Determining the frequency of tumor treating field (TT field) therapy based on tests performed on tumor cells

Abstract

Cancer treatment using TT fields (tumor treating electric fields) can be customized for individual subjects by extracting cancer cells from the subject's body. Alternating electric fields are then applied to the extracted cells at different frequencies, and voltage measurements are taken from the cells under two different conditions at each of the different frequencies. These voltage measurements can then be used to determine which frequency provides the greatest gradient at the cleavage furrow when similar cells divide, thereby increasing the effectiveness of the TT field. Treatment using the TT field can then proceed at the determined frequency.

Description

[Technical Field]

[0001] The present invention relates to determining the frequency of a Tumor Treating Field (TT Field) therapy based on tests performed on tumor cells. CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of U.S. Provisional Application No. 63 / 435,900, filed December 29, 2022, which is incorporated herein by reference in its entirety. [Background technology]

[0002] Tumor treating field (TT field) therapy is a proven approach to treating tumors using alternating current (AC) electric fields at frequencies between 50 kHz and 1 MHz (e.g., 150–250 kHz). TT field therapy has received FDA approval for treating glioblastoma (GBM) brain tumors and appears to be highly promising for many other tumor types. The Optune® system delivers TT fields to patients via four transducer arrays placed on the patient's skin near the tumor. The transducer arrays are arranged in pairs: one pair is placed on the left and right sides of the tumor, and the other pair is placed in front and behind the tumor. An AC signal generator (a) sends an AC current to the anterior / posterior pair of transducer arrays for one second to induce an electric field in the tumor in a first direction. Then, (b) sends an AC current to the left / right pair of arrays for one second to induce an electric field in the tumor in a second direction. Steps (a) and (b) are then repeated throughout the treatment period.

[0003] Traditionally, the frequency at which a TT field is applied to a subject has been based on the particular type of tumor being treated, for example, 200 kHz is the recommended frequency for a TT field when treating GBM, and 150 kHz is the recommended frequency for a TT field when treating gastric cancer. Summary of the Invention [Means for solving the problem]

[0004] One aspect of the present invention relates to a first method for determining a frequency at which to treat a specific subject using an AC electric field. The first method includes electrically connecting a first electrode to an interior of first cancer cells obtained from the subject, electrically connecting a second electrode to an interior of second cancer cells obtained from the subject, electrically connecting a third electrode to the interior of the first cancer cells, and electrically connecting a fourth electrode to the interior of the second cancer cells. The first method also includes applying an external AC electric field to the first and second cancer cells at a plurality of different frequencies for a plurality of different times, and measuring a first voltage between the third electrode and the fourth electrode while the external AC electric field is applied at each of the plurality of different frequencies during a period in which a low resistance path is established between the first electrode and the second electrode. The first method also includes measuring a respective second voltage between the third electrode and the fourth electrode while the external alternating current electric field is applied at each of the plurality of different frequencies during a period when a low resistance path is not established between the first electrode and the second electrode, and determining a frequency for treating the subject using an alternating current electric field based at least in part on the measured first voltage and the measured second voltage.

[0005] In some examples of the first method, the determining includes selecting a frequency at which a difference between the respective first voltages and the respective second voltages is greatest. In some examples of the first method, the determining includes selecting a frequency at which a difference between the respective first voltages and the respective second voltages is within 10% of the maximum difference. In some examples of the first method, the determining includes selecting a frequency at which a difference between the respective first voltages and the respective second voltages is within 25% of the maximum difference.

[0006] In some examples of the first method, each of the plurality of different frequencies is between 50 kHz and 1 MHz. In some examples of the first method, each of the plurality of different frequencies is between 75 kHz and 500 kHz. In some examples of the first method, the first electrode includes a hollow glass micropipette filled with a conductive liquid, and the second electrode includes a hollow glass micropipette filled with a conductive liquid.

[0007] In some examples of the first method, each of the plurality of different frequencies is between 75 kHz and 500 kHz, and the determining includes selecting a frequency at which a difference between the respective first voltage and the respective second voltage is within 25% of the maximum difference.

