Tumor Treating Fields (TTFields), and their concomitant application with FOLFOX, are effective for the treatment of gastric cancer cells

Einav ZeeviNaama Flint-BrodslyHila FishmanHila EneKerem Wainer-KatsirRoni Frechtel-GerziAntonia Martinez-CondeEyal Dor OnMijal MunsterYaara PoratTali VoloshinShiri DavidiGitit Lavy-ShahafAdi HaberMoshe Giladi^*Uri WeinbergYoram Palti

• Novocure (Israel), Haifa, Israel

Background: Tumor Treating Fields (TTFields) are electric fields that exert antimitotic effects and impair DNA damage repair in cancerous cells. Gastric cancer, one of the most common types of cancers worldwide, demonstrates poor long-term survival despite advances in therapeutic options. The goal of the current study was to examine in vitro the efficacy and mechanism of action of TTFields for treating gastric cancer, and the potential application of TTFields concomitant with FOLFOX, a standard gastric cancer treatment consisting of the therapeutic agents oxaliplatin and 5-fluorouracil [5-FU]. Methods: Human gastric cancer cell lines, AGS and KATOIII, were treated with TTFields using the inovitro system. To test the efficacy of TTFields (alone or together with oxaliplatin, 5-FU, or FOLFOX), cell count, colony formation, and apoptosis were examined. For mechanistic insight, control and TTFields-treated cells were examined by RNA sequencing. Immunofluorescent staining for phospho-histone H2AX (γH2AX), α-tubulin, phospho-histone 3 (pH3), and double stranded DNA was employed for detection of DNA damage, mitotic spindle defects, chromosome mislocalization, and micronuclei clusters, respectively. Expression of DNA damage repair proteins was measured using Western Blot. Results: Maximal cell count reduction following application of TTFields at various frequencies was identified at 150 kHz. Treatment also led to reduced colony formation, elevated apoptosis, and substantial changes in transcriptomic expression. TTFields-treated cells demonstrated increased DNA damage, elevated expression of DNA damage-related cyclin-dependent kinase (CDK) inhibitors, and reduced expression of proteins from the Fanconi Anemia-BRCA pathway for DNA repair. Treated cells further presented with abnormal mitotic figures, chromosome mislocalization, and micronuclei clusters, indicative of an antimitotic effect of TTFields. The application of TTFields concomitant with oxaliplatin, 5-FU or FOLFOX was more effective than each treatment alone. Conclusions: The study shows that TTFields induce an antimitotic effect and impair DNA damage repair in gastric cancer cells, and that concomitant treatment with FOLFOX has improved treatment efficacy in vitro, potentially due to the accumulation of DNA damage. Overall, TTFields treatment holds potential for treatment of gastric cancer alongside standard chemotherapy.