ABSTRACT
Provoked by sterile/nonsterile insults, prolonged monocyte mobilization and uncontrolled monocyte/macrophage activation can pose imminent or impending harm to the affected organs. Curiously, folate receptor beta (FRß), with subnanomolar affinity for the vitamin folic acid (FA), is upregulated during immune activation in hematopoietic cells of the myeloid lineage. This phenomenon has inspired a strong interest in exploring FRß-directed diagnostics/therapeutics. Previously, we have reported that FA-targeted aminopterin (AMT) therapy can modulate macrophage function and effectively treat animal models of inflammation. Our current investigation of a lead compound (EC2319) leads to discovery of a highly FR-specific mechanism of action independent of the root causes against inflammatory monocytes. We further show that EC2319 suppresses interleukin-6/interleukin-1ß release by FRß+ monocytes in a triple co-culture leukemic model of cytokine release syndrome with anti-CD19 chimeric antigen receptor T cells. Because of its chemical stability and metabolically activated linker, EC2319 demonstrates favorable pharmacokinetic characteristics and cross-species translatability to support future pre-clinical and clinical development.
Subject(s)
Aminopterin/pharmacology , Cytokine Release Syndrome/prevention & control , Folate Receptor 2/genetics , Folic Acid Antagonists/pharmacology , Folic Acid/metabolism , Macrophages/drug effects , Animals , Antigens, CD19/genetics , Antigens, CD19/immunology , CHO Cells , Cricetulus , Cytokine Release Syndrome/genetics , Cytokine Release Syndrome/immunology , Cytokine Release Syndrome/pathology , Female , Folate Receptor 1/antagonists & inhibitors , Folate Receptor 1/genetics , Folate Receptor 1/immunology , Folate Receptor 2/antagonists & inhibitors , Folate Receptor 2/immunology , Humans , Interleukin-1beta/genetics , Interleukin-1beta/immunology , Interleukin-6/genetics , Interleukin-6/immunology , Macrophage Activation/drug effects , Macrophages/immunology , Macrophages/pathology , Mice , Models, Biological , Monocytes/drug effects , Monocytes/immunology , Monocytes/pathology , RAW 264.7 Cells , Rats , Rats, Inbred Lew , Receptors, Chimeric Antigen/genetics , Receptors, Chimeric Antigen/immunology , T-Lymphocytes/drug effects , T-Lymphocytes/immunology , T-Lymphocytes/pathologyABSTRACT
Folate receptor (FR)-targeted small molecule drug conjugates (SMDCs) have shown promising results in early stage clinical trials with microtubule destabilizing agents, such as vintafolide and EC1456. In our effort to develop FR-targeted SMDCs with varying mechanisms of action, we synthesized EC2629, a folate conjugate of a DNA crosslinking agent based on a novel DNA-alkylating moiety. This agent was found to be extremely potent with an in vitro IC50 ~ 100× lower than folate SMDCs constructed with various microtubule inhibitors. EC2629 treatment of nude mice bearing FR-positive KB human xenografts led to cures in 100% of the test animals with very low dose levels (300 nmol/kg) following a convenient once a week schedule. The observed activity was not accompanied by any noticeable weight loss (up to 20 weeks post end of dosing). Complete responses were also observed against FR-positive paclitaxel (KB-PR) and cisplatin (KB-CR) resistant models. When evaluated against FR-positive patient derived xenograft (PDX) models of ovarian (ST070), endometrial (ST040) and triple negative breast cancers (ST502, ST738), EC2629 showed significantly greater anti-tumor activity compared to their corresponding standard of care treatments. Taken together, these studies thus demonstrated that EC2629, with its distinct DNA reacting mechanism, may be useful in treating FR-positive tumors, including those that are classified as drug resistant.
Subject(s)
Antineoplastic Agents/pharmacology , Cross-Linking Reagents/pharmacology , DNA/chemistry , Endometrial Neoplasms/drug therapy , Folate Receptors, GPI-Anchored/chemistry , Ovarian Neoplasms/drug therapy , Triple Negative Breast Neoplasms/drug therapy , Alkylating Agents/chemistry , Animals , Cattle , Cisplatin/administration & dosage , Dogs , Drug Delivery Systems , Drug Design , Drug Evaluation, Preclinical , Female , Folic Acid/analogs & derivatives , Folic Acid/pharmacology , Humans , Inhibitory Concentration 50 , KB Cells , Ligands , Mice , Mice, Inbred C57BL , Mice, Nude , Paclitaxel/administration & dosage , Rats , Vinca Alkaloids/pharmacology , Xenograft Model Antitumor AssaysABSTRACT
EC1456 is a folate-tubulysin conjugate constructed with an all-D enantiomeric spacer/linker configuration. When tested against folate receptor (FR)-positive cells, EC1456 demonstrated dose-responsive activity with an approximate 1000-fold level of specificity. Treatment of nude mice bearing FR-positive human xenografts (as large as 800 mm3) with non-toxic doses of EC1456 led to cures in 100% of the mice. Combinations of low dose EC1456 with standard of care agents such as platins, taxanes, topotecan and bevacizumab, safely and significantly augmented the growth inhibitory effects of these commonly used agents. When tested against FR-positive human tumor xenograft models having confirmed resistance to a folate-vinca alkaloid (vintafolide), cisplatin or paclitaxel, EC1456 was found to generate partial to curative responses. Taken together, these studies demonstrate that EC1456 has significant anti-proliferative activity against FR-positive tumors, including models which were anticancer drug resistant, thereby justifying a Phase 1 trial of this agent for the treatment of advanced human cancers.