[0008] Some examples of the first method further include obtaining the first cancer cells from the subject, obtaining the second cancer cells from the subject, and treating the subject using an alternating electric field at the determined frequency.

[0009] Another aspect of the present invention relates to a first device for determining a frequency at which to treat a specific subject using an alternating current electric field. The first device includes a first electrode configured to electrically connect with the interior of a first cell, a second electrode configured to electrically connect with the interior of a second cell, and a switch. The switch has a first terminal electrically connected to the first electrode and a second terminal electrically connected to the second electrode. The switch can operate in either (a) a closed state that establishes a low resistance path between the first electrode and the second electrode, or (b) an open state that does not establish a low resistance path between the first electrode and the second electrode.

[0010] In some embodiments of the first device, the first electrode comprises a hollow glass micropipette filled with a conductive liquid, and the second electrode comprises a hollow glass micropipette filled with a conductive liquid.

[0011] Some embodiments of the first device further comprise the first cell and the second cell, in which the first electrode is electrically connected to the interior of the first cell and the second electrode is electrically connected to the interior of the second cell.

[0012] Optionally, the embodiment described in the previous paragraph may further comprise a third electrode and a fourth electrode, wherein the third electrode is configured to electrically connect with the interior of the first cell, and the third electrode is configured to electrically connect with the interior of the first cell, and the fourth electrode is configured to electrically connect with the interior of the second cell, and the fourth electrode is configured to electrically connect with the interior of the second cell.

[0013] Optionally, the embodiments described in the previous paragraph may further include a controller configured to control the state of the electric field generator and the switch while the switch is in the closed state, such that the electric field generator applies an electric field to the first cell and the second cell at a plurality of different frequencies at respective first times, each of the plurality of different frequencies being between 50 kHz and 1 MHz. In these embodiments, the controller is also configured to input respective first voltage measurements obtained using the third electrode and the fourth electrode during each of the first times. The controller is also configured to control the state of the electric field generator and the switch while the switch is in the open state, such that the electric field generator applies an electric field to the first cell and the second cell at the plurality of different frequencies at respective second times. The controller is also configured to input respective second voltage measurements obtained using the third electrode and the fourth electrode during each of the second times.

[0014] Optionally, in the embodiment described in the previous paragraph, the controller may be further configured to determine a frequency that maximizes a difference between each first voltage measurement and each second voltage measurement.

[0015] Another aspect of the present invention relates to a second method for determining a frequency for treating a particular subject using an AC electric field, the second method including electrically connecting a first electrode to an interior of a first cell obtained from the subject, electrically connecting a second electrode to an interior of a second cell obtained from the subject, electrically connecting a third electrode to the interior of the first cell, and electrically connecting a fourth electrode to the interior of the second cell. The second method also includes applying an external AC electric field to the first and second cancer cells and measuring a first voltage between the third electrode and the fourth electrode while the external AC electric field is applied during at least one period during which a low resistance path is established between the first electrode and the second electrode. The second method also includes measuring a second voltage between the third electrode and the fourth electrode while the external alternating electric field is applied during at least one period when a low resistance path is not established between the first electrode and the second electrode, and determining a frequency for treating the subject using an alternating electric field based at least in part on the measured first voltage and the measured second voltage.

[0016] Some examples of the second method further include obtaining the first cell from the subject and obtaining the second cell from the subject.

[0017] Another aspect of the present invention relates to a third method for measuring characteristics of an electric field inside a cell, the third method comprising: electrically connecting a first electrode to the interior of the cell through a cell membrane; electrically connecting a second electrode to the interior of the cell through the cell membrane such that a distal end of the second electrode is spaced from a distal end of the first electrode; applying an external alternating electric field to the cell; and measuring a voltage between the first electrode and the second electrode while the external alternating electric field is applied to the cell.

[0018] In some examples of the third method, the external electric field has a frequency of 50 kHz to 1 MHz. In some examples of the third method, the external electric field has a frequency of 75 kHz to 500 kHz. In some examples of the first method, the first electrode includes a hollow glass micropipette filled with a conductive liquid, and the second electrode includes a hollow glass micropipette filled with a conductive liquid.

