BIOELECTRICITY

Volume 1, Number 4, 2019

ª Mary Ann Liebert, Inc.
DOI: 10.1089/bioe.2019.0028

Recent Advances in Electrochemotherapy

Maja Cemazar, PhD and Gregor Sersa, PhD

Abstract
Electrochemotherapy is gaining recognition as an effective local therapy that uses systemically or in-
tratumorally injected bleomycin or cisplatin with electroporation as a delivery system that brings drugs into the
cells to exert their cytotoxic effects. Preclinical work is still ongoing, testing new drugs, seeking the best
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treatment combination with other treatment modalities, and exploring new sets of pulses for effective tissue
electroporation. The applications of electrochemotherapy are being fully exploited in veterinary oncology,
where electrochemotherapy, because of its simple execution, has a relatively good cost–benefit ratio and is used
in the treatment of cutaneous tumors. In human oncology, electrochemotherapy is fully recognized as a local
therapy for cutaneous tumors and metastases. Its effectiveness is being explored in combination with immu-
nomodulatory drugs. However, the development of electrochemotherapy is directed into the treatment of deep-
seated tumors with a percutaneous approach. Because of the vast number of reports, this review discusses the
articles published in the past 5 years.

Keywords: electrochemotherapy, preclinical studies, veterinary oncology, human oncology

Introduction cytotoxicity, as the primary target of cisplatin in cells is DNA

T he term electrochemotherapy was coined in 1991
by Lluis Mir in a study describing the effect of local
application of noncytotoxic electric pulses on subcutaneous
tumors after intramuscular injection of bleomycin.1 This
treatment, which was named electrochemotherapy, led to a
reduction in size and even the eradication of induced tu-
mors in immunocompetent and nude mice. This work
stimulated other groups, namely, Heller in the United States
and Miklavcic and Sersa in Slovenia, to participate in ex-
perimental nonclinical work.2,3
For good antitumor response of electrochemotherapy, two
prerequisites must be fulfilled: first, the electrical parameters
of applied electric pulses should be selected in such way that
they will cause reversible electroporation of cell membranes,
preferably in all cells in tumors. Second, the drugs used in
combination with electroporation should have limited or no
transport available across the plasma membrane and should
be highly cytotoxic once inside the cells. Bleomycin, as a
hydrophilic antibiotic with endonuclease activity was a per-
fect candidate.4 However, also cisplatin, which was the sec-
ond drug that was introduced into electrochemotherapy,5
have properties that makes it suitable drug for combination
with electroporation. Cisplatin can enter the cells through
copper transporters, but in limited quantities, therefore
electroporation greatly enhances the uptake and consequently
molecule. Cisplatin forms DNA adducts that lead to cell
death, primarily apoptosis.6
Because of the remarkable antitumor effectiveness of this
treatment in preclinical studies, very soon clinical trials were
started in different types of tumors, including head and neck
squamous cell carcinoma and melanoma.7–9 Extensive work,
which was also performed in the frame of two European Union
funded projects, Cliniporator and European Standard Operating
Procedures of Electrochemotherapy, resulted in the publication
of results of clinical trials and the Standard Operating Proce-
dure for Electrochemotherapy.10,11 This publication, together
with the commercially availability of electroporators certified
for clinical use, represented a milestone in the development of
electrochemotherapy, because thereafter each year more clin-
ical centers in Europe started to use electrochemotherapy. In
addition, the number of publications describing the results of
clinical studies has steadily increased.
Another milestone in electrochemotherapy was its inclu-
sion in national and European Union guidelines for the treat-
ment of melanoma metastases, skin tumors, Kaposi’s sarcoma,
and liver metastases of colorectal cancer. The increasing
amounts of results in the field of electrochemotherapy, pre-
clinical and clinical, also made it necessary to standardize the
reporting of the methods and results; thus, two articles dealing
with recommendations on the preclinical as well as clinical use
of electroporation in electrochemotherapy were published. The

Department of Experimental Oncology, Institute of Oncology Ljubljana, Ljubljana, Slovenia.

1
 2 CEMAZAR AND SERSA
aim of these two publications was to provide guidelines for
authors as to what should be included in the publications,
which should ensure the reproducibility of the protocols and
provide the data for a comprehensive meta-analysis of clinical
data.12,13
Recently, a handbook of electroporation and several com-
prehensive review articles were published on electrochemo-
therapy14–16; thus, the aim of this review was to discuss the
recent literature, published in the past 5 years, on electro-
chemotherapy in the three main areas where electroche-
motherapy is studied and applied: in preclinical in vitro and
in vivo studies in laboratory animals, in veterinary oncol-
ogy, and in human clinical oncology. All studies on elec-
trochemotherapy in the past 5 years are included into this
review and grouped according to the specific topics or areas
of research.
Preclinical Studies
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Preclinical studies on electrochemotherapy can be divided
into three main research areas. The first area is the use of the
standard chemotherapeutics bleomycin and cisplatin in new
entities of cancer, not tested before in electrochemotherapy,
and in new combinations with other therapeutic modalities.
The second area is evaluating new antitumor compounds in
combination with electric pulses that are used for electro-
chemotherapy. The third area is the evaluation of new elec-
trical features (i.e., different types of pulses) and the
mathematical/numerical modeling of different effects perti-
nent to electrochemotherapy.
