An Overview of Sub-Cellular Mechanisms Involved in the Action of TTFields

May combine two claims.

Even if the force is essentially unchanged, Debye screening means that less than 5% of the charge remains exposed to the field, resulting in a net force of at most 0.5 pN, most likely insufficient to exert a major influence on the cytoskeleton.

“Even if the force is essentially unchanged, the Debye screening of electrostatic charges means that less than 5% of the charge remains exposed to the field resulting in a net force of at most 0.5 pN, most likely insufficient to exert a major influence on the cytoskeleton.”

May combine two claims.

At longer distances attractive forces prevail and the corresponding dipole-dipole attraction at 90 nm is only 0.08 pN.

“However, at longer distances attractive forces prevail and the corresponding dipole-dipole attraction at 90 nm is only 0.08 pN.”

May combine two claims.

A test charge of +5e a distance of 5 nm from the MT surface for a 10-μm MT experiences a force of 12 pN in water and 1 pN in ionic solution.

“a test charge of +5e a distance of 5 nm from the MT surface for a 10-µm MT, experiences a force of 12 pN in water and 1 pN in ionic solution.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Calculating the force due to a static electric field of 1 V/cm acting on a 10 μm-long microtubule, with q = 10−13 C for unscreened charges, results in F = 10 pN assuming the field is largely undiminished when penetrating a cell.

“calculating the force due to an electric field of a static electric field with a strength of 1 V/cm acting on a 10 µm-long microtubule, we find from F = qE, with q = 10−13 C for unscreened charges, that results in F = 10 pN assuming the field is largely undiminished when penetrating a cell”

• AssumptionThe authors state this as an assumption: The field of 1 V/cm is largely undiminished when penetrating a cell.

May combine two claims.

The noise coefficient is 50.2 m/s when dt ~ 5.0 × 10−13 s, which is much larger than the deterministic term.

“On the other hand, the noise coefficient is 2kBT/γ (dt)−1/2=50.2 m/s when the estimate obtained above is used: dt~5.0×10−13 s, which is much larger than the deterministic term.”

• AssumptionThe authors state this as an assumption: The time step dt can be estimated as dx/vH2O, where dx is the typical separation between two water molecules.
• AssumptionThe authors state this as an assumption: The outcome of a series of collisions with water molecules is approximately Gaussian.

May combine two claims.

The net force on the total charge in the cytoplasm with q = 4 × 10^11 e and E = 1 V/cm is approximately 6 μN, sufficient to cause major perturbations in the cell interior.

“Based on the above, we can estimate the net force on the total charge in the cytoplasm as F = qE, q = 4 × 1011 e and E = 1 V/cm, so the total force is approximately 6 µN, which is sufficient to cause major perturbations in the cell interior.”

May combine two claims.

In an oscillating electric field of amplitude 1 V/cm and frequency f = 200 kHz, an ion's position follows periodic motion x(t) = 0.1·A·sin(2πft), executing harmonic motion out of phase with the field, with the same frequency and an amplitude A approximately 10% of the radius.

“In an oscillating electric field of amplitude 1 V/cm and a frequency f = 200 kHz, an ion’s position will follow periodic motion given by: x(t) = 0.1·A·sin(2πft), i.e., will execute harmonic motion out of phase with the field, with the same frequency and an amplitude A approximately 10% of the radius.”

• AssumptionThe authors state this as an assumption: The friction coefficient for a spherical particle is γ = 6πηr.

May combine two claims.

About 10^7 filter state changes occur during a non-conducting state and about 10^10 switches per second (10 GHz).

“As the duration of the rather (stable) non-conducting state scales in the range 10−3–10−4 s and the within filter translocation time is on the order of 10−11 s, we can expect about 107 filter state changes during a non-conducting state and about 1010 switches per second (10 GHz).”

May combine two claims.

The typical time when particles start to be drifted away because of the electric field is t = 2(kBT)γ/(qE)^2; for a single ion t = 226.1 s, whereas for a typical globular protein t = 0.13 s, which is much larger than the period of an electric field oscillating at hundreds of kHz.

“From the above equation, a typical time when the particles start to be drifted away because of the electric field is t=2(kBT)γ/(qE)2. For a single ion (q=1 e, r=1.8 Å), t=226.1 s, whereas for a typical globular protein (q~100 e, r~1.0 nm), t=0.13 s, which is much larger than the period of an electric field oscillating at hundreds of kHz.”

• AssumptionThe authors state this as an assumption: The electric field is constant E for the ensemble distribution.

May combine two claims.

Including the conductance of both nanopores through an MT surface gives a conductance G0 of a ring as G0 = 10.7 nS and the corresponding resistivity as R = 93 MΩ.

“Including the conductance of both nanopores through an MT surface accounts for the leakage of IC cations into the lumen area and gives a conductance G0, of a ring as G0 = σ1 + σ2 = (2.93 + 7.8) nS = 10.7 nS and the corresponding resistivity as R = 1/G0 = 93 MΩ.”

May combine two claims.

For longitudinal effects, on the order of 50 rings, MTs only 400 nm long would respond to 100 kHz stimulation; for ionic flows pulsating radially around an MT, a 20-μm long MT would be required.

“Therefore, for longitudinal effects, on the order of 50 rings, MTs only 400 nm long would respond to 100 kHz stimulation. On the other hand, for ionic flows pulsating radially around an MT, a 20-µm long MT would be required.”

May combine two claims.

The power dissipated due to a current flowing along a 10 μm-long MT is in the 10−11 W range, comparable to the power generated by the cell in metabolic processes.

“Substituting the relevant numbers we obtain the power dissipated to be in the 10−11 W range which is comparable to the power generated by the cell in metabolic processes (100 W of power generation in the body/3 × 1013 cells in the body).”

May combine two claims.

For a tubulin dimer, CTD = 1.4 × 10−16 F and for an extended TT, CTT = 0.26 × 10−16 F; hence C0 = 1.92 × 10−16 F.

