrophil infiltration into tumor tissue, are the main causes of resistance to anticancer treatment. The development of drugs that reduce neutrophil infiltration into tumors is necessary to increase the anticancer effect of chemotherapy.Here, we show that the antitumor effect of the chemotherapy AC regimen (Adriamycin and cyclophosphamide) was increased by 1-palmitoyl-2-linoleoyl-3-acetyl-rac-glycerol (PLAG) cotreatment in the MDA-MB-231 triple-negative breast cancer xenograft mouse model. Tumor growth was inhibited up to 56% in mice treated with AC and inhibited up to 94% in mice cotreated with AC and PLAG. Side effects of chemotherapy, such as a reduction in body weight, were alleviated in mice cotreated with AC and PLAG. Excessive neutrophil infiltration caused by the AC regimen was successfully cleared in mice cotreated with AC and PLAG.We conclude that PLAG inhibits excessive neutrophil infiltration that aids tumor growth. Reduced neutrophils and increased lymphocytes in PLAG-treated mice can maximize the antitumor effect of the AC regimen and inhibit tumor growth.

Human Tumor Targeted Cytotoxic Mast Cells for Cancer Immunotherapy

The diversity of autologous cells being used and investigated for cancer therapy continues to increase. Mast cells (MCs) are tissue cells that contain a unique set of anti-cancer mediators and are found in and around tumors. We sought to exploit the anti-tumor mediators in MC granules to selectively target them to tumor cells using tumor specific immunoglobin E (IgE) and controllably trigger release of anti-tumor mediators upon tumor cell engagement. We used a human HER2/neu-specific IgE to arm human MCs through the high affinity IgE receptor (FcεRI). The ability of MCs to bind to and induce apoptosis of HER2/neu-positive cancer cells in vitro and in vivo was assessed. The interactions between MCs and cancer cells were investigated in real time using confocal microscopy. The mechanism of action using cytotoxic MCs was examined using gene array profiling. Genetically manipulating autologous MC to assess the effects of MC-specific mediators have on apoptosis of tumor cells was developed using siRNA. We found that HER2/neu tumor-specific IgE-sensitized MCs bound, penetrated, and killed HER2/neu-positive tumor masses in vitro. Tunneling nanotubes formed between MCs and tumor cells are described that parallel tumor cell apoptosis. In solid tumor, human breast cancer (BC) xenograft mouse models, infusion of HER2/neu IgE-sensitized human MCs co-localized to BC cells, decreased tumor burden, and prolonged overall survival without indications of toxicity. Gene microarray of tumor cells suggests a dependence on TNF and TGFβ signaling pathways leading to apoptosis. Knocking down MC-released tryptase did not affect apoptosis of cancer cells. These studies suggest MCs can be polarized from Type I hypersensitivity-mediating cells to cytotoxic cells that selectively target tumor cells and specifically triggered to release anti-tumor mediators. A strategy to investigate which MC mediators are responsible for the observed tumor killing is described so that rational decisions can be made in the future when selecting which mediators to target for deletion or those that could further polarize them to cytotoxic MC by adding other known anti-tumor agents. Using autologous human MC may provide further options for cancer therapeutics that offers a unique anti-cancer mechanism of action using tumor targeted IgE’s.

Dual-aptamer-engineered M1 macrophage with enhanced specific targeting and checkpoint blocking for solid-tumor immunotherapy

Chimeric antigen receptor T (CAR-T) cell therapy has faced a series of challenges and has shown very little efficacy in solid tumors to date. Although genetically engineered macrophages have achieved definite therapeutic effect in solid tumors, heterogeneous expression of engineered proteins and the potential for toxicity limit further applications. Herein, we propose a nongenetic and simple macrophage cell engineering strategy through glycan metabolic labeling and click reaction for the treatment of solid tumors. The aptamer-engineered M1 macrophage (ApEn-M1) showed enhanced active targeting ability for tumor cells invitro and invivo, resulting in significant cytotoxicity effects. Moreover, ApEn-M1 exhibited superior antitumor efficacy in a breast cancer xenograft mouse model and a lung metastasis mouse model of breast cancer. Interestingly, the ApEn-M1 could reprogram the immunity microenvironment by increasing Tcell infiltration and enhancing Tcell activity in the tumor region. Additionally, the administration of ApEn-M1 showed no obvious systemic side effects. With glycan metabolic labeling, the macrophages could be efficiently labeled with aptamers on the cell surface via click reaction without genetic alteration or cell damage. Hence, this study serves as a proof of concept for cell-surface anchor engineering and expands the range of nongenetic macrophage cell engineering strategies.

Photoacoustic Tomography Detects Response and Resistance to Bevacizumab in Breast Cancer Mouse Models.

Photoacoustic assessment of tumor oxygenation is a noninvasive early indicator of response to bevacizumab therapy, clearly distinguishing between control, responding, and resistant tumors within just a few weeks of treatment.

