Results from ongoing studies of two bispecific, hexavalent, antibodies, 1R-(E1)-(E1) and 1R-(15)-(15), for treating breast, pancreatic and other solid cancers, were presented during a recent scientific meeting. The bispecific, hexavalent, antibodies were generated using the patented Dock-and-Lock (DNL) conjugation technology developed by New Jersey-based biopharmaceutical company Immunomedics.
Both agents comprise intact hR1, a proprietary humanized antibody targeting the type I insulin-like growth factor receptor (IGF-1R). 1R-(E1)-(E1) was created by attaching four antibody fragments of hRS7, a humanized antibody that recognizes the trophoblast cell-surface marker (TROP-2), to hR1, while 1R-(15)-(15) utilized fragments from hMN-15, the Company’s proprietary antibody against the carcinoembryonic antigen cell adhesion molecule 6 (CEACAM6).
… these promising results attest to the potential of bispecific hexavalent antibodies for targeted therapy of solid cancers…
In diverse epithelial cancers, the expression of IGF-1R, TROP-2 and CEACAM6 are elevated, making them attractive targets for antibody-based cancer therapy. The use of two distinct monoclonal antibodies in combination to improve efficacy has produced promising results in some early clinical studies. However, combination therapy may be accomplished without increased toxicity with a single bispecific antibody.
The objective of this preclinical study was to explore the potential of 1R-(E1)-(E1) and 1R-(15)-(15) for treating breast and pancreatic cancers by evaluating their effects on three breast cancer cell lines of varying invasive activities and two pancreatic cancer cell lines.
All five cell lines were found to express IGF-1R, TROP-2, and CEACAM6 at various levels. When tested at 100 ug/mL, 1R-(E1)-(E1) reduced the invasion of a breast cancer cell line to less than 10% of the untreated control, whereas under the same conditions, a more invasive cell line appeared to be resistant and the parental antibodies showed no effect. 1R-(15)-(15) potently reduced the invasion of a pancreatic cancer cell line at the same concentration, but had little effect on breast cancer cells. Additionally, at 10 ug/mL, the anti-CEACAM6/anti-IGF-1R bispecific antibody reduced the invasion of pancreatic cancer cells to less than 10% of untreated cells.
The ability of the DNL-derived bispecific antibody to inhibit anchorage-independent growth was demonstrated, with a statistically significant difference when compared with samples treated with parental antibodies at the same concentrations. Furthermore, cells treated with 1R-(E1)-(E1)
produced fewer and much smaller cell colonies, the largest size of which was less than 1/10 of the untreated cells. “We believe these promising results, as exemplified by 1R-(E1)-(E1) and 1R-(15)-(15), attest to the potential of bispecific hexavalent antibodies for targeted therapy of solid cancers,” noted Cynthia L. Sullivan, President and Chief Executive Officer at Immunomedics.
Impact of Linker’s Stability on Efficacy of Antibody-SN-38 Conjugates
Immunomedics also presented a study examining the impact of linker chemistry on the stability and efficacy of antibody-SN-38 conjugates, using a variety of antibodies that differ in their internalization properties.
SN-38 is the active metabolite of irinotecan or CPT-11 (Camptosar®; Pfizer) an FDA-approved drug for metastatic colorectal cancer treatment. Irinotecan has also shown clinical activity in lung, breast and brain cancers. Due to its toxicity and poor solubility in water, SN-38 – which is about three orders of magnitude more potent than irinotecan, cannot be administered systemically to patients with cancer.
To increase the amount reaching the tumors and minimize the damage to normal tissues and organs, the Immunomedics has previously conjugated SN-38 to monoclonal antibodies for improved selective therapy of cancer. The antibody-drug conjugate (ADC), labetuzumab-SN-38, is currently in a Phase I trial for therapy of advanced colorectal cancer, whereas hRS7 (anti-TROP-2)-SN-38, will enter clinical testing for the potential treatment of certain solid cancers later this year.
The linker that connects SN-38 to the antibody is CL2A. This linker provides a practical approach for antibody-based targeted therapies with SN-38. Using CL2A, scientists at Immunomedics are able to produce SN-38 conjugates, including epratuzumab (anti-CD22), veltuzumab (anti-CD20), clivatuzumab (anti-mucin), labetuzumab (anti-CEACAM5) and hRS7 (anti-EGP-1, or TROP-2).
A new linker
In the current study, a new linker, CL2E, was evaluated and compared with CL2A. CL2E is more stable and only releases free SN-38 inside the tumor cell.
In cancer cell lines and animal models of human cancers, ADCs with the more inert CL2E linker were significantly less efficacious than those employing CL2A. These observations were independent of the internalization rates of carrier antibodies, indicating that purely cellular mechanisms of drug release were inadequate for delivering therapeutic levels of SN-38 to tumor cells. A more labile linker in CL2A provides a better release mechanism extracellularly as well as intracellularly, thereby increasing the bioavailability of the drug. This is the linker being used
in the ADC being tested clinically.
Mechanisms of Action of Epratuzumab
At the same cancer research meeting, the in vitro effects of immobilized epratuzumab on malignant B cells were also presented.
Epratuzumab is a humanized anti-CD22 antibody currently in advanced clinical trials in non-Hodgkin lymphoma (NHL) and systemic lupus erythematosus (SLE) patients. As a single agent, epratuzumab is well tolerated and depletes 30 to 50% of circulating B cells in patients with NHL and SLE. However, the mechanism of action of epratuzumab is still largely unknown.
In vitro, epratuzumab shows moderate antibody-dependent cell-mediated cytotoxicity, but no detectable complement-dependent toxicity, and displays cytotoxicity to CD22-expressing human lymphoma cancer cells only when immobilized onto plastic plates or combined with both an anti-immunoglobulin M antibody and a crosslinking secondary antibody. In this preclinical study, the molecular mechanism underlying the cytotoxic effects of immobilized epratuzumab was elucidated.
Using flow cytometry, immunofluorescence microscopy, and various cell-based assays, immobilized epratuzumab was found to induce many of the intracellular changes that were also observed upon crosslinking of anti-immunoglobulin M antibody, which appears to be crucial for cell death. The process also involved mitochondria by changing the mitochondrial membrane potential and the generation of reactive oxygen species. These findings indicate, for the first time, that immobilized epratuzumab and anti-immunoglobulin M antibody behave similarly in killing malignant B cells by affecting B-cell receptor-mediated signaling.