[0019] Another aspect of the present invention relates to a fourth method for determining the frequency of an AC electric field that maximizes an electrical gradient when cells in a given population divide, the fourth method comprising: electrically connecting a first electrode to an interior of a first cell from the population, electrically connecting a second electrode to an interior of a second cell from the population, electrically connecting a third electrode to the interior of the first cell, and electrically connecting a fourth electrode to the interior of the second cell. The fourth method also comprises applying an external AC electric field to the first and second cells at a plurality of different frequencies for a respective plurality of different times, and measuring a respective first voltage between the third electrode and the fourth electrode while the external AC electric field is applied at each of the plurality of different frequencies during a period during which a low resistance path is established between the first electrode and the second electrode. The fourth method also includes measuring a respective second voltage between the third electrode and the fourth electrode while the external alternating current electric field is applied at each of the plurality of different frequencies during a period when a low resistance path is not established between the first electrode and the second electrode, and determining a frequency for applying the alternating current electric field to the population of cells based at least in part on the measured first voltage and the measured second voltage.

[0020] In some examples of the fourth method, the determining includes selecting a frequency at which a difference between the respective first voltages and the respective second voltages is greatest. In some examples of the fourth method, the determining includes selecting a frequency at which a difference between the respective first voltages and the respective second voltages is within 10% of the maximum difference. In some examples of the fourth method, the determining includes selecting a frequency at which a difference between the respective first voltages and the respective second voltages is within 25% of the maximum difference.

[0021] In some examples of the fourth method, each of the plurality of different frequencies is between 50 kHz and 1 MHz. In some examples of the fourth method, each of the plurality of different frequencies is between 75 kHz and 500 kHz. In some examples of the first method, the first electrode includes a hollow glass micropipette filled with a conductive liquid, and the second electrode includes a hollow glass micropipette filled with a conductive liquid.

[0022] In some examples of the fourth method, each of the plurality of different frequencies is between 75 kHz and 500 kHz, and the determining includes selecting a frequency at which a difference between the respective first voltage and the respective second voltage is within 25% of the maximum difference. [Brief explanation of the drawings]

[0023] [Figure 1] FIG. 1 is a schematic diagram of a TT field applied to a dividing cell just before the original parent cell divides into two daughter cells. [Figure 2] FIG. 1 is a schematic diagram of an apparatus for determining frequencies for treating a particular subject using a TT field. [Figure 3] 3 is a flow chart for controlling the switches and field generators of FIG. 2 to determine the frequency at which to treat a particular subject using a TT field. [Figure 4] Shown are two separate cells present in a conductive medium. [Figure 5]We present a different approach to directly measure intracellular voltage gradients.

[0024] Various embodiments are described in detail below with reference to the accompanying drawings, in which like reference numerals represent like elements, and in which: DETAILED DESCRIPTION OF THE INVENTION

[0025] While the traditional approach of using the same frequency for all subjects with a given type of cancer is a viable solution, using a single frequency for all subjects may not provide the best results for each and every one of those subjects. For example, while 200 kHz may be the best frequency for most subjects with GBM, certain individuals with GBM may respond better to a different frequency (e.g., 175 kHz or 225 kHz).

[0026] TT fields do not damage non-dividing cells, affecting cells only while they are dividing, and because cancer cells divide much more rapidly than normal cells, TT fields selectively target cancer cells.

[0027] Figure 1 is a schematic diagram of a TT field E1 applied to a dividing cell just before the original parent cell divides into two daughter cells. At this stage, the dividing parent cell has an hourglass-like shape in that it has two parts connected by a relatively narrow neck (i.e., cleavage furrow). The TT field E1 is generated by applying a voltage across a pair of electrodes 10.

[0028] Without being bound by this theory, it is believed that increasing the electric field gradient at the cleavage furrow increases the effectiveness of the TT field. Two competing factors contribute to increasing the electric field gradient at the cleavage furrow of dividing cells. The first factor deals with the electric field entering the dividing cell. Because cell membranes have capacitance, the impedance of the cell membrane is inversely proportional to frequency. Therefore, when a dividing cell is placed within a conductive material and a TT field is applied to the conductive material, a TT field with a higher frequency will cross the cell membrane more easily than a TT field with a lower frequency. Therefore, the first factor favors the use of higher frequencies.