Abstract P5-16-16: A novel investigational nanoparticle therapeutic (DAN-222) for breast cancer and other solid tumors

Abstract Background: Here we introduce a novel nanoparticle therapeutic, DAN-222, which offers a variety of potential clinical benefits in delivery of chemotherapy versus native drug, including a broader therapeutic index. DAN-222 is a polymeric nanoparticle covalently conjugated with camptothecin (CPT), a topoisomerase I inhibitor, that demonstrates enhanced efficacy over native CPT at the same dose and has significantly reduced bone marrow exposure (limiting potential safety concerns), thus widening the therapeutic index with respect to both efficacy and safety. The nanoparticle therapeutic provides sustained drug exposure through conjugation of the chemotherapy payload to the novel biocompatible polymer scaffold that can be modulated by choice of linker, allowing for optimization of the release rate for a given payload and target. To illustrate the flexibility and capabilities of modulating payload release kinetics, we present a series of CPT-containing nanoparticles with different linkers encompassing a wide range of in vitro release rates. We also demonstrate the vastly enhanced efficacy of DAN-222 in multiple breast cancer xenograft mouse models. Methods: In vitro release: Release of CPT from nanoparticles formed with various linkers was evaluated by HPLC in pH 5.5 PBS (endosomal pH), pH 7.4 PBS (physiologic pH), mouse plasma, and human plasma. Efficacy: Breast cancer xenograft tumor models with human BT-474 and MDA-MB-231 were propagated in CB.17 SCID and NCr nu/nu mice respectively, with 10 animals per group. Results: All linkers demonstrate linear kinetics and release slowly at reduced pH, consistent with the mechanism of release. At physiologic pH, a diversity of in vitro release rates is achieved by modulating the sensitivity of the linker. Importantly, all linkers have similar release in mouse and human plasma, indicating strong cross-species translation. Results in the mouse xenograft models showed that DAN-222 had a superior and sustained efficacy compared to CPT and irinotecan (an approved topoisomerase-1 inhibitor) as demonstrated by tumor growth inhibition. Conclusions: We demonstrate the ability to modulate the payload release kinetics from the nanoparticle therapeutic by the choice of linker, with species-independent release rates. With an optimal linker selected, DAN-222 shows enhanced and sustained efficacy over native CPT. Since DAN-222 has previously been shown to have reduced bone marrow exposure of chemotherapy, it may provide a significant therapeutic advancement with a wider therapeutic index based on both enhanced efficacy and improved safety. DAN-222 is advancing to a clinical trial in HER2-negative metastatic breast cancer. In vitro release half-lives (t1/2) (hours)Linker 1Linker 2Linker 3Linker 4Linker 5PBS, pH 5.5>50>50>50>50>50PBS, pH 7.47.120.5>50>50>50Mouse Plasma1.74.817.237.7>50Human Plasma1.64.817.137.8>50 TreatmentBT-474MDA-MB-231Day 25 TGI(%)Day 36 TGI*(%)Day 25 TGI (%)Day 32 TGI* (%)Camptothecin (3 mg/kg)64404756Irinotecan (100 mg/kg)71594751DAN-222 (3 mg/kg CPT)93967475*Day the vehicle control group reached the study endpoint. Citation Format: Emily A Wyatt, Ashley P Wright, Carl M Blumenfeld, Robert J Lamm, Timothy Hagerty. A novel investigational nanoparticle therapeutic (DAN-222) for breast cancer and other solid tumors [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P5-16-16.

Abstract P2-13-09: An investigational next generation ADC (DAN-311) is highly effective in HER2-low breast cancer models