Electrochemotherapy: new tumor entities and new
combinations
In the first area, electrochemotherapy was combined with
another innovative technique, cold plasma, in a mouse mela-
noma model. The combined treatment resulted in a higher
number of cured mice than electrochemotherapy alone. Plas-
ma therapy alone was less effective than electrochemotherapy
with bleomycin; nevertheless, the authors propose that such
treatment can be used as an adjunct to electrochemotherapy or
even as a potential alternative in palliative intent because it can
be applied without anesthesia.17
In principle, owing to the physical properties of electro-
poration, electrochemotherapy with either cisplatin or bleo-
mycin should work on any type of tumor; however, increasing
evidence indicates that tumors respond with different response
rates to electrochemotherapy. This was demonstrated in pre-
clinical, and in veterinary and human clinical trials. The rea-
sons for difference in response are multifaceted and include
physical features pertinent to electrochemotherapy, such as
unavailability of the drug and insufficient coverage with
electric field and biological, pertinent to patient, such as pre-
vious treatments, difference in tumor microenvironment, mu-
tational status, and so on.2,14,18
Thus, electrochemotherapy is still evaluated in different
types of cancer, either in vitro or in vivo, including colon
carcinoma CT26 (in vitro)19; bladder cancer SW780 (in vitro
and in vivo)20; fibroblasts, human umbilical vein endothelial
cells (HUVEC; in vitro) and two squamous cell carcinomas,
CAL-27 and SCC-4 (in vitro)21; ovarian cell lines OvBH-1 and
SKOV-3 (in vitro)22; human neuroblastoma SH-SY5Y cells
(in vitro)23; BRAF-mutated melanoma cells (SK-MEL-28) and
their counterpart without mutation, CHL-1 cells (in vitro)24;
chemoresistant uveal melanoma cells (Mel 270, 92-1, OMM-1,
OMM-2) (in vitro)25; primary cells from metastatic pancreatic
tumors (in vitro)26; human papillomavirus (HPV)-positive head
and neck squamous cell carcinoma 2A3 (in vitro and in vivo)27;
Lewis lung carcinoma and CT 25 colorectal carcinoma
(in vivo)28; conjunctival melanoma CRMM1 and CRMM2 and
normal conjunctival epithelial cells HCjE-Gi (in vitro)29;
radioresistant head and neck squamous cell carcinoma Fa-
DuRR (in vitro and in vivo)30; and small cell lung cancer
H69AR cells (in vitro).31
In general, it was demonstrated that electrochemotherapy with
bleomycin or cisplatin causes immunogenic cell death in colon
carcinoma19,28; that human endothelial cells are more suscepti-
ble to electrochemotherapy than tumor cells,21 supporting the
antivascular effect in vivo; that electrochemotherapy is ef-
fective in ovarian carcinoma cells resistant to standard
therapy22; and that electrochemotherapy is also effective in
neuroblastoma and primary pancreatic tumor cells, which
was not demonstrated before.23,26 Furthermore, it was dem-
onstrated that electrochemotherapy with cisplatin is a prom-
ising therapeutic approach for bladder cancer20 and that it is
more effective in HPV-positive than HPV-negative head and
neck tumors.27 For electrochemotherapy with bleomycin, it
was demonstrated that it is more effective in melanoma cells
that harbor BRAF mutation24 and in melanoma cells that
are chemoresistant.25 In addition, electrochemotherapy with
bleomycin is more effective in radioresistant head and neck
tumors than electrochemotherapy with cisplatin,30 and it is
also effective against small cell lung cancer cells.31
Recently, it was shown that pancreatic tumor cells died from
necroptosis rather than from necrosis or apoptosis after elec-
trochemotherapy with bleomycin, cisplatin, and oxaliplatin.32
In addition, changes in the expression of stemness markers
(Nanog and Oct3/4) in pancreatic tumor cells that survived
electrochemotherapy were demonstrated. This change in
expression could potentially contribute to the initiation and/
or recurrence of pancreatic cancer after treatment and can
represent a new target for the development of new therapeutic
options.33
Many cancer studies are currently exploring immuno-
modulatory therapies in combination with standard cancer
treatments, but only two studies were on the preclinical level
exploring this combination. A synergistic effect was shown
for the combination of electrochemotherapy with cisplatin
and tumor necrosis factor-alpha, which has direct cytotoxic
effects on tumor cells and antiangiogenic effects, in a murine
sarcoma tumor model.34 Another immunomodulatory agent
combined with electrochemotherapy with cisplatin was an
anti-inducible T cell costimulatory (iCOS) agonist antibody.
The restriction of iCOS expression to T cells, which have
already recognized tumor antigen, results in the expansion
and activation of a range of antitumor T cell clones and thus
pronounced antitumor activity. The combination was tested
on CT-26 tumors and resulted in a 50% tumor cure rate with
100% resistance to tumor rechallenge.35
New candidate compounds for electrochemotherapy
The second area of preclinical research was elaborating on
combining electroporation with new candidate compounds to
achieve high and selective cytotoxicity toward tumor cells.
 RECENT ADVANCES IN ELECTROCHEMOTHERAPY
Among many compounds tested, that is, doxorubicin,36 ru-
thenium (III) compound,37 5-fluorouracil,38 phthalocya-
nines,39 photofrin,40 curcumin,41 the trans-platinum analogue
trans-[PtCl2(3-hmpy)2],42 oxaliplatin,43 betulinic acid,44,45
oat b-glucan,46 cyanine IR-775,47 catechin,48 and sodium
decahydrodecaborate,49 only calcium electroporation was
extensively studied.50–58 Calcium electroporation effectively
causes tumor necrosis, leaving the normal tissue less affected,
making it a very promising compound for further clinical
evaluation as it does not belong to the antineoplastic drugs,
thus precautions related to antineoplastic drugs regarding
storage, handling, disposal, and toxicity and mutagenicity are
not an issue for calcium electroporation.
Electrochemotherapy: new electrodes and numerical
modeling
The third large area of preclinical research is dealing with
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different types of electrodes that can be used in vitro, in vivo,
or in humans, that is, grid, single needle, or endoscopic,59–64
with different types (e.g., sine wave pulses, pulsed electro-
magnetic field, and high-frequency short bipolar pulses), and
parameters of electric pulses (amplitude and frequency).65–69
The overall aim of these studies is to provide new electrodes
that will be able to reach/treat the tumors that are not ac-
cessible with the current electrodes that are on the market and
to provide electrodes that will not induce pain, muscle con-
traction, and/or skin burns, that is, undesired side effects that
are commonly reported in clinical studies on electro-
chemotherapy. Another approach to foster and push forward
electrochemotherapy is advanced numerical modeling of
treatment, either in vitro or in vivo, to elucidate parameters
that can influence the treatment outcome or to provide fea-
sibility studies for future perspective electrochemotherapy
treatment of tumors.70–78
New developments
In addition to the abovementioned areas of research, the
Miklavcic group showed that magnetic resonance electric im-
pedance tomography can be used for determining the electric
field distribution during electrochemotherapy, which has im-
portant implications. Namely, this could enable corrective in-
tervention during the procedure to ensure better electric field
coverage of tumors and consequently better treatment out-
come.79 Furthermore, Golzio and colleagues developed a
fluorescence imaging technique for the electrochemotherapy
treatment of micrometastases in the peritoneal cavity during
open surgery, which could represent an important tool for the
treatment of inoperable metastases.80 Our group developed a
new, highly sensitive method for the identification and quan-
tification of bleomycin in tumors and serum. This method,
liquid chromatography coupled with high-resolution mass
spectrometry, enables the determination of bleomycin phar-
macology in different groups of patients, which will in turn lead
to adjustment (personalization) of the drug dose applied to a
specific patient, that is, elderly individuals, young individuals,
and individuals with impaired kidney function, and so on.81
Studies in Veterinary Oncology
Electrochemotherapy is gaining much attention in the
veterinary field. The main reason for this is that electro-
3
chemotherapy is a very effective treatment, for example, in
mast cell tumors in dogs, which are the most common skin
tumors in dogs. The complete response rate of mast cell tu-
mors after electrochemotherapy (70%) is comparable with or
even higher than that of surgery (50%).82 Furthermore, in
many cases, electrochemotherapy is less invasive than sur-
gery, causes fewer side effects than radiotherapy, and can be
repeated if needed, without the development of chemoresis-
tance. It is easy to perform and is relatively inexpensive,
especially in comparison with radiotherapy.83 The popularity
of electrochemotherapy among veterinarians is also demon-
strated by high attendance at the ‘‘Veterinary workshop on
electroporation-based treatments.’’84 In 2018, 21 participants
from Belgium, Brazil, Croatia, Denmark, Germany, Italy,
Romania, Slovenia, and the United Kingdom attended the
workshop, which also includes practical work on the electro-
chemotherapy treatment of dogs, cats, and exotic pets.
