Fázis II, randomizált, kettős Vak vizsgálat a bőr metasztázisok kezelésére kálcium elektroporációval
Phase: NA
Conditions: Cutaneous Melanoma
Sponsor: Szeged University

Calcium Electroporation for Treatment of Cutaneous 
Metastases; randomized double blind phase II study  
NCT03628417  
Primary Completion Date: 04/06/2019 
Last update: 03/10/2019 
Last release: 09/24/2019 
 
Introduction 
Background 
Electroporation is a method that can facilitate transport of molecules across the cell membrane and 
into the cell by means of electrical pulses. The method can be used with molecules that normally 
have difficulty passing the cell membrane such as chemotherapy (electrochemotherapy). 
Electrochemotherapy (ECT) is used in cancer therapy, where chemotherapy is administered 
intratumoral or intravenous, then followed by electrical pulses applied directly on the tumor. The 
chemotherapy accumulates in the cancer cells which results in an increased cytotoxic effect. The 
most used chemotherapeutic drug used in electrochemotherapy is bleomycin. Electrochemotherapy 
is a well-documented local treatment form for especially cutaneous tumors. Today, the treatment 
is used mostly in palliative care in more than 140 centres around Europe.  
 
In vitro and in vivo studies have shown that the combination of calcium and electroporation is an 
effective method in killing cancer cells without serious side effects. This new combination opens 
the possibility of replacing bleomycin with calcium in treatments with electroporation.   
 
Calcium electroporation is a local treatment where calcium is administered intratumoral and 
followed by electrical pulses applied on the tumor. 
 
Calcium electroporation would be possible to use on patients for whom chemotherapy is 
contradicted e.g. severe lung functions impairment, pregnant woman etc. Calcium electroporation 
is a simple and inexpensive cancer treatment that does not involve any administration of cytotoxic 
chemotherapy, and can be performed by surgeons, radiologists as well as oncologists. Both 
electroporation equipment and calcium are already being used in the clinic, so the treatment can 
easily be implemented. 
 
Method 
We wish to examine calcium electroporation on patients with cutaneous metastases from any 
histology and compare the effect with electrochemotherapy. On evaluations on tumor response we 
want to investigate if calcium can replace bleomycin in the treatment of cutaneous metastases.  

 
To investigate that we use a randomized double blinded phase II study on patients with small 
cutaneous metastases. The metastases will be randomized into two treatment arms: 
1. Intratumoral injection with calcium, followed by electrical pulses. 
2. Intratumoral injection with bleomycin, followed by electrical pulses. 
Randomization will be done separately on each metastasis 
 
Objectives 
Primary Objective  
The primary objective is to evaluate and compare response rate of calcium electroporation and 
electrochemotherapy on cutaneous metastases. Response will be evaluated individual on each 
treated metastasis by clinical examination and documented by clinical photo before and after 
treatment. Evaluation is performed according to RECIST-like guidelines. 
Secondary Objective 
The secondary objective is to register adverts events for calcium electroporation and 
electrochemotherapy respectively. Adverse Events (AE) and Serious Adverse Events (SAE) will 
be evaluated and graded according to Common Terminology Criteria for Adverse Events (CTCAE) 
version 4.0. 
Tertiary Objective 
The tertiary objective is to register if calcium affects the current strength in electroporation 
treatments. Maximum given current in metastases treated with calcium will be compared to 
maximum given current in metastases treated with bleomycin. 
 
Trial Design 
Trial Design for Treatment of Cutaneous Metastases 
It is a randomized double blinded phase II study (adapted from Denmark) comparing the effect of 
calcium electroporation with ECT with bleomycin on patients with cutaneous metastases of any 
histology. As calcium and bleomycin are administered intratumoral, only small metastases from 
0.5-3cm will be treated. The treatments will be compared in tumor response and adverse events. 
Treatment will be done in a palliative purpose. 
 
