A Novel Anti-Kv10.1 Nanobody Fused to Single-Chain TRAIL Enhances Apoptosis Induction in Cancer Cells

Abstract

Antibody-based therapies hold promise for a safe and efficient treatment of cancer. The identification of target tumor cells through a specific antigen enriched on their surface and the subsequent delivery of the therapeutic agent only to those cells requires, besides the efficacy of the therapeutic agent itself, the identification of an antigen enriched on the surface of tumor cells, the generation of high affinity antibodies against that antigen. We have generated single-domain antibodies (nanobodies) against the voltage-gated potassium channel Kv10.1, which outside of the brain is detectable almost exclusively in tumor cells. The nanobody with highest affinity was fused to an improved form of the tumor necrosis factor-related apoptosis inducing ligand TRAIL, to target this cytokine to the surface of tumor cells. The resulting construct, VHH-D9-scTRAIL, shows rapid and strong apoptosis induction in different tumor models in cell culture. The construct combines two sources of specificity, the expression of the antigen restricted to tumor cells and the tumor selectivity of TRAIL. Such specificity combined with the high affinity obtained through nanobodies make the novel agent a promising concept for cancer therapy.

Overcoming challenges of HERG potassium channel liability through rational design: Eag1 inhibitors for cancer treatment

“…19,[104][105][106][107][108][109][110][111][112][113][114][115][116][117] Therefore, Eag1 can be considered an almost universal tumor marker and has been used to target therapeutic agents to tumor cells, preserving the surrounding normal tissue. [118][119][120] Moreover, tumor cells expressing Eag1 obtain a selective advantage and become rapidly dependent on the channel to proliferate and survive, in a mechanism similar to the oncogene addiction described for classical oncogenes. This has two implications: On the one hand, patients with tumors that express Eag1 show in many cases shorter survival, like documented for fibrosarcoma, 117 ovary carcinoma, 121 glioblastoma, 116 acute lymphoid leukemia, 108 gastric, 105 head and neck, 111 or colon cancers.…”

“…Some of those constructs have shown efficacy in vivo as sensitizers to conventional chemotherapeutic agents. [118][119][120]…”

Ion Channels Orchestrate Pancreatic Ductal Adenocarcinoma Progression and Therapy

“…However, an increasing number of ion channel protein structures is becoming amenable for in silico drug design (e.g., Brömmel et al (2020b) ). Alternatively, peptide-based blockers or antibody targeting could be more specific alternatives ( Duranti and Arcangeli, 2019 ; Hartung et al, 2020 ; Tajti et al, 2020 ). Here, we list potential target ion channels and the existing knowledge about the consequence of their activation/inhibition in immune, stroma, and tumor cells as well.…”

Voltage-Gated Potassium Channels as Regulators of Cell Death

“…Additionally, specific small antibody fragments targeting the extracellular pore domain of Kv10.1 were exploited to selectively guide the Tumor necrosis factor-Related Apoptosis-Inducing Ligand (TRAIL) to tumor cells. These constructs sensitized breast cancer cells to chemotherapeutics they were otherwise resistant to Hartung and Pardo (2016) , or directly induced cell death in prostate and pancreatic cancer cells when fused to a TRAIL variant with enhanced pro-apoptotic activity ( Hartung et al, 2020 ). This antibody-based strategy was proven efficacious also in vivo ( Hartung and Pardo, 2016 ).…”

“…Inhibition with anti-Kv3.1 antibodies or downregulation reduces proliferation of oligodendrocyte progenitor cells -Tiwari-Woodruff et al, 2006 Kv3.4 -Increased expression under hypoxia increases proliferation of oral squamous cell carcinoma cells -Increased activity after radiotherapy arrests myeloid leukemia cells at G2/M due to hyperpolarization of the membrane -Qian et al, 2019 -Palme et al, 2013 Kv7.4 -Inhibition with 10 µM Linopirdine increases cell death of spiral ganglion neurons -Lv et al, 2010 Kv10.1 -Blockage with a monoclonal antibody decreases proliferation of different cancer cell lines and inhibits breast and pancreatic tumor growth -Inhibition with 10 µM Astemizole reduces cell proliferation and viability in a cervical cancer model -Inhibition with 1-3 µM Astemizole synergistically enhances the anti-proliferative effects of calcitriol on breast cancer cells -Simultaneous knockdown of Kv10.1 and overexpression of TRAIL induces apoptosis of MG-63 osteosarcoma cells and reduces tumor growth -Inhibition with 5 µM Astemizole reduces HepG2 and HuH-7 hepatocellular carcinoma cell viability and proliferation and tumor growth -TRAIL-ligated antibodies sensitize chemoresistant MDA-MB-435S breast cancer cells tochemotherapeutics and reduce tumor growth -Knockdown or inhibition with 5 µM Astemizole increases U-87MG glioblastoma sensitivity to Temozolomide -Knockdown or inhibition with 2.5-7.5 µM Astemizole increases the sensitivity of SH-SY5Y to rotenone-induced apoptosis -Inhibition with 7.5-9 µM Astemizole acts synergistically with Gefitinib to reduce proliferation and increase apoptosis of lung cancer cell lines -TRAIL-ligated nanobodies induce apoptosis in prostate and pancreatic cancer cells -Inhibition with 1-100 µM or 15 mg/kg Procyanidin B1 inhibits proliferation and migration of liver cancer cells and hepatoma growth -Gomez-Varela et alet al, 2017-Hartung et al, 2020-Na et al, 2020 …”