Lymphocytes including altered noxa activity

Abstract

Engineered cells include modified activity or expression of the protein Noxa from the PMAIP1 gene. In one or more embodiments, the engineered cell is a human cell. In one or more embodiments, the engineered cell includes one or more modifications to the PMAIP1 gene. In another aspect, methods of modulating the activity or expression of the protein Noxa include administering one or more nucleic acids to the cell. In one or more embodiments, the nucleic acids include a gene editing composition.

Description

[0001]PCT Application Attorney Docket No. 0110.000746WO01 LYMPHOCYTES INCLUDING ALTERED NOXA ACTIVITY CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of U.S. Provisional Application Serial No. 63 / 621,676, filed on January 17, 2024, which is incorporated by reference herein in its entirety. SEQUENCE LISTING This application contains a Sequence Listing electronically submitted via EFS-Web to the United States Patent and Trademark Office as an XML file entitled “0110.000746WO01.xml” having a size of 9,041 bytes and created on January 17, 2025. The information contained in the Sequence Listing is incorporated by reference herein. GOVERNMENT FUNDING This invention was made with government support under AI148876 awarded by the National Institutes of Health. The government has certain rights in the invention. SUMMARY This disclosure describes, in one aspect, an engineered cell including an altered level of Noxa protein. In another aspect, the present disclosure describes an engineered cell including an engineered Noxa protein having reduced pro-apoptotic activity relative to a non-engineered Noxa protein. The engineered cell may include one or more modifications to the PMAIP1 gene, such as a modification that decreases the expression or activity of the Noxa protein. In one or more embodiments, the modification encodes mutation of an aspartate at a position equivalent to aspartate 34 in SEQ ID NO:1 to alanine (D34A). In one or more embodiments, the modification encodes mutation of a serine at a position equivalent to serine 13 in SEQ ID NO:1 to glutamate (S13E) or aspartate (S13D). In another aspect, the present disclosure relates to methods of regulating Noxa expression and / or activity in a cell. A method may include administering a gene editing composition to the cell. The gene editing composition may introduce one or more modifications to the PMAIP1 gene. In one or more embodiments, the modification encodes mutation of an aspartate at a position equivalent to aspartate 34 in SEQ ID NO:1 to alanine (D34A).In one or more embodiments, the modification encodes mutation of a serine at a position equivalent to serine 13 in SEQ ID NO:1 to glutamate (S13E) or aspartate (S13D). In one or more embodiments, a method of the present disclosure increases the lifespan of a cell. In one or more embodiments, a method of the present disclosure decreases the pro- apoptotic activity of the Noxa protein. In one or more embodiments, a cell of the present disclosure is a human cell. The cell may be a T cell, such as a CD8+ T cell. The T cell may be a chimeric antigen receptor T cell (CAR T cell). The T cell may be a tumor infiltration lymphocyte (TIL). The cell may be an NK cell. The NK cell may be a CAR NK cell. The CAR NK cell may In another aspect, the present disclosure relates to an animal model including a nucleic acid encoding Noxa, such as hNoxa. The above summary is not intended to describe each disclosed embodiment or every implementation of the present invention. The description that follows more particularly exemplifies illustrative embodiments. In several places throughout the application, guidance is provided through lists of examples, which examples can be used in various combinations. In each instance, the recited list serves only as a representative group and should not be interpreted as an exclusive list. BRIEF DESCRIPTION