Cancer patients with cutaneous involvement have often several metastases. A maximum of ten 
metastases will be included per patient. One to six metastases (depending on the patient’s number 
of metastases) will be numbered 1-6 and randomized into one of two treatment arms: 
1. Intratumoral calcium followed by electroporation 
2. Intratumoral bleomycin followed by electroporation 
 
Randomization will be performed separately on each metastasis, so the patient may receive both 
treatments. Calcium and bleomycin will be mixed into syringes labeled with numbers according to 
the metastases and both treating doctor and patient will be blinded to the content of the syringes. 

 
If the patient has more than six metastases, then one to four metastases will be used for biopsy. The 
treatment of these metastases will be known, and biopsies will be performed before and after 
treatment. The metastases used for biopsy will not be evaluated on response. 
 
• One to six metastases: All metastases will be evaluated on response, no biopsies. 
• Seven metastases: six metastases will be evaluated on response and one metastasis will be 
used for biopsy before and/or after treatment with calcium. 
• Eight Metastases: six metastases will be evaluated on response and two metastases will be 
used for biopsy before and/or after treatment with calcium and bleomycin respectively. 
• Nine Metastases: six metastases will be evaluated on response and three metastases will be 
used for biopsy before and/or after treatment with calcium and bleomycin respectively. 
• Ten Metastases: six metastases will be evaluated on response and four metastases will be 
used for biopsy before and/or after treatment with calcium and bleomycin respectively. 
 
The treatment will be performed in local or general anesthesia depending on location and number 
of metastases. Bleomycin/calcium will be administered intratumorally and immediately after the 
electrode will be placed on the metastasis. The electric pulses are generated using a cliniporator 
according to ESOPE (European Standard Operating Procedure of Electrochemotherapy). It is a 
once only treatment and the patients will be followed up with regular clinical controls for six 
months. At follow up six months after treatment, the randomization code for the one patient will 
be revealed. If the patient agrees, biopsies will then be taken from an area treated with calcium and 
an area treated with bleomycin. Last visit will be one year after treatment. 
To identify the metastases at follow up visits, the metastases will be marked with a pen, numbered 
and documented with clinical photos at baseline. 
 
 
Randomization 
The metastases are randomized by block randomization and are done separately in each patient. 
The randomization is performed by an extern unit, using the computer program nQuery Adviser 
7.0.  The bleomycin and calcium are mixed and labeled by an extern unit, and since both calcium 
and bleomycin are transparent and identical in volume, it is possible to label the syringes in a 
manner to blind the treating doctor. The syringes are labeled with numbers according to the 
metastases so syringe number ‘1’ goes to metastasis number ‘1’ and so on. The randomization code 
is kept behind locked doors, and is not accessible to the treating doctor. The randomization code is 
revealed for each patient after follow up, 6 months after treatment. 
 
The randomization code can be revealed before completion of the investigation, if the investigator 
believes that the treatment causes so serious unexpected events or reactions, that a continuation of 
treatment is unacceptable. 
As the randomization is done separately in each patient, it is possible to break the code for the 
individual patient, without breaking the code of the other participants. 

 
Dose 
The dose of calcium chloride was amended to 220 mmol/L from 168 mmol/L  in this trial. 
 
Volume of calcium chloride is dependent on tumor volume. Smaller tumors should have bigger 
volume per cm3, as smaller tumors are expected to have a bigger loss of injected medicine into the 
surrounding tissue. The dose volume is calculated according to the 'European Standard Operating 
Procedure of the Electrochemotherapi (ESOPE)':  
 
Calcium 
Calcium chloride 220 mmol/L (9 mg/ml): 
• Tumor < 0,5 cm3 – 1 ml / cm3 tumor volume 
• Tumor >0,5 cm3 – 0,5 ml/ cm3 tumor volume 
Tumor volume = ab2π/6 (a=longest diameter, b= longest diameter perpendicular to a) 
 
Bleomycin 
Both dose and volume of bleomycin is standard according to ESOPE 
 
Bleomycin 1000 IU/ml:  
• Tumor < 0,5 cm3 – 1 ml / cm3 tumor volume 
• Tumor >0,5 cm3 – 0,5 ml/ cm3 tumor volume 
Tumor volume = ab2π/6 (a=longest diameter, b= longest diameter perpendicular to a) 
 
Maximum of injected bleomycin per tumor will be 1500 IU and total dose per treatment 7500 IU. 
Normal maximum limit for bleomycin is 15.000 IU/m2 body surface area. 
 
