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2-2017
Cold atmospheric plasma, a novel promising anti-
cancer treatment modality.
Dayun Yan
George Washington University
Jonathan H Sherman
George Washington University
Michael Keidar
George Washington University
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APA Citation
Yan, D., Sherman, J., & Keidar, M. (2017). Cold atmospheric plasma, a novel promising anti-cancer treatment modality.. Oncotarget, 8
(9). h?p://dx.doi.org/10.18632/oncotarget.13304
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www.impactjournals.com/oncotarget/ Oncotarget, 2017, Vol. 8, (No. 9), pp: 15977-15995
Cold atmospheric plasma, a novel promising anti-cancer
treatment modality
Dayun Yan
1
, Jonathan H. Sherman
2
and Michael Keidar
1
1
Department of Mechanical and Aerospace Engineering, The George Washington University, NW, Washington, DC, USA
2
Neurological Surgery, The George Washington University, NW, Washington, DC, USA
Correspondence to: Dayun Yan, email: ydy2012@gwmail.gwu.edu
Correspondence to: Michael Keidar, email: keidar@gwu.edu
Keywords: cold plasma, cancer treatment, reactive species, selectivity
Received: September 14, 2016 Accepted: October 29, 2016 Published: November 11, 2016
ABSTRACT
Over the past decade, cold atmospheric plasma (CAP), a near room temperature
ionized gas has shown its promising application in cancer therapy. Two CAP devices,
namely dielectric barrier discharge and plasma jet, show signicantly anti-cancer
capacity over dozens of cancer cell lines in vitro and several subcutaneous xenograft
tumors in vivo. In contrast to conventional anti-cancer approaches and drugs, CAP
is a selective anti-cancer treatment modality. Thus far establishing the chemical and
molecular mechanism of the anti-cancer capacity of CAP is far from complete. In
this review, we provide a comprehensive introduction of the basics of CAP, state of
the art research in this eld, the primary challenges, and future directions to cancer
biologists.
PLASMA AND COLD PLASMA
There are four fundamental states of matter: solid,
liquid, gas, and plasma (Figure 1a). As the energy exerting
on atoms increases, the thermal motion of atoms in the
solid aggravates and nally overcomes the restrictive
interaction between atoms in solid such as ionic bond
and forms liquid. Similarly, when the atoms in liquid
obtain adequately large energy to overcome the restrictive
Van der Waals force from surrounding atoms, these
liquid atoms will transfer into gas atoms. Obviously, the
translational energy of atoms in gas is much larger than
that in liquid and in solid. When the energy is large enough
for the electron to overcome the electrostatic potential
barrier, the electron will be stripped away creating a free
electron and a positive charged ion. This process is called
ionization. The plasma is on avarage a neutral ionized gas
composed of positive charged ions, electrons, and neutral
particles.
In general, temperature increases when matters
transform from solid to liquid to gas and to plasma. The
temperature of plasma is determined by thermal motions
of electrons and heavy particles such as atoms and ions.
In the case of a common thermal plasma, when the
density of particles is high, due to intensive collisions
between eletrons and heavy particles, all particles
approach thermal equilibrium [1]. The temperature in
such plasma is high, over several thousand degrees [1].
These plasmas are typically used under the atmospheric
pressure conditions. On the other hand, if atmospheric
pressure plasma discharge is fast, there is another class
of plasmas in which electrons and heavy particles are in
thermal non-equilibrium. In this case, temperature of the
heave particles is much lower than that of the electrons
(Figure 1b). We shall call these plasmas, cold atmospheric
plasmas (CAP). The heavy particle temperature of CAP
is between 25°C and 45°C [2]. Such plasmas can be used
in biomedicine [3]. Many reactive species including
oxygen-based radicals, nitrogen-based radicals, and other
components are generated in CAP [4-6]. This complicated
chemistry leads to a myriad of interaction between CAP
and biological systems including cells and tissues [7-9].
CAP DEVICES
Two main approaches have been widely used to
generate CAP, namely direct and indirect discharges.
In an indirect discharge, the active plasma species are
transported by a gas ow from the main discharge arc.