“For a tubulin dimer: CTD = 1.4 × 10−16 F and for an extended TT: CTT = 0.26 × 10−16 F. Hence: (7)C0=C0+2×C0=1.92×10−16 F”

• cites [100] Tuszynski, 2009, A nonlinear model of ionic wave propagation along microtubules, Eur. Biophys. J. sourceNot checked yet

  Not checked in the free run.

May combine two claims.

For a microtubule of length l, R1 scales with length, R2 is length independent, and capacitance scales with length, therefore τ1 scales with length squared while τ2 scales with length.

“Repeating these calculations for a microtubule of length l, we note that R1 scales with length of a microtubule, while R2 is length independent. The corresponding capacitance in both cases scales with length, therefore τ1 scales with length squared (l2) while τ2 scales with length.”

May combine two claims.

The characteristic charging time of an elementary unit capacitor C0 through the resistance R0 is T0 = R0C0 with an estimate for T0 = 1.2 × 10−8 s and the characteristic propagation velocity of the ionic wave v0 = 0.67 m/s.

“The characteristic charging (discharging) time of an elementary unit capacitor C0 through the resistance R0 is given by T0 = R0C0 with an estimate for T0 = 1.2 × 10−8 s and the characteristic propagation velocity of the ionic wave: v=l/T0 as v0 = 0.67 m/s.”

May combine two claims.

A characteristic time for this excitation is 1.2 × 10−5 s whose inverse, the frequency, f, is very close to the TTField value, i.e., 90 kHz.

“Interestingly, a characteristic time for this excitation can readily be estimated as 1.2 × 10−5 s whose inverse, the frequency, f, is very close to the TTField value, i.e., 90 kHz.”

May combine two claims.

A standard travelling-wave soliton preserves its width but its amplitude decays over the length of about 400 units corresponding to 3.2 μm, which is of the order of the MT length.

“A standard travelling-wave with speed v, for the normalized function u(x, t), can be used as a solution of the propagation equation, which is a soliton that preserves its width but its amplitude decays over the length of about 400 units corresponding to 3.2 µm, which is of the order of the MT length.”

May combine two claims.

The decay times are τ1 = 10−8 s along the MT length and τ2 = 10−9 s away from the MT surface.

“We obtain for decay times (τ = RC) the following values: (a) τ1 = 10−8 s along the MT length and (b) τ2 = 10−9 s away from the MT surface.”

May combine two claims.

For a single actin monomer, the time scale for LC oscillations is τ0 = 6 × 10−14 s; the decay time for longitudinal ionic waves is τ1 = 6 × 10−10 s while the corresponding time for radial waves is τ2 = 0.9 × 10−10 s.

“We readily find for a single actin monomer, that the time scale for LC oscillations is very fast, namely τ0 = (LC)1/2 and τ0 = 6 × 10−14 s. Secondly, the decay time for longitudinal ionic waves is τ1 = R1C = 6 × 10−10 s while the corresponding time for radial waves is τ2 = R2C = 0.9 × 10−10 s.”

May combine two claims.

For a 1-μm length of an AF, Reff = 1.2 × 109 Ω, Leff = 340 × 10−12 H and Ceff = 0.02 × 10−12 F.

“Hence, for a 1-µm length of an AF we find that Reff = 1.2 × 109 Ω, Leff = 340 × 10−12 H and Ceff = 0.02 × 10−12 F.”

May combine two claims.

For a 1-μm AF, τ0 = 10−11 s, which is still too short but τ1 = R1C = 2.4 × 10−5 s which is in the correct range of time for interactions with AC electric fields in the 100 kHz range.

“Taking as an example a 1-µm AF, we find τ0 = 10−11 s, which is still too short but τ1 = R1C = 2.4 × 10−5 s which is in the correct range of time for interactions with AC electric fields in the 100 kHz range.”

May combine two claims.

Treated cells remain stationary at metaphase for several hours, accompanied by abnormal mitotic figures as well as membrane rupture and blebbing leading to apoptosis.

“A significant prolongation of mitosis was predicted, where treated cells remain stationary at metaphase for several hours, which was accompanied by abnormal mitotic figures as well as membrane rupture and blebbing leading to apoptosis [2,3].”

• cites [2] Kirson, 2004, Disruption of cancer cell replication by alternating electric fields, Cancer Res. sourceNot checked yet

  Not checked in the free run.

• cites [3] Kirson, 2007, Alternating electric fields arrest cell proliferation in animal tumor models and human brain tumors, Proc. Natl. Acad. Sci. USA sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing).

A force of 1 pN applied to a tip of a microtubule may be used to bend it by as much as 1 μm.

“A force of 1 pN applied to a tip of a microtubule may be used to bend it by as much as 1 µm [24].”

• cites [24] Howard, J. (2001). Mechanics of Motor Proteins and the Cytoskeleton, Sinauer Associates.Not checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

A minimal amount of useful force at a nanometer scale is 1 pN.

“a minimal amount of useful force at a nanometer scale is 1 pN.”

May combine two claims.

For low frequencies, the intracellular space is shielded to a large degree from extracellular electric fields; the electric field strength inside a typical spherical cell is approximately five orders of magnitude lower than that outside the cell.

“For low frequencies, the intracellular space is shielded to a large degree from extracellular electric fields. For example, the electric field strength inside a typical spherical cell is approximately five orders of magnitude lower than that outside the cell [35,36].”

• cites [35] Hobbie, R.K., and Roth, B.J. (2007). Intermediate Physics for Medicine and Biology, Springer. [4th ed.].Not checked yet

  Not checked in the free run.

• cites [36] King, 1998, Electric field induced in cells in the human body when this is exposed to low-frequency electric fields, Phys. Rev. E Stat. Nonlinear Soft Matter Phys. sourceNot checked yet

  Not checked in the free run.

May combine two claims.

TTFields were approved by the FDA in 2011 for the treatment of recurrent glioblastoma multiforme and in late 2015 for newly diagnosed GBM in combination therapy with temozolomide.