Bleomycin and calcium chloride are mixed and labeled according to current guidelines. The 
mixtures will be used in a maximum of 6 hours after mixing. 
 
The Electrical Pulses  
The generation of electrical pulses is performed using a Cliniporator (IGEA, Italy). It is possible 
to use a plate or needle electrode. Needle electrodes are of different shapes e.g. linear or hexagonal, 
with different lengths and spacing between pins. The Cliniporator generates 8 pulses with duration 
of 0,1 ms and 400 V at a frequency of 5 kHz with linear electrodes, and 4 pulses with duration of 
0,1 ms and 730 V at a frequency of 5 kHz with hexagonal electrodes. The individual electrode is 
selected according to tumor size and localization. Cliniporator and electrodes are CE marked. 
 
Anesthesia  
The patients are anesthetized during treatment with local or general anesthesia. The anesthesia is 
planned in consultation between doctor and patient. 

 
Biopsy 
If the patient agrees biopsies will be performed from the tumor area before and after electroporation 
during local anaesthesia.  
 
Patients with more than 6 metastases: 
• One to two biopsies before treatment 
• On to two biopsies one week after treatment from metastases treated with calcium and 
bleomycin respectively. 
 
All patient’s regardless number of metastases will have done one biopsy from area treated with 
calcium and one from area treated with bleomycin, after the randomization code is revealed. 
 
All biopsies will be handled according to current guidelines and analyzed by a pathologist for 
amount of tumor tissue, inflammation, fibrosis and necrosis.  
 
Duration  
The treatment itself lasts a total of 30 to 60 minutes, depending if the patient receives local or 
general anesthesia. The patient will then be followed for up to 12 months with regular visits.  
 
Stop Rules 
The trial may be interrupted prematurely if: 
• Treatment causes such severe side effects that a continuation of treatment is unacceptable 
• Enrollment is too low to expect completion of the trial in its current form. 
 
The Research Ethics Committee and the Medicines Agency will be informed about the end of the 
trial. 
 
Statistical analysis  
This is a randomized non-inferiority study comparing calcium electroporation with 
electrochemotherapy with intratumoral injection of bleomycin. For dimensioning tumor number, 
analysis of non-inferiority study is used. Tumor response is evaluated at each follow up with 
surveying. Documentation is done with digital color photography, including a ruler to estimate 
tumor size. Primary evaluation of the response will be based on criteria similar RECIST guidelines 
1.1. Tumor response was analyzed using Fisher’s exact test on objectice response 6 months after 
treatment.  
  
Response rate of tumors ≤3cm treated with ECT with bleomycin is 85-100% (existing data). In this 
study, we have chosen not to put the expected response rate for ECT too high, but estimate it to 
85%. We have no clinical results for the treatment of calcium electroporation, but on the basis of 
preclinical studies, the response rate for calcium electroporation is estimated to 70%. In order to 
detect a clinical difference in the two treatments of 15%, with a significant level of 0.1, and a power 

of 80%, a minimum of 34 evaluable tumors need to be included. (Sample Size 
Calculator:  https://www2.ccrb.cuhk.edu.hk/stat/proportion/tspp_sup.htm )  
The metastases are evaluable 4 weeks after treatment. If a patient is excluded from the protocol 
before 4 weeks follow up, the patient will be replaced.  
  
The number of patients that are being pulled out of the trial will be listed and primary reason for 
the exclusion will be described.