Pulsed Field Assisted Chemotherapy

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Pulsed Field Assisted Chemotherapy

James E. Bare8005 Marble Ave. NE

Albuquerque, NM 87110

505-268-4272

[email protected]

This paper was originally written in 2004 and more published research has now been

located which is not referenced in this paper. A web page has been created with the full

abstracts from published research that demonstrates the synergism between pulsed EM

fields and chemotherapeutic medications .

Updated References and Abstracts

Research into the physiologic effects of low power, pulsed electromagnetic (EM) fields, has

produced a number of important discoveries. To date, with rare exception, these discoveries at

best are investigational, and have not been applied in a clinical manner. Much of this research

material is unknown to the general practitioner, and has not been correlated into a potentially

utilizable treatment method. This paper proposes the fusion of existing cancer chemotherapy

techniques with low power pulsed EM field research discoveries. Evidence is presented that

the sum of these combined effects far exceeds that of each method individually. A

transmitted, pulsed EM field, can be created which will safely produce whole body

permeation/saturation. Such saturation can create an interaction of the pulsed EM field with

chemotherapeutic medications, simultaneously, at all tumor sites throughout the body. By

creating a synergism of biochemical, electrochemical , and electronic principles, the

practitioner should be able to achieve a superior treatment outcome.

Present chemotherapy regimens are fraught with many shortcomings. The worst of these is

toxicity , which can include permanent damage to various vital organs including the heart,

lungs and kidneys. Even without such permanent damage, short term toxicity manifests as

severe nausea, repression of the hematopoietic system, alopecia, and many other types of

physical unpleasantness. Other major problems with chemotherapy are development of

cellular accommodation ( non responsive/refractory to treatment), patient/family mental

stress, and in many cases no clear cut outcome other than temporary tumor shrinkage. Actualprolongation of patient survival post treatment is all too often a secondary consideration in the

application of chemotherapy.

The great majority of cancer research is directed not to a solution of the these problems, but

rather towards a solution to cancer. For patients, the only reason they consent to such

abusive treatment, is that the consequences of untreated cancer outweigh the negative

repercussions of treatment. What is needed is a new chemotherapeutic medication which will

selectively affect cancer cells, and has minimal side effects/toxicity. This medication would

also have improved outcomes from treatment, and be highly resistant to cellular

accommodation . Primary importance would be directed to improved patient survival

percentages and overall length of survival. Such a medication does not now apparently exist,and next to an outright cure, could be considered as the Holy Grail of cancer treatment. It is

the contention of this paper that it may be possible to adapt at least a few of the presently
mailto:[email protected]://www.rifetechnologies.com/research.htmlhttp://www.rifetechnologies.com/research.htmlmailto:[email protected]


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utilized chemotherapeutic medications, to a new treatment protocol. A protocol that will

produce the effects that meet this idealized vision of cancer treatment.

This transformation of available chemotherapeutic medications and treatment is to be

accomplished via a synergistic combination of the medication with a low voltage and low

current, transmitted pulsed electromagnetic (EM) field. The ideal transmitted EM field is onethat can couple itself energetically to the entire body. That is, the field may be used to treat

the entire body at one time. This will ensure that wherever the medication can be delivered

within the body, treatment congruence between the field and the medication will occur. There

is considerable scientific supporting evidence for this proposal that will be discussed in the

following paragraphs.

The utilization of electrical pulses with chemotherapy medications is not new. This technique

is being investigated and employed by a variety of researchers and cancer treatment

specialists. The primary method utilized is some variant of what is known as

electropermeabilization or as it is also known, electroporation. An ultra short time duration,

but high intensity, electrical pulse , is applied to a tumor or group of cells to produce shortduration and reversible pores within the plasma membrane. Pores through which various ions

and molecules may be introduced into the interior of the cell. To use this technique, a tumor

site is saturated with a chemotherapy drug and then pulsed through implanted electrodes with

a very short and very intense electrical signal.

An associated technique being utilized by some clinicians and researchers is known as EChT

or Electro Chemical Treatment of tumors. In this method electrodes are inserted into the

cancer site along with a chemotherapeutic medication. In contrast to electropermeabilization,

relatively low currents and voltages are used on the tumor site. These voltages and currents

may be applied for many hours at a time. The migration of charged particles in the created

field is somewhat akin to electrophoresis and has produced some excellent responses to

treatment.

Many of the chemotherapeutic drugs chosen for EchT or electropermeabilization techniques

belong to the class known as Alkylating agents. These compounds are electrically polarized

and capable of being influenced to migrate in an electrical field vector. There are obvious and

non obvious reported problems to both of these electrically based methods. These include:

1. 1. The techniques can only be used on specific localized tumors.

2. 2. The patient must endure the implantation of electrodes and direct injection of the

chemotherapy medication. Often deep within the body.

3. The effects occur primarily between the electrodes, with very large tumors needingmultiple placement of needle electrodes. The technique cannot be utilized with

extremely small tumors. This allows small metastatic sites to escape treatment.

