Novel functional anti-HER3 monoclonal antibodies with potent anti-cancer effects on various human epithelial cancers

Abstract

Copyright: Okita et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License 3.0 (CC BY 3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. ABSTRACTResistance of progressive cancers against chemotherapy is a serious clinical problem. In this context, human epidermal growth factor receptor 3 (HER3) can play important roles in drug resistance to HER1-and HER2-targeted therapies. Since clinical testing of anti-HER3 monoclonal antibodies (mAbs) such as patritumab could not show remarkable effect compared with existing drugs, we generated novel mAbs against anti-HER3. Novel rat mAbs reacted with HEK293 cells expressing HER3, but not with cells expressing HER1, HER2 or HER4. Specificity of mAbs was substantiated by the loss of mAb binding with knockdown by siRNA and knockout of CRISPR/ Cas9-based genome-editing. Analyses of CDR sequence and germline segment have revealed that seven mAbs are classified to four groups, and the binding of patritumab was inhibited by one of seven mAbs. Seven mAbs have shown reactivity with various human epithelial cancer cells, strong internalization activity of cell-surface HER3, and inhibition of NRG1 binding, NRG1-dependent HER3 phosphorylation and cell growth. Anti-HER3 mAbs were also reactive with in vivo tumor tissues and cancer tissue-originated spheroid. Ab4 inhibited in vivo tumor growth of human colon cancer cells in nude mice. Present mAbs may be superior to existing anti-HER3 mAbs and support existing anti-cancer therapeutic mAbs.

Liver Endothelium Promotes HER3-mediated Cell Survival inKRASWild-type and Mutant Colorectal Cancer Cells

“…However, previous clinical studies showed that blocking HER3 had limited anti-tumor effect in several types of cancer by using either HER3 antibodies/inhibitors as monotherapies or in combination with EGFR-or HER2-targeted therapies such as cexucimab and trastuzumab [48][49][50] . The HER3 antibody used in this study, seribantumab, has been used in clinical studies for treating breast, lung, and ovarian cancer either alone or in combination with paclitaxel or EGFR/HER2 targeted therapies [51][52][53] .…”

Oncogenic transformation of NIH/3T3 cells by the overexpression of L-type amino acid transporter 1, a promising anti-cancer target

“…KO was performed as recently described [ 51 , 52 ] using pX330 and pCAG-EGxxFP [ 53 ] purchased from Addgene (Watertown, MA, USA). For CRISPR/Cas9-based CD98hc (SLC3A2) gene disruption, guide (g) RNA sequences (5′-GCCGCGTTGTCGCGAGCTAC-3′) corresponding to the CD98hc gene (318-bp ~ 337-bp from the initiation ATG site) were designed using CRISPR direct ( https://crispr.dbcls.jp/ ).…”

Antitumor effects of novel mAbs against cationic amino acid transporter 1 (CAT1) on human CRC with amplified CAT1 gene

“…Aseptic processing during cell culture was ideally controlled by a MediAir air purifier (Pieras Co., Ltd) equipped in the cell‐culture room. Cell growth was measured as previously described 13 . Briefly, a WST‐8–based cell counting kit (Dojin Chemicals) was used, and 450 nm was measured using a microplate reader.…”

“…Cell growth was measured as previously described. 13 Briefly, a WST-8-based cell counting kit (Dojin Chemicals) was used, and 450 nm was measured using a microplate reader.…”

“…For the selection of hybridomas, FCM was used. [12][13][14][15] For the first screening, HEK293F cells stably expressing CAT1-GFP were reacted with the culture supernatant of hybridoma cells for 45 minutes, followed by the incubation with PE-labeled donkey anti-rat IgG pAb (1:200; Jackson ImmunoResearch) for 30 minutes. For the second screening, reactivity of antibodies with HT29 cells (1.5 × 10 4 ) transfected with CAT1 or control siRNA (28.8 pmol; Dharmacon) was examined.…”