HER2‐targeted therapies effectively control systemic disease, but their efficacy against brain metastases is hindered by their low penetration of the blood‐brain and blood‐tumor barriers (BBB and BTB). We investigate brain uptake and antitumor efficacy of transferrin receptor (TfR)‐targeted, therapeutic nanoparticles designed to transcytose the BBB/BTB in three murine models. Two known models involving intracranial (IC) or intracardiac (ICD) injection of human breast cancer cells were employed, as was a third model developed here involving intravenous (IV) injection of the cells to form whole‐body tumors that eventually metastasize to the brain. We show the method of establishing brain metastases significantly affects therapeutic BBB/BTB penetration. Free drug accumulates and delays growth in IC‐ and ICD‐formed brain tumors, while non‐targeted nanoparticles show uptake and inhibition only in IC‐established metastases. TfR‐targeted nanoparticles accumulate and significantly delay growth in all three models, suggesting the IV model maintains a more intact BBB/BTB than the other models.

To date, ocular antibody therapies for the treatment of retinal diseases rely on injection of the drug into the vitreous chamber of the eye. Given the burden for patients undergoing this procedure, less frequent dosing through the use of long-acting delivery (LAD) technologies is highly desirable. These technologies usually require a highly concentrated formulation and the antibody must be stable against extended exposure to physiological conditions. Here we have increased the potential of a therapeutic antibody antigen-binding fragment (Fab) for LAD by using protein engineering to enhance the chemical and physical stability of the molecule. Structure-guided amino acid substitutions in a negatively charged complementarity determining region (CDR-L1) of an anti-factor D (AFD) Fab resulted in increased chemical stability and solubility. A variant of AFD (AFD.v8), which combines light chain substitutions (VL-D28S:D30E:D31S) with a substitution (VH-D61E) to stabilize a heavy chain isomerization site, retained complement factor D binding and inhibition potency and has properties suitable for LAD. This variant was amenable to high protein concentration (>250 mg/mL), low ionic strength formulation suitable for intravitreal injection. AFD.v8 had acceptable pharmacokinetic (PK) properties upon intravitreal injection in rabbits, and improved stability under both formulation and physiological conditions. Simulations of expected human PK behavior indicated greater exposure with a 25-mg dose enabled by the increased solubility of AFD.v8.

In women with human epidermal growth factor 2 (HER2)-positive breast cancer, improved control of systemic disease with new therapies has unmasked brain metastases that historically would have remained clinically silent. Efficacy of therapeutic agents against brain metastases is limited by their inability to permeate the blood-brain and blood-tumor barriers (BBB and BTB) in therapeutic amounts. Here, we investigate the potential of mucic acid-based, targeted nanoparticles designed to transcytose the BBB/BTB to deliver a small molecule drug, camptothecin (CPT), and therapeutic antibody, Herceptin, to brain metastases in mice. Treatment with BBB-targeted combination CPT/Herceptin nanoparticles significantly inhibits tumor growth compared to free CPT/Herceptin and BBB-targeted nanoparticles carrying CPT alone. Though not as efficacious, BBB-targeted nanoparticles carrying only Herceptin also elicit considerable antitumor activity. These results demonstrate the potential of the targeted nanoparticle system for the delivery of antibody alone or in combination with other drugs across the BBB/BTB to improve the therapeutic outcome.

1081 Background: Patients with BRCA-positive HER2-negative breast cancer benefit from PARP inhibitor therapy, but additional benefit is still desired. PARP inhibition alone does not prevent all mechanisms for repairing damage to DNA such as homologous recombination repair. An attractive combination for treating such patients would be combining a topoisomerase I inhibitor with a PARP inhibitor given the dual mechanism this would provide for DNA damage and inhibited repair, leading to tumor cell death. This combination has been tried in multiple phase 1 studies, but myelotoxicity prevented the combination from being evaluated further. DAN-222 is a novel investigational polymeric nanoparticle conjugated with camptothecin, a topoisomerase I inhibitor, that provides significant accumulation of drug in tumor tissues via the enhanced permeability and retention (EPR) effect and significantly reduced bone marrow exposure compared to native chemotherapy. These observations underscore the potential advantages of DAN-222 alone as well as in combination with other agents such as PARP inhibitors in solid tumors. Here, we report the effects of DAN-222 monotherapy and in combination with a PARP inhibitor on the growth inhibition in an HRD+ TNBC breast cancer (MDA-MB-436) and an HRD- ovarian (OVCAR3) xenograft mouse model. Methods: HRD+ breast cancer tumor cells (MDA-MB-436) were implanted into female NCr nu/nu mice and HRD- ovarian cancer tumor cells (OVCAR3) were implanted into female CB.17 SCID mice. Mice were randomized to vehicle or treatment arms until tumors reached 2000 mm3 or day 45 (MDA-MB-436) or 1000mm3 or day 45 (OVCAR3). The groups evaluated include multiple dose levels of DAN-222 as monotherapy and those also combined with niraparib. Results: Results were consistent in both the HRD+ and HRD- tumor models with profound dose-response of DAN-222 monotherapy inhibiting tumor growth. Additionally, synergy was demonstrated when DAN-222 was combined with niraparib, clearly evident with low doses of both products when used in combination. The table below highlights the synergy of the combination of DAN-222 at 0.3 mg/kg and niraparib at 25 mg/kg above each agent alone on the tumor growth inhibition in the MDA-MB-436 xenograft. Conclusions: Combining a PARP inhibitor with a topoisomerase I inhibitor delivered via this polymeric nanoparticle delivery system (DAN-222) has synergistic efficacy in both HRD+ and HRD- xenograft tumor models. These data support continued development of DAN-222 to treat solid tumors and its combination use with PARP inhibitors.[Table: see text]

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.