“TTFields have been approved by the United States Food and Drug Administration (FDA) in 2011 for the treatment of recurrent glioblastoma multiforme (GBM)”

• Also stated in: § 1. Introduction · plain

  “In late 2015, TTFields were also approved for newly diagnosed GBM patients in combination therapy with temozolomide [7,8] due to significantly increased survival times.”

• cites [5] Stupp, 2012, NovoTTF-100A versus physician’s choice chemotherapy in recurrent glioblastoma: A randomised phase III trial of a novel treatment modality, Eur. J. Cancer sourceNot checked yet

  Not checked in the free run.

• cites [7] Stupp, 2015, Maintenance therapy with tumor-treating fields plus temozolomide vs. temozolomide alone for glioblastoma: A randomized clinical trial, JAMA sourceNot checked yet

  Not checked in the free run.

• cites [8] Kirson, 2009, Chemotherapeutic treatment efficacy and sensitivity are increased by adjuvant alternating electric fields (TTFields), BMC Med. Phys. sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

Motor proteins generate forces on the order of several pN.

“Motor proteins generate forces on the order of several pN.”

May combine two claims.

TTFields are clinically applied to disrupt cell division, primarily for the treatment of glioblastoma multiforme.

“TTFields are clinically applied to disrupt cell division, primarily for the treatment of glioblastoma multiforme (GBM).”

• cites [5] Stupp, 2012, NovoTTF-100A versus physician’s choice chemotherapy in recurrent glioblastoma: A randomised phase III trial of a novel treatment modality, Eur. J. Cancer sourceNot checked yet

  Not checked in the free run.

• cites [7] Stupp, 2015, Maintenance therapy with tumor-treating fields plus temozolomide vs. temozolomide alone for glioblastoma: A randomized clinical trial, JAMA sourceNot checked yet

  Not checked in the free run.

• cites [8] Kirson, 2009, Chemotherapeutic treatment efficacy and sensitivity are increased by adjuvant alternating electric fields (TTFields), BMC Med. Phys. sourceNot checked yet

  Not checked in the free run.

May combine two claims.

The inhibitory effect of TTFields increases with increasing electric field intensity, resulting in complete proliferation arrest of rat glioma cells after 24 h exposure to a field intensity of 2.25 V/cm.

“the inhibitory effect increases with an increasing electric field intensity, resulting in a complete proliferation arrest of rat glioma cells after 24 h exposure to a field intensity of 2.25 V/cm.”

• cites [2] Kirson, 2004, Disruption of cancer cell replication by alternating electric fields, Cancer Res. sourceNot checked yet

  Not checked in the free run.

• cites [3] Kirson, 2007, Alternating electric fields arrest cell proliferation in animal tumor models and human brain tumors, Proc. Natl. Acad. Sci. USA sourceNot checked yet

  Not checked in the free run.

May combine two claims.

The induced electric field inside a spherical cell is uniform, whereas increasing non-uniformities are predicted for deviations of the regular shape.

“the induced electric field inside a spherical cell is uniform, whereas increasing non-uniformities are predicted for deviations of the regular shape.”

• cites [27] Bernhardt, 1973, On the generation of potential differences across the membranes of ellipsoidal cells in an alternating electrical field, Biophysik sourceNot checked yet

  Not checked in the free run.

• cites [28] Gimsa, 1999, A polarization model overcoming the geometric restrictions of the laplace solution for spheroidal cells: Obtaining new equations for field-induced forces and transmembrane potential, Biophys. J. sourceNot checked yet

  Not checked in the free run.

• cites [29] Gimsa, 2001, Analytical description of the transmembrane voltage induced on arbitrarily oriented ellipsoidal and cylindrical cells, Biophys. J. sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S4 (based).

An effect must not produce so much thermal energy as to seriously increase the temperature of the cell.

“Secondly, it must not produce so much thermal energy as to seriously increase the temperature of the cell.”

May combine two claims.

The cellular morphology during cytokinesis gives rise to a non-uniform intracellular electric field with a high density at the cleavage furrow, leading to dielectrophoretic forces acting on polarizable macromolecules.

“the cellular morphology during cytokinesis gives rise to a non-uniform intracellular electric field, with a high density at the cleavage furrow between the dividing cells. This non-uniform field leads to the development of dielectrophoretic (DEP) forces [17] acting on polarizable macromolecules”

• cites [17] Pohl, H.A. (1978). Dielectrophoresis, Cambridge University Press.Not checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S4 (based).

For an effect to be of significance at a molecular level, its interaction energy must exceed thermal energy, i.e., kT per degree of freedom (4 × 10−21 J).

“for an effect to be of significance at a molecular level, its interaction energy must exceed thermal energy, i.e., kT per degree of freedom (i.e., 4 × 10−21 J).”

May combine two claims.

The net charge on a tubulin dimer depends on pH and changes from +5 at pH 4.5 to 0 at pH 5 and drops to −30 at pH 8.

“The net charge on a tubulin dimer depends on pH and changes from +5 at pH 4.5 to 0 at pH 5 and drops to −30 at pH 8 [68].”

• cites [68] Brown, 2005, Molecular dynamics simulations of tubulin structure and calculations of electrostatic properties of microtubules, Math. Comput. Model. sourceNot checked yet

  Not checked in the free run.

May combine two claims.

For a spherical cell during metaphase, the modeling results predict that for frequencies lower than 10 kHz only small changes of the field are detected and the intracellular field strength Ei almost equals zero; a first significant increase of Ei is observed at approximately 200 kHz, and Ei increases rapidly as the frequencies increase above this value.

“For a spherical cell during metaphase, the modeling results predict that for frequencies lower than 10 kHz only small changes of the field are detected and the intracellular field strength, Ei, almost equals zero. A first significant increase of Ei is observed at approximately 200 kHz, and Ei increases rapidly as the frequencies increase above this value.”

• cites [37] Wenger, C., Giladi, M., Bomzon, Z., Salvador, R., Basser, P.J., and Miranda, P.C. (2015, January 25–29). Modeling Tumor Treating Fields (TTFields) application in single cells during metaphase and telophase. In Proceedings of the 2015 37th Annual International Conference of the IEEE Engineering in Medicine and Biology Society (EMBC), Milan, Italy. sourceNot checked yet

  Not checked in the free run.

