Humanized anti-DEspR IgG4S228P antibody increases overall survival in a pancreatic cancer stem cell-xenograft peritoneal carcinomatosis ratnu/nu model.
BACKGROUND
Pancreatic peritoneal carcinomatosis (PPC), with the worst median overall-survival (mOS), epitomizes the incurability of metastatic cancer. Cancer stem cells (CSCs) underpin this incurability. However, inhibitors of CSC-stemness fail to increase mOS in cancer patients despite preclinical tumor-reduction. This shortfall reinforces that preclinical efficacy should be defined by increased mOS in the presence of cancer comorbidities, CSC-heterogeneity and plasticity. The primary objectives of this study are: to test the dual endothelin-1/signal peptide receptor, DEspR, as a nodal therapeutic target in PPC, given DEspR induction in anoikis-resistant pancreatic CSCs, and to validate humanized anti-DEspR antibody, hu-6g8, as a potential therapeutic for PPC.


METHODS
We used heterogeneous pools of CSCs selected for anoikis resistance from reprogrammed Panc1 and MiaPaCa2 tumor cells (TCs), and adherent TCs reprogrammed from CSCs (cscTCs). We used multiple anti-DEspR blocking antibodies (mAbs) with different epitopes, and a humanized anti-DEspR recombinant mAb cross-reactive in rodents and humans, to test DEspR inhibition effects. We measured DEspR-inhibition efficacy on multiple prometastatic CSC-functions in vitro, and on tumorigenesis and overall survival in a CSC-derived xenograft (CDX) nude rat model of PPC with comorbidities.


RESULTS
Here we show that DEspR, a stress-survival receptor, is present on subsets of PDAC Panc1-TCs, TC-derived CSCs, and CSC-differentiated TCs (cscTCs), and that DESpR-inhibition decreases apoptosis-resistance and pro-metastatic mesenchymal functions of CSCs and cscTCs in vitro. We resolve the DNA-sequence/protein-function discordance by confirming ADAR1-RNA editing-dependent DEspR-protein expression in Panc1 and MiaPaCa2 TCs. To advance DEspR-inhibition as a nodal therapeutic approach for PPC, we developed and show improved functionality of a recombinant, humanized anti-DEspR IgG4S228P antibody, hu-6g8, over murine precursor anti-DEspR mabs. Hu-6g8 internalizes and translocates to the nucleus colocalized with cyto-nuclear shuttling galectins-1/3, and induces apoptotic cell changes. DEspR-inhibition blocks transperitoneal dissemination and progression to peritoneal carcinomatosis of heterogeneous DEspR±/CD133 ± Panc1-derived CSCs in xenografted nude rats, improving mOS without chemotherapy-like adverse effects. Lastly, we show DEspR expression in Stage II-IV primary and invasive TCs in the stroma in PDAC-patient tumor arrays.


CONCLUSION
Collectively, the data support humanized anti-DEspR hu-6g8 as a potential targeted antibody-therapeutic with promising efficacy, safety and prevalence profiles for PPC patients.
['Gromisch|Christopher M|CM|', 'Tan|Glaiza L A|GLA|', 'Pasion|Khristine Amber|KA|', 'Moran|Ann-Marie|AM|', 'Gromisch|Matthew S|MS|', 'Grinstaff|Mark W|MW|', 'Carr|Francis J|FJ|', 'Herrera|Victoria L M|VLM|', 'Ruiz-Opazo|Nelson|N|']
D000818:Animals D061067:Antibodies, Monoclonal, Humanized D000074322:Antineoplastic Agents, Immunological D017209:Apoptosis D045744:Cell Line, Tumor D002465:Cell Movement D002470:Cell Survival D002471:Cell Transformation, Neoplastic D004195:Disease Models, Animal D019008:Drug Resistance, Neoplasm D006801:Humans D007074:Immunoglobulin G D007150:Immunohistochemistry D016130:Immunophenotyping D014411:Neoplastic Stem Cells D010190:Pancreatic Neoplasms D010534:Peritoneal Neoplasms D051381:Rats D044022:Receptor, Endothelin A D040262:Receptors, Vascular Endothelial Growth Factor D023041:Xenograft Model Antitumor Assays
Pancreatic cancer Peritoneal carcinomatosis Cancer stem cells Dual endothelin-1/signal peptide receptor DEspR IgG4 antibody therapy Nude rat xenograft tumor model