OF THE FIGURES FIG. 1. Noxa is induced in human CD8+ T cells following co-stimulation and remains highly expressed through the immune response in vitro. (A) Representative Western blot showing Noxa protein levels at the indicated times after in vitro co-stimulation of naïve CD8+ T cells. (B) A schematic depicting a simplified immune response model marking phases that correlate established markers with Noxa protein levels. (C) Flow cytometric analysis showing the percentage of CD45RA+ and / or CD45RO+ cells following co-stimulation (n=10). FIG. 2. Percentage of CD69+ cells, CD107α+ cells, and fold change in mean fluorescence intensity (MFI, an indicator of protein expression level) of HLA-DR+ CD8+ T cellsin population (error bars indicate the mean ± s.e.m. n=10; *p≤0.05 **p≤0.01 (d=donor). FIG. 3. Noxa induction upon TCR engagement is transcriptional. (A) Naïve human CD8+ T cells were co-stimulated with anti-CD3 / CD28 and Noxa protein expression was analyzed by Western blot. (B) mRNA expression was analyzed by RT-PCR. (C) Open chromatin profiles of promoter / enhancer regions surrounding the Noxa gene determined by bioinformatic analysis of ATAC-Seq data from genomic DNA of naïve and 48-hour and 7-day stimulated CD8T cells (memory & re-stimulated cell data not shown). Black arrows mark noted differences. FIG. 4. Noxa is phosphorylated on serine 13 in activated, proliferating human CD 8+ T cells. Western blot of lysates (40 μg) from naïve CD8+ T cells stimulated until day 19, showing pS13-Noxa expression by the phospho-specific Rb mAb pNoxa (Cell Signaling Technology); membranes were stripped and re-probed for total Noxa with Rb mAb, D8L7Ua (Cell Signaling Technology), also utilized for most Western blots shown under preliminary studies. Mcl-1 levels were detected with Rb mAb D2W9E (Cell Signaling Technology). FIG. 5. Noxa intracellular localization is regulated during an immune response. Noxa expression in whole cell lysates (WCL) and mitochondrial (mito) and cytosolic (cyto) fractions from CD8+ T cells five days (d5), seven days (d7), and nine days (9d) post-stimulation. The antibody that detects total Noxa is rabbit mAb, from Cell Signaling Technology. Tubulin and mitochondrial cytochrome oxidase (COX) IV were used as controls for the fractionation protocol. FIG. 6. Glutamine is required for induction of Noxa protein in CD8+ T cells and for increased mitochondrial respiration upon activation. Naïve human CD8+ T cells were co- stimulated anti-CD3 / CD28 Abs in complete media with 2 mM glutamine (HQ) or without glutamine (-Q). (A) Quantification of Noxa protein by Western blot in unstimulated and 24-hour stimulated cells. (B) Quantification of Noxa transcript levels by RT-PCR analysis in unstimulated and 24-hour stimulated cells. (C, D) OCR and spare respiratory capacity using the Mito Stress Test in unstimulated cells and CD8T cells activated for 40 hours with or without glutamine. N= 5 (Error bars indicate the mean ± s.e.m. **p≤0.01). US: unstimulated; HQ: 2 mM glutamine; -Q: no glutamine. FIG. 7. Glutamine, not glutaminolysis, is required for upregulation of Noxa in CD8+ T cells following stimulation. (A, B) Human CD8+ T cells were co-stimulated with anti- CD3 / CD28 Abs in medium with 2 mM glutamine (HQ), 0.2 mM glutamine (LQ), no glutamine (NQ), or in HW medium including CB839 or DON. Noxa protein levels a representative donor an SRC in response to treatments 40 hours post stimulation data (data are from 5 technical replicates; error bars indicate the mean±s.e.m. *p≤0.01. (C, D) Activation level on day 2 post- stimulation is shown as fold change in CD69 and CD107a positively relative to HQ, representative of three independent donors (error bars indicate the mean±s.e.m., n=3). FIG. 8. Proliferation rates of CD8+ T cells labeled with Cell Trace Violet and subjected to above treatments on day 4 post-stimulation. (A) Quantification of Noxa in cells treated with CB-839, a