• AssumptionRests on an assumption: The computational model’s predictions for isolated cells apply to cells in tissue during TTFields treatment.

May combine two claims.

Within the dividing cell the intracellular electric field distribution is non-uniform with highest field intensities at the cleavage plane; these maximum intensities are much higher than the applied field and appear for frequencies in the range 100-500 kHz depending on the stage of cytokinesis.

“The model further showed that within the dividing cell the intracellular electric field distribution is non-uniform with highest field intensities at the cleavage plane (black lines in the left panel of Figure 1). These maximum intensities are much higher than the applied field and appear for frequencies in the range 100–500 kHz depending on the stage of cytokinesis”

• cites [37] Wenger, C., Giladi, M., Bomzon, Z., Salvador, R., Basser, P.J., and Miranda, P.C. (2015, January 25–29). Modeling Tumor Treating Fields (TTFields) application in single cells during metaphase and telophase. In Proceedings of the 2015 37th Annual International Conference of the IEEE Engineering in Medicine and Biology Society (EMBC), Milan, Italy. sourceNot checked yet

  Not checked in the free run.

• AssumptionRests on an assumption: The computational model’s predictions for isolated cells apply to cells in tissue during TTFields treatment.

May combine two claims.

Frequency-, cell size-, and field-intensity dependences were confirmed.

“Frequency-, cell size-, and field-intensity dependences were confirmed.”

• cites [37] Wenger, C., Giladi, M., Bomzon, Z., Salvador, R., Basser, P.J., and Miranda, P.C. (2015, January 25–29). Modeling Tumor Treating Fields (TTFields) application in single cells during metaphase and telophase. In Proceedings of the 2015 37th Annual International Conference of the IEEE Engineering in Medicine and Biology Society (EMBC), Milan, Italy. sourceNot checked yet

  Not checked in the free run.

• AssumptionRests on an assumption: The computational model’s predictions for isolated cells apply to cells in tissue during TTFields treatment.

May combine two claims.

Above 1 MHz electric current is shunted across the membrane and the impedance is dominated by the cytoplasm; at 1 GHz the cellular structure becomes electrically invisible.

“above 1 MHz electric current is shunted across the membrane and the impedance is dominated by the cytoplasm. Thus, for an increasing frequency, the electric field inside the cell is augmented and at 1 GHz the cellular structure becomes ‘‘electrically invisible’’ as previously reported [33].”

• cites [33] Stewart, 2005, Cylindrical cell membranes in uniform applied electric fields: Validation of a transport lattice method, IEEE Trans. Biomed. Eng. sourceNot checked yet

  Not checked in the free run.

May combine two claims.

During metaphase a uniform non-zero Ei is induced; during cytokinesis, a non-uniform Ei is induced with a substantially increased strength at the cleavage furrow.

“During metaphase a uniform non-zero Ei is induced. Depending on cell properties, the frequency window of the predicted transition range might be shifted. During cytokinesis, a non-uniform Ei is induced with a substantially increased strength at the cleavage furrow.”

• cites [37] Wenger, C., Giladi, M., Bomzon, Z., Salvador, R., Basser, P.J., and Miranda, P.C. (2015, January 25–29). Modeling Tumor Treating Fields (TTFields) application in single cells during metaphase and telophase. In Proceedings of the 2015 37th Annual International Conference of the IEEE Engineering in Medicine and Biology Society (EMBC), Milan, Italy. sourceNot checked yet

  Not checked in the free run.

• AssumptionRests on an assumption: The computational model’s predictions for isolated cells apply to cells in tissue during TTFields treatment.

May combine two claims.

This interaction energy is 4000 times smaller than thermal energy kT, too small to affect the dynamics of an individual tubulin dimer.

“This is clearly too small (4000 times smaller than thermal energy kT) to affect the dynamics of an individual tubulin dimer.”

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

MTs have also been theorized to have special conducting properties involving the lumen but there has been no direct experimental determination of the electric properties of the MT lumen.

“MTs have also been theorized to have special conducting properties involving the lumen [55] but there has been no direct experimental determination of the electric properties of the MT lumen.”

• cites [55] Sahu, 2013, Atomic water channel controlling remarkable properties of a single brain microtubule: Correlating single protein to its supramolecular assembly, Biosens. Bioelectron. sourceNot checked yet

  Not checked in the free run.

May combine two claims.

There is a small non-cancellation effect along the MT axis but this amounts to less than 10% of the next dipole moment, hence it is doubtful that an entire MT can be aligned in electric fields with intensities lower than 10 V/cm.

“There is a small non-cancellation effect along the MT axis but this amounts to less than 10% of the next dipole moment, hence it is doubtful that an entire MT can be aligned in electric fields with intensities lower than 10 V/cm.”

May combine two claims.

MTs have been predicted to exhibit intrinsic electronic conductivity as well as ionic conductivity along their length.

“MTs have been predicted to exhibit intrinsic electronic conductivity as well as ionic conductivity along their length [72].”

• cites [72] Lyshevski, S.E. (2007). Nano and Molecular Electronics Handbook, CRC Press. sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Due to the inhomogeneity in the electric field distribution, significant dielectrophoretic forces are expected to develop within the cell and these DEP forces are thought to be important factors in the mechanism of action of TTFields.

“Due to the inhomogeneity in the electric field distribution, significant dielectrophoretic (DEP) forces are expected to develop within the cell and these DEP forces are thought to be important factors in the mechanism of action of TTFields [41].”

• cites [41] Giladi, 2008, Microbial growth inhibition by alternating electric fields, Antimicrob. Agents Chemother. sourceNot checked yet

  Not checked in the free run.

• AssumptionRests on an assumption: The computational model’s predictions for isolated cells apply to cells in tissue during TTFields treatment.

May combine two claims.

MTs have a highly electro-negatively charged outer surface as well as C-terminal tails, resulting in a cloud of counter-ions surrounding them.

