DETAILED ACTION
Notice of Pre-AIA or AIA Status
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Information Disclosure Statement
The information disclosure statements (IDS) submitted on 11/13/2024 are in compliance with the provisions of 37 CFR 1.97. Accordingly, the information disclosure statement is being considered by the examiner.
Drawings
The drawings received on 8/07/2024 are objected. Drawings are unacceptable due to poor image quality and legibility. Specifically, the drawings are blurry, pixelated, and contain excessively small fonts. Additionally, data labels and axis lines on graphs (e.g., Figures 1G, 1J, 2A, and 2C) are illegible. New corrected drawings in compliance with 37 CFR 1.121(d) are required in this application because the drawings are blurry, pixelated, and contain excessively small fonts and illegible. Corrected drawing sheets in compliance with 37 CFR 1.121(d) are required in reply to the Office action to avoid abandonment of the application. Any amended replacement drawing sheet should include all of the figures appearing on the immediate prior version of the sheet, even if only one figure is being amended. The figure or figure number of an amended drawing should not be labeled as “amended.” If a drawing figure is to be canceled, the appropriate figure must be removed from the replacement sheet, and where necessary, the remaining figures must be renumbered and appropriate changes made to the brief description of the several views of the drawings for consistency. Additional replacement sheets may be necessary to show the renumbering of the remaining figures. Each drawing sheet submitted after the filing date of an application must be labeled in the top margin as either “Replacement Sheet” or “New Sheet” pursuant to 37 CFR 1.121(d). If the changes are not accepted by the examiner, the applicant will be notified and informed of any required corrective action in the next Office action. The objection to the drawings will not be held in abeyance.
Claim Rejections - 35 USC § 102
The following is a quotation of the appropriate paragraphs of 35 U.S.C. 102 that form the basis for the rejections under this section made in this Office action:
A person shall be entitled to a patent unless –
(a)(1) the claimed invention was patented, described in a printed publication, or in public use, on sale, or otherwise available to the public before the effective filing date of the claimed invention.
Claims 8 -20 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by TEITELL et al. (US 2016/0103118).
Regarding claim 8, Teitell teaches a method for determining treatment response parameters with a microscope imaging system (refer to US 2016/0103118), the method comprising: receiving, by an electronic controller (relative position of the microscope objective and observation chamber can be controlled by computer and can be translatable in three-dimensions allowing for rapid, automated image acquisition, [0051], Fig. 1), a plurality of images of a plurality of therapy treatment samples over a period of time from a camera (Live cell interferometry … a lateral shearing interferometric camera connected to an ordinary microscope with live cell imaging capability, screening is performed on target cells disposed on a substrate…. screening can be performed on arrays of microwells, e.g., arrays comprising greater than 100, or greater than 1000, or greater than 10,000 microwells, [0069]; Target cells were imaged in standard culture media for 1.5 h prior to the start of each experiment to confirm the live cell culture imaging platform maintains viability of target cells in the absence of CTLs. M202 target cells showed a positive mass accumulation rate, indicating a healthy population and the maintenance of cell viability. (FIG. 3, panels D-E; FIG. 7, panel B). Control experiments demonstrated maintenance of both T and target cell viability during extended imaging periods (FIGS. 7 and 8), [0113]);
determining, via the electronic controller, (observation chamber can be controlled by computer .. for rapid, automated image acquisition, [0051]; a cell mass for each therapy treatment sample of the plurality of therapy treatment samples based on each image of the plurality of images (live cell interferometer (LCI; An advanced optical technique that uses wave interference to image and measure microscopic, label-free changes in biological cells) is capable of measuring the mass of both adherent and non-adherent cells, [0051]; the methods utilize label-free optical imaging to identify changes in mass of cells (e.g., an increase in mass of a T-cell and/or a decrease in mass of a target cell) as an indicator of T cell activation when T cells are presented with target cells bearing a cognate antigen, [0064]; the change in cell mass is determined using various interferometric and/or quantitative phase imaging microscopy techniques, [0068]);
tracking, via the electronic controller, the cell mass over the period of time for each therapy treatment sample of the plurality of therapy treatment samples (target cells comprise cells of a cancer selected from the group consisting of breast cancer, central nervous system cancer, cervical cancer, colorectal cancer, testicular cancer, ovarian cancer, leukemia, a lymphoma, a melanoma, a soft tissue sarcoma, testicular cancer, and thyroid cancer, [0045]; By tracking T and target cell mass changes using label-free optical imaging methods, e.g., the methods permit direct measurements of the target and responding T cell during T cell mediated cytotoxicity to facilitate TCR cloning for use in adoptive immunotherapy against cancer. These methods can similarly be used to identify T-cells that are activated by cells infected with pathogens (e.g., cells infected with virus, bacteria, fungus, etc.), to identify cells that express (e.g., recombinantly express) particular peptides or proteins, and the like, [0066]); and