[0029] The second factor relates to the path taken by the TT field line after it enters the dividing cell. In the state shown in Figure 1, the path between the two daughter cells is a low-resistance impedance path, with essentially zero capacitive component. At lower frequencies, the material within the dividing cell has a lower impedance than the cell membrane, and in this case, the current preferentially follows the lower impedance path through the cleavage furrow. However, as the frequency of the TT field increases to a point where the impedance of the cell membrane drops sufficiently, the current preference for following the path through the cleavage furrow disappears. Therefore, the second factor favors the use of lower frequencies.

[0030] *IF* It is easy to directly measure the gradient of the electric field at the cleavage furrow of a dividing cell, and it is straightforward to balance the two factors mentioned above and determine the frequency that provides the maximum gradient. *IF* It is easy to directly measure the gradient; dividing cells can be placed in a conductive material (e.g., saline), and an alternating electric field can be applied to the conductive material at many different frequencies. Then, for each of the different frequencies, the resulting gradient at the cleavage furrow can be measured, and the maximum gradient can be identified. However, in practice, directly measuring the gradient of the electric field at the cleavage furrow of a dividing cell is very difficult, if not impossible.

[0031] The present application discloses an indirect approach to determining a frequency that balances the two factors described above to maximize the gradient at the cleavage furrow of dividing cells (and thus enhance the effectiveness of the TT field). This approach can be advantageously used to select an individualized frequency of TT field treatment for each individual subject. In particular, it relies on testing cancer cells extracted from each individual subject before TT field therapy begins for that same individual subject.

[0032] 2 is a schematic diagram of an apparatus for determining a frequency for treating a particular subject using an alternating electric field (e.g., a TT field). A first cancer cell C1 and a second cancer cell C2 are extracted (e.g., during a biopsy) from the body of the particular subject to be ultimately treated.

[0033] The first electrode 21 and the second electrode 22 are configured to electrically connect to the interior of the first cell C1 and the second cell C2, respectively. Each of these electrodes 21, 22 can be implemented, for example, using a hollow glass micropipette filled with a conductive liquid (e.g., saline), similar to conventional patch clamp electrodes. Alternative approaches for implementing these electrodes 21, 22 can also be used. For example, the first electrode 21 can be electrically connected to the interior of the first cell C1, and the second electrode 22 can be electrically connected to the interior of the second cell C2, using techniques similar to conventional patch clamp electrodes.

[0034] Switch S1 has one terminal electrically connected to first electrode 21 and a second terminal electrically connected to second electrode 22. Switch S1 can operate in either (a) a closed state in which a low resistance path is established between the first electrode and the second electrode, or (b) an open state in which a low resistance path is not established between the first electrode and the second electrode.

[0035] The third electrode 23 and the fourth electrode 24 are configured to be electrically connected to the interior of the first cell C1 and the second cell C2, respectively. Each of these electrodes 23, 24 can be implemented as described above for the first electrode 21 and the second electrode 22. The third electrode 23 is electrically connected to the interior of the first cell C1, and the fourth electrode 24 is electrically connected to the interior of the second cell C2 (e.g., using the same techniques as described above for the first electrode 21 and the second electrode 22).

[0036] Controller 50 is configured to control the state of switch S1 and the electric field generator (which in the exemplary embodiment shown in FIG. 2 includes an AC signal generator 60 connected to electrodes 10 arranged and positioned to apply an AC field E2 to the medium in which cells C1, C2 reside). The states of switch S1 and the electric field generator are controlled as described below in connection with FIG. 3. The embodiment of FIG. 2 also includes voltage measurement circuitry 70 configured to measure AC voltages induced across third electrode 23 and fourth electrode 24 when electric field E2 is applied to cells C1, C2 under a variety of different conditions. Voltage measurement circuitry 70 can be, for example, a voltmeter, an analog front end configured to measure voltage, an oscilloscope, or any of a variety of other voltage measurement circuits.

