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 .
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
Restriction election
Applicant’s election without traverse of invention Group I (claims 1-2, 7-8, 12, 20, 30, 42-43, 46-47, 52-54, 58, 60-61, 66-67, 69-70, 73-76, 80-85, 90-91, 93, 95-96, 98, 100-101, 106-108, and 110-114), applicant’s election without traverse of species Group I (In vitro method; claims 46-47), applicant’s election without traverse of species Group II (Z-scores ; claim 70), applicant’s election without traverse of species Group III (Percentage of expression levels; claim 83), applicant’s election without traverse of species Group IV (Bulk RNA; claim 95), and applicant’s election without traverse of species Group V (Autologous; claim 113) in the reply filed on 12/15/2025 is acknowledged.
Claim 3 is withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a non-elected invention.
Claim Status
Claims 1-3, 7-8, 12, 20, 30, 42-43, 46-47, 52-54, 58, 60-61, 66-67, 69-70, 73-76, 80-85, 90-91, 93, 95-96, 98, 100-101, 106-108, and110-114 are pending.
Claims 4-6, 9-11, 13-19, 21-29, 31-41, 44-45, 48-51, 55-57, 59, 62-65, 68, 71-72, 77-79, 59-89, 92, 94, 97, 99, 102-105, 109, and 116-117 are canceled.
Claims 3, 42-43, 67, 69, 74-76, 80-82, 85, 90-91, 96, and 114 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a non-elected invention, as described above.
Claims 1-2, 7-8, 12, 20, 30, 46-47, 52-54, 58, 60-61, 66, 70, 73, 83-84, 93, 95, 98, 100-101, 106-108, and 110-113 are under examination.
Claims 1-2, 7, 12, 47, 60-61, 83-84, 101, and 112 are objected to.
Claims 1-2, 7-8, 12, 20, 30, 46-47, 52-54, 58, 60-61, 66, 70, 73, 83-84, 93, 95, 98, 100-101, 106-108, and 110-113 are rejected.
Priority
Applicant's claim for the benefit of a prior-filed application, PCT/US2020/043627, filed 07/24/2020, which claims benefit of domestic benefit to US provisional application 62/878701, filed 07/25/2019 is acknowledged.
Information Disclosure Statement
The information disclosure statements (IDS) filed on 11/04/2022 and 10/17/2023 are in compliance with the provisions of 37 CFR 1.97 and have therefore been considered. Signed copies of the IDS documents are included with this Office Action.
Drawings
The Drawings submitted 01/24/2022 are accepted.
Specification
The disclosure is objected to for the following informalities. It is noted that for purposes of the instant Office Action, any reference to the specification pertains to the clean copy of the substitute specification as originally filed on 01/24/2022.
Hyperlinks
The disclosure is objected to because it contains an embedded hyperlink and/or other form of browser-executable code. Applicant is required to delete the embedded hyperlink and/or other form of browser-executable code; references to websites should be limited to the top-level domain name without any prefix such as http:// or other browser-executable code. See MPEP § 608.01. Non-limiting examples include paragraphs [00303] and [00385]. Applicant will note that this is exemplary and other instances may exist. It is requested that all instances be corrected.
Trade Names
The use of the terms Ultimate™, Trizol®, Sybr®, as example, which are trade names or marks used in commerce, have been noted in this application. Each term should be accompanied by corresponding generic terminology; furthermore the terms should be capitalized wherever they appear or, where appropriate, include a proper symbol indicating use in commerce such as ™, ™ , or ® following the terms. Applicant is requested to review the Specification and Drawings for all instances. For example Trizol [0088].
Although the use of trade names and marks used in commerce (i.e., trademarks, service marks, certification marks, and collective marks) is permissible in patent applications, the proprietary nature of the marks should be respected and every effort made to prevent their use in any manner which might adversely affect their validity as commercial marks.
Appropriate correction for all objections to the specification is required.
Claim Rejections - 35 USC § 101
35 U.S.C. 101 reads as follows:
Whoever invents or discovers any new and useful process, machine, manufacture, or composition of matter, or any new and useful improvement thereof, may obtain a patent therefor, subject to the conditions and requirements of this title.
A. Claims 1-2, 7-8, 12, 20, 30, 46-47, 52-54, 58, 60-61, 66, 70, 73, 83-84, 93, 95, 98, 101, and 110-111 are rejected under 35 U.S.C. 101 because the claimed invention is directed to one or more judicial exceptions without significantly more.
MPEP 2106 organizes judicial exception analysis into Steps 1, 2A (Prongs One and Two) and 2B as follows below. MPEP 2106 and the following USPTO website provide further explanation and case law citations: uspto.gov/patent/laws-and-regulations/examination-policy/examination-guidance-and-training-materials.
Framework with which to Evaluate Subject Matter Eligibility:
Step 1: Are the claims directed to a process, machine, manufacture, or composition of matter;
Step 2A, Prong One: Do the claims recite a judicially recognized exception, i.e. a law of nature, a natural phenomenon, or an abstract idea;
Step 2A, Prong Two: If the claims recite a judicial exception under Prong One, then is the judicial exception integrated into a practical application (Prong Two); and
Step 2B: If the claims do not integrate the judicial exception, do the claims provide an inventive concept.
Framework Analysis as Pertains to the Instant Claims:
Step 1
With respect to Step 1: yes, the claims are directed to methods and product, i.e., a process, machine, or manufacture within the above 101 categories [Step 1: YES; See MPEP § 2106.03].
Step 2A, Prong One
With respect to Step 2A, Prong One, the claims recite judicial exceptions in the form of abstract ideas. The MPEP at 2106.04(a)(2) further explains that abstract ideas are defined as:
mathematical concepts (mathematical formulas or equations, mathematical relationships and mathematical calculations);
certain methods of organizing human activity (fundamental economic practices or principles, managing personal behavior or relationships or interactions between people); and/or
mental processes (procedures for observing, evaluating, analyzing/ judging and organizing information).