“MTs have a highly electro-negatively charged outer surface as well as C-terminal tails (TTs), resulting in a cloud of counter-ions surrounding them.”

May combine two claims.

Uppalapati et al. work covers the range of frequencies overlapping with TTFields, although the amplitudes are much larger due to the voltage bias of 40 V across a 20-μm gap giving an electric field of 2 × 104 V/cm as opposed to 1 V/cm.

“The work of Uppalapati et al. [79] covers the range of frequencies overlapping with TTFields, although the amplitudes are much larger due to the voltage bias of 40 V across a 20-µm gap giving an electric field of 2 × 104 V/cm as opposed to 1 V/cm).”

• cites [79] Uppalapati, 2008, Microtubule alignment and manipulation using AC electrokinetics, Small sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Above f = 5 MHz, electro-hydrodynamic flows were virtually eliminated, and the conductivity of MTs was estimated at 0.25 S/m.

“above f = 5 MHz, electro-hydrodynamic flows were virtually eliminated, and the conductivity of MTs was estimated at 0.25 S/m.”

• cites [79] Uppalapati, 2008, Microtubule alignment and manipulation using AC electrokinetics, Small sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing).

Ion transition occurs through a sequence of stable multi-ion configurations through the filter region of the channels, which allows rapid and ion-selective conduction.

“Ion transition occurs through a sequence of stable multi-ion configurations through the filter region of the channels, which allows rapid and ion-selective conduction [85].”

• cites [85] Roux, 2001, Energetics of ion conduction through the K+ channel, Nature sourceNot checked yet

  Not checked in the free run.

May combine two claims.

The electric field across the membrane is on the order of 105 V/cm, which is 4-5 orders of magnitude greater than TTFields' amplitude.

“The electric field across the membrane is on the order of 105 V/cm (0.1 V over 8–10 nm), which is 4–5 orders of magnitude greater than TTFields’ amplitude.”

May combine two claims.

Time for translocations between two filter sites separated by ~0.3 nm is 5 × 10−10 s and 5 × 10−11 s.

“Time scales for the processes in ion channels can be estimated by the time for translocations (ttr) between two filter sites separated by ~0.3 nm, i.e., 5 × 10−10 s [88] and 5 × 10−11 s [87].”

• cites [88] Shrivastava, 2002, K+ versus Na+ ions in a K channels selectivity filter: A simulation study, Biophys. J. sourceNot checked yet

  Not checked in the free run.

• cites [87] Guidoni, 2002, Potassium permeation through the KcsA channel: A density functional study, Biochim. Biophys. Acta sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Below 500 kHz, MTs flow toward the centerline of electrodes; the electro-osmotic force causes the movement of the fluid in a vortex-like manner.

“Below 500 kHz, MTs flow toward the centerline of electrodes. The electro-osmotic force causes the movement of the fluid in a vortex-like manner.”

• cites [79] Uppalapati, 2008, Microtubule alignment and manipulation using AC electrokinetics, Small sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Even with less frequent Brownian collisions, e.g., dt ~ 10−6 s, the noise coefficient is 0.035 m/s, still much larger than the coupling with the electric field, meaning that an electric field of amplitude 1 V/cm has an exceedingly small probability to influence the diffusion of a single Brownian particle even if the net charge q is 100-1000 times larger as in the case of a protein.

“Even in the case of less frequent Brownian collisions, e.g., dt~10−6 s, the noise coefficient is 0.035 m/s which is still much larger than the coupling with the electric field, meaning that an electric field of amplitude 1 V/cm has an exceedingly small probability to influence the diffusion of a single Brownian particle even if the net charge q is 100–1000 times larger as in the case of a protein.”

• AssumptionThe authors state this as an assumption: There is no interaction other than the coupling with an external electric field E(Xt) for the particle.

May combine two claims.

MTs exposed to low frequency AC fields (f < 10 kHz) exhibit a flow motion due to ionic convection; for frequencies above 10 kHz this convection effect is absent.

“MTs exposed to low frequency AC fields (f < 10 kHz) exhibit a flow motion due to ionic convection. However, for frequencies above 10 kHz this convection effect is absent.”

• cites [77] Minoura, 2006, Dielectric measurement of individual microtubules using the electroorientation method, Biophys. J. sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Liu et al. reported activation of a Na+ pumping mode with an oscillating electric field with a strength of 20 V/cm, which is comparable to the fields of interest in this review, but at a much higher frequency (1.0 MHz) than those of interest.

“Liu et al. [80] reported activation of a Na+ pumping mode with an oscillating electric field with a strength of 20 V/cm, which is comparable to the fields of interest in this review, but at a much higher frequency (1.0 MHz) than those of interest.”

• cites [80] Liu, 1990, Activation of Na+ and K+ pumping modes of (Na, K)-ATPase by an oscillating electric field, J. Biol. Chem. sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Uppalapati et al. exposed taxol-stabilized MTs in solution to an AC field, which exhibited electro-osmotic and electro-thermal flow, in addition to MT dielectrophoresis effects.

“Uppalapati et al. [79] exposed taxol-stabilized MT’s in solution to an AC field, which exhibited electro-osmotic and electro-thermal flow, in addition to MT dielectrophoresis effects.”

• cites [79] Uppalapati, 2008, Microtubule alignment and manipulation using AC electrokinetics, Small sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Lowering the pH of the solution to the isoelectric point of MTs around pH 5 should substantially reduce the DEP effect and additionally lowering the frequency will reduce it further.

“We predict that lowering the pH of the solution to the isoelectric point of MTs around pH 5 should substantially reduce this effect and additionally lowering the frequency will reduce it further due to the dependence of the first term on the square of the frequency.”

Probably not a substantial claim: may not pass S2 (checkable), S3 (load-bearing).

The multiple mechanisms of MT conductance provide ample possibility to explain the varied reports on MT conductivity in the literature.

“The multiple mechanisms of MT conductance provide ample possibility to explain the varied reports on MT conductivity in the literature.”