[0037] 3 is a flow chart illustrating how controller 50 controls the state of switch S1 and electric field generators 60+10 to determine the frequency at which to treat a particular subject using an AC field (e.g., a TT field). Processing begins at S10, where the controller initializes a loop counter variable i to 1.

[0038] In S20, the controller 50 commands the AC signal generator 60 to output a first frequency F1, which applies an AC electric field to the first and second cells via the electrodes 10 for the first time.

[0039] In S22, the controller 50 commands the switch S1 to close and then inputs a first voltage measurement FV1 via the voltage measurement circuit 70. This first voltage measurement FV1 represents the voltage between the third electrode 23 and the fourth electrode 24 while an AC electric field is applied to the cells C1, C2 at the first frequency while the switch S1 is closed. Next, in S24, the controller 50 commands the switch S1 to open and then inputs a second voltage measurement SV1 via the voltage measurement circuit 70. This second voltage measurement SV1 represents the voltage between the third electrode 23 and the fourth electrode 24 while an AC electric field is applied to the cells C1, C2 at the first frequency while the switch S1 is open.

[0040] Steps S30 and S32 collectively determine whether additional passes through the loop are needed and initiate any such additional passes through the loop. More specifically, in S30, the controller 50 checks whether the loop counter i has reached its maximum value n. If the loop counter has not reached its maximum value, the loop counter i is incremented in S32, and processing returns to S20 for an additional pass through the loop S20-S24 at the next frequency (e.g., a second, third, fourth, ..., nth pass). Alternatively, if the loop counter i has reached its maximum value n, processing proceeds to step S40. Using this loop structure, the controller 50 obtains a first voltage measurement FVi (while S1 is closed) and a second voltage measurement SVi (while S1 is open) at each of the different frequencies (i.e., F1-Fn).

[0041] In the special case where the maximum value of the loop counter is 1 (i.e., n=1), there is only a single path through the loop, and therefore only a single first voltage and a single second voltage.

[0042] In particular, closing switch S1 establishes a low impedance path between the first electrode 21 and the second electrode 22 (connected to the interiors of the first cell C1 and the second cell C2, respectively), thus simulating the situation shown in Figure 1 where two daughter cells are connected via a low resistance impedance path, while opening switch S1 simulates the situation where two separate cells reside in a conductive medium, as shown in Figure 4.

[0043] After collecting the voltage measurements FVi and SVi (i=1 to n) obtained as described above in connection with steps S22 and S24 (performed at n different frequencies), those measurements can be used to predict the gradient at the interconnection between two daughter cells during mitosis. More specifically, the magnitude of that gradient relates to the difference between (a) the voltage measured using the third and fourth electrodes 23 and 24 of FIG. 2 when an AC field is applied at a given frequency while switch S1 is closed, and (b) the voltage measured using those same electrodes 23, 24 when an AC field is applied at the same given frequency while switch S1 is open.

[0044] Thus, the measured first voltages (FV1-FVn) and the measured second voltages (SV1-SVn) can be used in S40 to determine a frequency for treating the subject using an AC electric field. This can be achieved, for example, by selecting a frequency at which the difference between each first voltage (FVi) and each second voltage (SVi), for i=1-n, is greatest. Alternatively, it can be achieved by selecting a frequency at which the difference between each first voltage (FVi) and each second voltage (SVi) is within 1%, 2%, 5%, 10%, or 25% of the maximum difference.

[0045] The number of frequencies tested (i.e., n) can range from 2 to 100 or more. For example, if frequencies from 75 kHz to 500 kHz were tested and the tested frequencies were spaced 25 kHz apart, n would be 18. In another example, if frequencies from 190 kHz to 210 kHz were tested and the tested frequencies were spaced 1 kHz apart, n would be 21. Each of the multiple different frequencies could be, for example, 50 kHz to 1 MHz, 75 kHz to 500 kHz, 75 kHz to 300 kHz, etc. Narrower ranges (e.g., 175 kHz to 225 kHz for glioblastoma patients) can be used when the expected frequency range can be predicted based on the type of cancer being treated.