With respect to the instant claims, under the Step 2A, Prong One evaluation, the claims are found to recite abstract ideas that fall into the grouping of mental processes (in particular procedures for observing, analyzing and organizing information) and mathematical concepts (in particular mathematical relationships and formulas) are as follows:
Independent claim 1:
applying the expression levels of the one or more metagenes as input to a process configured to determine a probability of the cell or the plurality of cells having metagene expression levels of a determined dopaminergic precursor cell;
determining a deviation score for the cell or the plurality of cells, wherein the deviation score indicates the degree to which the gene expression levels in the test dataset deviate from gene expression levels in one or more reference cells in the reference database, wherein the one or more reference cells are at a stage of differentiation indicating a determined dopaminergic precursor cell;
Dependent claim 53:
determined using regression analysis based on (i) the one or more metagenes determined from the one or more reference cells in the reference database and (ii) the gene expression levels in the test dataset.
Dependent claim 54:
determined by merging the gene expression levels in the test dataset with the reference database to create an updated reference database
applying the dimensionality reduction technique on the updated reference database.
Dependent claim 101:
applying the expression levels of the one or more metagenes as input to a process configured to determine a probability of the cell or the plurality of cells having metagene expression levels of a determined dopaminergic precursor cell;
determining a deviation score for the cell or the plurality of cells, wherein the deviation score indicates the degree to which the gene expression levels in the test dataset deviate from gene expression levels in one or more reference cells in the reference database, wherein the one or more reference cells are at a stage of differentiation indicating a determined dopaminergic precursor cell
Dependent claims 2, 7-8, 20, 46, 52, 60-61, 66, 70, 73, 83-84, 93, 95, 98, 110-111, and 113 recite further steps that limit the judicial exceptions in independent claim 1 and, as such, also are directed to those abstract ideas. For example, claim 2 further limits the process of claim 1, claim 7 further limits the reference cells of claim 1, claim 8 further limits the in vitro population of claim 1, claim 20 further limits the reference database of claim 1, claim 46 further limits the class label of claim 2, claim 52 further limits expression levels of claim 1, claim 60 further limits the label classification, claim 61 further limits the threshold of claim 60, claim 66 further limits deviation score of claim 1, claim 70 further limits the single-gene deviation, claim 73 further limits the gene expression levels of claim 1, claim 83 further limits label classification of claim 1, claim 84 further limits the label classification of claim 1, claim 93 further limits gene expression levels of claim 1, claim 95 further limits the RNA sequencing of claim 93, claim 95 further limits the receiving of the test dataset of claim 1, claims 110-111 further limit the population of determined cells of claim 107, claim 113 further limit the population of determined cells of claim 1.
The abstract ideas recited in the claims are evaluated under the Broadest Reasonable Interpretation (BRI) and determined to each cover performance either in the mind and/or by mathematical operation because the method only requires a user to manually determine and merge. Without further detail as to the methodology involved in “determine a probability of the cell or the plurality of cells having metagene expression levels of a determined dopaminergic precursor cell”, “determining a deviation score for the cell or the plurality of cells”, “determined using regression analysis”, “merging the gene expression levels”, “applying the dimensionality reduction technique”, “determine a probability of the cell or the plurality of cells”, and “determining a deviation score for the cell or the plurality of cells”, under the BRI, one may simply, for example, use pen and paper to identifying dopaminergic neurons and progenitor cells.
Therefore, 1-2, 7-8, 12, 20, 30, 46-47, 52-54, 58, 60-61, 66, 70, 73, 83-84, 93, 95, 98, 100-101, 106, and 110-111 recite an abstract idea [Step 2A, Prong 1: YES; See MPEP § 2106.04].
Step 2A, Prong Two
Because the claims do recite judicial exceptions, direction under Step 2A, Prong Two, provides that the claims must be examined further to determine whether they integrate the judicial exceptions into a practical application (MPEP 2106.04(d)). A claim can be said to integrate a judicial exception into a practical application when it applies, relies on, or uses the judicial exception in a manner that imposes a meaningful limit on the judicial exception. This is performed by analyzing the additional elements of the claim to determine if the judicial exceptions are integrated into a practical application (MPEP 2106.04(d).I.; MPEP 2106.05(a-h)). If the claim contains no additional elements beyond the judicial exceptions, the claim is said to fail to integrate the judicial exceptions into a practical application (MPEP 2106.04(d).III).
Additional elements, Step 2A, Prong Two
With respect to the instant recitations, the claims recite the following additional elements:
Independent claim 1 and dependent claim 101:
receiving a test dataset comprising (a)gene expression levels, and (b) expression levels of one or more metagenes for a cell or a plurality of cells comprised in an in vitro population of neuronal progenitor cells, wherein the one or more metagenes are determined based on correlated gene expression levels of reference cells in a reference database, wherein the reference cells are neuronal cells at one or more different stages of differentiation;
outputting, based on the probability and the deviation score, a computed label classification comprising an indication of whether said cell or said plurality of cells from the in vitro population of neuronal progenitor cells is a determined dopaminergic precursor cell.
Dependent claim 46:
using an in vitro method
Dependent claims 12, 47 recite steps that further limit the recited additional elements in the claims. For example, claim 12 further limits culturing of claim 8, claim 47 further limits in vitro method of claim 46;
Considerations under Step 2A, Prong Two
With respect to Step 2A, Prong Two, the additional elements of the claims do not integrate the judicial exceptions into a practical application for the following reasons. Those steps directed to data gathering, such as “receiving”, and to data outputting, such as “outputting”, perform functions of collecting the data needed to carry out the judicial exceptions. Data gathering and outputting do not impose any meaningful limitation on the judicial exceptions, or on how the judicial exceptions are performed. Data gathering and outputting steps are not sufficient to integrate judicial exceptions into a practical application (MPEP 2106.05(g)).