In a direct discharge, living tissue or cells is one of the
electrodes and is an active part of the discharge. Based
Review
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these principles, two CAP devices, the plasma jet [6, 10,
11] and the dielectric barrier discharge (DBD) [12, 13],
have been developed and widely used in plasma medicine
[14-22]. The plasma jet is also called the plasma pencil
[23, 24], the plasma needle [2, 25, 26], or the plasma gun
[27] in some references. As shown in Figure 2, the plasma
jet device and the DBD device share similar physical
principles, components, and materials. In these two
devices, a violet plasma is generated between an annode
and a cathode. Either anode or cathode is covered by a
layer of dielectric materials such as quartz [19, 28]. In
many plasma jet devices, the quartz hollow tube is used
as the dielectric layer on the cathode [2, 29]. The metal
cathode such as copper surrounds the quartz tube. In
addition, the plasma jet device needs a carrying gas such
as helium [18, 26, 30] or argon [25, 31, 32] to sustain the
formation of CAP while the DBD device can generate the
plasma directly in the air [20-22, 33]. In some applications,
oxygen [4, 34] and nitrogen [35, 36] have been added
in the carrying gas to achieve the specic chemical
composition. Due to the continuous ow of the carrying
gas, a CAP jet forms. On the other hand, the DBD device
tends to generate a short but a wide plasma. Moreover, the
functions of samples in the two cold plasma devices are
also different. In the plasma jet device, the sample is just
treated by the plasma jet [4, 18, 37]. On the other hand, in
DBD device, the sample is a part of discharge [20, 22, 38].
The CAP in DBD will not be generated if the sample is not
Figure 1: The physical description of plasmas. a. Schematic illustration of the four fundamental states of matter. The triangular tails
represent the thermal motion strength of particles. b. Schematic illustration of the thermal plasma and the cold plasma. Brown balls, violet
balls, and iridescent balls represent the neutral atoms, the positively charged ions, and electrons, respectively.
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adequately close to the second electrode. Based on these
properties and features, the plasma jet device may be more
suitable for gentlly treating a small area on a sample. In
contrast, the DBD may be more suitable for a more intense
treatment on a large area of sample. A recent review
written by X. Lu, et al., comprehensively introduced the
physical foundation of the reactive species generation in
different CAP devices [39].
RESEARCH STATUS
There exist a number of published reviews that
discuss the application of CAP on cancer treatment [40-
44] [45]. In this review, we focused on introducing the
basic concept of CAP and the biological basis of the anti-
cancer mechanism of CAP. Since the rst report about the
killing effect of DBD on melanoma in 2007, the eld of
CAP application in cancer treatment experienced a fast
growth (Figure 3a). Through a comprehensive survey
of all publications by September 2016, it is found that
about 75% papers in this eld were published in the
multidisciplinary journals such as PLoS ONE, the applied
physics-related journals such as Applied Physics Letters,
as well as the plasma-related journals such as Plasma
Processes and Polymers (Figure 3b). On the other hand,
only about 25% papers were published in the life science
& medicine-related journals. As a result, the research in
this eld is mainly focused on describing the anti-cancer
effect of CAP treatment on different cancer cell lines [41,
44] and tumors in animal models [20, 30, 46]. To date,
the CAP treatment has demonstrated its signicant anti-
cancer capacity over approximately 20 cancers types in
vitro. Among these cancer cell lines, brain cancer [27,
47, 48], skin cancer [2, 19, 49], breast cancer [50-52],
colorectal cancer [15, 53, 54], lung cancer [18, 46, 55],
cervical cancer [56-58], leukemia [23, 59, 60], hepatoma
[25, 37, 58], as well as head & neck cancer [61-63] have
been intensively investigated (Figure 3c). Moreover, about
70% of the entire publications employed the plasma jet
devices as the anti-cancer tools (Figure 3d).
In addition to resisting the growth of cancer cells,
CAP is also able to restore the sensitivity of chemo-
resistant cancer cells to specic drugs. One example is that
CAP restored temozolomide (TMZ)-resistant glioblastoma
cells to TMZ therapy [64]. Another example is that CAP
made tumor necrosis factor-related apoptosis-inducing
ligand (TRAIL)-resistant colorectal cancer cells sensitive
to the TRAIL treatment [65].
