Association of Left Ventricular Remodeling Assessment by Cardiac Magnetic Resonance With Outcomes in Patients With Chronic Aortic Regurgitation.

Importance: Chronic aortic regurgitation (AR) causes left ventricular (LV) volume overload, which results in progressive LV remodeling negatively affecting outcomes. Whether cardiac magnetic resonance (CMR) volumetric quantification can provide incremental risk stratification over standard clinical and echocardiographic evaluation in patients with chronic moderate or severe AR is unknown. Objective: To compare LV remodeling measurements by CMR and echocardiography between patients with and without heart failure symptoms and to verify the association of remodeling measurements of patients with chronic moderate or severe AR but no or minimal symptoms with clinical outcomes receiving medical management. Design, Setting, and Participants: This multicenter retrospective cohort study included consecutive patients with at least moderate chronic native AR evaluated by 2-dimensional transthoracic echocardiography and CMR examination within 90 days from each other between January 2012 and February 2020 at Allina Health System. Data were analyzed from June 2021 to January 2022. Exposures: Clinical evaluation and risk stratification by CMR. Main Outcomes and Measures: The end point was a composite of death, heart failure hospitalization, or progression of New York Heart Association functional class while receiving medical management, censoring patients at the time of aortic valve replacement (when performed) or at the end of follow-up. Results: Of the 178 included patients, 119 (66.9%) were male, 158 (88.8%) presented with no or minimal symptoms (New York Heart Association class I or II), and the median (IQR) age was 58 (44-69) years. Compared with patients with no or minimal symptoms, symptomatic patients had greater LV end-systolic volume index (LVESVi) by CMR (median [IQR], 66 [46-85] mL/m2 vs 42 [30-58] mL/m2; P < .001), while there were no significant differences by echocardiography (LVESVi: median [IQR], 38 [30-58] mL/m2 vs 27 [20-42] mL/m2; P = .07; LV end-systolic diameter index: median [IQR], 21 [17-25] mm/m2 vs 18 [15-22] mm/m2; P = .17). During the median (IQR) follow-up of 3.3 (1.6-5.8) years, 50 patients with no or minimal symptoms receiving medical management developed the composite end point, which, in multivariate analysis adjusted for age and EuroSCORE II, was independently associated with LVESVi of 45 mL/m2 or greater and aortic regurgitant fraction of 32% or greater, the latter adding incremental prognostic value to CMR volumetric assessment. Conclusions and Relevance: In patients with chronic moderate or severe AR, patients presenting with heart failure symptoms have greater LVESVi by CMR than those with no or minimal symptoms. In patients with no or minimal symptoms, CMR quantification of LVESVi and AR severity may identify those at risk of death or incident heart failure and therefore should be considered in the clinical evaluation and decision-making of these patients.

A HER2 Tri-Specific NK Cell Engager Mediates Efficient Targeting of Human Ovarian Cancer.

Clinical studies validated antibodies directed against HER2, trastuzumab, and pertuzumab, as useful methodology to target breast cancer cases where HER2 is expressed. The hope was that HER2 targeting using these antibodies in ovarian cancer patients would prove useful as well, but clinical studies have shown lackluster results in this setting, indicating a need for a more comprehensive approach. Immunotherapy approaches stimulating the innate immune system show great promise, although enhancing natural killer (NK) function is not an established mainstream immunotherapy. This study focused on a new nanobody platform technology in which the bispecific antibody was altered to incorporate a cytokine. Herein we describe bioengineered CAM1615HER2 consisting of a camelid VHH antibody fragment recognizing CD16 and a single chain variable fragment (scFv) recognizing HER2 cross-linked by the human interleukin-15 (IL-15) cytokine. This tri-specific killer engager (TriKETM) showed in vitro prowess in its ability to kill ovarian cancer human cell lines. In addition, we demonstrated its efficacy in inducing potent anti-cancer effects in an in vivo xenograft model of human ovarian cancer engrafting both cancer cells and human NK cells. While previous approaches with trastuzumab and pertuzumab faltered in ovarian cancer, the hope is incorporating targeting and cytokine priming within the same molecule will enhance efficacy in this setting.

Bispecific Targeting of EGFR and Urokinase Receptor (uPAR) Using Ligand-Targeted Toxins in Solid Tumors.

Ligand-targeted toxins (LTTs) are bioengineered molecules which are composed of a targeting component linked to a toxin that induces cell death once the LTT binds its target. Bispecific targeting allows for the simultaneous targeting of two receptors. In this review, we mostly focus on the epidermal growth factor receptor (EGFR) as a target. We discuss the development and testing of a bispecific LTT targeting EGFR and urokinase-type plasminogen activator receptor (uPAR) as two attractive targets implicated in tumor growth and in the regulation of the tumor microvasculature in solid tumors. In vitro and mouse xenograft studies have shown that EGFR-targeted bispecific angiotoxin (eBAT) is effective against human solid tumors. Canine studies have shown that eBAT is both safe and effective against canine hemangiosarcoma, which is physiologically similar to human angiosarcoma. Finding the appropriate dosing strategy and sequencing of eBAT administration, in combination with other therapeutics, are among important factors for future directions. Together, the data indicate that eBAT targets cancer stem cells, it may have a role in inhibiting human tumor vasculature, and its bispecific conformation may have a role in reducing toxicity in comparative oncologic trials in dogs.