With respect to claim 46, the additional elements of the claims do not integrate the judicial exceptions into a practical application for the following reasons. Those steps directed to culturing one or more iPSC in vitro do not impose any meaningful limitations on the abstract idea, or on how the abstract idea is performed. These steps are insignificant extra-solution activity to the judicial exception (MPEP 2106.05(g)(2)).
Thus, none of the claims recite additional elements which would integrate a judicial exception into a practical application, and the claims are directed to one or more judicial exceptions [Step 2A, Prong 2: NO; See MPEP § 2106.04(d)].
Step 2B (MPEP 2106.05.A i-vi)
According to analysis so far, the additional elements described above do not provide significantly more than the judicial exception. A determination of whether additional elements provide significantly more also rests on whether the additional elements or a combination of elements represents other than what is well-understood, routine, and conventional. Conventionality is a question of fact and may be evidenced as: a citation to an express statement in the specification or to a statement made by an applicant during prosecution that demonstrates a well-understood, routine or conventional nature of the additional element(s); a citation to one or more of the court decisions as discussed in MPEP 2106(d)(II) as noting the well-understood, routine, conventional nature of the additional element(s); a citation to a publication that demonstrates the well-understood, routine, conventional nature of the additional element(s); and/or a statement that the examiner is taking official notice with respect to the well-understood, routine, conventional nature of the additional element(s).
With respect to the instant claims, the courts have found that receiving and outputting data are well-understood, routine, and conventional functions of a computer when claimed in a merely generic manner or as insignificant extra-solution activity (see Symantec, 838 F.3d at 1321, 120 USPQ2d at 1362 (utilizing an intermediary computer to forward information), buySAFE, Inc. v. Google, Inc., 765 F.3d 1350, 1355, 112 USPQ2d 1093, 1096 (Fed. Cir. 2014) (computer receives and sends information over a network), Versata Dev. Group, Inc. v. SAP Am., Inc., 793 F.3d 1306, 1334, 115 USPQ2d 1681, 1701 (Fed. Cir. 2015), and OIP Techs., 788 F.3d at 1363, 115 USPQ2d at 1092-93, as discussed in MPEP 2106.05(d)(II)(i)).
As such, the claims simply append well-understood, routine, conventional activities previously known to the industry, specified at a high level of generality, to the judicial exception (MPEP2106.05(d)). The data gathering steps as recited in the instant claims constitute a general link to a technological environment which is insufficient to constitute an inventive concept which would render the claims significantly more than the judicial exception (MPEP2106.05(g)&(h)).
With respect to claims 46, the methods of culturing one or more iPSC in vitro are well-understood, routine, and conventional in the art. The specification discloses methods for generating iPSCs are known [00169] and the in vitro cell population of neuronal progenitor cells provided herein may be formed by methods commonly known and used in the art to differentiate dopaminergic neurons from iPSCs [0067], and any available and known method for inducing differentiation of the cells, e.g., pluripotent stem cells, into floor plate midbrain progenitor cells, determined dopaminergic precursor cells, and/or dopamine (DA) neurons can be used [00170].
Taken alone, the additional elements do not amount to significantly more than the above-identified judicial exception(s). Even when viewed as a combination, the additional elements fail to transform the exception into a patent-eligible application of that exception. Thus, the claims as a whole do not amount to significantly more than the exception itself [Step 2B: NO; See MPEP § 2106.05].
Therefore, the instant claims are not drawn to eligible subject matter as they are directed to one or more judicial exceptions without significantly more. For additional guidance, applicant is directed generally to the MPEP § 2106.
B. With respect to claims 100 and 106, the claimed invention is further directed to non-statutory subject matter. The claim(s) does/do not fall within at least one of the four categories of patent eligible subject matter because they recite a natural product and a storage media. Therefore, the claims read on carrier waves and include transitory propagating signals. (In re Nuijten, Federal Circuit, 2006). It is noted that the recitation of a "non-transitory computer-readable medium" would overcome the rejection with respect to this issue under 101.
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.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
A. Claim(s) 1-2, 7-8, 12, 20, 30, 46, 52, 54, 58, 60, 66, 70, 83-84, 93, 95, 98, 100-101, 106-108, and 111-112 is/are rejected under 35 U.S.C. 103 as being unpatentable over Loring et al. (US Pub 2011/118130 A1, published 05/19/2011, cited on IDS 10/17/2023) in view of Qi et al.( WO 2017/132596 A1, published 03/17/2017, cited on IDS dated 11/04/2022) .
Claim 1 is directed to a computer implemented method of classifying an in vitro population of neuronal progenitor cells, the method comprising: receiving a test dataset comprising (a)gene expression levels, and (b) expression levels of one or more metagenes for a cell or a plurality of cells comprised in an in vitro population of neuronal progenitor cells, wherein the one or more metagenes are determined based on correlated gene expression levels of reference cells in a reference database, wherein the reference cells are neuronal cells at one or more different stages of differentiation; applying the expression levels of the one or more metagenes as input to a process configured to determine a probability of the cell or the plurality of cells having metagene expression levels of a determined dopaminergic precursor cell; determining a deviation score for the cell or the plurality of cells, wherein the deviation score indicates the degree to which the gene expression levels in the test dataset deviate from gene expression levels in one or more reference cells in the reference database, wherein the one or more reference cells are at a stage of differentiation indicating a determined dopaminergic precursor cell; and outputting, based on the probability and the deviation score, a computed label classification comprising an indication of whether said cell or said plurality of cells from the in vitro population of neuronal progenitor cells is a determined dopaminergic precursor cell.
Loring discloses compositions and methods for defining cells [title]. Loring further discloses a strategy for classifying stem cells using a computer implemented method. The method is comprising several steps for assaying a cell such as: receiving a reference database, obtaining and associating the data with labeled associated biological classes of the cells, receiving a test dataset, and obtaining and merging the test dataset into the database.