Electrochemotherapy is also being used in the treatment of
exotic pets, such as ferrets, cockatiels, turtles, rats, and hedge-
hogs.85–91 Especially in turtles, where fibropapillomatosis is an
important cause of morbidity and mortality of sea turtles, elec-
trochemotherapy is a very promising treatment option. Com-
plete responses were obtained after electrochemotherapy with
intralesional bleomycin in two treated turtles presenting fi-
bropapillomatosis and one with squamous cell carcinoma, with
no healing complications and no recurrence up to 1 year.85,91 In
cats, because of the severe toxicity of cisplatin, electro-
chemotherapy with bleomycin is being used, mainly for the
treatment of squamous cell carcinoma of the head.92,93 Elec-
trochemotherapy was well tolerated, with no evident local or
systemic side effects. In both studies, the complete response rate
was *80%. Therefore, electrochemotherapy should be con-
sidered as an alternative treatment option to other ablative lo-
coregional therapies, especially in cases where tumors are
located in sensitive regions of the body and other therapies are
not accepted by owners because of their invasiveness, mutila-
tion, or high cost.92,93
The main role of electrochemotherapy in veterinary on-
cology is in the treatment of various tumors in dogs. The
owners of dogs are increasingly prone to treat their pets, and
electrochemotherapy is a viable option for these cases, es-
pecially skin tumors. Electrochemotherapy is becoming a
standard therapy for mast cell tumors and an adjuvant treat-
ment to surgery for soft tissue sarcomas. The results of
electrochemotherapy treatment of two large cohorts of dogs
with soft tissue sarcoma were recently published. In a study
by Torrigiani et al.,94 52 dogs with 54 soft tissue sarcomas
were treated either with electrochemotherapy with bleomy-
cin alone, during open surgery, or with electrochemotherapy
as an adjuvant to surgery. The recurrence rate in groups that
were treated with electrochemotherapy during or after sur-
gery was similar (23% and 25%).94 In another study, in ad-
dition to an intravenous administration of bleomycin, the
tumor bed was also infiltrated with cisplatin. This combina-
tion was very effective, 26 of 30 dogs were without evidence
of recurrence, and the median estimated disease-free interval
was 857 days.95 As mentioned, electrochemotherapy can be
used as a treatment of choice for smaller mast cell tumors
(median 1 cm), or in cases of larger tumors, it can be com-
bined with surgery, especially in locations (extremities and
the head) where surgery with curative intent would involve
functional mutilation.96
 4 CEMAZAR AND SERSA

In mast cell tumors, electrochemotherapy was also com-
bined with immune gene therapy with interleukin-12. The main
aim of the study was, in addition to treatment response, the
evaluation of the safety of the gene therapy, thus horizontal
gene transfer to bacteria isolated from the dogs’ skin and the
presence of plasmid on the site of injection. After 1 week, the
plasmid was not detected at the site of injection, and no hori-
zontal gene transfer was obtained. The efficacy of the therapy
was pronounced: 13 of 18 (72% dogs) achieved complete re-
sponse, 2 of 18 (11%) achieved partial response, and 1 (6%)
achieved stable disease at the end of the observation period,
which was 40 months. Again, the treatment was more effective
on smaller tumors (<2 cm). Compared with the literature data
for recurrence rate after surgery or electrochemotherapy alone,
it could be concluded that electrochemotherapy combined with
interleukin-12 reduced the recurrence rate and prevented the
development of distant metastases.82,97 Histologically, 8 weeks
after treatment these tumors were infiltrated with macrophages,
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and one as an adjuvant to surgery. The objective response rate
was higher than for melanoma, that is, 91% (10/11), with a
median survival time of 3.5 months for dogs that responded
partially (2/11) and 28 months for dogs that were killed
owing to tumor-unrelated causes.100 A novel approach for the
treatment of oral tumors of different histologies was proposed
by Maglietti et al.,18 who performed the combined systemic
and local administration of bleomycin. Sixty days after the
treatment, the complete response rate to combined systemic
and local bleomycin and electrochemotherapy was 54%,
whereas in the control group receiving electrochemotherapy
with systemic bleomycin only, partial responses were ob-
tained in 19% dogs.18 The same group also proposed a single
electrode approach for the treatment of tumors in the nasal
cavity in dogs, which are difficult to reach using standard
electrodes. The new approach was compared with standard
treatment using surgery with adjuvant chemotherapy. After 1
year, 60% dogs that were treated with electrochemotherapy

and a significant reduction in microvessels was observed.98
This treatment combination was also evaluated in oral malig-
nant melanoma, one of the most common tumors in the oral
cavity of elderly dogs, with a 2-month survival time if left
untreated.99 Nine dogs were included, and 1 month after the
treatment, the objective response rate was 67% and the median
survival time was 6 months. Although the results were not as
good as those in other tumor types and other locations, elec-
trochemotherapy with interleukin-12 gene therapy may be
beneficial when other modalities are not acceptable because of
invasiveness or cost.
Another common tumor in the oral cavity of dogs is non-
tonsillar squamous cell carcinoma. Eleven dogs were treated
with electrochemotherapy with intravenous bleomycin alone
were still alive, whereas only 10% were alive in the surgical
group, demonstrating that such an approach can be safely
applied to the treatment of nasal duct tumors.101
Yet another application of electrochemotherapy in veteri-
nary medicine is the treatment of sarcoids in horses. Although
benign, sarcoids recur very quickly after surgery and are
usually located in sensitive locations, such as the eyelids.
Cisplatin in sesame oil is commonly used to treat such tumors;
therefore, electrochemotherapy was also explored. Recently, a
new study on 31 horses and 1 donkey was published, con-
firming the excellent results obtained previously, with a 100%
complete response rate.102,103 Electrochemotherapy with cis-
platin can be used alone, when tumors are small, or it can be
used together with debulking surgery in cases of larger tumors.

FIG. 1. A typical response of melanoma skin metastases to electrochemotherapy treatment. The patient had a primary
melanoma excised. Two years after the excision, metastases were present, and electrochemotherapy with intravenous
bleomycin was performed using needle row electrodes. Ten nodules were treated with electrochemotherapy, and one was
excised for the determination of BRAF mutation. (A) Tumor nodules before electrochemotherapy; (B) immediately after the
excision of one tumor nodule and electrochemotherapy of all other tumors; (C) 14 days after treatment; (D) 3 months after
treatment; (E) 9 months after treatment; (F) 17 months after treatment.
 RECENT ADVANCES IN ELECTROCHEMOTHERAPY
Calcium electroporation was also evaluated recently for the
treatment of sarcoids. Tumors were treated with calcium
electroporation and then surgically excised at different time
points to determine the extent of necrosis, which was present in
>50% of the tumor area in 9 of 13 treated sarcoids.104
Clinical Studies in Human Oncology
The most vivid field of electrochemotherapy is clinical.