4. 4. Some tissues do not tolerate the direct application of chemotherapeutic

medications, and there may be collateral damage to healthy tissue.

5. 5. With EChT, there is also metallic ion migration off the electrodes into the

surrounding tissues. This can create localized areas of metal toxicity.

6. 6. The patient must endure the bursts of electrical current into their body from either

EChT or electropermeabilization. In EChT, the electricity is much akin to that

delivered by a muscle stimulator. An electropermeabilization pulse is more akin to the

spark derived from an automotive ignition coil. To use a more graphic example of

what a patient experiences, imagine having a muscle stimulator or car ignition coilattached directly to your colon.


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In defense of electropermeabilization and EChT, both methods overcome some major

problems in traditional oral or IV chemotherapy. Through direct injection into a tumor, the

medication can be concentrated at the site of the tumor without creating whole body toxicity.

Electropermeabilization and EChT offer a method to increase drug delivery into the cancer

cells, which has brought about some very significant responses to treatment. Further, the

amount of medication necessary to produce a toxic response within the cancer cells, is attimes, significantly reduced.

EChT and Electropermeabilization are not the methods to be utilized in this proposal . They

have been discussed only to demonstrate that there are existing synergies between electrical

fields and chemotherapeutic medications.

Concept Discussion:

The human body ( or for that matter any mammals body) is very complex electrically . Each

cell within the body has a particular set of electrical parameters within which it functions. The

cells of the body operate and maintain their homeostasis at least partially through electro

chemical processes. Electrically charged particles such as ions, and proteins, accumulate on

both sides of the various membrane interfaces of a cell, and create an electrical potential.

Normal cells and cancer cells have many different physiologic qualities, which include those

of their plasma membrane potential and electrochemistry. A normal cell has a plasma

membrane potential of about 70 to 100 mv (Cone, 1970, 1975, 1985). This is equal to an

electrical potential of between 10 and 20 million volts per meter ( Brown, 1999). The

mitochondrial membranes of a normal cell maintain an electrical potential of almost 40

million volts per meter (Brown, 1999).

When a cell becomes cancerous, plasma membranes degenerate and depolarize ( Marino et.al. 1994 ) .More succinctly, the electrical potential across the membrane drops drastically.

The degeneration and loss of electrical potential of the plasma membrane in cancer allows it

to become more permeable to water, and to certain ions such as sodium that normally are

found in abundance on the outside of a normal cell. In cancer, sodium is allowed through the

cells plasma membrane and accumulates within the cytosol. Meanwhile the ionic elements of

potassium, magnesium , calcium, and zinc, which are normally found inside of the cell

plasma membrane, tend to migrate out of the cytosol (Seeger and Wolz, 1990). As the

distribution of ions shift, cancer cells tend to become very electronegative, and the tissue

areas surrounding the cancer cells become quite electropositive. This electropositivity has

been used for detection of cancer (Marino, et. al, 1994). The combined effects of the low

plasma membrane electrical potential and ionic imbalance, assists in the conversion of anormal cells aerobic based metabolism to that of a cancer cell with an anaerobic based

metabolism. Conversion to anaerobic glycolysis (fermentation) as a primary mechanism for

energy production results in excessive accumulation of organic acids and acidic pH alterations

in cancerous tissues (Seeger and Wolz, 1990).

The loss of electrical potential across a cancer cells plasma membrane is the foundation upon

which this proposal is based. The application of an artificially created electrical field of a

few tens of millivolts at the plasma membrane of a cancer cell will create an

electrochemical imbalance of the cell. An imbalance which this proposal intends to exploit.

Ions and molecules ( chemotherapy medications) enter cells through a variety of differentmethods. These methods include ion gating, osmosis, and endocytosis. In a normal cell, these

mechanisms are all driven by cellular energetic pathways. Ion Gating and Osmotic methods


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are capable of only passing small ions through the cell plasma membrane. Endocytosis, can

pass large macromolecules such as sugars, and most importantly medications. These three

methods of transport can be artificially mediated and elicited by the presence of an external

pulsed electrical field. (Teissie and Tsong 1981, Petrov and Mircevova, 1986 , Rosemberg

and Korenstein ,1997 ) When so accomplished they are known as Voltage Dependent Ion

Gating, Electro Osmosis, and Electro Endocytosis. By inducing these electrically drivenmethods of transport, pulsed electrical fields are capable of producing disruption of the

cancer cells electro chemical balance and function ( Panagopoulos, Karabarbounis and

Margaritis, 2002).

Electro endocytosis is of significant interest to this proposal. Endocytosis is a process

whereby the cell plasma membrane invaginates and surrounds a large macro molecule. This

piece of plasma membrane then closes off to form a vesicle which transports the molecule

within the cell. The reverse process is known as exocytosis, whereby the empty vesicle is

transported back to the plasma membrane. The vesicle then opens, and rejoins the plasma

membrane. The process of endocytosis must be balanced to some degree with the process of

exocytosis. If an extreme excess of endocytosis in relation to exocytosis occurs, a possible

compromise of the cells plasma membrane may ensue.