Loring discloses an example with brain-derived cells from sixty-two of the samples which were described as neural stem or progenitor cells based on their source, culture methods and classical markers [0193]. Loring further discloses most of the designated neural stem cells were distributed among multiple clusters, indicating a great deal of diversity in neural stem cell preparations [0193]. Loring also discloses that one group of the brain-derived lines, those derived from surgical specimens from living patients (HANSE cells, see below), remained together throughout the iterative clustering (FIG. 2, cluster 6; see also FIG. 4) [0193]. Loring further discloses the HANSE cell group consisted of transcriptional profiles that were derived from neurosurgical specimens following published protocols for multipotent neural progenitor derivation and propagation [0193]. Loring also discloses machine learning process as disclosed in [0193]: the method is comprising ways to sort the cell types with an unsupervised machine learning approach to cluster transcriptional profiles of the cell preparations into stable distinct groups. Loring also discloses a method of characterizing cells comprising: a. global profiling of known and unknown samples; b. collecting reference data from known samples into a matrix database; c. merging the unknown sample data with the matrix database; d. grouping the cells with a machine learning algorithm; e. determining the identity of the unknown sample based on its similarity to the known sample matrix [claim 21].
Claim 1 differs from this known method of Loring in that it deals with a population of in vitro neuronal progenitor cells in order to identify those which might differentiate into a determined dopaminergic precursor cell (for a later medical use for treating neuro-degenerative diseases).
However, Qi discloses differentiation of cortical neurons from human pluripotent stem cells [title]. Qi further discloses an in vitro method for inducing differentiation of human stem cells into cortical neurons (and proximate precursors thereof) comprises contacting a population of human stem cells with an effective amount of one or more selected compounds in order to express detectable levels of PAX6 at least 6 days after initiation [p. 2, par. 3] which read on neuronal progenitor cells as disclosed in the specification [p. 3, par. 5].
Claim 2 is directed to the computer implemented method of claim 1, wherein: the process comprises a supervised classification model trained using (i) expression levels of the one or more metagenes of the reference cells in the reference database; and (ii) class labels indicating each of the one or more different stages of differentiation for reference cells in the reference database, to determine a probability of a cell or a plurality of cells having metagene expression levels of a determined dopaminergic precursor cell.
Loring discloses methods, wherein the sample is a cell line, primary cell or tissue; wherein the global profile is a transcriptional profile; wherein the matrix consists of known class labels; wherein the machine learning algorithm is an unsupervised classification; wherein the unsupervised machine learning algorithm comprises a bootstrapping sparse non-negative matrix factorization; wherein the machine learning algorithm is a supervised classification; wherein the method is a computer implemented method; further comprising the step of outputting results from the unsupervised classification; and/or any limitation or characteristic disclosed herein alone or in combination [0171].
Claim 7 is directed to the computer implemented method of claim 1 of, wherein the reference cells are an in vitro population of neuronal progenitor cells.
Claim 7 differs from the known method of Loring in that it deals with a population of in vitro neuronal progenitor cells in order to identify those which might differentiate into a determined dopaminergic precursor cell (for a later medical use for treating neuro-degenerative diseases).
However, Qi discloses differentiation of cortical neurons from human pluripotent stem cells [title]. Qi further discloses an in vitro method for inducing differentiation of human stem cells into cortical neurons (and proximate precursors thereof) comprises contacting a population of human stem cells with an effective amount of one or more selected compounds in order to express detectable levels of PAX6 at least 6 days after initiation [p. 2, par. 3] which read on neuronal progenitor cells as disclosed in the specification [p. 3, par. 5].
Claim 8 is directed to the computer implemented method of claim 1, wherein said in vitro population of neuronal progenitor cells is formed by culturing one or more induced pluripotent stem cells (iPSC) in vitro for a period of time under conditions capable of differentiating the one or more iPSCs to a neuronal progenitor cell, optionally wherein the neuronal progenitor cell is one or more of a floor plate midbrain progenitor cells, determined dopaminergic precursor cells, or dopamine (DA) neurons.
Loring in that it deals with a population of in vitro neuronal progenitor cells in order to identify those which might differentiate into a determined dopaminergic precursor cell (for a later medical use for treating neuro-degenerative diseases) but Loring is silent on the culturing one or more induced pluripotent stem cells (iPSC) in vitro for a period of time under conditions capable of differentiating the one or more iPSCs to a neuronal progenitor cell.
However, Qi discloses the in vitro method for inducing differentiation of human stem cells into cortical neurons and precursors thereof comprises contacting a population of human stem cells [p. 5, par. 2]. Qi further discloses subjecting said population of differentiated cells to conditions favoring maturation of said differentiated cells into a population of cortical neurons [p. 5, par. 3]. Qi also discloses said conditions favoring maturation comprise culturing said population of differentiated cells in a suitable cell culture medium [p. 5, par. 3]. Qi further discloses present disclosure also provides for a population of in vitro differentiated cells expressing one or more neuronal marker, for example, a cortical neuron marker, or precursor cells thereof, prepared according to the methods described herein [p. 5, par. 4].
Claim 12 is directed to the computer implemented method of claim 8, wherein the culturing is for period of time that is between at or about 2 and at or about 25 days.
Loring in that it deals with a population of in vitro neuronal progenitor cells in order to identify those which might differentiate into a determined dopaminergic precursor cell (for a later medical use for treating neuro-degenerative diseases) but Loring is silent on culturing is for period of time that is between at or about 2 and at or about 25 days.
However, Qi discloses the in vitro method for inducing differentiation of human stem cells into cortical neurons and precursors where the cells are cultured for at least between 4 and 20 days, or at least between 6 and 16 days, at least between about 8 and 14 days, or at least between about 10 and 12 days [p. 4, par. 6-p. 5, par. 1].