The results from almost 90 clinical trials or case reports were
published in the past 5 years. A typical response of skin
metastases to electrochemotherapy is the formation of a crust
that eventually falls off, leaving a very good cosmetic effect
(Fig. 1). Electrochemotherapy is becoming an established,
well-recognized treatment option in Europe. Recently,
electrochemotherapy was listed as a treatment option for
localized lesions of Kaposi’s sarcoma in European guide-
lines.105 Many of the publications are review articles and
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meta-analyses, which will further help to position electro-
chemotherapy among other local ablative therapies.106–108
Different aims were ascribed in these meta-analyses, including
the determination of predictive factors in nonmelanoma skin
cancer, cost-effectiveness, treatment efficacy, pain control,
quality of life, and the role in the palliative management of
cutaneous metastases.14,15,109–111
In multivariate analysis of predictive factors regarding
clinical pathological and instrumental predictors, only the
tumor’s site and appearance reached statistical significance,
showing that this treatment can still be refined to achieve better
treatment outcomes.111 All these excellent reviews present
valuable data about the usefulness, effectiveness, safety, and
patient compliance of electrochemotherapy. However, many
clinical studies could not be included in systematic reviews
because of the lack of information (data) in the published ar-
ticles. To avoid this, recommendations for improving the
quality of reporting clinical electrochemotherapy study results
FIG. 2. Schematic representation of the timeline of the electrochemotherapy procedure and the response of tumor cells.
5
based on qualitative systematic review were published and
should be implemented by the authors.12 Here, we will focus on
new developments in electrochemotherapy that will lead to
further development and dissemination of the therapy.
First, for subcutaneous as well as for deep-seated tumors, it
is very important that the electric field coverage is adequate
because this is a prerequisite for the permeabilization of cell
membranes and the entry of the drug. Therefore, specifically
for deep-seated tumors, treatment planning and radiological
monitoring during the procedure to determine complete
coverage of the treated tumor are of utmost importance.112,113
Namely, monitoring of changes during and after electro-
chemotherapy is important for evaluation of the progress of
the procedure that is based on pretreatment plan and its re-
sults. It is important that tumor is not over treated as this will
cause irreversible electroporation, or under treated (not en-
ough cells exposed to electric field above threshold value) as
this restricts the access of drugs into the cells and conse-
quently their action on target, that is, DNA.112,113 For tumors
in the head and neck region, the treatment plan can be cou-
pled to the navigation system for accurate positioning of
single long-needle electrodes into and around tumors.114
Furthermore, the new, sensitive method for the determi-
nation of bleomycin in serum and tumors enables a redefi-
nition of the optimal therapeutic window for the application
of electric pulses after bleomycin intravenous injection.
Based on the pharmacokinetic data obtained in elderly pa-
tients (>65 years), the therapeutic window for bleomycin can
be extended to up to 40 min.115 Furthermore, because of
prolonged therapeutic concentrations in the serum, a lower
dose of bleomycin can also be used, especially in elderly
patients. Preliminary results demonstrated comparable ef-
fectiveness in nonmelanoma skin metastases in head and
neck cancer,116 which was also confirmed for other types of
skin tumors, that is, melanoma, breast cancer, and Kaposi’s
sarcoma.117
 6 CEMAZAR AND SERSA
In addition to the standard chemotherapeutic drugs used in
clinical electrochemotherapy, bleomycin and cisplatin, cal-
cium chloride was evaluated in a clinical setting based on
very promising preclinical results. A double-blinded phase II
study was performed comparing the effect of calcium elec-
troporation with electrochemotherapy. The treatment was
performed on 47 cutaneous metastases from breast cancer
and melanoma. The objective response rate 6 months after the
treatment was 13 of 18 (72%) for calcium electroporation and
16 of 19 (84%) for electrochemotherapy, without a signifi-
cant difference between the rates. Thus, calcium electro-
poration is feasible and effective in patients with cutaneous
metastases.118 This finding is especially important in clinical
environments, where the acquisition, handling, and waste
disposal of antineoplastic drugs can present a challenge.
Recently, an updated, simplified version of standard op-
eration procedures was published based on vast experience
that was gained after the publication of the first standard
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operating procedures in 2006.11,119 The updated standard
operating procedures are based on data from the treatment of
many different histologies and increasing sizes (volume) of
tumors that was originally thought to not be possible. The
specialists from different fields, that is, oncology, general
surgery, neck surgery, plastic surgery, and dermatology,
prepared the recommendations for indications for electro-
chemotherapy, pretreatment information and evaluation,
treatment choices and follow-up.119 These new standard
operating procedures should enable further implementation
of electrochemotherapy into national and European guide-
lines for the treatment of different types of tumors.
Future Perspectives and Conclusion
The main difference between electrochemotherapy and
other local ablative therapies is that electrochemotherapy
combines two modalities, chemotherapy and the application
of electric pulses (Fig. 2). Thus, the tumor cells are dying not
directly owing to the application of physical energy, such as
in the case of other local ablative therapies, such as irre-
versible electroporation, radiofrequency ablation, microwave
ablation, and cryoablation, but because of the action of che-
motherapeutic drugs. This further means that the cells are
dying in a more controllable way, either by apoptosis or other
programmed types of cell death, resulting in a slower
shrinkage of the tumor without the development of massive
necrosis, which represents a major burden for patients and
possesses a risk of further complications, such as infections.
At present, at the preclinical level, much effort is devoted to
evaluation of new combination of electrochemotherapy with
other types of therapy and in new cancer entities and to de-
velopment of so-called ‘‘painless electroporation.’’ In vet-
erinary field, electrochemotherapy is gaining recognition in
small animal practice, and is increasingly used for treatment
of exotic companion animals, such as turtles, ferrets, and
birds. At the clinical level, the most important advancement
is the publication of results of bigger cohorts of patients with
specific tumors, treated with electrochemotherapy. This is
possible because of the clinical data repositories of several
groups of institutions practicing electrochemotherapy. Future
studies in preclinical and clinical electrochemotherapy will
focus on deciphering the molecular mechanism that would
identify tumors/patients that will benefit the most from
electrochemotherapy and combinations either with different
systemic treatments (immune, small molecule, or hormonal
treatments) or with other local ablative therapies. Another
very important field that still needs to be developed is per-
cutaneous treatment by electrochemotherapy. First reports on
the treatment of liver metastases from pancreatic cancer and
for spine metastases have already been presented with en-
couraging results.120–122
Author Contributions
M.C. designed the concept of the article and drafted it. G.S.
critically revised it. Both coauthors have reviewed and ap-
proved the manuscript because submission.
Disclaimer
The funder had no role in the design, decision to publish, or
preparation of the article. Language editing services for this
article were provided by American Journal Experts. The ar-
ticle has been submitted solely to this journal and is not
published, in press, or submitted elsewhere.
Author Disclosure Statement
No competing financial interests exist.
Funding Information
This study was supported by the Slovenian Research
Agency (ARRS), grant No. P3-0003.
References
1. Mir LM, Orlowski S, Belehradek J, et al. Electro-
chemotherapy potentiation of antitumour effect of bleomy-
cin by local electric pulses. Eur J Cancer 1991;27:68–72.
2. Sersa G, Cemazar M, Miklavcic D, et al. Electro-
chemotherapy—variable antitumor effect on different tumor
models. Bioelectrochem Bioenerg 1994;35:23–27.
