Canady Cold Helios Plasma Reduces Soft Tissue Sarcoma Viability by Inhibiting Proliferation, Disrupting Cell Cycle, and Inducing Apoptosis: A Preliminary Report

Abstract

Soft tissue sarcomas (STS) are a rare and highly heterogeneous group of solid tumors, originating from various types of connective tissue. Complete removal of STS by surgery is challenging due to the anatomical location of the tumor, which results in tumor recurrence. Additionally, current polychemotherapeutic regimens are highly toxic with no rational survival benefit. Cold atmospheric plasma (CAP) is a novel technology that has demonstrated immense cancer therapeutic potential. Canady Cold Helios Plasma (CHCP) is a device that sprays CAP along the surgical margins to eradicate residual cancer cells after tumor resection. This preliminary study was conducted in vitro prior to in vivo testing in a humanitarian compassionate use case study and an FDA-approved phase 1 clinical trial (IDE G190165). In this study, the authors evaluate the efficacy of CHCP across multiple STS cell lines. CHCP treatment reduced the viability of four different STS cell lines (i.e., fibrosarcoma, synovial sarcoma, rhabdomyosarcoma, and liposarcoma) in a dose-dependent manner by inhibiting proliferation, disrupting cell cycle, and inducing apoptosis-like cell death.

Keywords: cancer treatment; cold atmospheric plasma; cold plasma device; fibrosarcoma; liposarcoma; rhabdomyosarcoma; soft tissue sarcoma; synovial sarcoma.

Figure 1

Bar graph showing the cell viability of HT-1080, SW982, and RD cells 48-h post-CHCP treatment compared to mock controls. Cells were seeded in 96-well plates at a concentration of 5 × 10³ cells/well and treated with (A) 1 or (B) 3 LPM at 20–120 P for 1–2 min. Helium alone (0 P) did not significantly impact HT-1080 or SW982 cell viability. CHCP significantly reduced HT-1080 and SW982 viability at all 3 LPM tested doses and RD viability at most 3 LPM tested doses compared to mock controls. Statistical significance for CHCP versus mock controls (^a p ≤ 0.05) and 1 LPM versus 3 LPM CHCP treatment (^b p ≤ 0.05) were considered.

Figure 2

Bar graph showing the cell viability of 94T778 cells 48-h post-CHCP treatment compared to mock controls. Liposarcoma cells were seeded in 12-well plates at a concentration of 2.5 × 10⁴ cells/well and treated with 3 LPM at 120 P for 1–7 min. Helium alone (0 P) did not significantly impact cell viability. All treatment durations lasting at least 3 min significantly reduced viability of 94T778 cells compared to mock controls. Statistical significance for CHCP versus mock controls (^a p ≤ 0.05) were considered.

Figure 3

Representative confocal microscopy images showing Ki67 (green) expression in DAPI-stained (blue) (A) mock control and CHCP-treated (120 P for 5 min) liposarcoma cells after (B) 6, (C) 24, and (D) 48 h. Most mock control cells expressed Ki67, indicating normal cancer proliferation. CHCP-treated cells showed an initial spike in Ki67 expression after 6 h, followed by a sharp decrease in Ki67 expression after 24 and 48 h.

Figure 4

Representative IncuCyte images showing mock control and CHCP-treated cells in G1 (green), S/G2/M (red) and G2-S (yellow) phases over 48 h. Mock control had a relatively even population distribution of all cell cycle phases. Cells treated with 120 P for 5 min arrested in S/G2/M (6 h) before recovering (24–36 h). Cells treated with 120 P for 7 min immediately arrested in S/G2/M (0–6 h) before shrinking (12 h) and undergoing apoptosis (24 h), eventually resulting in zero visible live cells (36–48 h).

Figure 5

Distribution of cell population in live, early apoptotic, or late apoptotic/dead stages 24- or 48-h post-CHCP treatment presented in (A) flow cytometry images and (B) a bar graph. Statistical significance for CHCP versus mock controls (^a p ≤ 0.05) and 24- versus 48-h post-CHCP treatment (^b p ≤ 0.05) were considered.