Claim 20 is directed to the computer implemented method of claim 1, wherein the reference database comprises gene expression levels determined from one or more reference cell populations, wherein each of the one or more reference cell populations are formed by culturing one or more iPSC in vitro for a different period of time each under conditions capable of differentiating the one or more iPSCs to a neuronal progenitor cell, optionally wherein the neuronal progenitor cell is one or more of a floor plate midbrain progenitor cells, determined dopaminergic precursor cells, or dopamine (DA) neuron.
Loring also discloses a method of characterizing cells comprising: a. global profiling of known and unknown samples; b. collecting reference data from known samples into a matrix database; c. merging the unknown sample data with the matrix database; d. grouping the cells with a machine learning algorithm; e. determining the identity of the unknown sample based on its similarity to the known sample matrix [claim 21] but is silent on the culturing one or more induced pluripotent stem cells (iPSC) in vitro for a period of time under conditions capable of differentiating the one or more iPSCs to a neuronal progenitor cell.
However, Qi discloses the in vitro method for inducing differentiation of human stem cells into cortical neurons and precursors thereof comprises contacting a population of human stem cells [p. 5, par. 2]. Qi further discloses subjecting said population of differentiated cells to conditions favoring maturation of said differentiated cells into a population of cortical neurons [p. 5, par. 3]. Qi also discloses said conditions favoring maturation comprise culturing said population of differentiated cells in a suitable cell culture medium [p. 5, par. 3]. Qi further discloses present disclosure also provides for a population of in vitro differentiated cells expressing one or more neuronal marker, for example, a cortical neuron marker, or precursor cells thereof, prepared according to the methods described herein [p. 5, par. 4].
Claim 30 is directed to the computer implemented method of claim 1, wherein the one or more metagenes and the expression levels of the one or more metagenes are determined by using a dimensionality reduction technique on one or more reference cells of the one or more reference database.
Loring discloses the analysis of cellular RNA consists of microarray analysis; wherein the unsupervised machine learning approach comprises a bootstrapping sparse non-negative matrix factorization; wherein the transcriptional profile is a global gene expression profile called a stem cell matrix [0169] which reads on expression levels determined by using a dimensionality reduction technique.
Claim 46 is directed to the computer implemented method of claim 2, wherein the class label indicating each of the one or more different stages of differentiation of the reference cells is determined using an in vitro method.
Loring discloses identifying computationally derived class labels only based on biological characteristics; further comprising identifying differences in at least one dataset for at least one label between at least two samples in at least two clusters; further comprising filtering within a cluster for samples within having a similar label profile, such as common regulatory biochemical or metabolic activity; further comprising defining differentially regulated networks; further comprising using the networks to define a class membership, manipulate class membership, or define biological function of an unknown cell; and/or any limitation or characteristic disclosed herein alone or in combination [0167].
Claim 52 is directed to the computer implemented method of claim 1, wherein the expression levels of the one or more metagenes in the test dataset is determined based on (i) the one or more metagenes determined from the one or more reference cells in the reference database and (ii) the gene expression levels in the test dataset.
Loring discloses methods of assaying a cell comprising, receiving a reference database, wherein the reference database comprises cell datasets produced from characteristic data for at least two known cell lines, tissues, or primary cells, wherein the data was obtained by global profiling, wherein the data was associated with one or more labeled associated biological classes of the cells, receiving a test dataset, wherein the test dataset comprises data produced for an unknown cell line, tissue, or primary cell, wherein the test dataset was obtained by a similar global profiling as for the reference database, merging the test dataset into the database producing an updated reference database [0165]. Loring further discloses the test dataset contains several undifferentiated ePSC lines were new examples of the cell line that was included in the core dataset, and the other lines were not represented by samples in the core dataset, included were 7 samples from five different germ tumor cell lines, which had been previously reported to have pluripotent features [0195].
Claim 54 is directed to the computer implemented method of claim 30, wherein the expression levels of the one or more metagenes in the test dataset is determined by merging the gene expression levels in the test dataset with the reference database to create an updated reference database and applying the dimensionality reduction technique on the updated reference database.
Loring discloses methods of assaying a cell comprising, receiving a reference database, wherein the reference database comprises cell datasets produced from characteristic data for at least two known cell lines, tissues, or primary cells, wherein the data was obtained by global profiling, wherein the data was associated with one or more labeled associated biological classes of the cells, receiving a test dataset, wherein the test dataset comprises data produced for an unknown cell line, tissue, or primary cell, wherein the test dataset was obtained by a similar global profiling as for the reference database, merging the test dataset into the database producing an updated reference database [0165].
Claim 58 is directed to the computer implemented method of claim 30, wherein the number of the one or more metagenes is chosen based on evaluating one or more metrics determined from performing the dimensionality reduction technique using multiple candidate numbers of metagenes.
Loring discloses to sort the cell types an unsupervised machine learning approach to cluster transcriptional profiles of the cell preparations into stable distinct groups was used. Sparse nonnegative matrix factorization (sNMF) was adjusted for this task by implementing a bootstrapping algorithm to find the most stable groupings, or clusters [0193]. Loring further discloses in sparse NMF the matrix-decomposition process is forced by a "sparseness" factor lambda to reconstruct the data matrix with fewer features in the W-matrix [0265] which reads on a metric.
Claim 60 is directed to the computer implemented method of claim 1, wherein the computed label classification indicates that said cell or plurality of cells from the in vitro population of neuronal progenitor cells is a determined dopaminergic precursor cell if the probability of the cell or the plurality of cells having metagene expression levels of the determined dopaminergic precursor cell is greater than a threshold probability value.
Loring discloses the resulting genes and their expression values in each sample were subjected to the Prediction Analysis for Microarrays (PAM) algorithm by Tibshirani and colleagues (leave-one-out-cross-validation, categories: no pluripotence vs. pluripotence vs. germ-line-competence; class probabilities were re-computed 10 000 times [0026].