3. Heller R, Jaroszeski M, Messina J, et al. Treatment of
B16 mouse melanoma with the combination of electro-
permeabilization and chemotherapy. Bioelectrochem
Bioenerg 1995;36:83–87.
4. Mir LM, Tounekti O, Orlowski S. Bleomycin: Revival of
an old drug. Gen Pharmacol 1996;27:745–748.
5. Sersa G, Cemazar M, Miklavcic D. Antitumor effectiveness of
electrochemotherapy with cis-diamminedichloroplatinum(II)
in mice. Cancer Res 1995;55:3450–3455.
6. Kilari D, Guancial E, Kim ES. Role of copper transporters in
platinum resistance. World J Clin Oncol 2016;7:106–113.
7. Belehradek J, Orlowski S, Poddevin B, et al. Electro-
chemotherapy of spontaneous mammary tumours in mice.
Eur J Cancer 1991;27:73–76.
8. Rudolf Z, Stabuc B, Cemazar M, et al. Electrochemotherapy
with bleomycin. The first clinical experience in malignant
melanoma patients. Radiol Oncol 1995;29:229–235.
9. Heller R, Jaroszeski MJ, Glass LF, et al. Phase I/II trial for
the treatment of cutaneous and subcutaneous tumors using
electrochemotherapy. Cancer 1996;77:964–971.
10. Marty M, Sersa G, Garbay JR, et al. Electrochemotherapy—
an easy, highly effective and safe treatment of cutaneous
and subcutaneous metastases: Results of ESOPE (European
Standard Operating Procedures of Electrochemotherapy)
study. Eur J Cancer Suppl 2006;4:3–13.
 RECENT ADVANCES IN ELECTROCHEMOTHERAPY
11. Mir LM, Gehl J, Sersa G, et al. Standard operating pro-
cedures of the electrochemotherapy: Instructions for the
use of bleomycin or cisplatin administered either sys-
temically or locally and electric pulses delivered by the
Cliniporator (TM) by means of invasive or non-invasive
electrodes. Eur J Cancer Suppl 2006;4:14–25.
12. Campana LG, Clover AJP, Valpione S, et al. Re-
commendations for improving the quality of reporting
clinical electrochemotherapy studies based on qualitative
systematic review. Radiol Oncol 2016;50:1–13.
13. Cemazar M, Sersa G, Frey W, et al. Recommendations
and requirements for reporting on applications of electric
pulse delivery for electroporation of biological samples.
Bioelectrochemistry 2018;122:69–76.
14. Campana LG, Miklavčič D, Bertino G, et al. Electro-
chemotherapy of superficial tumors—current status: Basic
principles, operating procedures, shared indications, and
emerging applications. Semin Oncol 2019;46:173–191.
15. Campana LG, Edhemovic I, Soden D, et al.
Downloaded by East Carolina University from www.liebertpub.com at 12/07/19. For personal use only.
Electrochemotherapy—emerging applications technical
advances, new indications, combined approaches, and
multi-institutional collaboration. Eur J Surg Oncol 2019;
45:92–102.
16. Miklavcic D. Handbook of Electroporation. 1st edition.
New York, NY: Springer, 2017.
17. Daeschlein G, Scholz S, Lutze S, et al. Comparison be-
tween cold plasma, electrochemotherapy and combined
therapy in a melanoma mouse model. Exp Dermatol 2013;
22:582–586.
18. Maglietti F, Tellado M, Olaiz N, et al. Combined local and
systemic bleomycin administration in electrochemotherapy
to reduce the number of treatment sessions. Radiol Oncol
2016;50:58–63.
19. Calvet CY, Famin D, André FM, et al. Electro-
chemotherapy with bleomycin induces hallmarks of im-
munogenic cell death in murine colon cancer cells.
Oncoimmunology 2014;3:e28131.
20. Vásquez JL, Ibsen P, Lindberg H, et al. In vitro and
in vivo experiments on electrochemotherapy for bladder
cancer. J Urol 2015;193:1009–1015.
21. Landström F, Ivarsson M, Von Sydow AK, et al.
Electrochemotherapy—evidence for cell-type selectivity
in vitro. Anticancer Res 2015;35:5813–5820.
22. Saczko J, Pilat J, Choromanska A, et al. The effectiveness
of chemotherapy and electrochemotherapy on ovarian cell
lines in vitro. Neoplasma 2016;63:450–455.
23. Esmekaya MA, Kayhan H, Coskun A, et al. Effects of
cisplatin electrochemotherapy on human neuroblastoma
cells. J Membr Biol 2016;249:601–610.
24. Dolinsek T, Prosen L, Cemazar M, et al. Electro-
chemotherapy with bleomycin is effective in BRAF mu-
tated melanoma cells and interacts with BRAF inhibitors.
Radiol Oncol 2016;50:274–279.
25. Fiorentzis M, Kalirai H, Katopodis P, et al. Electro-
chemotherapy with bleomycin and cisplatin enhances
cytotoxicity in primary and metastatic uveal melanoma
cell lines in vitro. Neoplasma 2018;65:210–215.
26. Michel O, Kulbacka J, Saczko J, et al. Electroporation
with cisplatin against metastatic pancreatic cancer:
In vitro study on human primary cell culture. Biomed Res
Int 2018;2018:1–12.
27. Prevc A, Niksic Zakelj M, Kranjc S, et al. Electro-
chemotherapy with cisplatin or bleomycin in head and
neck squamous cell carcinoma: Improved effectiveness of
7
cisplatin in HPV-positive tumors. Bioelectrochemistry
2018;123:248–254.
28. Tremble LF, O’Brien MA, Soden DM, et al. Electro-
chemotherapy with cisplatin increases survival and in-
duces immunogenic responses in murine models of lung
cancer and colorectal cancer. Cancer Lett 2019;442:475–
482.
29. Fiorentzis M, Katopodis P, Kalirai H, et al. Conjunctival
melanoma and electrochemotherapy: Preliminary results
using 2D and 3D cell culture models in vitro. Acta Oph-
thalmol 2019;97:e632–e640.
30. Zakelj MN, Prevc A, Kranjc S, et al. Electrochemotherapy
of radioresistant head and neck squamous cell carcinoma
cells and tumor xenografts. Oncol Rep 2019;41:1658–
1668.
31. Dra˛g-Zalesińska M, Saczko J, Choromańska A, et al. Cis-
platin and vinorelbine -mediated electrochemotherapeutic
approach against multidrug resistant small cell lung cancer
(H69AR) in vitro. Anticancer Res 2019;39:3711–3718.
32. Fernandes P, O’Donovan TR, McKenna SL, et al. Elec-
trochemotherapy causes caspase-independent necrotic-
like death in pancreatic cancer cells. Cancers (Basel)
2019;11:1177.
33. Ali MS, Gill KS, Saglio G, et al. Expressional changes in
stemness markers post electrochemotherapy in pancreatic
cancer cells. Bioelectrochemistry 2018;122:84–92.