glutaminase inhibitor, DON, a glutamine agonist, NQ, HQ, or unstimulated (US). (B) Noxa levels in CD8+ T cells activated with anti-CD3 / CD28 Abs in HQ, LQ, NQ medium or NQ containing α-KG or glutamate. (C) Degranulation marker CD107a levels relative to HQ in day 4 post-stimulation US= unstimulated, HQ= 2 mM glutamine, LQ= 0.2 mM glutamine, NQ= 0 mM glutamine, CB839= 2 μM glutaminase inhibitor, DON= 200 μM glutamine agonist, 6-Diaxo-5- oxo-L-norleucine, α-KG= 4 mM dimethyl-alpha-ketoglutarate, E= 4 mM dimethyl glutamate. FIG. 9. Noxa is required for conversion of glutamine to glutamate but not for mitochondrial respiration in activated CD8+ T cells. (A) Noxa protein levels in 12-hour activated CD3+ (pan) T cells transfected with control or Noxa siRNA (B) LCMS analysis of TCA metabolites. Enrichment of M+5 labeled glutamine and glutamate in derivatized cell extracts of transfected stimulated T cells incubated with medium containing [U-13C5] glutamine for 12 hours, n=3, ***p<0.001. (C, D) Maximum oxygen consumption rate (OCR) and OCR / extracellular acidification rate (ECAR) ratio of uncoupled mitochondria in control and Noxa siRNA are shown. FIG. 10. Noxa is required for conversion of glutamine to glutamate but not for mitochondrial respiration in activated CD8+ T cells. (A) Western blot showing Noxa protein levels in unstimulated, control (siCON) and Noxa-silenced (siNoxa) CD8+ T cells and control naive cells treated with CD839 (2 μM and 5 μM) 40 hours after co-stimulation. (B) A bar graph showing intracellular glutamate concentrations in unstimulated, control (siCON) and Noxa- silenced (siNoxa) CD8+ T cells and control naive cells treated with CD839 (2 μM and 5 μM) 40 hours after co-stimulation. (C) A schematic depicting the point of action of CB839 and the hypothesized direct / indirect target of Noxa in activated T cells. (D) OCR of siCON and siNoxa CD8+ T cells. (E) Quantification of spare respiratory capacity (SRC) of siCON and siNoxa CD8+ T cells. (F) OCR of an independent experiment with untransfected cells exposed to CD839 (2 μM and 5 μM) 40 hours after stimulation. (G) An independent experiment showing SRC of untransfected CD8+ T cells exposed to CD839 (2 μM and 5 μM) 40 hours after stimulation. FIG. 11. Noxa knockout (KO) CD8+ T cells show reduced apoptosis during the contraction phase. NoxaKO cells were generated using CRISPR / Cas9 and sgRNA. (A) A simplified schematic model of regulation of hNoxa protein expression by glutamine and mTORC1 during the CD8+ T cell immune response V9302= transmembrane glutamine flux antagonist; JPH203= LAT 1 inhibitor, glutamine-leucine antiporter antagonist. (B) Western blot of NoxaKO and Cas9-control cells 48 hours after co-stimulation, early passage (EP) and late passage (LP). (C) Percent live cells in population of control and NoxaKO cells. Error bars indicate the mean±s.e.m. n=3. FIG. 12. Memory cells generated in vitro display robust respiratory capacity and dependence on FAO before restimulation. (A) SEAHORSE mitochondrial stress test profile showing OCR of unstimulated (resting) cells 40 hours after primary stimulation, unstimulated memory cells, and 40-hour recall stimulated memory cells. (B) SEAHORSE mitochondrial stress test profile showing SRC of the cells in (A). (C) Representative OCR of unstimulated memory and memory cells 16 hours after recall stimulation in the presence of FAO inhibitor etomoxir (5 μM) or vehicle control, (error bars indicate the mean±s.e.m. n=5). (D) SEAHORSE mitochondrial stress test profile showing SRC of the cells in (C) FIG. 13. Human Noxa (hNoxa) is regulated normally in CD8+ T cells from hNoxa GR mice (A) Western blot analysis of protein extracts from unstimulated and 40-hour antiCD3 / CD28 stimulated hNoxa CD8+ T cells in medium with or in without glutamine. (B, C) SCENITH- based metabolic profiling of unstimulated and 16-hour stimulated hNoxa CD8+ T cells isolated from spleen.2DG= 