Claim 66 is directed to the computer implemented method of claim 1, wherein the deviation score for the cell or the plurality of cells is determined using a single-gene deviation score for each of one or more genes in the test dataset.
Loring discloses rkp is the Pearson correlation coefficient between xk and p, rkp is close to 1 if the corresponding transcript is strongly upregulated in A compared to the other conditions, and close to -1 if it is strongly downregulated in A, this measure has been suggested as an aparametric differential expression score [0280]. Loring further discloses the Pearson correlation is invariant under normalization of the patterns to zero mean and standard deviation of 1 [0280].
Claim 70 is directed to the computer implemented method of claim 66, wherein the single-gene deviation scores are z-scores determined using: differences between the gene expression levels of the test dataset and the gene expression levels in the one or more reference cells in the reference database; and standard deviations of gene expression levels in one or more of the one or more reference cells of the reference database.
Loring discloses rkp is the Pearson correlation coefficient between xk and p, rkp is close to 1 if the corresponding transcript is strongly upregulated in A compared to the other conditions, and close to -1 if it is strongly downregulated in A, this measure has been suggested as an aparametric differential expression score [0280]. Loring further discloses the Pearson correlation is invariant under normalization of the patterns to zero mean and standard deviation of 1 [0280].
Claim 83 is directed to the computer implemented method of claim 1, wherein the computed label classification indicates that said cell or plurality of cells from the in vitro population of neuronal progenitor cells is a determined dopaminergic precursor cell if the deviation score indicates that at least or at least about 50%,50%, 70%, 80%, 90%, or 95% of gene expression levels in the test dataset are no more than five standard deviations away from gene expression levels of the one or more reference cells in the reference database.
Loring discloses probes representing the 299 PluriNet genes were extracted from the quantile-normalized Stem Cell Matrix Core and Test Data Set, log 2-transformed for variance stabilization gene-wise ("row-") normalized over all 219 [0015]. As quantile-normalized is a method that makes the statistical distributions of gene expression identical across all samples, the mean Z-score for any given gene across all samples will be zero and 100% of the gene expression are no more than five standard deviations away from gene expression levels of the one or more reference cells in the reference database.
Claim 84 is directed to the computer implemented method of claim 1, wherein the computed label classification indicates that said cell or plurality of cells from the in vitro population of neuronal progenitor cells is a determined dopaminergic precursor cell if the deviation score indicates that at least or at least about 95% of gene expression levels in the test dataset are no more than 10, 9, 8, 7, 6, or 5 standard deviations away from the gene expression levels of the one or more reference cells in the reference database.
Loring discloses probes representing the 299 PluriNet genes were extracted from the quantile-normalized Stem Cell Matrix Core and Test Data Set, log 2-transformed for variance stabilization gene-wise ("row-") normalized over all 219 [0015]. As quantile-normalized is a method that makes the statistical distributions of gene expression identical across all samples, the mean Z-score for any given gene across all samples will be zero and 100% of the gene expression are no more than five standard deviations away from gene expression levels of the one or more reference cells in the reference database.
Claim 93 is directed to the computer implemented method of claim 1, wherein said gene expression levels are obtained from microarray analysis of cellular RNA, RNA sequencing, or both.
Loring discloses methods, wherein the stem cells are embryonic stem cells; wherein the stem cells are induced pluripotent stem cells; wherein the stem cells are somatic multipotent cells, such as neural stem cells, mesenchymal stem cells, or cardiac stem cells; wherein the analysis of cellular RNA consists of microarray analysis [0169].
Claim 95 is directed to the computer implemented method of claim 93, wherein the RNA sequencing is performed on bulk RNA from the plurality of cells or a plurality of reference cells.
Loring discloses wherein the transcriptional profile is a global gene expression profile called a stem cell matrix [0169] which reads on bulk RNA.
Claim 98 is directed to the computer implemented method of claim 1, wherein receiving said test dataset comprises receiving input from an array analysis system.
Loring discloses wherein receiving the updated reference database comprises receiving the record from an array analysis system.
Claim 100 is directed to the computer implemented method of claim 1, wherein said one or more reference databases forms part of a storage medium.
Loring discloses wherein receiving the updated reference database comprises receiving the updated reference database from a storage medium.
Claim 101 is directed to the computer implemented method of claim 1 a, comprising repeating the receiving, applying, determining, and outputting steps if the computed label classification indicates that said cell or plurality of cells is not a determined dopaminergic neuronal cell, optionally wherein the steps are repeated using the same or a different in vitro population of neuronal progenitor cells.
As there are many statistical methods that can be used when analysis is done this way it would be obvious to reevaluate the data if results are not what are expected.
Claim 106 is directed to a population of determined dopaminergic precursor cells identified by the method of claim 1.
Loring is silent on dopaminergic precursor cells.
However, Qi discloses a population of in vitro differentiated cells expressing one or more neuronal marker, for example, a cortical neuron marker, or precursor cells p. 5, par. 4]. Qi further discloses the assay could also serve as a tool to define neurons of related lineages but distinct projection patterns such as midbrain dopamine neurons of A9 [p. 52, par. 1].
Claim 107 is directed to a method of treatment, the method comprising administering to a subject having Parkinson's disease the population of determined dopaminergic precursor cells of claim 106.
Loring is silent on administering to a subject having Parkinson's disease the population of determined dopaminergic precursor cells.
However, Qi discloses method of treating a neurodegenerative disorder in a subject, comprising administering an effective amount of the population of in vitro differentiated cells [claim 25].
Claim 108 is directed to the method of claim 107, wherein the administering is by implanting the population of determined dopaminergic precursor cells into one or more brain regions of the subject.
Loring is silent on administering to a subject having Parkinson's disease the population of determined dopaminergic precursor cells.