determining, via the electronic controller, a plurality of response parameters for each therapy treatment sample of the plurality of therapy treatment samples based on the cell mass over the period of time (observation chamber can be controlled by computer .. … for rapid, automated image acquisition, [0051]; target cells comprise cells of a cancer selected from the group consisting of breast cancer, central nervous system cancer, cervical cancer, colorectal cancer, testicular cancer, ovarian cancer, leukemia, a lymphoma, a melanoma, a soft tissue sarcoma, testicular cancer, and thyroid cancer; [9945], Paragraphs [0045-0050] mentioned different cells; FIG. 5, panels A-F, illustrate LCI measurement of CTL mass and mass accumulation rate during T cell mediated cytotoxicity. Panel A: Mass versus time of an activated CTL and corresponding target cell. t=0 h is the point at which the target cell detaches from the substrate at the beginning of cell death. CTL+target cell refers to total mass of both cells in frames where they could not be measured separately. Panel B: Normalized mass versus time of 10 CTL-mediated cytotoxicity events. CTL mass is normalized relative to the mass at the time of target cell morphology change, which is used as the t=0 h point for all traces. Gray lines show best fit lines used for determining mass accumulation rates. Panel C: Average mass accumulation rate of CTLs before a cytotoxic event, during the first 100 minutes of a cytotoxic event, and after the first 100 minutes of a cytotoxic event demonstrating an approximately 4-fold increase in mass accumulation during the first 100 minutes of a cytotoxic event. Panel D: LCI image of 9 unresponsive and 1 cytotoxic T cell illustrating an approximately 3-fold difference in mass. The white arrow indicates the activated T cell, as determined by tracking this cell after persistent contact with target cell and subsequent target cell death. Panel E: The average mass of 116 activated CTLs is approximately 2.8-fold greater than the average mass of unresponsive controls. Panel F: Average area of activated CTLs is only approximately 1.4-fold greater than non-activated controls and not significant at the 95% confidence level, illustrating the utility of LCI mass measurements for determining CTL activation, [0055]; FIG. 7, panels A-D, shows averaged, normalized mass versus time plots for control target cell growth conditions showing robust growth on the LCI stage, and specificity of T cell mediated cytotoxicity. Panel A: Unaffected M202 cells (n=632) during treatment with F5 TCR transduced, CD8+ T cells. Panel B: M202 cells (n=117) prior to treatment with F5 TCR transduced, CD8+ T cells. Panel C: M202 cells (n=2058) treated with F5 TCR negative, CD8+ T cells. Panel D: Antigen-irrelevant, PC-3 prostate cancer cells treated with F5 TCR transduced, CD8+ T cells. Line shows mean normalized mass versus time (normalized relative to mass at first time point). Shaded region shows the mean +/−SD, [0057]).
Regarding claim 9, Teitell teaches the method according to claim 8 (see above), further comprising: determining, via the electronic controller, a treatment response for each therapy treatment sample of the plurality of therapy treatment samples based on the plurality of response parameters, (Live cell interferometry (LCI) is a label-free optical microscopy technique that measures whole cell responses. In certain embodiments LCI uses a Michelson-type interferometer, [0080]; Based on this interaction, cell mass can be related to the measured phase retardation of light passing through each cell with 2% precision in total cell mass (Reed et al. (2011), [0080]).
Regarding claim 10, Teitell teaches the method according to claim 8 (see above), wherein the plurality of response parameters includes at least one selected from the group consisting of a specific growth rate, a half maximal effective concentration, a depth of response, a time of response, and a standard deviation of response (Teitell teaches the response parameters includes a depth of response: “the phase information is proportional to the optical path length (optical thickness) a depth profile and/or size/mass of sample can be calculated” [0088], “providing a substrate bearing a plurality of target cells”, [0007], “a sample chamber (imaging chamber) containing said substrate including a plurality of microwells [0039], while the light through the sample speeds up or slows down and brings a corresponding phase change. The phase delay or advance depends on the relation of the refraction index between the sample and surrounding environment. Since the phase information is proportional to the optical path length (optical thickness) a depth profile and/or size/mass of sample can be calculated, [0088]; which teaches the plurality of response parameters including at least a depth of response; Teitell also teaches a specific growth rate: “LCI provides quantitative maps of the mass distribution within target cells. These mass distributions from successive image frames can be integrated to yield measurements of target cell mass over time [0115], the measurements of target cell mass over time, gives a specific growth rate).