34. Cemazar M, Todorovic V, Scancar J, et al. Adjuvant TNF-
a therapy to electrochemotherapy with intravenous cis-
platin in murine sarcoma exerts synergistic antitumor
effectiveness. Radiol Oncol 2015;49:32–40.
35. Tremble LF, O’Brien MA, Forde PF, et al. ICOS activa-
tion in combination with electrochemotherapy generates
effective anti-cancer immunological responses in murine
models of primary, secondary and metastatic disease.
Cancer Lett 2018;420:109–115.
36. Kulbacka J, Daczewska M, Dubińska-Magiera M, et al.
Doxorubicin delivery enhanced by electroporation to
gastrointestinal adenocarcinoma cells with P-gp over-
expression. Bioelectrochemistry 2014;100:96–104.
37. Hudej R, Miklavcic D, Cemazar M, et al. Modulation of
activity of known cytotoxic ruthenium(III) compound
(KP418) with hampered transmembrane transport in
electrochemotherapy in vitro and in vivo. J Membr Biol
2014;247:1239–1251.
38. Saczko J, Kamińska I, Kotulska M, et al. Combination of
therapy with 5-fluorouracil and cisplatin with electro-
poration in human ovarian carcinoma model in vitro.
Biomed Pharmacother 2014;68:573–580.
39. Zielichowska A, Saczko J, Garbiec A, et al. The pho-
todynamic effect of far-red range phthalocyanines (AlPc
and Pc green) supported by electropermeabilization in
human gastric adenocarcinoma cells of sensitive and
resistant type. Biomed Pharmacother 2015;69:145–
152.
40. Choromanska A, Kulbacka J, Rembialkowska N, et al.
Effects of electrophotodynamic therapy in vitro on human
melanoma cells—melanotic (MeWo) and amelanotic
(C32). Melanoma Res 2015;25:210–224.
41. Mittal L, Raman V, Camarillo IG, et al. Ultra-microsecond
pulsed curcumin for effective treatment of triple negative
breast cancers. Biochem Biophys Res Commun 2017;491:
1015–1020.
42. Kranjc S, Cemazar M, Sersa G, et al. In vitro and in vivo
evaluation of electrochemotherapy with trans-platinum
 8 CEMAZAR AND SERSA
analogue trans-[PtCl2(3-Hmpy)2]. Radiol Oncol 2017;51:
295–306.
43. Ursic K, Kos S, Kamensek U, et al. Comparable effec-
tiveness and immunomodulatory actions of oxaliplatin and
cisplatin in electrochemotherapy of murine melanoma.
Bioelectrochemistry 2018;119:161–171.
44. Hartmann P, Butt E, Fehér Á, et al. Electroporation-
enhanced transdermal diclofenac sodium delivery into the
knee joint in a rat model of acute arthritis. Drug Des Devel
Ther 2018;12:1917–1930.
45. Ma˛czyńska J, Choromańska A, Kutkowska J, et al. Effect
of electrochemotherapy with betulinic acid or cisplatin on
regulation of heat shock proteins in metastatic human
carcinoma cells in vitro. Oncol Rep 2019;41:3444–3454.
46. Choromanska A, Kulbacka J, Harasym J, et al. Anticancer
activity of oat b-glucan in combination with electropora-
tion on uman cancer cells. Acta Pol Pharm 2017;74:616–
623.
47. We_zgowiec J, Kulbacka J, Saczko J, et al. Biological ef-
Downloaded by East Carolina University from www.liebertpub.com at 12/07/19. For personal use only.
fects in photodynamic treatment combined with electro-
permeabilization in wild and drug resistant breast cancer
cells. Bioelectrochemistry 2018;123:9–18.
48. Michel O, Przystupski D, Saczko J, et al. The favourable
effect of catechin in electrochemotherapy in human pan-
creatic cancer cells. Acta Biochim Pol 2018;65:173–184.
49. Garabalino MA, Olaiz N, Portu A, et al. Electroporation
optimizes the uptake of boron-10 by tumor for boron
neutron capture therapy (BNCT) mediated by GB-10: A
boron biodistribution study in the hamster cheek pouch
oral cancer model. Radiat Environ Biophys 2019;58:455–
467.
50. Staresinic B, Jesenko T, Kamensek U, et al. Effect of
calcium electroporation on tumour vasculature. Sci Rep
2018;8:9412.
51. Frandsen SK, Gissel H, Hojman P, et al. Calcium elec-
troporation in three cell lines: A comparison of bleomycin
and calcium, calcium compounds, and pulsing conditions.
Biochim Biophys Acta Gen Subj 2014;1840:1204–1208.
52. Frandsen SK, Gibot L, Madi M, et al. Calcium electro-
poration: Evidence for differential effects in normal and
malignant cell lines, evaluated in a 3D spheroid model.
PLoS One 2015;10:e0144028.
53. Falk H, Forde PF, Bay ML, et al. Calcium electroporation
induces tumor eradication, long-lasting immunity and
cytokine responses in the CT26 colon cancer mouse
model. Oncoimmunology 2017;6:e1301332.
54. Romeo S, Sannino A, Scarfı̀ MR, et al. ESOPE-equivalent
pulsing protocols for calcium electroporation: An in vitro
optimization study on 2 cancer cell models. Technol
Cancer Res Treat 2018;17:153303381878807.
55. Hoejholt KL, Mužić T, Jensen SD, et al. Calcium elec-
troporation and electrochemotherapy for cancer treatment:
Importance of cell membrane composition investigated by
lipidomics, calorimetry and in vitro efficacy. Sci Rep
2019;9:4758.
56. Frandsen SK, Gehl J. Effect of calcium electroporation in
combination with metformin in vivo and correlation be-
tween viability and intracellular ATP level after calcium
electroporation in vitro. PLoS One 2017;12:e0181839.
57. Frandsen SK, Krüger MB, Mangalanathan UM, et al.
Normal and malignant cells exhibit differential responses
to calcium electroporation. Cancer Res 2017;77:4389–
4401.
58. Hansen EL, Sozer EB, Romeo S, et al. Dose-dependent
ATP depletion and cancer cell death following calcium
electroporation, relative effect of calcium concentration
and electric field strength. PLoS One 2015;10:e0122973.
59. Pedersoli F, Ritter A, Zimmermann M, et al. Single-needle
electroporation and interstitial electrochemotherapy: In vivo
safety and efficacy evaluation of a new system. Eur Radiol
2019;29:6300–6308.
60. Vera-Tizatl AL, Vera-Tizatl CE, Vera-Hernández A, et al.
Computational feasibility analysis of electrochemotherapy
with novel needle-electrode arrays for the treatment of
invasive breast ductal carcinoma. Technol Cancer Res
Treat 2018;17:153303381879493.
61. Campana LG, Dughiero F, Forzan M, et al. A prototype of
a flexible grid electrode to treat widespread superficial
tumors by means of electrochemotherapy. Radiol Oncol
2016;50:49–57.
62. Forde P, Sadadcharam M, Bourke M, et al. Preclinical
evaluation of an endoscopic electroporation system. En-
doscopy 2016;48:477–483.