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

The ability of the electric field to penetrate into the cell's interior is strongly depended on cell shape, being easier in non-spherical cells.

“As discussed above, this is strongly depended on the ability of the electric field to penetrate into the cell’s interior, which is easier in the case of non-spherical cells.”

May combine two claims.

Stracke et al. applied electric fields in the range of 3 V/cm to suspended MTs, which moved at pH 6.8 from the negative electrode to the positive one indicating a negative net charge, and an electrophoretic mobility of about 2.6 × 10−4 cm2·V−1·s−1.

“Electric fields in the range of 3 V/cm were applied by Stracke et al. [75] to suspended MTs, which moved at pH 6.8 from the negative electrode to the positive one indicating a negative net charge, and an electrophoretic mobility of about 2.6 × 10−4 cm2·V−1·s−1.”

• cites [75] Stracke, 2002, Analysis of the migration behaviour of single microtubules in electric fields, Biochem. Biophys. Res. Commun. sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing).

In cell division, coherent polarization waves have been implicated as playing the key role in chromosome alignment and their subsequent separation.

“In cell division, coherent polarization waves have been implicated as playing the key role in chromosome alignment and their subsequent separation [18,19].”

• cites [18] Cooper, 1981, Coherent polarization waves in cell division and cancer, Collect. Phenom.Not checked yet

  Not checked in the free run.

• cites [19] Pohl, 1981, Life cycle alterations of the micro-dielectrophoretic effects of cell, J. Biol. Phys. sourceNot checked yet

  Not checked in the free run.

May combine two claims.

van den Heuvel et al. with electric field strengths of 40 V/cm yielded MT electrophoretic mobility in the range of 2.6 × 10−8 m2·V−1·s−1, in line with previous reports, and found the effective charge of a tubulin dimer to be approximately 23 e−.

“Electrophoresis experiments were also performed by van den Heuvel et al. [96], with electric field strengths of 40 V/cm, yielding MT electrophoretic mobility in the range of 2.6 × 10−8 m2·V−1·s−1, in line with previous reports. They found the effective charge of a tubulin dimer to be approximately 23 e−.”

• cites [96] Lemay, 2007, Electrophoresis of individual microtubules in microchannels, Proc. Natl. Acad. Sci. USA sourceNot checked yet

  Not checked in the free run.

May combine two claims.

For a single dimer, C = 6.6 × 10−16 F, R1 = 6 × 106 Ω (along the MT), R2 = 1.2 × 106 Ω (perpendicular to the MT) and L = 2 × 10−12 H.

“For a single dimer: C = 6.6 × 10−16 F, R1 = 6 × 106 Ω (along the MT), R2 = 1.2 × 106 Ω (perpendicular to the MT) and L = 2 × 10−12 H.”

• cites [60] Priel, 2006, A biopolymer transistor: Electrical amplification by microtubules, Biophys. J. sourceNot checked yet

  Not checked in the free run.

May combine two claims.

These frequencies are much too high to be affected by TTFields; therefore, we do not expect TTFields to be capable of disrupting the MT structure.

“Again, these frequencies are much too high to be affected by TTFields. Therefore, we do not expect TTFields to be capable of disrupting the MT structure.”

Classified as probably not a substantial claim. May combine two claims.

Preto et al. concluded that classical electromagnetic dipole-dipole interactions at high enough frequencies can lead to attraction between oscillating dipoles over distances comparable to the size of the cell; even including a coherently coupled layer of water molecules around a protein, this would require frequencies in the THz range or higher.

“Finally, Preto et al. [92] re-evaluated the Froehlich mechanism for long-range interactions and concluded that classical electromagnetic dipole-dipole interactions at high enough frequencies can lead to attraction between oscillating dipoles over distances comparable to the size of the cell. However, even including a coherently coupled layer of water molecules around a protein, this would require”

• cites [92] Preto, 2012, Experimental assessment of the contribution of electrodynamic interactions to long-distance recruitment of biomolecular partners: Theoretical basis, Phys. Rev. E Stat. Nonlinear Soft Matter Phys. sourceNot checked yet

  Not checked in the free run.

Classified as probably not a substantial claim.

Counter-ion condensation occurs when the average distance between charges, b, is such that λB/b = S > 1.

“Counter-ion condensation occurs when the average distance between charges, b, is such that λB/b = S > 1.”

• cites [98] Manning, 1978, The molecular theory of polyelectrolyte solutions with applications to the electrostatic properties of polynucleotides, Q. Rev. Biophys. sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

They proposed the resonant recognition model (RRM) based on the distribution of energy of delocalized proteins and charge transfer under resonance with a velocity of 7.87 × 105 m/s and covering distances of 3.8 Å between amino acids, giving a characteristic frequency between 1013 and 1015 Hz.

“They proposed the so-called resonant recognition model (RRM) based on the distribution of energy of delocalized proteins in a biological system and charge transfer under resonance with a velocity of 7.87 × 105 m/s and covering distances of 3.8 Å between amino acids, giving a characteristic frequency between 1013 and 1015 Hz.”

• cites [103] Cosic, 2014, Prediction of Tubulin resonant frequencies using the Resonant Recognition Model (RRM), IEEE Trans Nanobiosci. sourceNot checked yet

  Not checked in the free run.

• cites [104] Cosic, 2015, Is it possible to predict electromagnetic resonances in proteins, DNA and RNA?, EPJ Nonlinear Biomed. Phys. sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Isozaki et al. found for control MTs that the electrophoretic mobility is approximately 2 × 10−8 m2·V−1·s−1, which is consistent with Stracke et al.

“It was found for control MTs that the electrophoretic mobility is approximately: 2 × 10−8 m2·V−1·s−1 which is consistent with Stracke et al. [75].”

• cites [95] Isozaki, 2015, Control of microtubule trajectory within an electric field by altering surface charge density, Sci. Rep. sourceNot checked yet

  Not checked in the free run.

• cites [75] Stracke, 2002, Analysis of the migration behaviour of single microtubules in electric fields, Biochem. Biophys. Res. Commun. sourceNot checked yet

  Not checked in the free run.