It is well known that at very high electrical field strengths, a process known as vesicle

electroformation occurs. This process can create extremely large or macro size vesicles out of

bilayer lipid membranes. A high electrical field strength dependent process is not the

mechanism this proposal seeks to utilize.

Rosemberg and Korenstein ( 1997) have shown that the process of electro endocytosis can

occur at very low field strengths. They found it possible to incorporate molecules in the 1-

2000 kD range into 85% of the cells used in their test. Other researches have supported this

finding. The necessary field strength can be as low as 20 V/cm or a transmembrane potential

of 6mv ( Teissie & Tsong, 1981). Electrically polar and nonpolar chemotherapeutic

molecules have been utilized with very low electrical field strengths to produce highly

significant positive treatment outcomes. Entin, et. al. (2003) , used fields of 40V/cm (12 mv

added transmembrane potential ) with Bleomycin, Taxol , and Cisplatin in treatment of mice

inoculated with melanoma cells. A low voltage enhancement was also reported by Miyazaki,

et. al. (2003). Mice inoculated with Colon cancer cells were given intratumor injections of

Bleomycin and exposed to fields of from 50 to 150V/cm . The use of chemotherapeutic

medications with electrical fields can result in enhanced cellular sensitivity to the medication.

Gray , et. al. (2000) found severe over dosage reactions to Adriamycin (ADM) occurred

when ADM was administered to animals kept in a static but very intense electrical field .

Cisplatin and its immediate family of molecules acts directly upon DNA. It has been used

successfully in the combined treatment of B-16 melanoma bearing mice with low ( 20-

100V/cm) field strength ( Entin et. al., 2003). It seems apparent that not only will the

presence of cis-platinum type molecules within a cancer cell be advantageous, but so would a

simultaneous stimulation of the activity level of the cells DNA. Cancer cells go through

periods of rest and activity and most chemotherapeutic drugs are primarily effective against

rapidly dividing cells. Application of chemotherapeutic medications is often timed to a

perceived interval of genetic/cellular activity. The usage of low voltage pulsed fields seems

to create an artificial window of activity. Low voltage pulsed electrical fields act not just

upon the ability of a cell to aid molecular & ionic transport, they also act upon the cells DNA.

Binderman, et. al.(1985) found that at between 13 to 50 V/cm , cell cultures of skeletal originwould immediately show changes in cyclic AMP levels and enhanced DNA synthesis . Blank

and Soo ( 1997 ) reported a frequency dependent effect on Na, K-ATPase enzyme activation


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in fields of from 3- 3000 Hz. Pulsed EM fields have been linked not just to enzyme reactions

but also to increased transcription rates for specific genes. Pulsed EM fields act directly on

signal transduction pathways and with electrons in DNA to stimulate biosynthesis ( Goodman

and Blank 2002).

Chemotherapy cellular resistance is the most common cause of treatment failure and hasseveral different etiologies. The most ubiquitous method of resistance to treatment is that of

activation and expression of energy dependent transporters that literally remove the

medications from the cells ( Gottesman 2002). The medication, due to the action of

transporters, is not allowed to accumulate to a toxic level and cannot perform its assigned

task. Even though small amounts of chemotherapeutic drugs do enter resistant cancer cells,

the drugs biochemical action is thwarted. It is thought that the primary action of

chemotherapeutic drugs is the initiation of apoptosis. Apoptosis is at least partially mediated

by the behavior and chemical signals produced by the mitochondria. The mitochondria of

malignant cells are electrochemically different from those of normal cells. This

electrochemical difference acts to block mitochondrial response to the medication, and

thwarts the apoptotic cascade.

The plasma membrane potential of mitochondria in cancers cells has been found to be

elevated, in one case it was approximately 60 mv higher than that of control epithelial cells (

Modica-Napolitano and Aprille 1987 ). Elevated mitochondrial membrane potential coincides

with Cisplatin resistance in some cancer cell lines ( Dorward and Singh 1996). The high

transmembrane potentials of mitochondria are created by a relative electropositivity of the

outer membrane and a high negativity of the inner membrane ( Johnson , et. al. 1981).

Mitochondrial based apoptotic mechanisms require that the plasma membranes of

mitochondria depolarize, resulting in diminution of the transmembrane potential ( Mayer and

Oberbauer 2003). Due to their elevated electropotential, mitochondria of cancer cells are

going to have to undergo a much greater fall in electrical potential than a normal cell to

initiate apoptosis. Further, formation of mitochondrial permeability transition pores ( a key

factor in the initiation of mitochondrial based apoptosis) is inhibited as pH decreases (Nicolli ,

et. al. 1994).

There are other inhibitors of treatment response within cancer cells. The majority of solid

tumors show some if not total resistance to chemotherapy. This resistance is mediated by a

local stress response to the microenvironment. When solid tumors are subjected to local

conditions of hypoxia, acidic pH and low levels of glucose they react by stopping division.