63. Ongaro A, Campana LG, De Mattei M, et al. Effect of
electrode distance in grid electrode: Numerical models
and in vitro tests. Technol Cancer Res Treat 2018;17:
153303381876449.
64. Ongaro A, Campana LG, De Mattei M, et al. Evaluation
of the electroporation efficiency of a grid electrode for
electrochemotherapy: From numerical model to in vitro
tests. Technol Cancer Res Treat 2016;15:296–307.
65. Shankayi Z, Firoozabadi SM, Hassan ZS. Optimization of
electric pulse amplitude and frequency in vitro for low
voltage and high frequency electrochemotherapy. J
Membr Biol 2014;247:147–154.
66. Spugnini EP, Melillo A, Quagliuolo L, et al. Definition of
novel electrochemotherapy parameters and validation of
their in vitro and in vivo effectiveness. J Cell Physiol 2014;
221:1177–1181.
67. Kranjc S, Kranjc M, Scancar J, et al. Electrochemotherapy
by pulsed electromagnetic field treatment (PEMF) in
mouse melanoma B16F10 in vivo. Radiol Oncol 2016;50:
39–48.
68. Scuderi M, Rebersek M, Miklavcic D, et al. The use of
high-frequency short bipolar pulses in cisplatin electro-
chemotherapy in vitro. Radiol Oncol 2019;53:194–205.
69. Garcia-Sanchez T, Mercadal B, Polrot M, et al. Successful
tumor electrochemotherapy using sine waves. IEEE Trans
Biomed Eng 2019; [Epub ahead of print]; DOI: 10.1109/
TBME.2019.2928645.
70. Mahna A, Firoozabadi SMP, Shankayi Z. The effect of
ELF magnetic field on tumor growth after electro-
chemotherapy. J Membr Biol 2014;247:9–15.
71. Dermol J, Miklavčič D. Mathematical models describing
chinese hamster ovary cell death due to electroporation
in vitro. J Membr Biol 2015;248:865–881.
72. Forjanič T, Miklavčič D. Mathematical model of tumor
volume dynamics in mice treated with electro-
chemotherapy. Med Biol Eng Comput 2017;55:1085–
1096.
73. Cindrič H, Kos B, Tedesco G, et al. Electrochemotherapy
of spinal metastases using transpedicular approach—a
numerical feasibility study. Technol Cancer Res Treat
2018;17:153303461877025.
74. Bhonsle S, Lorenzo MF, Safaai-Jazi A, et al. Characteriza-
tion of nonlinearity and dispersion in tissue impedance
 RECENT ADVANCES IN ELECTROCHEMOTHERAPY
during high-frequency electroporation. IEEE Trans Biomed
Eng 2018;65:2190–2201.
75. Goldberg E, Suárez C, Alfonso M, et al. Cell membrane
electroporation modeling: A multiphysics approach.
Bioelectrochemistry 2018;124:28–39.
76. Dermol-Černe J, Vidmar J, Ščančar J, et al. Connecting the
in vitro and in vivo experiments in electrochemotherapy—
a feasibility study modeling cisplatin transport in mouse
melanoma using the dual-porosity model. J Control Re-
lease 2018;286:33–45.
77. Berkenbrock JA, Machado RG, Suzuki DOH. Electro-
chemotherapy effectiveness loss due to electric field in-
dentation between needle electrodes: A numerical study. J
Healthc Eng 2018;2018:1–8.
78. Pintar M, Langus J, Edhemović I, et al. Time-dependent finite
element analysis of in vivo electrochemotherapy treatment.
Technol Cancer Res Treat 2018;17:153303381879051.
79. Kranjc M, Markelc B, Bajd F, et al. In situ monitoring of
electric field distribution in mouse tumor during electro-
Downloaded by East Carolina University from www.liebertpub.com at 12/07/19. For personal use only.
poration. Radiology 2015;274:115–123.
80. Josserand V, Kéramidas M, Lavaud J, et al. Electro-
chemotherapy guided by intraoperative fluorescence im-
aging for the treatment of inoperable peritoneal micro-
metastases. J Control Release 2016;233:81–87.
81. Kosjek T, Krajnc A, Gornik T, et al. Identification and
quantification of bleomycin in serum and tumor tissue by
liquid chromatography coupled to high resolution mass
spectrometry. Talanta 2016;160:164–171.
82. Kodre V, Cemazar M, Pecar J, et al. Electrochemotherapy
compared to surgery for treatment of canine mast cell
tumours. In Vivo 2009;23:55–62.
83. Tozon N, Lampreht Tratar U, Znidar K, et al. Operating
procedures of the electrochemotherapy for treatment of
tumor in dogs and cats. J Vis Exp 2016;16:54760.
84. Tozon N, Milevoj N, Lampreht Tratar U, et al. Veterinary
workshop on electroporation-based treatments. In: 3rd
World Congress on Electroporation and Pulsed Electric
Fields in Biology, Medicine, and Food and Environmental
Technologies, Toulouse, France. 2019, p. 95.
85. Brunner CHM, Dutra G, Silva CB, et al. Electro-
chemotherapy for the treatment of fibropapillomas in
Chelonia mydas. J Zoo Wildl Med 2014;45:213–218.
86. Lanza A, Baldi A, Spugnini EP. Surgery and electro-
chemotherapy for the treatment of cutaneous squamous
cell carcinoma in a yellow-bellied slider (Trachemys
scripta scripta). J Am Vet Med Assoc 2015;246:455–
457.
87. Lanza A, Pettorali M, Baldi A, et al. Surgery and elec-
trochemotherapy treatment of incompletely excised
mammary carcinoma in two male pet rats (Rattus norve-
gicus). J Vet Med Sci 2017;79:623–625.
88. Racnik J, Svara T, Zadravec M, et al. Electro-
chemotherapy with bleomycin of different types of cuta-
neous tumours in a ferret (Mustela putorius furo). Radiol
Oncol 2017;52:98–104.
89. Racnik J, Svara T, Zadravec M, et al. Electro-
chemotherapy with cisplatin for the treatment of a non-
operable cutaneous fibroma in a cockatiel (Nymphicus
hollandicus). N Z Vet J 2019;67:155–158.
90. Spugnini EP, Lanza A, Sebasti S, et al. Electro-
chemotherapy palliation of an oral squamous cell carci-
noma in an African hedgehog (Atelerix albiventris). Vet
Res Forum 2018;9:379–381.
9
91. Donnelly KA, Papich MG, Zirkelbach B, et al. Plasma
bleomycin concentrations during electrochemotherapeutic
treatment of fibropapillomas in green turtles Chelonia
mydas. J Aquat Anim Health 2019;31:186–192.
92. Tozon N, Pavlin D, Sersa G, et al. Electrochemotherapy
with intravenous bleomycin injection: An observational
study in superficial squamous cell carcinoma in cats. J
Feline Med Surg 2014;16:291–299.
93. Spugnini EP, Pizzuto M, Filipponi M, et al. Electropora-
tion enhances bleomycin efficacy in cats with periocular
carcinoma and advanced squamous cell carcinoma of the
head. J Vet Intern Med 2015;29:1368–1375.