May combine two claims.

This effect can cause MT instability as well as interference with motor protein transport.

“This effect can cause MT instability as well as interference with motor protein transport as discussed below.”

May combine two claims.

Short MTs would be more sensitive to the longitudinal wave generation by TTFields while long MTs should lead to perpendicular wave generation.

“It appears that short MTs would be more sensitive to the longitudinal wave generation by TTFields while long MTs should lead to perpendicular wave generation.”

May combine two claims.

Additional heat generated by these processes may be disruptive to living cells although there is no experimentally detected thermal effect of TTFields.

“Consequently, additional heat generated by these processes may be disruptive to living cells although there is no experimentally detected thermal effect of TTFields.”

May combine two claims.

C-termini being very flexible and highly charged (with approximately 40% of the tubulin's charge located there) are likely to dynamically respond to electric fields as local changes of pH are correlated with positive and negative electric field's polarities.

“However, C-termini being very flexible and highly charged (with approximately 40% of the tubulin’s charge located there) are likely to dynamically respond to electric fields as local changes of pH are correlated with positive and negative electric field’s polarities, respectively.”

Classified as probably not a substantial claim.

Manning postulated that polyelectrolytes may have condensed ions in their surroundings if a sufficiently high linear charge density is present on the polymer's surface.

“Manning [98] postulated that polyelectrolytes may have condensed ions in their surroundings if a sufficiently high linear charge density is present on the polymer’s surface [99].”

• cites [98] Manning, 1978, The molecular theory of polyelectrolyte solutions with applications to the electrostatic properties of polynucleotides, Q. Rev. Biophys. sourceNot checked yet

  Not checked in the free run.

• cites [99] Zimm, 1984, Distribution of counterions around a cylindrical polyelectrolyte and Manning’s condensation theory, Biopolymers sourceNot checked yet

  Not checked in the free run.

May combine two claims.

For tubulin and MTs, three specific ranges of resonant frequencies have been predicted by the RRM approach: 97-101 THz, 340-350 THz and 445-470 THz, none of which overlaps with TTField frequencies.

“For tubulin and MTs, three specific ranges of resonant frequencies have been predicted by the RRM approach: 97–101 THz, 340–350 THz and 445–470 THz, none of which overlaps with TTField frequencies.”

• cites [103] Cosic, 2014, Prediction of Tubulin resonant frequencies using the Resonant Recognition Model (RRM), IEEE Trans Nanobiosci. sourceNot checked yet

  Not checked in the free run.

• cites [104] Cosic, 2015, Is it possible to predict electromagnetic resonances in proteins, DNA and RNA?, EPJ Nonlinear Biomed. Phys. sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Metabolic oscillations in cells with a period of approximately 10 to 12 s were measured in vivo, which is many orders of magnitude slower than any AC electric field effects discussed here.

“It is also worth mentioning that recently metabolic oscillations in cells with a period of approximately 10 to 12 s, were measured in vivo [102] which is many orders of magnitude slower than any AC electric field effects discussed here.”

• cites [102] Chen, 2014, In vivo tissue-wide synchronization of mitochondrial metabolic oscillations, Cell Rep. sourceNot checked yet

  Not checked in the free run.

May combine two claims.

A typical time scale for C-termini motion is 100 ps, which is too fast for the 100 kHz frequency range of TTFields.

“A typical time scale for C-termini motion is 100 ps, which is too fast for the 100 kHz frequency range of TTFields.”

May combine two claims.

Isozaki et al. estimated the effective charges of the TAMRA- and AlexaFluor 488-tagged tubulin dimer as 10 e− and 9.7 e−, approximately 20%-30% of the vacuum values but much larger than earlier experimental estimates.

“Importantly, they estimated the effective charges of the TAMRA- and AlexaFluor 488-tagged tubulin dimer as 10 e− and 9.7 e−, which obviously is only a fraction (approximately 20%–30%) of the vacuum values but much larger than earlier experimental estimates.”

• cites [95] Isozaki, 2015, Control of microtubule trajectory within an electric field by altering surface charge density, Sci. Rep. sourceNot checked yet

  Not checked in the free run.

May combine two claims.

The velocity v = 0.0005 m/s produces EMF in the range of 108-325 kHz for TERT, TERT mRNA and Telomere; this velocity corresponds to the propagation of solitons on α-helices.

“Of particular interest to this review is the velocity v = 0.0005 m/s which produces EMF in the range of 108–325 kHz for TERT, TERT mRNA and Telomere. This velocity corresponds the propagation of solitons on α-helices.”

• cites [103] Cosic, 2014, Prediction of Tubulin resonant frequencies using the Resonant Recognition Model (RRM), IEEE Trans Nanobiosci. sourceNot checked yet

  Not checked in the free run.

• cites [104] Cosic, 2015, Is it possible to predict electromagnetic resonances in proteins, DNA and RNA?, EPJ Nonlinear Biomed. Phys. sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing).

Cosic et al. reported EM resonances in biological molecules (proteins, DNA and RNA) in THz, GHz, MHz and kHz ranges.

“Cosic et al. [103,104] reported EM resonances in biological molecules (proteins, DNA and RNA) in THz, GHz, MHz and kHz ranges.”

• cites [103] Cosic, 2014, Prediction of Tubulin resonant frequencies using the Resonant Recognition Model (RRM), IEEE Trans Nanobiosci. sourceNot checked yet

  Not checked in the free run.

• cites [104] Cosic, 2015, Is it possible to predict electromagnetic resonances in proteins, DNA and RNA?, EPJ Nonlinear Biomed. Phys. sourceNot checked yet

  Not checked in the free run.

May combine two claims.

Endogenous current densities in cells range from 0.2 to 60 μA/cm2, translating into 0.002 < j < 0.6 A/m2.

“Current densities should also be briefly discussed in relation to previously reported endogenous current densities, j, in cells, which range from 0.2 to 60 µA/ cm2 [101]. This translates into 0.002 < j < 0.6 A/m2.”