When cell division is arrested in this manner, chemotherapeutic medications become

ineffective, that is they fail to induce apoptosis. ( Tomida and Tsuruo 1999 ). In summary,cancer cells are at least partially self protected against chemotherapy initiation of apoptosis

through their electrochemistry.

Pulsed Field Correction of Drug Resistance:

Correction of cellular stress factors, inhibition of intercellular drug transporter mechanisms,

excess acid production, and excess mitochondrial membrane potentials, may be possible

through the application of external pulsed EM fields.

Rosemberg and Korenstein ( 1997 ) found that low voltage induced electro endocytosis could

be used to incorporate polysaccharides and - galactosides into cells. A confirmation of this

effect was presented by Rols, et. al.( 1995) . When stressed by hypoxic conditions, cancer

cells produce compounds that create neoangiogenesis. They also produce more hypoxia


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inducible factor 1 (HIF1) , increase expression of hypoxia regulated genes , and

metabolically shift to utilize the oxidizing ability within the fatty acid synthesis pathway

( Hochachka, et. al. 2002 ) . Angiogenic responses to hypoxia can lead to increased tumor

growth , increased metastasis and poor treatment outcomes ( Duffy, et. al. 2003). Through

the application of pulsed EM fields it may be possible to circumvent these stress responses to

hypoxia. Di Carlo, et.al. ( 2000), found that EM field exposures protected chick embryosfrom hypoxic insult. More definitively it was found that EM fields could be used to induce

increased heat shock protein 70 (hsp70) levels ( Han,et. al. 1998, Carmody, et. al. 2000 ).

Hsp70 plays an important part in the protection of tissues to hypoxia ( Rafiee, et. al. 2003 ).

Overproduction of hsp70 has been found to protect cancer cells from apoptosis following

irradiation and chemotherapy (Witkin, 2001). This must be judged against the consequences

of a hypoxic response. The expression of hsp 70 is key to preventing some of the sequences

that lead to hypoxic response. More importantly, hsp70 has been linked to anti cancer

immune system responses. Apoptosis can result in an immunogenic or non immunogenic

response. The immunogenic response is dependent upon the presence of hsp70. Stressed

cancer cells express hsp 70 on their plasma membranes , and can initiate an antitumor

response by the immune system ( Feng, et. al. 2003). This response can be significant. Micewere inoculated with Colon 26 cells and then treated with EChT and Bleomycin. An 80 to 100

% response rate occurred. When the same mice were reinoculated with Colon 26 cells, the

mice rejected the cells and no tumor growth was noted. Injection of a different cancer

produced tumors in these same mice (Miyazaki et. al. 2003 ). It has been found that

immunogenic response to necrotic and apoptotic tumor cells can be equivalent (Kotera et. al.

2001)

To clarify, the presence of pulsed EM fields may inhibit many of the undesirable cellular

metabolic responses of cancer cells to hypoxia. These responses include; generation of hsp70,

inhibition of expression of HIF1, diminish the production of angiogenic proteins, initiation

of an anti cancer immune response, and possibly inhibit the activity of the FAS to reduce

cellular growth rates. This proposed outcome from the use of pulsed EM fields to inhibit

angiogenesis and tumor growth is supported in the literature. Pulsed EM fields were found to

inhibit tumor growth with a reduction in the extent of vascularization and increased areas of

tumor necrosis compared to controls ( Williams, et. al. 2001).

In a cancer cell, Na+ ions accumulate inside the cell and K+ ions accumulate outside the cell.

Modification and reversal of the local ionic concentrations and can be created by an external

AC field. ( Teissie and Tsong 1981). Further , elevated pHi will increase the delta pH acrossthe plasma membrane. It has been found that as a Multi Drug Resistant (MDR) protein is

expressed, the delta psi or plasma membrane potential decreases ( Roepe, et. al. 1993). The

presence of a pulsed field will circumvent the fall in plasma membrane potential. The energy

within a pulsed EM field is capable of being absorbed by plasma membranes at least partially

through the process of electro conformational coupling (Tsong,, et. al.,1989) Once absorbed,

this energy may be converted to the chemical bond energy of ATP or to the potential energy

of concentration gradients ( Tsong, et. al., 1989, Timashev 1981) . EM pulse responses of

cellular membrane systems are frequency dependent. This response has been shown in

several membrane enzyme systems ( Markin and Tsong 1991, Luchian, et. al., 2002, Ruiz-

Gomez, et.al., 2002, Gluck et.al. 2001). Frequencies can also be utilized to affect cell

division and growth. Kirson et al.,2004 ,found that low intensity frequencies in the 100 KHzto 300 KHz produced an inhibitory effect on a variety of human and rodent cell tumor lines.


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This effect was non thermal, and acted through both arrest of cell division and destruction of

cells undergoing division.

Movement of ions and molecules across the mitochondrial membranes is normally

accomplished by a chemiosmotic mechanism that is free electron dependent. It is possible that

the presence of a pulsed field could supplement, drive, or supplant this mechanism and affectthe electropotential gradient of mitochondria. This may result in a diminution of the outer

membranes electropositivity, allowing the Voltage Dependent Anion Channels ( VDAC) to

open. As the electropotential of the mitochondrial membranes become depolarized the

probability of Mitochondrial Permeability Transition Pore ( MPTP ) formation increases.