Regarding claim 11, Teitell teaches the method according to claim 8 (see above), wherein each therapy treatment sample of the plurality of therapy treatment samples includes live cancer cells exposed to a different therapeutic drug (a substrate is provided on which are disposed a plurality of target cells (e.g., cancer cells, cells infected with a pathogen, cells expressing a characteristic marker, cells transfected with a construct to recombinantly express a protein or peptide, etc.). The target cells are contacted with ctytotoxic T lymphocytes (CTLs) and those CTLs bearding a T cell receptor that recognizes/is activated by an antigen presented by the target cell(s) increase their mass. As the target cell is killed that cell shows a decrease in mass. Thus, an increase in mass of a T cell and/or a decrease in mass of a target cell is an indicator that the T cell bears a T cell receptor activated by antigens presented on the target cell, [0065];. mass of the T cell(s) and/or target cell(s) are detected using live cell interferometry (LCI). Live cell interferometry (LCI) is a label-free, quantitative phase microscopy technique that quantifies whole cell mass response within several hours and is uniquely suited to working with patient samples to identify TCRs against known or unknown antigens. [0077]).
Regarding claim 12, Teitell teaches the method according to claim 11 (see above), wherein the cell mass is a mass of the live cancer cells exposed to the different therapeutic drugs (identification of T cell receptors (TCRs) against known or unknown antigens is a major bottleneck in the development of cancer immune therapies .. the low frequency of TCRs directed against self-antigens, the low affinity of desired TCRs, and the small amount of tissue available per patient, [0063]).
Regarding claim 13, Teitell teaches the method according to claim 8 (see above), further comprising: determining, via the electronic controller, a phase shift of the amount of illumination passing through each therapy treatment sample of the plurality of therapy treatment samples (observation chamber can be controlled by computer .. for rapid, automated image acquisition, [0051]; changes in mass of the T cell(s) and/or target cell(s) are detected using live cell interferometry (LCI). Live cell interferometry (LCI) is a label-free, quantitative phase microscopy technique that quantifies whole cell mass response within several hours and is uniquely suited to working with patient samples to identify TCRs against known or unknown antigens. Briefly, the interaction of light with matter slows light as it passes through a cell, resulting in a measurable shift in phase. By quantifying this phase shift across the entire cell, the mass of the cell can be determined very precisely. It has been shown that LCI can be used to profile the mass response or mass accumulation rate of hundreds or thousands of cells simultaneously under controlled culture conditions, [0077]).
Regarding claim 14, Teitell teaches the method according to claim 8 (see above), further comprising: determining, via the electronic controller, a mass accumulation rate for each therapy treatment sample of the plurality of therapy treatment samples based on the determined cell mass and the period of time (The relative position of the microscope objective and observation chamber can be controlled by computer and can be translatable in three-dimensions allowing for rapid, automated image acquisition, [0051]; LCI can be used to profile the mass response or mass accumulation rate of hundreds or thousands of cells simultaneously under controlled culture conditions, [0077]).
Regarding claim 15, Teitell teaches a quantitative phase imaging microscope (refer to US 2016/0103118), comprising: a camera configured to capture images of a plurality of therapy treatment samples over a period of time; (Target cells are imaged by the LCI, [0083]; camera connected to an ordinary microscope with live cell imaging capability [0069], interference pattern is recorded, e.g., with a CCD camera, [0086]; Fig. 1 shows a CCD camera; the LCI comprises a Michelson-type interference microscope that compares the optical thickness of a reference cell to the optical thickness of samples placed in the observation chamber. Suspended in the observation chamber is a mirrored substrate, allowing the LCI to make measurements of optical thickness on transparent cells. The relative position of the microscope objective and observation chamber can be controlled by computer and can be translatable in three-dimensions allowing for rapid, automated image acquisition., [0051]) and
an electronic controller communicatively coupled to the quantitative phase imaging microscope (observation chamber can be controlled by computer, [0051], Fig. 1 shows objective control); the electronic controller configured to:
receive a plurality of images of the plurality of therapy treatment samples over the period of time from the camera (Live cell interferometry … a lateral shearing interferometric camera connected to an ordinary microscope with live cell imaging capability, screening can be performed on arrays of microwells … screening is performed on target cells disposed on a substrate…. screening can be performed on arrays of microwells, e.g., arrays comprising greater than 100, or greater than 1000, or greater than 10,000 microwells, [0069]; Target cells were imaged in standard culture media for 1.5 h prior to the start of each experiment to confirm the live cell culture imaging platform maintains viability of target cells in the absence of CTLs. M202 target cells showed a positive mass accumulation rate, indicating a healthy population and the maintenance of cell viability. (FIG. 3, panels D-E; FIG. 7, panel B). Control experiments demonstrated maintenance of both T and target cell viability during extended imaging periods (FIGS. 7 and 8), [0113]);
determine a cell mass for each therapy treatment sample of the plurality of therapy treatment samples based on each image of the plurality of images (live cell interferometer is capable of measuring the mass of both adherent and non-adherent cells, [0051]; the methods utilize label-free optical imaging to identify changes in mass of cells (e.g., an increase in mass of a T-cell and/or a decrease in mass of a target cell) as an indicator of T cell activation when T cells are presented with target cells bearing a cognate antigen, [0064]; the change in cell mass is determined using various interferometric and/or quantitative phase imaging microscopy techniques, [0068]);
track the cell mass over the period of time for each therapy treatment sample of the plurality of therapy treatment samples (target cells comprise cells of a cancer selected from the group consisting of breast cancer, central nervous system cancer, cervical cancer, colorectal cancer, testicular cancer, ovarian cancer, leukemia, a lymphoma, a melanoma, a soft tissue sarcoma, testicular cancer, and thyroid cancer, [0045]; By tracking T and target cell mass changes using label-free optical imaging methods, e.g., the methods permit direct measurements of the target and responding T cell during T cell mediated cytotoxicity to facilitate TCR cloning for use in adoptive immunotherapy against cancer. These methods can similarly be used to identify T-cells that are activated by cells infected with pathogens (e.g., cells infected with virus, bacteria, fungus, etc.), to identify cells that express (e.g., recombinantly express) particular peptides or proteins, and the like, [0066]); and
determine a plurality of response parameters for each therapy treatment sample of the plurality of therapy treatment samples based on the cell mass over the period of time (target cells comprise cells of a cancer selected from the group consisting of breast cancer, central nervous system cancer, cervical cancer, colorectal cancer, testicular cancer, ovarian cancer, leukemia, a lymphoma, a melanoma, a soft tissue sarcoma, testicular cancer, and thyroid cancer; [9945], Paragraphs [0045-0050] mentioned different cells; FIG. 5, panels A-F, illustrate LCI measurement of CTL mass and mass accumulation rate during T cell mediated cytotoxicity. Panel A: Mass versus time of an activated CTL and corresponding target cell. t=0 h is the point at which the target cell detaches from the substrate at the beginning of cell death. CTL+target cell refers to total mass of both cells in frames where they could not be measured separately. Panel B: Normalized mass versus time of 10 CTL-mediated cytotoxicity events. CTL mass is normalized relative to the mass at the time of target cell morphology change, which is used as the t=0 h point for all traces. Gray lines show best fit lines used for determining mass accumulation rates. Panel C: Average mass accumulation rate of CTLs before a cytotoxic event, during the first 100 minutes of a cytotoxic event, and after the first 100 minutes of a cytotoxic event demonstrating an approximately 4-fold increase in mass accumulation during the first 100 minutes of a cytotoxic event. Panel D: LCI image of 9 unresponsive and 1 cytotoxic T cell illustrating an approximately 3-fold difference in mass. The white arrow indicates the activated T cell, as determined by tracking this cell after persistent contact with target cell and subsequent target cell death. Panel E: The average mass of 116 activated CTLs is approximately 2.8-fold greater than the average mass of unresponsive controls. Panel F: Average area of activated CTLs is only approximately 1.4-fold greater than non-activated controls and not significant at the 95% confidence level, illustrating the utility of LCI mass measurements for determining CTL activation, [0055]; FIG. 7, panels A-D, shows averaged, normalized mass versus time plots for control target cell growth conditions showing robust growth on the LCI stage, and specificity of T cell mediated cytotoxicity. Panel A: Unaffected M202 cells (n=632) during treatment with F5 TCR transduced, CD8+ T cells. Panel B: M202 cells (n=117) prior to treatment with F5 TCR transduced, CD8+ T cells. Panel C: M202 cells (n=2058) treated with F5 TCR negative, CD8+ T cells. Panel D: Antigen-irrelevant, PC-3 prostate cancer cells treated with F5 TCR transduced, CD8+ T cells. Line shows mean normalized mass versus time (normalized relative to mass at first time point). Shaded region shows the mean +/−SD, [0057]).