Targeting EGFR and uPAR on human rhabdomyosarcoma, osteosarcoma, and ovarian adenocarcinoma with a bispecific ligand-directed toxin.

PURPOSE: Human sarcomas are rare and difficult to treat cancerous tumors typically arising from soft tissue or bone. Conversely, carcinomas are the most common cancer subtype in humans and the primary cause of mortality across all cancer patients. While conventional therapeutic modalities can prolong disease-free intervals and survival in some cases, treatment of refractory or recurrent solid tumors is challenging, and tumor-related mortality remains unacceptably high. The identification of overexpressed cell surface receptors on sarcoma and carcinoma cells has provided a valuable tool to develop targeted toxins as an alternative anticancer strategy. Recent investigation of recombinant protein-linked toxins that specifically target these cancer receptors has led to the development of highly specific, cytotoxic, and deimmunized drugs that can kill cancer cells. METHODS: This study investigated a recombinant protein called epidermal growth factor bispecific angiotoxin (eBAT), which is designed to target the epidermal growth factor receptor (EGFR) on cancer cells and the urokinase plasminogen activator receptor (uPAR) on cancer cells and associated tumor vasculature. Both receptors are expressed by a variety of human sarcomas and carcinomas. Flow cytometry techniques were used to determine binding affinity of eBAT to cancer cells, and proliferation assays were performed to calculate tumor killing ability based on half-maximal inhibitory concentrations. RESULTS: eBAT demonstrated cytotoxicity against a variety of sarcoma and carcinoma cells that overexpress EGFR and uPAR in vitro and showed greater cell killing ability and binding affinity to cancer cells compared with its monospecific counterparts. CONCLUSION: The results of our study are promising, and further studies will be necessary to confirm the applicability of eBAT as a supplementary therapy for a variety of sarcomas, carcinomas, and possibly other refractory malignancies that express EGFR and uPAR.

Engineering of Anti-CD133 Trispecific Molecule Capable of Inducing NK Expansion and Driving Antibody-Dependent Cell-Mediated Cytotoxicity.

PURPOSE: The selective elimination of cancer stem cells (CSCs) in tumor patients is a crucial goal because CSCs cause drug refractory relapse. To improve the current conventional bispecific immune-engager platform, a 16133 bispecific natural killer (NK) cell engager (BiKE), consisting of scFvs binding FcγRIII (CD16) on NK cells and CD133 on carcinoma cells, was first synthesized and a modified interleukin (IL)-15 crosslinker capable of stimulating NK effector cells was introduced. MATERIALS AND METHODS: DNA shuffling and ligation techniques were used to assemble and synthesize the 1615133 trispecific NK cell engager (TriKE). The construct was tested for its specificity using flow cytometry, cytotoxic determinations using chromium release assays, and lytic degranulation. IL-15-mediated expansion was measured using flow-based proliferation assays. The level of interferon (IFN)-γ release was measured because of its importance in the anti-cancer response. RESULTS: 1615133 TriKE induced NK cell-mediated cytotoxicity and NK expansion far greater than that achieved with BiKE devoid of IL-15. The drug binding and induction of cytotoxic degranulation was CD133+ specific and the anti-cancer activity was improved by integrating the IL-15 cross linker. The NK cell-related cytokine release measured by IFN-γ detection was higher than that of BiKE. NK cytokine release studies showed that although the IFN-γ levels were elevated, they did not approach the levels achieved with IL-12/IL-18, indicating that release was not at the supraphysiologic level. CONCLUSION: 1615133 TriKE enhances the NK cell anti-cancer activity and provides a self-sustaining mechanism via IL-15 signaling. By improving the NK cell performance, the new TriKE represents a highly active drug against drug refractory relapse mediated by CSCs.

Safe and Effective Sarcoma Therapy through Bispecific Targeting of EGFR and uPAR.

Sarcomas differ from carcinomas in their mesenchymal origin. Therapeutic advancements have come slowly, so alternative drugs and models are urgently needed. These studies report a new drug for sarcomas that simultaneously targets both tumor and tumor neovasculature. eBAT is a bispecific angiotoxin consisting of truncated, deimmunized Pseudomonas exotoxin fused to EGF and the amino terminal fragment of urokinase. Here, we study the drug in an in vivo "ontarget" companion dog trial as eBAT effectively kills canine hemangiosarcoma and human sarcoma cells in vitro We reasoned the model has value due to the common occurrence of spontaneous sarcomas in dogs and a limited lifespan allowing for rapid accrual and data collection. Splenectomized dogs with minimal residual disease were given one cycle of eBAT followed by adjuvant doxorubicin in an adaptive dose-finding, phase I-II study of 23 dogs with spontaneous, stage I-II, splenic hemangiosarcoma. eBAT improved 6-month survival from <40% in a comparison population to approximately 70% in dogs treated at a biologically active dose (50 µg/kg). Six dogs were long-term survivors, living >450 days. eBAT abated expected toxicity associated with EGFR targeting, a finding supported by mouse studies. Urokinase plasminogen activator receptor and EGFR are targets for human sarcomas, so thorough evaluation is crucial for validation of the dog model. Thus, we validated these markers for human sarcoma targeting in the study of 212 human and 97 canine sarcoma samples. Our results support further translation of eBAT for human patients with sarcomas and perhaps other EGFR-expressing malignancies. Mol Cancer Ther; 16(5); 956-65. ©2017 AACR.