[0046] Advantageously, the technique described above in connection with Figures 2 and 3 does not require precise alignment of the distal ends of the first, second, third, and fourth electrodes 21-24, nor does it require a specific spacing between them. Indeed, the technique described above is independent of the spacing between electrode elements and can be used to determine frequencies for treatment even when the electrode spacing varies widely. Furthermore, the technique described above is independent of the amplitude of the AC electric field applied to cells C1 and C2 (as long as the same amplitude of the applied electric field E2 is used for all frequencies tested). This is because the voltage difference technique described above does not provide a direct measurement of the gradient at the cleavage furrow. Instead, it provides indirect data, from which information regarding the frequency resulting in the maximum gradient can be inferred.

[0047] After the frequency is determined in S40, the subject is treated with a TT field at that frequency in S50.

[0048] Figure 5 shows a different approach to directly measuring intracellular voltage gradients. In this approach, a first electrode 31 is electrically connected to the interior of a cell C1 through the cell membrane, and a second electrode 32 is electrically connected to the interior of the cell through the cell membrane so that the distal end of the second electrode is spaced from the distal end of the first electrode. For example, the distal end of the second electrode can be spaced 1 μm to 5 μm or 1 μm to 3 μm from the distal end of the first electrode. For example, the distal end of the second electrode can be approximately 1 μm, approximately 2 μm, approximately 3 μm, approximately 4 μm, or approximately 5 μm from the distal end of the first electrode. The first and second electrodes can each be hollow glass micropipettes filled with a conductive liquid. An external AC electric field E3 is applied to the cell, and then the voltage between the first electrode 31 and the second electrode 32 is measured while the external AC electric field E3 is applied. The external electric field can have a frequency of 50 kHz to 1 MHz or 75 kHz to 500 kHz. This approach differs from the approach described above in connection with Figures 2 and 3 because electrode spacing and positioning in the approach of Figure 5 are critical.

[0049] While the present invention has been disclosed with reference to particular embodiments, numerous modifications, variations, and variations can be made to the described embodiments without departing from the scope and spirit of the invention, as defined in the appended claims. Accordingly, it is intended that the present invention not be limited to the described embodiments, but rather have the full scope defined by the language of the following claims and their equivalents.

Claims

1. 1. A method for determining a frequency for treating a particular subject using an alternating electric field, the method comprising: electrically connecting a first electrode to the interior of a first cancer cell obtained from the subject; electrically connecting a second electrode to the interior of a second cancer cell obtained from the subject; electrically connecting a third electrode to the interior of the first cancer cell; electrically connecting a fourth electrode to the interior of the second cancer cell; applying an external alternating electric field to the first and second cancer cells at a plurality of different frequencies for a respective plurality of different times; measuring a first voltage between the third electrode and the fourth electrode while applying the external alternating electric field at each of the plurality of different frequencies during a period in which a low resistance path is established between the first electrode and the second electrode; measuring a second voltage between the third electrode and the fourth electrode while applying the external alternating electric field at each of the plurality of different frequencies during a period when a low resistance path is not established between the first electrode and the second electrode; determining a frequency for treating the subject using an alternating electric field based at least in part on the measured first voltage and the measured second voltage.

2. The method of claim 1 , wherein the determining comprises selecting a frequency at which a difference between the respective first voltage and the respective second voltage is greatest.

3. 2. The method of claim 1, wherein said determining comprises selecting a frequency at which a difference between said respective first voltage and said respective second voltage is within 10% of said maximum difference.

4. 2. The method of claim 1, wherein said determining comprises selecting a frequency at which a difference between said respective first voltage and said respective second voltage is within 25% of said maximum difference.

5. The method of claim 1 , wherein each of the plurality of different frequencies is between 50 kHz and 1 MHz.

6. The method of claim 1 , wherein each of the plurality of different frequencies is between 75 kHz and 500 kHz.

7. each of the plurality of different frequencies is between 75 kHz and 500 kHz; 2. The method of claim 1, wherein said determining comprises selecting a frequency at which a difference between said respective first voltage and said respective second voltage is within 25% of said maximum difference.