Regarding claim 16, Teitell teaches the quantitative phase imaging microscope according to claim 15 (see above), wherein the electronic controller is configured to: determine a treatment response for each therapy treatment sample of the plurality of therapy treatment samples based on the plurality of response parameters (observation chamber can be controlled by computer … for rapid, automated image acquisition, [0051]; Live cell interferometry (LCI) is a label-free optical microscopy technique that measures whole cell responses. In certain embodiments LCI uses a Michelson-type interferometer, [0080]; Based on this interaction, cell mass can be related to the measured phase retardation of light passing through each cell with 2% precision in total cell mass (Reed et al. (2011), [0080]).
Regarding claim 17, Teitell teaches the quantitative phase imaging microscope according to claim 15 (see above), wherein the electronic controller is configured to: determine the plurality of response parameters for each therapy treatment sample of the plurality of therapy treatment samples in cells from a patient derived organoid (Target cells were imaged in standard culture media for 1.5 h prior to the start of each experiment to confirm the live cell culture imaging platform maintains viability of target cells in the absence of CTLs. M202 target cells showed a positive mass accumulation rate, indicating a healthy population and the maintenance of cell viability. [FIG. 3, panels D-E; FIG. 7, panel B], [0113]. With respect to therapy treatment samples in cells from a patient derived organoid: Teitell teaches “target cells (e.g., mammalian cells), [Abstract]; Mammalian cells are the fundamental building blocks of all mammals, including humans. “a substrate is provided on which are disposed a plurality of target cells (e.g., cancer cells, cells infected with a pathogen ..)”, [0065]; Teitell teaches Live cell interferometry (LCI) is a label-free, quantitative phase microscopy technique that quantifies whole cell mass response within several hours and is uniquely suited to working with patient samples to identify TCRs against known or unknown antigens, [0077]; introduce a quantitative method to track cells using live cell interferometry (LCI), a label-free microscopy technique … T and target cell mass changes provides a kinetic, quantitative assessment of T cell activation and a relatively rapid approach to identify specific, activated patient-derived T cells for applications in cancer immunotherapy, [0094]).
Regarding claim 18, Teitell teaches the quantitative phase imaging microscope according to claim 15 (see above), wherein the electronic controller is configured to: determine a difference in the plurality of response parameters between each therapy treatment sample of the plurality of treatment samples (When passing through a relatively transparent sample, the intensity of the light changes very little, while the light through the sample speeds up or slows down and brings a corresponding phase change, [0088]); Fig. 1 a sample assembly comprising an observation chamber adapted to contain the cell, a reference assembly comprising a reference chamber adapted to contain the reference cell, and a beam splitter for splitting a light beam from a light source into a test beam and a reference beam; [0085]);
Regarding claim 19, Teitell teaches the quantitative phase imaging microscope according to claim 18 (see above), wherein the difference in the plurality of response parameters is determined from a tumor organoid (Live cell interferometry can be used to optically profile the mass response of a co-culture of target cells and candidate CTLs, screening can be performed on arrays of microwells [0069]; growing body of work has focused on the identification of tumor infiltrating T lymphocytes (TILs) bearing TCR recognition of autologous tumor cells … LCI may therefore provide a viable alternative for the identification and isolation of rare effector T cells killing autologous tumor cells, [0123]; [0080]; [abstract]; target cells comprise cells of a cancer selected from the group consisting of astrocytoma’s, atypical teratoid/rhabdoid tumor, pituitary tumor, trophoblastic tumor, Wilm's tumor …, [claim 39]).
Regarding claim 20, Teitell teaches the quantitative phase imaging microscope according to claim 15 (see above), wherein the electronic controller (controlled by a computer, [0051]) is configured to: determine a tumor cell from a non-tumor cell for each therapy treatment sample of the plurality of therapy treatment samples based on the cell mass over the period of time, (potential application of the LCI technique presented here is for the identification and isolation of single and potentially rare CTLs. A growing body of work has focused on the identification of tumor infiltrating T lymphocytes (TILs) bearing TCR recognition of autologous tumor cells, LCI imaging platform is fundamentally compatible with a segmented culture system that will allow for isolation of rare cells that may be lost in the current open perfusion cell culture system. LCI may therefore provide a viable alternative for the identification and isolation of rare effector T cells killing autologous tumor cells or HLA-matched cancer cell lines, [0123]; The capacity to measure the mass of a single CTL opens several potential downstream applications including T cell biological studies pertaining to metabolic or differentiation states in addition to the analysis of CTLs for potential use in adoptive immunotherapy protocols, [0084]).
Claim Rejections - 35 USC § 103
The following is a quotation of 35 U.S.C.103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
Claims 1-7 are rejected under 35 U.S.C. 103 as being unpatentable over
TEITELL et al. (US 2016/0103118), in view of NPL Lei Tian et al. “Quantitative differential phase contrast imaging in an LED array microscope”, OPTICS EXPRESS 11394, Vol. 23, No. 9, May 2015, (of record; copy attached), and further in view of Pattison et al. (US 2021/0157114).