However, Qi discloses the composition further comprises a biocompatible scaffold or matrix, for example, a biocompatible three-dimensional scaffold that facilitates tissue regeneration when the cells are implanted or grafted to a subject [p. 38, par. 5]. Qi further discloses neurons transplanted at day 8 of differentiation into the postnatal mouse cortex are functional and establish long-distance projections as illustrated using iDISCO-based whole brain imaging [p. 43, par. 1].
Claim 111 is directed to the method of claim 107, wherein the population of determined dopaminergic precursor cells is allogeneic to the subject.
Loring is silent on administering to a subject having Parkinson's disease the population of determined dopaminergic precursor cells.
However, Qi further discloses neurons transplanted at day 8 of differentiation into the postnatal mouse cortex are functional and establish long-distance projections as illustrated using iDISCO-based whole brain imaging [p. 43, par. 1].
Claim 112 is directed to a method of treating a subject having Parkinson's disease, the method comprising: implanting a population of determined dopaminergic precursor cells into a brain region of a subject having Parkinson's disease, wherein the population of determined dopaminergic precursor cells has been identified using the computer implemented method of claim 1.
Loring is silent on administering to a subject having Parkinson's disease the population of determined dopaminergic precursor cells.
However, Qi discloses method of treating a neurodegenerative disorder in a subject, comprising administering an effective amount of the population of in vitro differentiated cells [claim 25]. Qi further discloses the composition further comprises a biocompatible scaffold or matrix, for example, a biocompatible three-dimensional scaffold that facilitates tissue regeneration when the cells are implanted or grafted to a subject [p. 38, par. 5].
In regards to claim(s) 1-2, 7-8, 12, 20, 30, 46, 52, 54, 58, 60, 66, 70, 73, 83-94, 93, 95, 98, 100-101, 106-108, and 111-112, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to combine Loring with Qi as they both disclose in vitro methods of inducing differentiation of human stem cells into cortical neurons, and cortical neurons generated by such methods. The motivation would have been to modify the neural cells of Loring with the proximate precursors and cortical neurons including PAX 6 of Qi in order to express detectable levels of PAX6 at least 6 days after initiation as disclosed by Qi [p. 3, par. 5]. The same biomarker PAX6 is used in present specification and the skilled person in the art would use this metagene expression level in the computer implemented method to solve the technical problem posed. One could have therefore combined the elements as claimed by the known methods of Loring and Qi, and that in combination, each element merely would have performed the same function as it did separately for the predictable result. Furthermore, using known markers and metagenes in cells for the identification and sorting of neuronal progenitor cells combined with known computer implemented method for classifying a cell population (as exemplified in Loring) would appear obvious to the skilled person in the art of neuronal progenitor cell and their medical use. Additionally, the use of known iPSC, their period of culture, presence of inhibitor or activators in the cell incubation medium, or statistical tools and reference database are all obvious optimizations in their respective technical fields (cell culture and computer based method for classifying.
B. Claims 47, 110 and 113 are rejected under 35 U.S.C. 103 as being unpatentable over Loring in view of Qi, as applied to claims 1, 106-107, and 112 as above, and in further view of Kan et al. (Kan, Inna, et al. "Dopaminergic differentiation of human mesenchymal stem cells—utilization of bioassay for tyrosine hydroxylase expression." Neuroscience letters 419.1 (2007): 28-33., cited on IDS dated 10/04/2022).
Claim 47 is directed to the computer implemented method of claim 46, wherein: the in vitro method comprises assessing dopamine production levels of a reference cell population; and the class label is designated as a determined dopaminergic precursor cell if the dopamine production levels are increased relative to a pluripotent stem cell.
Loring discloses identifying computationally derived class labels only based on biological characteristics; further comprising identifying differences in at least one dataset for at least one label between at least two samples in at least two clusters; further comprising filtering within a cluster for samples within having a similar label profile, such as common regulatory biochemical or metabolic activity; further comprising defining differentially regulated networks; further comprising using the networks to define a class membership, manipulate class membership, or define biological function of an unknown cell; and/or any limitation or characteristic disclosed herein alone or in combination [0167] but is silent on the class label is designated as a determined dopaminergic precursor cell if the dopamine production levels are increased relative to a pluripotent stem cell.
However, Kan discloses high-performance liquid chromatography (HPLC) analysis, following 48 h incubation in differentiation medium I, cells were exposed to differentiation medium II supplemented with 10 ng/ml human glial cell line-derived neurotrophic factor (GDNF; R&D systems) for 96 h [p. 29, col. 2, par. 1]. Kan further discloses Reverse-phase HPLC coupled with an electrochemical detector (ECD) was used to measure dopamine levels [p. 29, col. 2, par. 3]. It would be obvious to select those classify cells as a candidate if it increase the dopamine production in relation to a pluripotent stem cell as the purpose of theses cells is to mimic the effect of dopamine.
Claims 110 and 113 are directed to the method of claims 107 and 112, wherein the population of determined dopaminergic precursor cells is autologous to the subject.
Loring and Qi are silent on the population of determined dopaminergic precursor cells is autologous to the subject.
However, Kan discloses dopaminergic differentiation of human mesenchymal stem cells—Utilization of bioassay for tyrosine hydroxylase expression [title]. Kan further discloses our study implies that MSCs may be employed for autologous therapy of PD, avoiding the risks of obtaining neural stem cells from the brain [p. 32, col. 2 par. 2].
In regards to claim(s) 47, 110, and 113, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to combine Loring and Qi with Kan as they all are directed to differentiation of human stem cells. The motivation would have been to combine the methods of Loring and Qi with the HPLC analysis and autologous therapy of PD of Kan so that MSCs may be primed in vitro towards a dopaminergic fate offering the promise of innovative therapy for currently incurable human disorders, including PD as disclosed by Kan [abstract], a finding that one of ordinary skill in the art would have recognized that the results of the combination were predictable.
