Inhibition of Ovarian Cancer Cell Spheroid Formation by Synthetic Peptides Derived from Nectin-4.

Kristin L.M Boylan,Rory D Manion,Amy P N Skubitz,Heena Shah,Keith M Skubitz

doi:10.3390/ijms21134637

Abstract

The formation of 3D multicellular spheroids in the ascites fluid of ovarian cancer patients is an understudied component of the disease progression. Spheroids are less sensitive to chemotherapy, in part due to the protection afforded by their structure, but also due to their slower proliferation rate. Previous studies suggest that the cell adhesion molecule Nectin-4 plays a key role in the formation of ovarian cancer spheroids. In this study, we further examined the role of Nectin-4 at early time points in spheroid formation using real-time digital photography. Human NIH:OVCAR5 ovarian cancer cells formed aggregates within 8 h, which further contracted into compact spheroids over 24 h. In contrast, Nectin-4 knockdown cells did not form tightly compacted spheroids. Synthetic peptides derived from Nectin-4 were tested for their ability to alter spheroid formation in two ovarian cancer cell lines. Nectin-4 peptide 10 (N4-P10) had an immediate effect on disrupting ovarian cancer spheroid formation, which continued for over 24 h, while a scrambled version of the peptide had no effect. N4-P10 inhibited spheroid formation in a concentration-dependent manner and was not cytotoxic; suggesting that N4-P10 treatment could maintain the cancer cells as single cells which may be more sensitive to chemotherapy.

Highlights

With an overall 5 year survival rate of only 50%, ovarian cancer is regarded as the deadliest gynecological malignancy [1]
Previous investigations into the role of Nectin-4 revealed a disparity in spheroid formation potential between parental NIH:OVCAR5 cells and Nectin-4 shRNA knockdown cells (N4-KD-VB9) when the cells were monitored at a few selected time points over 3–5 days [15]
Peptide Nectin-4 peptide 10 (N4-P10) Inhibits a Second Ovarian Cancer Cell Line from Forming Spheroids In previous studies, we examined the role of Nectin-4 in the biological functions of ovarian canIcnerpcreelvlsio[u1s4,s1t5u]d. iWese, swheowexeadmtihnaetdNtIhHe:rOoVleCoAf RN5eccteilnls-4foinrmthteigbhitoslopghiecraolifduangcgtiroengsatoefsoovnarfliaatn-bcoatntocemr cells [1l4o,1w5-]a.tWtaechsmhoewntepdlathteast,NwIhHil:eOaVsCeAcoRn5dchelulsmfaonrmovtiagrhiatnspcahnecroeridcealglglirneeg,aCteAsOonV3fl,afto-brmotetodmmloorwe -laototaseclhyment plaagtegsr,ewgahtieledaspsehceornodidhsu[m15a].n Ionvathriiasnstcuadnyce, rwceelsloliungeh, tCtAoOmVo3r,efocrommepdremhoenresilvoeolsyelaynaaglygzreegtahteedroslpehoefroids [1N5]e. cItnint-h4isinstsupdhye,rwoied sfoourmghattitoonm

Summary

Introduction

With an overall 5 year survival rate of only 50%, ovarian cancer is regarded as the deadliest gynecological malignancy [1]. Ascites fluid serves as a rich source of nutrients and is a suitable microenvironment for ovarian cancer cell proliferation and metastasis [4]. Both single cells and spheroids are shed from tumors and metastasize to sites within the peritoneal cavity through the ascites fluid. It is important to consider the physiology of spheroids, as they are more resistant to a wide range of chemotherapies [5,6]. When floating in ascites fluid, spheroids exhibit a decreased cell proliferation. They are able to regain their proliferative abilities upon attaching to tissue [7]. Cell–cell adhesion enables cell signaling to take place and plays an important role in spheroid formation

Methods

Results

Conclusion