Regarding claim 1, Teitell teaches a microscope imaging system (refer to US 2016/0103118) comprising:
a quantitative phase imaging microscope (the change in cell mass is determined using various interferometric and/or quantitative phase imaging microscopy techniques, [0068]) including
a stage (observation chamber, [0031], Fig. 1) configured to hold and move a well plate (The relative position of the microscope objective and observation chamber can .. be translatable in three-dimensions [0051]; cells are disposed in microwells, [0017], [0069]; the chamber configured to hold the microwells, Fig. 1; the observation chamber comprises at least one perfusion conduit adapted to circulate a cell media within the chamber, [0085]) having a plurality of therapy treatment samples (samples placed in the observation chamber, [0051]; T cell(s) and/or target cell(s) are detected, is uniquely suited to working with patient samples to identify TCRs against known or unknown antigens, [0077], Fig. 1);
a light emitting diodes (LED light source, see Fig. 1) configured to illuminate the plurality of therapy treatment samples (light passing through the tested object. When passing through a relatively transparent sample, the intensity of the light changes very little, while the light through the sample speeds up or slows down and brings a corresponding phase change, [0088]); Fig. 1 a sample assembly comprising an observation chamber adapted to contain the cell, a reference assembly comprising a reference chamber adapted to contain the reference cell, and a beam splitter for splitting a light beam from a light source into a test beam and a reference beam; [0085]);
a camera (Target cells are imaged by the LCI, [0083]; camera connected to an ordinary microscope with live cell imaging capability [0069], interference pattern is recorded, e.g., with a CCD camera, [0086]; Fig. 1 shows a CCD camera) configured to capture images of the plurality of therapy treatment samples over a period of time using an amount of illumination provided by the LEDs (FIG. 9, panels A-H, shows intensity images of cells on the interferometer stage after 18 h of imaging showing typical target cell conditions, Left column shows the full image frame, [0059]; Target cells were imaged in standard culture media for 1.5 h prior to the start of each experiment to confirm the live cell culture imaging platform maintains viability of target cells in the absence of CTLs, [0113], observe how the mass of the cell(s) changes over a period of time, [0039]); and an electronic controller (observation chamber can be controlled by computer and … for rapid, automated image acquisition, [0051]; communicatively coupled to the quantitative phase imaging microscope (observation chamber can be controlled by computer, [0051], Fig. 1 shows objective control); the electronic controller configured to:
receive a plurality of images of the plurality of therapy treatment samples over the period of time from the camera (Live cell interferometry … a lateral shearing interferometric camera connected to an ordinary microscope with live cell imaging capability, screening is performed on target cells disposed on a substrate…. screening can be performed on arrays of microwells, e.g., arrays comprising greater than 100, or greater than 1000, or greater than 10,000 microwells, [0069]; Target cells were imaged in standard culture media for 1.5 h prior to the start of each experiment to confirm the live cell culture imaging platform maintains viability of target cells in the absence of CTLs. M202 target cells showed a positive mass accumulation rate, indicating a healthy population and the maintenance of cell viability. (FIG. 3, panels D-E; FIG. 7, panel B). Control experiments demonstrated maintenance of both T and target cell viability during extended imaging periods (FIGS. 7 and 8), [0113]);
determine a cell mass for each therapy treatment sample of the plurality of therapy treatment samples based on each image of the plurality of images (live cell interferometer is capable of measuring the mass of both adherent and non-adherent cells, [0051]; the methods utilize label-free optical imaging to identify changes in mass of cells (e.g., an increase in mass of a T-cell and/or a decrease in mass of a target cell) as an indicator of T cell activation when T cells are presented with target cells bearing a cognate antigen, [0064]; the change in cell mass is determined using various interferometric and/or quantitative phase imaging microscopy techniques, [0068]);
track the cell mass over the period of time for each therapy treatment sample of the plurality of therapy treatment samples (target cells comprise cells of a cancer selected from the group consisting of breast cancer, central nervous system cancer, cervical cancer, colorectal cancer, testicular cancer, ovarian cancer, leukemia, a lymphoma, a melanoma, a soft tissue sarcoma, testicular cancer, and thyroid cancer, [0045]; By tracking T and target cell mass changes using label-free optical imaging methods, e.g., the methods permit direct measurements of the target and responding T cell during T cell mediated cytotoxicity to facilitate TCR cloning for use in adoptive immunotherapy against cancer. These methods can similarly be used to identify T-cells that are activated by cells infected with pathogens (e.g., cells infected with virus, bacteria, fungus, etc.), to identify cells that express (e.g., recombinantly express) particular peptides or proteins, and the like, [0066]);