2-deoxyglucose, Oligo= Oligomycin, ODG= both drugs. FIG. 14. A schematic depicting the Rapid Expansion Protocol (REP) and genetic engineering of TIL. TIL were activated using plate-bound anti-CD3 and soluble anti-CD28 antibodies for 4 days and electroporated with CRISPR reagents, following which, TIL were transferred to REP culture in the presence of 100:1 irradiated PBMC and 3000 IU / mL IL-2 for 14 days. FIG. 15. A Western blot showing Noxa expression in stimulated human NK cells. NK cells, isolated from three independent donors (D11, D14 and D16), were cultured in the described medium in the presence of IL-2 (100 IU / mL) and activated through co-culture with C2, K562 feeder cells. FIG. 16. Knocking out Noxa extends the lifespan of MSLN-CAR CD8+T cells without affecting function. (A) Western blotting of NoxaKO and Cas9-control in WT T cells. (B) Western blotting of NoxaKO and Cas9-control in MSLN-CAR T cells. (C) Representative flow cytometry plots of non-transduced, unsorted T cells transduced with RQR8 MSLN-CAR T. Transduced T cells with RQR8 constructs were stained with the QBEND / 10 mAb to demonstrate co-expression of CAR and marker genes. (D) A representative schematic of the serial co-culture used to generate (E). (E) Serial killing of MSLN-CAR T cells against A1847 cells. In the first repetitive culture, WT or MSLN-CAR T cells were co-cultured with A1847 tumor at the E:T ratio=3:1 for 48 hours. For the second and third co-cultures, MSLN-CAR T cells were collected from the previous co-culture. FIG. 17. Knocking out Noxa expands lifespan of MSLN-CAR CD8+T cells without affecting function. Intracellular cytokine analysis of MSLN-CAR T cells co-cultured with A1847 cells. (A-C) Expression levels of CD107a, IFN-γ and TNF-α in wild-type (WT) and MSLN-CAR T cells after co-culture with A1847 cells at an effector-to-target (E:T) ratio of 1:1 for 6 hours . (D-G) WT Cas9 control and NoxaKO cell populations were evaluated at the indicated time points after stimulation with anti-CD3 and anti-CD28 for CD69 (D), GZMB (E) cleaved- caspase3 (F) expression and for viability (G). FIG. 18. Knocking out Noxa expands lifespan of MSLN-CAR CD8+T cells without affecting function. MSLN-CAR T Cas9-control and NoxaKO cells were evaluated after stimulation with anti-CD3 and anti-CD28 at the indicated time points for percent CD69+ (A), GZMB+ (B) cleaved-caspase-3+ (C) and viable (D) cells in the population. FIG. 19. Characterization of Noxa knockout (NoxaKO) in wild-type (WT) CD8+ T cells. (A) Western blot analysis of Noxa expression in WT Cas9-control and NoxaKO CD8+ T cells after 48 hours of stimulation with anti-CD3 / CD28. (B) A histogram of the percent of CD69+ WT Cas9-control and NoxaKO CD8+ T cells quantified using flow cytometry at various time points post-stimulation. (C) A histogram of the percent of GZMB+ cells from (B). (C) A histogram of the percent of cleaved-caspase 3+ cells from (B). (D) A histogram of the percent of cleaved- viable cells from (B). FIG. 20. Characterization of NoxaKO in B7H3-CAR CD8+ T cells. (A) Western blot analysis of Noxa expression in B7H3-CAR Cas9-control and NoxaKO CD8+ T cells after 48 hours of stimulation with anti-CD3 / CD28. (B-E) Flow cytometric analysis of B7H3-CAR Cas9- control and NoxaKO CD8+ T cells at various time points post-stimulation. Shown are percent CD69+ (B), GZMB+ (C), cleaved-caspase 3+ (D), and viable cells (E) in the population. FIG. 21. NoxaKO in CD19-CAR CD8+ T cells. (A) Western blot analysis of Noxa expression in CD19-CAR Cas9-control and NoxaKO CD8+ T cells after 48 hours of stimulation with anti-CD3 / CD28. (B-E) Flow cytometric analysis of CD19-CAR Cas9-control and NoxaKO CD8+ T cells at various time points post-stimulation. (B-E) show percent CD69+, GZMB+, cleaved-caspase 3+ and viable cells. FIG. 22. Characterization of NoxaKO in MSLN-CAR CD8+ T cells. (A) Western blot analysis of Noxa expression in MSLN-CAR Cas9-control and