(Petronilli, et. al., 1994). Formation of MPTPs is a necessary precursor to the initiation of

mitochondrial based apoptotic mechanisms. Similar pulsed fields gradients should act to

inhibit the expulsion of drug molecules from the cell. The transporters will have to work

against an artificially created concentration gradient that seeks to actively bring ions and

molecules into the cell.

Pulsed EM Field Generation:

From a practitioners viewpoint any instrument that might generate a pulsed EM field must

conform to several criteria. Primarily, the device must be safe for the patient, and application

must be consistent from patient to patient. Secondarily the device must be easy for the

practitioner to utilize. Presently, there is not such a clinical device approved for use by the US

FDA. Countries such as Canada have approved pulsed EM field devices that meet this

criteria. Below is a picture of a type of device that will produce the necessary non thermal

pulsed EM field. A variant of this instrument is presently approved by Health Canada as a

transmitted field TENS device for the control of pain. Effective range is approximately 6

meters, typical treatment distance of the patient from the device is 2 meters.

300 Watt PEPpulsedfield transmitter. 27.125MHzcarrier frequency.Pulserate capability 225 KHz.


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TransmittedPulse. 10 KHz pulserate ( 10,000pulsesper second)

Summary :

The application of a transmitted pulsed EM field of low potential makes it possible to modify

the electrochemistry and physiology of cancer cells. Cellular membrane systems and cellular

mitosis are frequency responsive, and can be influenced by pulsed EM fields of varying

frequency sequences and duration. Frequency sequences can be created to selectively

encourage different metabolic responses. The presence of the external field will produce a

change in transmembrane potentials . This change in potentials will create osmotic, ion

gating, and endocytotic effects at the cells plasma membranes. Internal and external actions

of the field will result in ion/molecular transport, reversal of cellular stress factors, interfere

with angioneogeneis, increase DNA transcription rates, produce an immune response, and

cause inhibition of drug resistance mechanisms

It is the hypothesis of this paper that the totality of these effects will be enhanced transport of

chemotherapeutic medications into the cells, retention of the medications to achieve high

concentration levels, and increased responsiveness to the utilized medication . As the

concentration of the medication increases in conjunction with the pulsed field altered cancer

cell metabolism, apoptotic effects should become predominant. Transmitted pulsed EM

fields when used in conjunction with chemotherapeutic medications will; decrease the

treatment dosage substantially, produce an enhanced response to treatment, and have

minimal to no toxic side effects.

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38.Teissie J. , Tsong T. Voltage Modulation of Na+/K+ Transport in Human Erythrocytes.

Journal of Physiology ( Paris ) , 1981 May;77(9):1043-1053 PMID: 6286955

39. Timashev SF, Effect of Electrical Fields on the Kinetics of Biological Processes,

Biofizika 1981 Jul-Aug;26(4):642-646

40.Tomida A, Tsuruo T, Drug Resistance Mediated by Cellular Stress Response to theMicroenvironment of Solid Tumors. Anticancer Drug Des. 1999 Apr;14(2):169-177

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41.Tsong TY, Liu DS, Chauvin F, Astumian RD,Resonance Electroconformational

Coupling: A Proposed Mechanism for Energy and Signal Transductions by Membrane

Proteins. Biosci. Rep. 1989 Feb;9(1):13-26 PMID: 2655737

42.Tsong TY, Liu DS, Chauvin F,Electroconformational Coupling (ECC): An Electric

Field Induced Enzyme Oscillation for Cellular Energy and Signal Transductions.

Bioelectrochemistry and Bioenergetics Vol 21 Issue 3, June 1989 319-33143.Williams CD, Markov MS, Hardman WE, Cameron IL. Therapeutic Electromagnetic

Field Effects on Angiogenesis and Tumor Growth. Anticancer Res. 2001 Nov-

Dec;21(6A):3887-91

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44. Witkin SS, Heat Shock Protein Expression and Immunity: Relevance to Gynecologic

Oncology Eur J. Gynaecol. Oncol. 2001;22(4):249-56 PMID 11695802

Research Abstracts Showing the Synergistic Effects

Between Pulsed Fields and Chemotherapy

Since the paper was written in 2004, more supporting published references to the effect have

been discovered. Some have been published as recently as March of 2006. As more references

are discovered or published, they will be added to this web page.

BMC Cancer. 2006 Mar 17;6:72.

Alternating current electrical stimulation enhanced chemotherapy: a novel

strategy to bypass multidrug resistance in tumor cells.

Janigro D, Perju C, Fazio V, Hallene K, Dini G, Agarwal MK, Cucullo L.