C. Claims 53 and 73 are rejected under 35 U.S.C. 103 as being unpatentable over Loring in view of Qi, as applied to claims 1 and 52 as above, and in further view of Urgard et al. (Urgard, Egon, et al. "Metagenes associated with survival in non-small cell lung cancer." Cancer informatics 10 (2011): CIN-S7135, newly cited).
Claim 53 is directed to the computer implemented method of claim 52, wherein the expression levels of the one or more metagenes in the test dataset is determined using regression analysis based on (i) the one or more metagenes determined from the one or more reference cells in the reference database and (ii) the gene expression levels in the test dataset.
Loring discloses MATISSE used the Pearson correlation coefficient as a measure of similarity between the expression patterns of gene pairs [0278]. Loring further discloses these similarity values serve as a starting point for the computation of pair-wise weights using a probabilistic model [0278]. Loring also discloses the Pearson correlation between a pair of genes captures a global similarity trend between their expression patterns [0278]. Loring further disclose groups of genes were extracted that are not only similar across the experimental conditions, but also show significantly high or significantly low expression values in a specific subset of the samples, identified using the sNMF clustering scheme [0287], but is silent on metagenes in the test dataset is determined using regression analysis.
However, Urgard discloses Bayesian regression analysis and Kaplan-Meier survival analysis were performed to determine metagenes associated with survival [abstract]. Urgard further discloses identifying 599 genes which were down-regulated and 402 genes which were up-regulated in NSCLC compared to the normal lung tissue and 112 genes which were up-regulated and 101 genes which were down-regulated in AC/BC compared to the SCC [abstract]. Urgard also discloses, for stage Ib patients the metagenes potentially associated with survival were identified [abstract] which reads on wherein the expression levels of the one or more metagenes in the test dataset is determined using regression analysis based on (i) the one or more metagenes determined from the one or more reference cells in the reference database and (ii) the gene expression levels in the test dataset.
Claim 73 is directed to the computer implemented method of claim 1, wherein the gene expression levels in the one or more reference cells in the reference database are determined using regression analysis based on (i) the expression levels of the one or more metagenes in the test dataset and (ii) the gene expression levels in the test dataset.
Loring discloses MATISSE used the Pearson correlation coefficient as a measure of similarity between the expression patterns of gene pairs [0278]. Loring further discloses these similarity values serve as a starting point for the computation of pair-wise weights using a probabilistic model [0278]. Loring also discloses the Pearson correlation between a pair of genes captures a global similarity trend between their expression patterns [0278]. Loring further disclose groups of genes were extracted that are not only similar across the experimental conditions, but also show significantly high or significantly low expression values in a specific subset of the samples, identified using the sNMF clustering scheme [0287].
However, Urgard discloses Bayesian regression analysis and Kaplan-Meier survival analysis were performed to determine metagenes associated with survival [abstract]. Urgard further discloses identifying 599 genes which were down-regulated and 402 genes which were up-regulated in NSCLC compared to the normal lung tissue and 112 genes which were up-regulated and 101 genes which were down-regulated in AC/BC compared to the SCC [abstract]. Urgard also discloses, for stage Ib patients the metagenes potentially associated with survival were identified [abstract] which reads on wherein the expression levels of the one or more metagenes in the test dataset is determined using regression analysis based on (i) the one or more metagenes determined from the one or more reference cells in the reference database and (ii) the gene expression levels in the test dataset.
In regards to claim(s) 53 and 73, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to combine Loring and Qi with Urgard as they all are directed to identify differentially expressed genes between all tumor samples and cancer-free control tissue. The motivation would have been to combine the methods of Loring and Qi with the regression analysis of Urgard so that accurately classify early-stage NSCLC patients into different prognostic groups may be helpful in selecting patients who should receive specific therapies as disclosed by Urgard [abstract], a finding that one of ordinary skill in the art would have recognized that the results of the combination were predictable.
D. Claims 61, are rejected under 35 U.S.C. 103 as being unpatentable over Loring in view of Qi, as applied to claims 1 and 60 as above, and in further view of Muller et al. (Müller, FJ., Schuldt, B., Williams, R. et al. A bioinformatic assay for pluripotency in human cells. Nat Methods 8, 315–317 (2011), cited on IDS dated 11/04/2022).
Claim 61 is directed to the computer implemented method of claim 60, wherein: the threshold probability value is set such that a determined dopaminergic precursor cell is identified with greater than or greater than about 75%, 80%, 85%, 90%, or 95% sensitivity; and/or the threshold probability value is set such that a determined dopaminergic precursor cell is identified with greater than or greater than about 75%, 80%, 85%, 90%, or 95% specificity.
Loring and Qi are silent on determined dopaminergic precursor cell is identified with greater than or greater than about 75%, 80%, 85%, 90%, or 95% sensitivity; and/or the threshold probability value is set such that a determined dopaminergic precursor cell is identified with greater than or greater than about 75%, 80%, 85%, 90%, or 95% specificity.
However, Muller discloses PluriTest, a robust open-access bioinformatic assay of pluripotency in human cells based on their gene expression profiles [abstract]. Muller further discloses set thresholds that separated pluripotent from non-pluripotent samples in HT12v3 test datasets with 98% sensitivity and 100% specificity [p. 316, col. 1par. 3].
In regards to claim(s) 61, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to combine Loring and Qi with Muller as they all are directed to bioinformatic assay for pluripotency in human cells. The motivation would have been to combine the methods of Loring and Qi with the sensitivity and specificity thresholds of Muller as using all of the expression information available provides much higher discriminatory power and the ability to identify deviations from known patterns that may lead to additional insights into cellular phenotypes as disclosed by Muller [p. 317, col. 2, par. 1], a finding that one of ordinary skill in the art would have recognized that the results of the combination were predictable.
Conclusion
No claims are allowed.
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/D.M.B./Examiner, Art Unit 1685
/Soren Harward/Primary Examiner, TC 1600