8. 10. The method of claim 1, wherein the first electrode comprises a hollow glass micropipette filled with a conductive liquid, and the second electrode comprises a hollow glass micropipette filled with a conductive liquid.

9. obtaining the first cancer cells from the subject; obtaining the second cancer cells from the subject; 10. The method of claim 1, further comprising treating the subject with an alternating electric field at the determined frequency.

10. 1. An apparatus for determining a frequency for treating a particular subject using an alternating electric field, the apparatus comprising: a first electrode configured to be in electrical communication with an interior of the first cell; a second electrode configured to be in electrical communication with the interior of the second cell; a switch having a first terminal and a second terminal, the first terminal electrically connected to the first electrode and the second terminal electrically connected to the second electrode, the switch operable in either (a) a closed state in which a low resistance path is established between the first electrode and the second electrode, or (b) an open state in which a low resistance path is not established between the first electrode and the second electrode.

11. 10. The method of claim 1, wherein the tenth electrode comprises a hollow glass micropipette filled with a conductive liquid, and the second electrode comprises a hollow glass micropipette filled with a conductive liquid.

12. the first cell, wherein the first electrode is electrically connected to the interior of the first cell; The device of claim 10, further comprising: the second cell, wherein the second electrode is electrically connected to the interior of the second cell.

13. a third electrode configured to be in electrical communication with the interior of the first cell, wherein the third electrode is configured to be in electrical communication with the interior of the first cell; 13. The apparatus of claim 12, comprising: a fourth electrode configured to be in electrical communication with the interior of the second cell, wherein the fourth electrode is configured to be in electrical communication with the interior of the second cell.

14. controlling a state of the electric field generator and the switch such that, while the switch is in the closed state, the electric field generator applies an electric field to the first cell and the second cell at a plurality of different frequencies at respective different first times, wherein each of the plurality of different frequencies is between 50 kHz and 1 MHz; inputting respective first voltage measurements obtained using the third electrode and the fourth electrode during each of the first periods of time; controlling a state of the electric field generator and the switch such that, while the switch is in the open state, the electric field generator applies an electric field to the first cell and the second cell at the plurality of different frequencies at respective different second times; and inputting respective second voltage measurements obtained using the third electrode and the fourth electrode during each of the second periods of time.

15. 15. The apparatus of claim 14, wherein the controller is further configured to determine a frequency that maximizes a difference between each first voltage measurement and each second voltage measurement.

16. 1. A method for determining the frequency of an alternating electric field that maximizes the electrical gradient at which cells in a given population divide, the method comprising: electrically connecting a first electrode to an interior of a first cell from the population; electrically connecting a second electrode to the interior of a second cell from the population; electrically connecting a third electrode to the interior of the first cell; electrically connecting a fourth electrode to the interior of the second cell; applying an external alternating electric field to the first and second cells at a plurality of different frequencies for a respective plurality of different times; measuring a first voltage between the third electrode and the fourth electrode while applying the external alternating electric field at each of the plurality of different frequencies during a period in which a low resistance path is established between the first electrode and the second electrode; measuring a second voltage between the third electrode and the fourth electrode while applying the external alternating electric field at each of the plurality of different frequencies during a period when a low resistance path is not established between the first electrode and the second electrode; determining a frequency for applying an alternating electric field to the population of cells based at least in part on the measured first voltage and the measured second voltage.

17. 17. The method of claim 16, wherein said determining comprises selecting a frequency at which a difference between said respective first voltage and said respective second voltage is greatest.

18. 17. The method of claim 16, wherein said determining comprises selecting a frequency at which a difference between said respective first voltage and said respective second voltage is within 10% of said maximum difference.

19. 17. The method of claim 16, wherein said determining comprises selecting a frequency at which a difference between said respective first voltage and said respective second voltage is within 25% of said maximum difference.

20. each of the plurality of different frequencies is between 75 kHz and 500 kHz; 17. The method of claim 16, wherein said determining comprises selecting a frequency at which a difference between said respective first voltage and said respective second voltage is within 25% of said maximum difference.

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