94. Torrigiani F, Pierini A, Lowe R, et al. Soft tissue sarcoma
in dogs: A treatment review and a novel approach using
electrochemotherapy in a case series. Vet Comp Oncol
2019;17:234–241.
95. Spugnini EP, Vincenzi B, Amadio B, et al. Adjuvant
electrochemotherapy with bleomycin and cisplatin com-
bination for canine soft tissue sarcomas: A study of 30
cases. Open Vet J 2019;9:88–93.
96. Lowe R, Gavazza A, Impellizeri JA, et al. The treatment
of canine mast cell tumours with electrochemotherapy
with or without surgical excision. Vet Comp Oncol 2017;
15:775–784.
97. Cemazar M, Ambrozic Avgustin J, Pavlin D, et al. Effi-
cacy and safety of electrochemotherapy combined with
peritumoral IL-12 gene electrotransfer of canine mast cell
tumours. Vet Comp Oncol 2017;15:641–654.
98. Salvadori C, Svara T, Rocchigiani G, et al. Effects of
electrochemotherapy with cisplatin and peritumoral IL-12
gene electrotransfer on canine mast cell tumors: A histo-
pathologic and immunohistochemical study. Radiol Oncol
2017;51:286–294.
99. Harvey HJ, MacEwen EG, Braun D, et al. Prognostic
criteria for dogs with oral melanoma. J Am Vet Med
Assoc 1981;178:580–582.
100. Simčič P, Lowe R, Granziera V, et al. Electro-
chemotherapy in treatment of canine oral non-tonsillar
squamous cell carcinoma. a case series report. Vet Comp
Oncol 2019;[Epub ahead of print]; DOI: 10.1111/
vco.12530.
101. Maglietti F, Tellado M, Olaiz N, et al. Minimally invasive
electrochemotherapy procedure for treating nasal duct
tumors in dogs using a single needle electrode. Radiol
Oncol 2017;51:422–430.
102. Tozon N, Kramaric P, Kos Kadunc V, et al. Electro-
chemotherapy as a single treatment or adjuvant treatment
to surgery of cutaneous sarcoid tumours in horses: A 31-
case retrospective study. Vet Rec 2016;179:627.
103. Tamzali Y, Borde L, Rols MP, et al. Successful treatment
of equine sarcoids with cisplatin electrochemotherapy: A
retrospective study of 48 cases. Equine Vet J 2012;44:
214–220.
104. Galant L, Delverdier M, Lucas M-N, et al. Calcium
electroporation: The bioelectrochemical treatment of
spontaneous equine skin tumors results in a local necrosis.
Bioelectrochemistry 2019;129:251–258.
105. Lebbe C, Garbe C, Stratigos AJ, et al. Diagnosis and
treatment of Kaposi’s sarcoma: European consensus-
based interdisciplinary guideline (EDF/EADO/EORTC).
Eur J Cancer 2019;114:117–127.
106. Cadossi R, Ronchetti M, Cadossi M. Locally enhanced
chemotherapy by electroporation: Clinical experiences
 10
and perspective of use of electrochemotherapy. Futur
Oncol 2014;10:877–890.
107. Campana LG, Testori A, Mozzillo N, et al. Treatment of
metastatic melanoma with electrochemotherapy. J Surg
Oncol 2014;109:301–307.
108. De Virgilio A, Fusconi M, Greco A, et al. The role of
electrochemotherapy in the treatment of metastatic head
and neck cancer. Tumori 2013;99:634.
109. De Virgilio A, Ralli M, Longo L, et al. Electro-
chemotherapy in head and neck cancer: A review of an
emerging cancer treatment (Review). Oncol Lett 2019;17:
1253–1256.
110. Quaglino P, Matthiessen LW, Curatolo P, et al. Predicting
patients at risk for pain associated with electrochemotherapy.
Acta Oncol 2015;54:298–306.
111. Rotunno R, Paolino G, Cantisani C, et al. Predictive fac-
tors in non-melanoma skin cancers treated with electro-
chemotherapy. G Ital Dermatol Venereol 2018;153:11–18.
112. Marčan M, Pavliha D, Kos B, et al. Web-based tool for
Downloaded by East Carolina University from www.liebertpub.com at 12/07/19. For personal use only.
visualization of electric field distribution in deep-seated
body structures and planning of electroporation-based
treatments. Biomed Eng Online 2015;14:S4.
113. Boc N, Edhemovic I, Kos B, et al. Ultrasonographic
changes in the liver tumors as indicators of adequate tu-
mor coverage with electric field for effective electro-
chemotherapy. Radiol Oncol 2018;52:383–391.
114. Groselj A, Kos B, Cemazar M, et al. Coupling treatment
planning with navigation system: A new technological
approach in treatment of head and neck tumors by elec-
trochemotherapy. Biomed Eng Online 2015;14:S2.
115. Groselj A, Krzan M, Kosjek T, et al. Bleomycin phar-
macokinetics of bolus bleomycin dose in elderly cancer
patients treated with electrochemotherapy. Cancer Che-
mother Pharmacol 2016;77:939–947.
116. Groselj A, Bosnjak M, Strojan P, et al. Efficiency of
electrochemotherapy with reduced bleomycin dose in the
treatment of nonmelanoma head and neck skin cancer:
Preliminary results. Head Neck 2018;40:120–125.
CEMAZAR AND SERSA
117. Rotunno R, Campana LG, Quaglino P, et al. Electro-
chemotherapy of unresectable cutaneous tumours with
reduced dosages of intravenous bleomycin: Analysis of 57
patients from the International Network for Sharing
Practices of Electrochemotherapy registry. J Eur Acad
Dermatol Venereol 2018;32:1147–1154.
118. Falk H, Matthiessen LW, Wooler G, et al. Calcium elec-
troporation for treatment of cutaneous metastases; a ran-
domized double-blinded phase II study, comparing the effect
of calcium electroporation with electrochemotherapy. Acta
Oncol 2018;57:311–319.
119. Gehl J, Sersa G, Matthiessen LW, et al. Updated standard
operating procedures for electrochemotherapy of cutane-
ous tumours and skin metastases. Acta Oncol 2018;57:
874–882.
120. Cornelis FH, Korenbaum C, Ben Ammar M, et al. Mul-
timodal image-guided electrochemotherapy of un-
resectable liver metastasis from renal cell cancer. Diagn
Interv Imaging 2019;100:309–311.
121. Cornelis FH, Ben Ammar M, Nouri-Neuville M, et al.
Percutaneous image-guided electrochemotherapy of spine
metastases: Initial experience. Cardiovasc Intervent
Radiol 2019;42:1806–1809.
122. Djokic M, Badovinac D, Petric M, et al. Treatment of
hepatocellular carcinoma with electrochemotherapy. In:
3rd World Congress on Electroporation and Pulsed Elec-
tric Fields in Biology, Medicine, and Food and Environ-
mental Technologies, Toulouse, France. 2019, p. 143.
Address correspondence to:
Maja Cemazar, PhD
Institute of Oncology Ljubljana
Ljubljana 1000
Slovenia
Email: mcemazar@onko-i.si