determine a plurality of response parameters for each therapy treatment sample of the plurality of therapy treatment samples based on the cell mass over the period of time; and determine a treatment response for each therapy treatment sample of the plurality of therapy treatment samples based on the plurality of response parameters (target cells comprise cells of a cancer selected from the group consisting of breast cancer, central nervous system cancer, cervical cancer, colorectal cancer, testicular cancer, ovarian cancer, leukemia, a lymphoma, a melanoma, a soft tissue sarcoma, testicular cancer, and thyroid cancer; [9945], Paragraphs [0045-0050] mentioned different cells; FIG. 5, panels A-F, illustrate LCI measurement of CTL mass and mass accumulation rate during T cell mediated cytotoxicity. Panel A: Mass versus time of an activated CTL and corresponding target cell. t=0 h is the point at which the target cell detaches from the substrate at the beginning of cell death. CTL+target cell refers to total mass of both cells in frames where they could not be measured separately. Panel B: Normalized mass versus time of 10 CTL-mediated cytotoxicity events. CTL mass is normalized relative to the mass at the time of target cell morphology change, which is used as the t=0 h point for all traces. Gray lines show best fit lines used for determining mass accumulation rates. Panel C: Average mass accumulation rate of CTLs before a cytotoxic event, during the first 100 minutes of a cytotoxic event, and after the first 100 minutes of a cytotoxic event demonstrating an approximately 4-fold increase in mass accumulation during the first 100 minutes of a cytotoxic event. Panel D: LCI image of 9 unresponsive and 1 cytotoxic T cell illustrating an approximately 3-fold difference in mass. The white arrow indicates the activated T cell, as determined by tracking this cell after persistent contact with target cell and subsequent target cell death. Panel E: The average mass of 116 activated CTLs is approximately 2.8-fold greater than the average mass of unresponsive controls. Panel F: Average area of activated CTLs is only approximately 1.4-fold greater than non-activated controls and not significant at the 95% confidence level, illustrating the utility of LCI mass measurements for determining CTL activation, [0055]; FIG. 7, panels A-D, shows averaged, normalized mass versus time plots for control target cell growth conditions showing robust growth on the LCI stage, and specificity of T cell mediated cytotoxicity. Panel A: Unaffected M202 cells (n=632) during treatment with F5 TCR transduced, CD8+ T cells. Panel B: M202 cells (n=117) prior to treatment with F5 TCR transduced, CD8+ T cells. Panel C: M202 cells (n=2058) treated with F5 TCR negative, CD8+ T cells. Panel D: Antigen-irrelevant, PC-3 prostate cancer cells treated with F5 TCR transduced, CD8+ T cells. Line shows mean normalized mass versus time (normalized relative to mass at first time point). Shaded region shows the mean +/−SD, [0057]).
Teitell doesn’t explicitly teach using an array of light emitting diodes and microscope stage holds and move.
Teitell and Tian are related as quantitative phase imaging microscope.
Tian teaches using an array of light emitting diodes (Quantitative differential phase contrast imaging in an LED array microscope, with an LED array microscope, which uses computational illumination for flexibly achieving a diverse array of imaging capabilities, Each LED can be controlled individually to illuminate the sample from a unique angle [in para: introduction]). It would have been obvious to one of ordinary skill in the art at the time the application was filed to modify the microscope imaging system of Teitell to replace the microscope’s illumination unit with an LED array as taught by Tian for the predictable advantage of computational illumination for flexibly achieving a diverse array of imaging capabilities, phase contrast, flexible patterning of the LED array will be able to implement DPC measurements in real-time and along arbitrary axes of asymmetry, without any mechanical parts, Each LED can be controlled individually to illuminate the sample from a unique angle [in para: introduction, [in introduction] as taught by Tian [in para introduction].
The modified Teitell doesn’t explicitly teach that the stage holds and move.
Teitell and Pattison are related as imaging microscope.
Pattison teaches the microscope stage holds and move, (either one or both of the objective 7 and the stage 20 may be movable, [0045]; a controller 50 for controlling movement of the objective 7 and/or of the stage 20 as applicable, [0046]).
It would have been obvious to one of ordinary skill in the art at the time the application was filed to modify the modified microscope imaging system of Teitell to configure the stage movable, as taught by Pattison for the predictable advantage of change adjust the focus of the apparatus accurately.