Division of Cerebrovascular Research, Cleveland Clinic Lerner College of Medicine,

Cleveland, OH 44106, USA. [email protected]

BACKGROUND: Tumor burden can be pharmacologically controlled by inhibiting cell

division and by direct, specific toxicity to the cancerous tissue. Unfortunately, tumors often

develop intrinsic pharmacoresistance mediated by specialized drug extrusion mechanisms

such as P-glycoprotein. As a consequence, malignant cells may become insensitive to variousanti-cancer drugs. Recent studies have shown that low intensity very low frequency electrical

stimulation by alternating current (AC) reduces the proliferation of different tumor cell lines

by a mechanism affecting potassium channels while at intermediate frequencies interfere with

cytoskeletal mechanisms of cell division. The aim of the present study is to test the hypothesis

that permeability of several MDR1 over-expressing tumor cell lines to the chemotherapic

agent doxorubicin is enhanced by low frequency, low intensity AC stimulation. METHODS:

We grew human and rodent cells (C6, HT-1080, H-1299, SKOV-3 and PC-3) which over-

expressed MDR1 in 24-well Petri dishes equipped with an array of stainless steel electrodes

connected to a computer via a programmable I/O board. We used a dedicated program to

generate and monitor the electrical stimulation protocol. Parallel cultures were exposed for 3

hours to increasing concentrations (1, 2, 4, and 8 microM) of doxorubicin followingstimulation to 50 Hz AC (7.5 microA) or MDR1 inhibitor XR9576. Cell viability was

assessed by determination of adenylate kinase (AK) release. The relationship between MDR1


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expression and the intracellular accumulation of doxorubicin as well as the cellular

distribution of MDR1 was investigated by computerized image analysis

immunohistochemistry and Western blot techniques. RESULTS: By the use of a variety of

tumor cell lines, we show that low frequency, low intensity AC stimulation enhances

chemotherapeutic efficacy. This effect was due to an altered expression of intrinsic cellular

drug resistance mechanisms. Immunohistochemical, Western blot and fluorescence analysisrevealed that AC not only decreases MDR1 expression but also changes its cellular

distribution from the plasma membrane to the cytosol. These effects synergistically

contributed to the loss of drug extrusion ability and increased chemo-sensitivity.

CONCLUSION: In the present study, we demonstrate that low frequency, low intensity

alternating current electrical stimulation drastically enhances chemotherapeutic efficacy in

MDR1 drug resistant malignant tumors. This effect is due to an altered expression of intrinsic

cellular drug resistance mechanisms. Our data strongly support a potential clinical application

of electrical stimulation to enhance the efficacy of currently available chemotherapeutic

protocols.

PMID: 16545134

1: Anticancer Res. 2001 Jan-Feb;21(1A):317-20.

Drug resistance modification using pulsing electromagnetic field

stimulation for multidrug resistant mouse osteosarcoma cell line.

Hirata M, Kusuzaki K, Takeshita H, Hashiguchi S, Hirasawa Y, Ashihara T.

Department of Orthopaedic Surgery, Kyoto Prefectural University of Medicine,

Kawaramachi-Hirokoji, Kamigyo-ku, Kyoto, Japan.

Multidrug resistance (MDR) is one of the major problems in osteosarcoma

chemotherapy. Therefore, methods of overcoming MDR are urgently needed. In this

study, we investigated the effects of pulsing electromagnetic field stimulation

(PEMFs) on a MDR murine osteosarcoma cell line which strongly expresses P-

glycoprotein (P-gp). To assess the reversal effects of PEMFs on doxorubicin (DOX)

resistance, MTT assay was applied. Viable cells were assessed by the trypan blue

exclusion test. Fluorescence intensity of DOX binding to nuclear DNA of each cell

was measured using a cytofluorometer. Changes in P-gp expression in each cell were

detected by the indirect immunofluorescence method using an antibody to Pgp.

PEMFs increased DOX binding ability to nuclear DNA and inhibited cell growth,although it had no significant effect on P-gp expression. These findings indicated that

PEMFs reversed the DOX resistance of the MOS/ADR1 cells by inhibiting P-gp

function. The results suggested that PEMFs may be useful as a local treatment for

MDR osteosarcoma.

PMID: 11299755

1: Radiats Biol Radioecol. 2003 May-Jun;43(3):351-4.

[Antitumor effect of joint action of low intensity electromagnetic fields

and ultra low doses of doxorubicin]

[Article in Russian]


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Ostrovskaia LA, Budnik MI, Korman DB, Bliukhterova NV, Fomina MM,

Rykova VA, Burlakova EB.

Emanuel Institute of Biochemical Physics, Russian Academy of Sciences, Moscow,

119991 Russia. [email protected]

Combined action of a low intensive physical factor and a chemotherapeutic agent in

ultralow doses against Lewis lung carcinoma was studied. Antitumor activity of low

intensiwe electromagnetic field was expressed as inhibition of tumor growth at 60%

compare to control. Ultra low doses of doxorubicin as well as its standard dose

resulted in inhibition of tumor growth by 60-70% in comparison with control. Joint

action of both factors leaded to increasing in the antitumor effect that reached such

level of tumor growth inhibition as 85% relative to control.

PMID: 12881995

Cancer Biochem Biophys. 1999 Jul;17(1-2):89-98.

