Nanoelectroablation control and vaccination

• US 10,695,127 B2
• Filed: 04/15/2019
• Issued: 06/30/2020
• Est. Priority Date: 12/01/2014
• Status: Active Grant

• Current Assignee: Pulse Biosciences, Inc
• Original Assignee: Pulse Biosciences, Inc
• Inventors: Nuccitelli, Richard Lee, Berridge, Jon Casey, Mallon, Zachary, Kreis, Mark, Athos, Brian, Nuccitelli, Pamela
• Primary Examiner(s): Koharski, Christopher
• Assistant Examiner(s): Moss, James
• Application Number:
  US16/384,676
• Publication Number:
  US 20190239949A1
• Time in Patent Office: 442 Days
• CPC Class Codes: A61B 1/00045 Display arrangement
• CPC Class Codes: A61B 1/043 for fluorescence imaging
• CPC Class Codes: A61B 1/05 characterised by the image sensor, e.g. camera, being in the distal end portion
• CPC Class Codes: A61B 1/06 with illuminating arrangements
• CPC Class Codes: A61B 18/1477 Needle-like probes
• CPC Class Codes: A61B 18/1492 having a flexible, catheter-like structure, e.g. for heart ablation A61B18/1477 takes precedence
• CPC Class Codes: A61B 2018/00273 Anchoring means for temporary attachment of a device to tissue
• CPC Class Codes: A61B 2018/00529 Liver
• CPC Class Codes: A61B 2018/00577 Ablation
• CPC Class Codes: A61B 2018/00613 Irreversible electroporation
• CPC Class Codes: A61B 2018/00642 with feedback, i.e. closed loop control
• CPC Class Codes: A61B 2018/00892 Voltage
• CPC Class Codes: A61B 2018/00904 Automatic detection of target tissue
• CPC Class Codes: A61B 2018/00982 combined with or comprising means for visual or photographic inspections inside the body, e.g. endoscopes
• CPC Class Codes: A61B 2034/101 Computer-aided simulation of surgical operations
• CPC Class Codes: A61B 2034/107 Visualisation of planned trajectories or target regions
• CPC Class Codes: A61B 34/20 Surgical navigation systems; Devices for tracking or guiding surgical instruments, e.g. for frameless stereotaxis
• CPC Class Codes: A61B 6/03 Computed tomography [CT] echo-tomography A61B8/14
• CPC Class Codes: A61B 6/12 Arrangements for detecting or locating foreign bodies
• CPC Class Codes: A61B 90/37 Surgical systems with images on a monitor during operation
• CPC Class Codes: A61M 2037/0007 : having means for enhancing the permeation of substances through the epidermis, e.g. using suction or depression, electric or magnetic fields, sound waves or chemical agents
• CPC Class Codes: A61N 1/326 : for promoting growth of cells, e.g. bone cells
• CPC Class Codes: A61N 1/40 : Applying electric fields by inductive or capacitive coupling microwave apparatus A61N5/00; Applying radio-frequency signals

Abstract

Techniques for treating a tumor and vaccinating against cancer are described. The techniques include treating the tumor by positioning electrodes over an interface between the tumor and non-tumor tissue and applying sub-microsecond pulsed electric fields. The positioning is facilitated by an imaginary contour line of a threshold value of the electric field. In an example, the imaginary contour line is overlaid over images that include the tumor such that the electrodes are properly positioned over the tumor. The techniques also include vaccinating against cancer by passing sub-microsecond pulsed electric fields through tumor cells of a subject sufficient to cause the tumor cells to express calreticulin on surface membranes. The tumor cells are extracted and introduced with the expressed calreticulin into the subject or another subject, thereby providing a vaccination.

Claims

1. A method of vaccinating a subject against a tumor, the method comprising

passing multiple treatments of sub-microsecond pulsed electric fields with different parameters through tumor cells of a first subject;

selecting, based on the multiple treatments, optimized parameters to formulate an effective sub-microsecond pulsed electric fields treatment which increases or maximizes apoptosis of the tumor cells of the first subject; and

using the formulated sub-microsecond pulsed electric fields treatment with the optimized parameters to create a vaccine with the increased or maximized apoptotic tumor cells to treat the tumor cells or other tumor cells of the first subject or a different subject with a same type of tumor cells.