Moreover, CAP can obtain a stronger anti-cancer
capacity through the synergistic application with
nanoparticles technologies. M.G. Kong, et al. gave a
detailed illustration in a review to describe the potential
synergistic application of CAP and nanoparticles in
medicine [66]. The enhanced anti-melanoma effect was
rst achieved using CAP to treat melanoma cancer cells
which had been pretreated with the anti-FAK antibody-
conjugated gold nanoparticles [49]. Clearly, such surface-
modied nanoparticles weaken the normal function of
FAK, which may intensify the CAP-triggering detachment
of melanoma cells from the substrate. It was further
demonstrated that the pretreatment of gold nanoparticles
without the specic antibody modication also enhanced
the anti-glioblastoma effect of the plasma jet [67]. A
recent study demonstrated that the combined treatment
of polyethylene glycol (PEG)-coated gold nanoparticles
and CAP increased cancer cells death in solid tumors
Figure 2: The plasma jet device and dielectric barrier discharge (DBD) device are two main CAP devices used in
plasma medicine. The same components in the plasma jet and DBD are drawn with the same colors. The left inset is reproduced
with permission from Alan Siu, et al., PLoS ONE, 10(6), e0126313 (2015). Copyright 2015 Public Library of Science. The right inset is
reproduced with permission from Sameer Kalghatgi, et al., PLoS ONE, 6(1), e16270 (2011). Copyright 2011 Public Library of Science.
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and decreased epithelial-mesenchymal transition (EMT)
[68]. In addition to using nanoparticles, drug encapsulated
core-shell nanoparticles synthesized via co-axial
electrospraying has also shown its synergistic anti-cancer
potential with CAP on breast cancer cells [69]. These
studies indicate that nanoparticles may weaken or damage
the normal function of specic proteins or pathways,
which resist the intracellular change due to the CAP-
originated reactive species.
Two basic strategies of using CAP have been
developed. One is to employ the plasma jet [25] or DBD
[19] to directly treat the cells seeded in a petri dish or
a multi-wells plate or the subcutaneous tumors in mice
(Figure 4a). Another approach is to use the cold plasma-
stimulated solutions (PSA) mainly the cold plasma-
stimulated medium (PSM) to inhibit the growth of cancer
cells during the standard cell culture process [47, 70] or
to inhibit the growth of tumor tissues by injecting PSM
into the tumor tissues of mice [71] (Figure 4b). PSM is
also named cold plasma-activated medium (PAM). Most
studies utilized the rst method. Over past 3 years, the
second approach is gradually becoming a hot topic [66,
72-80]. In this review, we just discussed the former one.
INTERACTION BETWEEN CAP AND
BIOLOGICAL SYSTEM
The interaction between CAP and cells in tissue or
cells grown in a petri dish builds the foundation of the anti-
cancer effect of CAP. Such interaction is a combination
of physical and chemical factors as shown schematically
in Figure 5. Ultraviolet, heat, and electromagnetic eld
are physical factors in CAP. Chemical factors include
dozens of reactive species that are generated in the gas
phase of CAP. Among them, oxygen-based species such
as hydroxyl (OH.) [5, 18, 50], singlet oxygen (
1
O
2
) [5, 35,
50], superoxide (O
2
.
-
) [5], hydrogen peroxide (H
2
O
2
) [5],
ozone (O
3
) [5, 60], as well as nitrogen-based species such
as nitric oxide (NO) [5, 18, 36], nitrogen dioxide (NO
2
)
[60], nitrogen trioxide (NO
3
) [60], nitrous oxide (N
2
O)
[60], and dinitrogen tetroxide (N
2
O
4
) [60] have been
observed in CAP. In addition, positive charged ions such
as N
2
+
[18, 37, 50] and electrons [5] are also generated by
CAP.
To date, most in vitro studies have focused on the
anti-cancer effect of CAP on cancer cells cultured in a petri
Figure 3: The research status of the application of CAP on cancer treatment by 2016. a. Publication number. *: by the end
of September. b. The journal types of articles. c. Cancers in articles. d. Plasma devices in articles.