• cites [101] Ussing, H.H., and Thorn, N.A. (1973). Transport Mechanisms in Epithelia, Academic Press.Not checked yet

  Not checked in the free run.

May combine two claims.

The oscillation times are too short for potential effects with 100 kHz-range fields (the time of TTFields oscillations is on the order of 5-10 μs).

“Clearly, the oscillation times are too short for potential effects with 100 kHz-range fields (the time of TTFields oscillations is on the order of 5–10 µs).”

May combine two claims.

The condensate thickness λ for the tubulin dimer (λTD) and C-termini (λTT) is λTD = 2.5 nm and λTT = 1.1 nm.

“An estimate of the respective condensate thickness λ of the counter-ion sheath for the tubulin dimer (λTD) and C-termini (λTT) is λTD = 2.5 nm and λTT = 1.1 nm, as analyzed in [61].”

• cites [61] 2011, Nonlinear ionic pulses along microtubules, Eur. Phys. J. E Soft Matter sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

TTFields are not expected to affect permanent dipoles of proteins such as tubulin and actin.

“These fields are not expected to affect permanent dipoles of proteins such as tubulin and actin.”

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

TTField-generated forces on cytoplasmic organelles may reach up to 60 pN, causing their movement toward the cleavage furrow.

“As a consequence, the TTField-generated forces acting on these organelles may reach values up to 60 pN resulting in their movement toward the cleavage furrow.”

• cites [2] Kirson, 2004, Disruption of cancer cell replication by alternating electric fields, Cancer Res. sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

AC-field-generated oscillating ionic flows may in general affect the net charge of the cytoplasm.

“May in general affect the net charge of the cytoplasm.”

Probably not a substantial claim: may not pass S4 (based).

Depending on field orientation relative to the microtubule axis, three types of ionic waves can be generated: longitudinal, helical, and radial.

“Depending on the orientation of the electric fields with the cell axis and in particular with the MT axis (however, they fan out from centrosomes in mitotic cells, so there will be at different angles to any field), there could in general be three types of ionic waves generated: Longitudinal waves propagating along the MT surface.”

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

AFs are approximately 7 nm in diameter, with a periodic helical structure repeating every 37 nm.

“AFs are approximately 7 nm in diameter, with a periodic helical structure repeating every 37 nm.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Cytoplasmic organelles are electrically polarized by TTFields within dividing cells.

“It was determined that cytoplasmic organelles are electrically polarized by the field within dividing cells.”

• cites [2] Kirson, 2004, Disruption of cancer cell replication by alternating electric fields, Cancer Res. sourceNot checked yet

  Not checked in the free run.

May combine two claims.

AFs are characterized by a high electrostatic charge density resulting in ionic current conductivity involving the counter-ions surrounding them, which is very similar to the effects observed for MTs.

“They are characterized by a high electrostatic charge density [108,109] resulting in ionic current conductivity involving the counter-ions surrounding them [109], which is very similar to the effects observed for MTs [60].”

• cites [108] Cantiello, 1991, Osmotically induced electrical signals from actin filaments, Biophys. J. sourceNot checked yet

  Not checked in the free run.

• cites [109] Lin, 1993, A novel method to study the electrodynamic behavior of actin filaments. Evidence for cable-like properties of actin, Biophys. J. sourceNot checked yet

  Not checked in the free run.

• cites [60] Priel, 2006, A biopolymer transistor: Electrical amplification by microtubules, Biophys. J. sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

AC-field-generated oscillating ionic flows may to a lesser degree affect ion channel dynamics.

“May to a lesser degree affect ion channel dynamics.”

May combine two claims.

For an AF with n monomers, R1,i = 6.11 × 106 Ω, and R2,i = 0.9 × 106 Ω, such that R1,i = 7R2,i.

“where R1,i = 6.11 × 106 Ω, and R2,i = 0.9 × 106 Ω, such that R1,i = 7R2,i [112].”

• cites [112] Portet, 2004, Ionic wave propagation along actin filaments, Biophys. J. sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Cytoplasmic organelles can move at velocities up to 30 μm/s and could pile up at the cleavage furrow within a few minutes, interfering with cytokinesis and potentially leading to cell destruction.

“These organelles can move at velocities up to 30 μm/s and, as a result, they could pile up at the cleavage furrow within a few minutes, interfering with cytokinesis, which may lead to cell destruction.”

• cites [2] Kirson, 2004, Disruption of cancer cell replication by alternating electric fields, Cancer Res. sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing).

If a field is oriented at an angle to the microtubule axis, all three wave types may be generated simultaneously.

“If a field is oriented at an angle to the MT axis, it is expected that all these wave types may be generated simultaneously.”

May combine two claims.

In experiments, an input voltage pulse was applied with an amplitude of 200 mV for a duration of 800 ms; electrical signals measured at the opposite end demonstrated that AFs support axial non-linear ionic currents.

“In these experiments [30,42], an input voltage pulse was applied with an amplitude of 200 mV for a duration of 800 ms. Electrical signals were measured at the opposite end of the AF demonstrating that AFs support axial non-linear ionic currents.”

• cites [30] Kotnik, 2000, Second-order model of membrane electric field induced by alternating external electric fields, IEEE Trans. Biomed. Eng. sourceNot checked yet

  Not checked in the free run.

• cites [42] Sun, 2007, Analytical solutions of AC electrokinetics in interdigitated electrode arrays: Electric field, dielectrophoretic and traveling-wave dielectrophoretic forces, Phys. Rev. E Stat. Nonlinear Soft Matter Phys. sourceNot checked yet

  Not checked in the free run.

Probably not a substantial claim: may not pass S3 (load-bearing).

Kinesin transport may be very strongly disrupted by rapid oscillations of microtubule C-termini.

“However, kinesin binds to MTs through C-termini, which are very sensitive to electric field fluctuations and hence it is possible that kinesin transport would be very strongly disrupted by these rapid oscillations of C-termini.”

• AssumptionRests on an assumption: Microtubule C-termini are very sensitive to electric field fluctuations.