2. The method of claim 1, wherein the passing multiple treatments is performed on different cells of the tumor cells, such that different electric field intensities, number of pulses, pulse widths, or frequencies are tried on the different cells of the tumor cells.

3. The method of claim 1, the method comprising storing the optimized parameters with an association to at least one of an identifier of the first subject or a type of the tumor cells.

4. The method of claim 1, wherein the passing multiple treatments is performed on the tumor cells extracted from the first subject.

5. The method of claim 4, wherein the step of using the formulated sub-microsecond pulsed electric fields treatment with the optimized parameters comprises reintroducing to the first subject at least a portion of the extracted tumor cells that has been treated with the optimized parameters.

6. The method of claim 4, wherein the step of using the formulated sub-microsecond pulsed electric fields treatment with the optimized parameters comprises applying the formulated sub-microsecond pulsed electric fields treatment with the optimized parameters to a second sample of tumor cells extracted from the first subject or to a sample extracted from the different subject with the same type of tumor cells.

7. The method of claim 1, wherein the sub-microsecond pulsed electric fields are pulsed at 100 ns pulse duration.

8. The method of claim 1, wherein the apoptosis of the tumor cells is detected using one or more of the following techniques:

detecting cleaved caspase-3 or cleaved caspase-9 within treated tumor cells, detecting pyknosis, detecting calreticulin on outer surface walls of the tumor cells, detecting cytochrome c release from mitochondria within the tumor cells or Damage-Associated Molecular Pattern molecules (DAMPs).

9. The method of claim 1, wherein the tumor cells with the increased or maximized apoptosis are administered subdermally, intraperitoneally or into a bloodstream.

10. The method of claim 1, the method comprising detecting apoptotic tumor cells in the treated tumor cells by:

introducing a fluorescent label for cleaved caspase-3 or cleaved caspase-9 into the treated tumor cells;

illuminating the fluorescent label; and

separating the apoptotic tumor cells from the treated tumor cells based on the fluorescent label.

11. The method of claim 1, the method comprising detecting apoptotic tumor cells in the treated tumor cells by:

introducing calreticulin antibodies connected to magnetic beads to the treated tumor cells, the calreticulin antibodies binding with calreticulin on surfaces of the apoptotic tumor cells;

applying a magnet to the magnetic beads; and

separating the apoptotic tumor cells from the treated tumor cells based on the magnetic beads.

12. The method of claim 1, the method comprising detecting apoptotic tumor cells in the treated tumor cells by:

measuring a temperature increase of the tumor cells caused by the sub-microsecond pulsed electric fields; and

adjusting an intensity, pulse widths, or frequency of the sub-microsecond pulsed electric fields based on the measured temperature increase.

13. The method of claim 1, wherein at least the step of passing multiple treatments of sub-microsecond pulsed electric fields is performed using a device that stores instructions, that upon execution on the device, configure the device to at least perform the step.

14. The method of claim 1, wherein at least a portion of the sub-microsecond pulsed electric fields has an intensity greater than 10 kV/cm and pulse widths between 0.1 and 1000 nanoseconds.

15. The method of claim 1, the method comprising utilizing a user interface to input or select from stored options an identifier of the subject and/or a type of the tumor cells.

16. The method of claim 1, wherein the optimized parameters comprise one or more of the following:

electric field intensities, number of pulses, amplitude, pulse widths, or frequencies.

17. The method of claim 1, the method comprising transferring the treated tumor cells from a pulse generator to a flow cytometry system.

18. The method of claim 1, the method comprising

generating the vaccine that includes apoptotic cells from the treated tumor cells.

19. The method of claim 18, wherein the vaccine is generated by further including necrotic tumor cells in the vaccine.

20. The method of claim 1, the method further comprising sorting the treated tumor cells between the apoptotic cells and non-apoptotic cells.