Magnetic field induced inhibition of human osteosarcoma cells

treated with adriamycin.

Chakkalakal DA, Mollner TJ, Bogard MR, Fritz ED, Novak JR, McGuire MH.

Creighton University Biomedical Engineering Center, Creighton University School of

Medicine, Omaha, NE 68105, USA.

Morbidity resulting from the toxicity of chemotherapeutic drugs suggests that novel

approaches are worthy of investigation. Development of the use of low intensity

magnetic fields as an adjuvant to current treatment regimens to prevent metastatic

disease may prove to be efficacious. Using a cell culture model, we have developed a

magnetic field (MF) treatment that offers the possibility of lowering the therapeutic

dose of these drugs and thereby reducing morbidity. Our studies have found that a low

intensity (approximately 2 gauss) MF signal and a relatively low dose (0.1 microg/ml)

of Adriamycin (ADR) inhibited proliferation of human osteosarcoma cells by 82%,

whereas the MF and ADR acting individually caused only 19% and 44% inhibition,

respectively.

PMID: 10738905

Bioelectromagnetics. 2002 Dec;23(8):578-85.

Influence of 1 and 25 Hz, 1.5 mT magnetic fields on antitumor drug

potency in a human adenocarcinoma cell line.

Ruiz-Gomez MJ, de la Pena L, Prieto-Barcia MI, Pastor JM, Gil L, Martinez-

Morillo M.

Laboratory of Radiobiology, Department of Radiology and Physical Medicine, Faculty

of Medicine, University of Malaga, Teatinos, Malaga, Spain.


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The resistance of tumor cells to antineoplastic agents is a major obstacle during cancer

chemotherapy. Many authors have observed that some exposure protocols to pulsed

electromagnetic fields (PEMF) can alter the efficacy of anticancer drugs; nevertheless,

the observations are not clear. We have evaluated whether a group of PEMF pulses

(1.5 mT peak, repeated at 1 and 25 Hz) produces alterations of drug potency on a

multidrug resistant human colon adenocarcinoma (HCA) cell line, HCA-2/1(cch). Theexperiments were performed including (a) exposures to drug and PEMF exposure for

1 h at the same time, (b) drug exposure for 1 h, and then exposure to PEMF for the

next 2 days (2 h/day). Drugs used were vincristine (VCR), mitomycin C (MMC), and

cisplatin. Cell viability was measured by the neutral red stain cytotoxicity test. The

results obtained were: (a) The 1 Hz PEMF increased VCR cytotoxicity (P < 0.01),

exhibiting 6.1% of survival at 47.5 microg/ml, the highest dose for which sham

exposed groups showed a 19.8% of survival. For MMC at 47.5 microg/ml, the % of

survival changed significantly from 19.2% in sham exposed groups to 5.3% using 25

Hz (P < 0.001). Cisplatin showed a significant reduction in the % of survival (44.2-

39.1%, P < 0.05) at 25 Hz and 47.5 microg/ml, and (b) Minor significant alterations

were observed after nonsimultaneous exposure of cells to PEMF and drug. The dataindicate that PEMF can induce modulation of cytostatic agents in HCA-2/1(cch), with

an increased effect when PEMF was applied at the same time as the drug. The type of

drug, dose, frequency, and duration of PEMF exposure could influence this

modulation. Copyright 2002 Wiley-Liss, Inc.

PMID: 12395412

1: J Environ Pathol Toxicol Oncol. 1993 Oct-Dec;12(4):193-7.

Biological effects of PEMF (pulsing electromagnetic field): an attemptto modify cell resistance to anticancer agents.

Pasquinelli P, Petrini M, Mattii L, Galimberti S, Saviozzi M, Malvaldi G.

C.R.E.S.A.M., Pisa, Italy.

Pulsing Electromagnetic Field (PEMF) effects lead to a modification of the multidrug

resistance (MDR) of cells in vitro and in vivo. The murine leukemic doxorubicin-

resistant cell line, P388/Dx, subjected to PEMF irradiation in vitro, showed a

significant difference in thymidine incorporation when the concentration of

doxorubicin reached a level of 1 microgram/mL, which corresponds to the inhibitiondose 50 (ID50). The human lymphoblastic leukemia vinblastine-resistant cell line,

CEM/VLB100, also showed a significant modification under the same experimental

conditions at the in vitro ID50 corresponding to a vinblastine concentration of 100

ng/mL. BDF1 mice transplanted with P388/Dx cells also had an increase in their life

span when doxorubicin was injected intraperitoneally in fractionated doses, while

being subjected to PEMF irradiation.

PMID: 8189374

1: Pharmacol Res. 2003 Jul;48(1):83-90.

Static and ELF magnetic fields enhance the in vivo anti-tumor efficacy

of cis-platin against lewis lung carcinoma, but not of cyclophosphamide


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against B16 melanotic melanoma.

Tofani S, Barone D, Berardelli M, Berno E, Cintorino M, Foglia L, Ossola P,

Ronchetto F, Toso E, Eandi M.