Abstract Background: Nanoparticles (NPs) have been developed to enhance the pharmacokinetic (PK) and biodistribution characteristics of molecules. Many early NPs, including liposomes, dendrimers, and polymeric micelles, were limited in their ability to access target tissue and lacked uniform release rates. A promising nanoparticle therapeutic has been developed utilizing biocompatible polymer chemistry. By covalently attaching chemotherapy to the polymer through linker strategies that allow for modulation of release and PK, polymeric nanoparticles provide advantages over encapsulation nanoparticle strategies. Additionally, a targeting moiety can be added to make a next-generation ADC with the drug-to-antibody-ratio (DAR) well controlled and significantly greater (e.g. DAR=60) compared to traditional ADCs. DAN-311 is a Next-Generation Antibody Drug Conjugate (NG-ADC) with a HER2-targeting agent (trastuzumab) conjugated to a therapeutic nanoparticle with camptothecin (CPT), a topoisomerase I inhibitor. The advantages that DAN-311 offers to the 50% of patients with breast cancer have tumors with low HER2 expression is two-fold: 1) the larger DAR can lead to an enhanced bystander effect where the chemotherapy is effective against the targeted cells and also the neighboring cells, and 2) the nanoparticle is also active against HER2 non-expressing cells through the enhanced permeability and retention (EPR) effect. Here, we present the control and consistency of key DAN-311 NP characteristics that allow for efficient and large scale-manufacture, including particle size, chemotherapy load (DAR), and payload release rate. We also report the enhanced efficacy of DAN-311 in a HER2-low breast cancer (JIMT-1) xenograft mouse model. Methods: Release of CPT from nanoparticles across range of combined properties was evaluated by HPLC in pH 5.5 PBS (endosomal pH), pH 7.4 PBS (physiologic pH), mouse plasma, and human plasma. HER2-low breast cancer tumor cells (JIMT-1) were implanted into female CB.17 SCID mice. Mice were randomized to vehicle or treatment arms until tumors reached 2000 mm3 or day 46. The groups evaluated included trastuzumab, the non-targeted core nanoparticle of DAN-311, and DAN-311. Results: NPs across a wide range in polymer molecular weight and drug loading demonstrate striking consistency in particle size (ca. 30-45 nm), DAR (ca. 60-70 CPT molecules), and demonstrate linear release kinetics in various physiologically-relevant media. DAN-311 exhibited significantly greater tumor growth inhibition compared to vehicle, core particle, and trastuzumab. The non-targeted core particle also demonstrated significant tumor growth inhibition compared to vehicle and trastuzumab, which was sustained for at least 3 weeks after the final dose. Core particle = same DAN-311 nanoparticle without HER2 targeting. Conclusions: DAN-311, a Next-Generation ADC with a HER2-targeting agent (trastuzumab) on a polymeric nanoparticle conjugated with CPT demonstrated a significant ability to inhibit tumor growth in a HER2-low xenograft mouse model. Importantly, the nanoparticle targeted delivery of CPT for cells with low expression of receptors is highly efficient in cell killing and the large DAR provides greater bystander cell killing capability; and non-HER2-expressing lesions or regions of tumors will additionally be treated by the nanoparticle through EPR. DAN-311 is advancing to clinical study to treat HER2-low metastatic breast cancer patients. TreatmentDose (mg/kg)Day 22 TGI (%)Day 46 TGI (%)Trastuzumab10 mg/kg Tras629Core particle1 mg/kg CPT3955DAN-31110 mg/kg Tras/1 mg/kg CPT8777 Citation Format: Ashley P Wright, Emily A Wyatt, Robert J Lamm, Carl M Blumenfeld, Jodi D Bradley, Gayatri Shrikhande, Timothy Hagerty. An investigational next generation ADC (DAN-311) is highly effective in HER2-low breast cancer models [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P2-13-09.

Self-assembled tetrahedral framework nucleic acid mediates tumor-associated macrophage reprogramming and restores antitumor immunity

There is increasing interest in depleting or repolarizing tumor-associated macrophages (TAMs) to generate a proinflammatory effect. However, TAMs usually display an immunosuppressive M2-like phenotype in the tumor microenvironment. Apparently, developing a macrophage-targeting delivery system with immunomodulatory agents is urgent. In this study, an efficient siRNA and CpG ODNs delivery system (CpG-siRNA-tFNA) was prepared with nucleic acid stepwise self-assembled. The tFNA composed of CpG ODNs and siRNA showed a higher stability and an enhanced cellular uptake efficiency. Moreover, the CpG-siRNA-tFNA effectively reprogrammed TAMs toward M1 phenotype polarization with increased proinflammatory cytokine secretion and NF-κB signal pathway activation, which triggers dramatic antitumor immune responses. Additionally, the CpG-siRNA-tFNA exhibited superior antitumor efficacy in a breast cancer xenograft mouse model without obvious systemic side effects. Taken together, CpG-siRNA-tFNA displayed greatly antitumor effect by facilitating TAM polarization toward M1 phenotypes in favor of immunotherapy. Hence, we have developed an efficient therapeutic strategy with immunomodulatory agents for clinical applications.

Defective Signalling of the BRCA1 Neighbouring gene, NBR2, leads to ER-? Negative tumours in breast cancer xenograft mouse models

Majority of breast and ovarian cancer treatment modalities are based on endocrine therapies like antiestrogens which are dependent on the hormone receptor status of tumours. Although BRCA1 is a major regulator of Estrogen Receptor (ER-α), BRCA1 mutations are largely limited to hereditary cases. Here, we show the role of NBR2 (neighbour of BRCA1) in regulating ER-α. We demonstrate a positive correlation between NBR2 & ESR1 in TCGA datasets. Further, the study revealed that shRNA-mediated knockdown of NBR2 led to the downregulation of ER-α in breast cancer cells, MCF7. Finally, the downregulation of ER-α in NBR2 knockdown xenograft tumours (in female NSG mice), which showed higher invasive properties than wild type tumours was demonstrated. Thus, we concluded that in ER-α negative tumours with NBR2 deficiency, biguanides such as metformin and phenformin, which are reported to have a better efficacy under NBR2 deficient conditions, could serve as more suitable alternatives to antiestrogens.