Regarding claim 2, the modified Teitell teaches the microscope imaging system according to claim 1 (see above), wherein the plurality of response parameters include at least one selected from the group consisting of a specific growth rate, a half maximal effective concentration, a depth of response, a time of response, and a standard deviation of response (Since the phase information is proportional to the optical path length (optical thickness) a depth profile and/or size/mass of sample can be calculated, [0088]).
Regarding claim 3, the modified Teitell teaches the microscope imaging system according to claim 1 (see above), wherein each therapy treatment sample of the plurality of therapy treatment samples includes live cancer cells exposed to a different therapeutic drug (a substrate is provided on which are disposed a plurality of target cells (e.g., cancer cells, cells infected with a pathogen, cells expressing a characteristic marker, cells transfected with a construct to recombinatnly express a protein or peptide, etc.). The target cells are contacted with ctytotoxic T lymphocytes (CTLs) and those CTLs bearding a T cell receptor that recognizes/is activated by an antigen presented by the target cell(s) increase their mass. As the target cell is killed that cell shows a decrease in mass. Thus, an increase in mass of a T cell and/or a decrease in mass of a target cell is an indicator that the T cell bears a T cell receptor activated by antigens presented on the target cell, [0065];. mass of the T cell(s) and/or target cell(s) are detected using live cell interferometry (LCI). Live cell interferometry (LCI) is a label-free, quantitative phase microscopy technique that quantifies whole cell mass response within several hours and is uniquely suited to working with patient samples to identify TCRs against known or unknown antigens. [0077]).
Regarding claim 4, the modified Teitell teaches the microscope imaging system according to claim 1 (see above), wherein the cell mass is a mass of the live cancer cells exposed to the different therapeutic drugs (identification of T cell receptors (TCRs) against known or unknown antigens is a major bottleneck in the development of cancer immune therapies .. the low frequency of TCRs directed against self-antigens, the low affinity of desired TCRs, and the small amount of tissue available per patient, [0063]).
Regarding claim 5, the modified Teitell teaches the microscope imaging system according to claim 1 (see above), wherein the quantitative phase imaging microscope further includes: a lens configured to focus the camera on the plurality of therapy treatment samples (Live cell interferometry (or another quantitative phase imaging technique, including digital holographic microscopy, a lateral shearing interferometric camera connected to a ordinary microscope with live cell imaging capability, and the like) can be used to optically profile the mass response of a co-culture of target cells and candidate CTLs. [0069]).
Regarding claim 6, the modified Teitell teaches the microscope imaging system according to claim 1 (see above), wherein when determining the cell mass for each therapy treatment sample of the plurality of therapy treatment samples, the electronic controller (observation chamber can be controlled by computer for rapid, automated image acquisition, [0051];) is further configured to: determine a phase shift of the amount of illumination passing through each therapy treatment sample of the plurality of therapy treatment samples (changes in mass of the T cell(s) and/or target cell(s) are detected using live cell interferometry (LCI). Live cell interferometry (LCI) is a label-free, quantitative phase microscopy technique that quantifies whole cell mass response within several hours and is uniquely suited to working with patient samples to identify TCRs against known or unknown antigens. Briefly, the interaction of light with matter slows light as it passes through a cell, resulting in a measurable shift in phase. By quantifying this phase shift across the entire cell, the mass of the cell can be determined very precisely. It has been shown that LCI can be used to profile the mass response or mass accumulation rate of hundreds or thousands of cells simultaneously under controlled culture conditions, [0077]).
Regarding claim 7, the modified Teitell teaches the microscope imaging system according to claim 1 (see above), wherein when tracking the cell mass over the period of time for each therapy treatment sample of the plurality of therapy treatment samples, the electronic controller is further configured to: determine a mass accumulation rate for each therapy treatment sample of the plurality of therapy treatment samples based on the determined cell mass and the period of time. (The relative position of the microscope objective and observation chamber can be controlled by computer and can be translatable in three-dimensions allowing for rapid, automated image acquisition, [0051]; LCI can be used to profile the mass response or mass accumulation rate of hundreds or thousands of cells simultaneously under controlled culture conditions, [0077]).
Conclusion
The prior art made of record and not relied upon is considered pertinent to applicant's disclosure. 1. Gimzewski et al. (US 9810683). 2. Wagner (US 20130252237), 3. Wong et al (US 20110092762). These prior arts teach Live Cell Interferometry, microscopy, quantitative measurements, Observation Chamber or stage, response, result or feedback, capturing image of data, controller, track, cell size and shape and period of time.
Changing claim 2 and claim 10 claim language from “include at least one selected from the group consisting of” to “include the group consisting of” will overcome the prior art in record.
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/R.A/Examiner, Art Unit 2872
/BUMSUK WON/Supervisory Patent Examiner, Art Unit 2872
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