Department of Medical Physics, Ivrea Hospital, ASL 9, 10015 (TO), Ivrea, Italy.

Previous works showed that exposure to static and extremely low frequency (ELF)

magnetic fields (MF) over 3 mT slows down the growth kinetics of human tumors

engrafted s.c. in immunodeficient mice, reducing their metastatizing power and

prolonging mouse survival. In the experiments reported here, immunocompetent mice

bearing murine Lewis Lung carcinomas (LLCs) or B16 melanotic melanomas were

exposed to MF and treated respectively with two commonly used anti-cancer drugs:

cis-diamminedichloroplatinum (cis-platin) and N,N-bis (2-chloroethyl)tetra-hydro-2H-

1,3,2-oxazaphosphorin-2-amine 2-oxide (cyclophosphamide). The experiment

endpoint was survival time. The survival time of mice treated with cis-platin (3mg/kg

i.p.) and exposed to MF was significantly (P


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Infection of orthopaedic implants is a significant problem, with increased antibiotic

resistance of adherent 'biofilm' bacteria causing difficulties in treatment. We have

investigated the in vitro effect of a pulsed electromagnetic field (PEMF) on the

efficacy of antibiotics in the treatment of infection of implants. Five-day biofilms of

Staphylococcus epidermidis were grown on the tips of stainless-steel pegs.They were

exposed for 12 hours to varying concentrations of gentamicin or vancomycin inmicrotitre trays at 37 degrees C and 5% CO2. The test group were exposed to a

PEMF. The control tray was not exposed to a PEMF. After exposure to antibiotic the

pegs were incubated overnight, before standard plating onto blood agar for colony

counting. Exposure to a PEMF increased the effectiveness of gentamicin against the

five-day biofilms of Staphylococcus epidermidis. In three of five experiments there

was reduction of at least 50% in the minimum biofilm inhibitory concentration. In a

fourth experiment there was a two-log difference in colony count at 160 mg/l of

gentamicin. Analysis of variance (ANOVA) confirmed an effect by a PEMF on the

efficacy of gentamicin which was significant at p < 0.05. There was no significant

effect with vancomycin.

PMID: 12793569

Antimicrob Agents Chemother. 1996 Sep;40(9):2012-4.

Bacterial biofilms and the bioelectric effect.

Wellman N, Fortun SM, McLeod BR.

Engineering Research Center, Department of Electrical Engineering, Montana State

University, Bozeman 59717-0378, USA.

Bacterial biofilms are acknowledged to be a major factor in problems of ineffective

sterilization often encountered in clinics, hospitals, and industrial processes. There

have been indications that the addition of a relatively small direct current electric field

with the sterilant used to combat the biofilm greatly increases the efficacy of the

sterilization process. The results of the experiments reported in this paper support the

concept of the "bioelectric effect" as reported by J.W. Costerton, B. Ellis, K. Lam, F.

Johnson, and A.E. Khoury (Antimicrob. Agents Chemother, 38:2803-2809, 1994).

With a current of 1 mA flowing through the chamber containing the biofilm, an

increase in the killing of the bacteria of about 8 log orders was observed at the end of

24 h (compared with the control with the same amount of antibacterial agent but nocurrent). We also confirmed that the current alone does not affect the biofilm and that

there appear to be optimum levels of both the current and the sterilant that are needed

to obtain the maximum effect.

PMID: 8878572 [PubMed - indexed for MEDLINE]

Antimicrob Agents Chemother. 2004 Dec;48(12):4662-4.

A radio frequency electric current enhances antibiotic efficacy

against bacterial biofilms.

Caubet R, Pedarros-Caubet F, Chu M, Freye E, de Belem Rodrigues M, Moreau


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JM, Ellison WJ.

Unite Securite Microbiologique des Aliments, Institut des Sciences et Techniques des

Aliments de Bordeaux, Universite de Bordeaux 1, Talence, France. [email protected]

bordeaux1.fr

Bacterial biofilms are notably resistant to antibiotic prophylaxis. The concentration of

antibiotic necessary to significantly reduce the number of bacteria in the biofilm

matrix can be several hundred times the MIC for the same bacteria in a planktonic

phase. It has been observed that the addition of a weak continuous direct electric

current to the liquid surrounding the biofilm can dramatically increase the efficacy of

the antibiotic. This phenomenon, known as the bioelectric effect, has only been

partially elucidated, and it is not certain that the electrical parameters are optimal. We

confirm here the bioelectric effect for Escherichia coli biofilms treated with

gentamicin and with oxytetracycline, and we report a new bioelectric effect with a

radio frequency alternating electric current (10 MHz) instead of the usual direct

current. None of the proposed explanations (transport of ions within the biofilm,production of additional biocides by electrolysis, etc.) of the direct current bioelectric

effect are applicable to the radio frequency bioelectric effect. We suggest that this new

phenomenon may be due to a specific action of the radio frequency electromagnetic

field upon the polar parts of the molecules forming the biofilm matrix.

PMID: 15561841

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