NoxaKO CD8+ T cells after 48 hours of stimulation with anti-CD3 / CD28. (B-E) Flow cytometric analysis of MSLN-CAR Cas9- control and NoxaKO CD8+ T cells at various time points post-stimulation. (B-E) show percent CD69+, GZMB+, cleaved-caspase 3+ and viable cells. FIG. 23. Target cell-specific activation of NoxaKO CAR T cells. (A) A schematic representation of WT or CAR T cell stimulation with tumor target cells. (B-D) Flow cytometric analysis of WT Cas9-control and NoxaKO CD8+ T cells before (unstimulated, US) and after stimulation with antiCD3 / CD28 or target cell lines. Cells were analyzed for: CD69 (B) on day 2 post-stimulation, GZMB (C) on day 4 post-stimulation and HLA-DR (D) on day 4 post- stimulation. FIG. 24. B7H3-CAR Cas9-control and NoxaKO CD8+ T cells were co-cultured with RH30 tumor cells and analyzed at various time points post-stimulation. Parameters measured include: CD69 (A), GZMB (B), cleaved caspase-3 (C) and cell viability (D). FIG. 25. CD19-CAR Cas9-control and NoxaKO CD8+ T cells were co-cultured with Raji lymphoma cells and analyzed at various time points post-stimulation. Parameters measured include: CD69 (A), GZMB (B), cleaved caspase-3 (C) and cell viability (D). FIG. 26. MSLN-CAR Cas9-control and NoxaKO CD8+ T cells were co-cultured with A1847 ovarian cancer cells and analyzed at various time points post-stimulation. Parameters measured include: CD69 (A), GZMB (B), cleaved caspase-3 (C) and cell viability (D). FIG. 27. Targeting strategy underlying the gene replacement mouse model of hNoxa (encoded by the gene PMIAP). Murine genomic and coding regions are in black and corresponding Noxa promoter and coding regions from the human genome are in gray. An hNoxa DA strategy (not shown) was used to incorporate the DA mutation within the donor vector. FIG. 28. In vitro activation of splenic CD8+T cells from control and gene replacement hNoxa mice. (A) Western blot analysis of Noxa expression in CD8+ T cells isolated from the spleens of WT mice. Cells were analyzed in unstimulated conditions (US) and at various time points following stimulation with anti-CD3 / CD28 Dynabeads. (B) Western blot analysis of Noxa expression in CD8+ T cells isolated from the spleens of hNoxa mice. Cells were analyzed like in (A). (C). A histogram showing the percentage of CD69+ isolated WT and hNoxa CD8+ T cells measured using flow cytometry at various time points post-stimulation. CD69 was used as an activation marker. (D) A histogram showing the percentage of Granzyme B (GZMB)+ isolated WT and hNoxa CD8+ T cells measured using flow cytometry at various time points post-stimulation. GZMB was used as a measure of cytotoxicity. FIG. 29. In vivo infection pilot in controls and gene replacement hNoxa and hNoxa DA mice. Mice were infected with lymphocytic choriomeningitis virus (LCMV) and analyzed 12 weeks post-infection. Spleen, liver, and small intestine intraepithelial lymphocytes (SI IELs) were harvested and processed for flow cytometric analysis of live, unfixed cells. (A) Percentages of CD8+ tetramer+ cells. (B) Specific CD8+ tetramer+ cells and resident memory T cells in the spleen of infected mice: Long-lived effector cells (LLEC); central memory T cells (Tcm); and effector memory T cells (Tem). (C) ++non-lymphoid tissues (NLT). (D) small intestine intraepithelial lymphocytes (SI IELs). (E) liver. FIG. 29. In vivo infection pilot in controls and gene replacement hNoxa and hNoxa DA mice. Mice were infected with lymphocytic choriomeningitis virus (LCMV) and analyzed 12 weeks post-infection. spleen, liver, and small intestine intraepithelial lymphocytes (SI IELs) were harvested and processed for flow cytometric analysis of live, unfixed cells. Percentages of CD8⁺ tetramer⁺ cells (A) and specific CD8⁺ T cell subsets (B) in the spleen of infected mice. Long-lived effector cells (LLEC), central memory T cells (Tcm), and effector memory T cells (Tem). Percentages of CD8⁺ tetramer⁺ cells (C) and tissue-resident memory T cells (Trm)(D) in non-lymphoid tissues (NLT), including small intestine intraepithelial lymphocytes (SI IELs) and liver (E). FIG. 30. CD8+ TILs from NSCLC patient sample show higher viability in the absence of Noxa. (A) Tumor-infiltrating lymphocytes (TILs) were cultured from non-small cell lung cancer (NSCLC) patient sample. TILs were stimulated with anti-CD3 / CD28 and supplemented with IL- 2 to promote expansion. The cells were cultured for 20 days in G-Rex plates to ensure robust T cell growth and viability. Following the expansion phase, CD8+ T cells were isolated from the expanded TIL population for downstream analyses. (B) Flow cytometric analysis of the CD8+ population in TILs before and after CD8+ T cell isolation. (C) Western blot analysis of Noxa expression in Cas9 control and Noxa knockout (NoxaKO) CD8+ T cells isolated from TILs. (D- F) Flow cytometric analysis of Cas9-control and NoxaKO CD8+ T cells at various time points post-stimulation with anti-CD3 / CD28 evaluated CD69 (D), GZMB (E) expression and cell viability (F). DETAILED DESCRIPTION OF ILLUSTRATIVE EMBODIMENTS One of the current challenges in cell-based therapies is early cell death and, consequently, a lack of therapeutic persistence in the patient. Cell death is of particular concern in hematopoietic cell therapies, such as chimeric antigen receptor (CAR) T cell therapy. Accordingly, methods of extending the lifespan of hematopoietic cells, particularly lymphoid cells such as T cells, B cells, and NK cells, are of interest. Each phase of a hematopoietic cell’s response to immune challenge, including activation, expansion, functional differentiation, and cell death, is closely linked to changes in cellular metabolic programs. Metabolic rearrangements often facilitate the transition from a non- proliferative to proliferative state, or from a proliferative to a differentiated state. For example, in response to stimulation by a cognate antigen, naïve or memory T cells increase their glucose uptake, limit catabolic fatty acid β-oxidation (FAO) and pyruvate oxidation and rely on aerobic glycolysis for generating biomass essential for proliferation. Cells also switch to glutamine as a fuel source for energy via mitochondrial oxidative phosphorylation (OxPhos). Nutrient availability is, thus, linked to both lymphocyte and cancer cell expansion, and is particularly important in microenvironments where tumor suppressive effector T cells compete for resources with rapidly growing tumors. A deeper understanding of metabolic adaptations and their regulation in T cells is critical for designing therapies aimed at enhancing their tumor suppressor role. Additionally, manipulating the availability of nutrients can influence the proliferative ability and / or function of T cells through metabolic reprogramming. Similar observations have been made with other hematopoietic cells, such as NK cells and B cells. Direct manipulation of signaling molecules involved in metabolic reprogramming may be a more effective method of influencing cell activity. However, the signaling molecules involved in metabolic reprogramming have historically been poorly understood. The present application describes identification of a signaling molecule involved in metabolic reprogramming (the Bcl-2 family protein Noxa) and methods of engineering cells via directly manipulating its expression. In particular, the present application relates to methods of extending lymphocyte lifespan by modifying cellular levels of signaling molecules such as Noxa. Phorbol-12-myristate-13-acetate-induced protein 1, also referred to as “PMAIP1” or “Noxa,” is a member of the Bcl-2 protein family. Human Noxa (hNoxa), the smallest member of the Bcl-2 family, has been identified as promoter of apoptosis. However, as is described herein, Noxa not only promotes apoptosis following an immune response but also promotes growth, facilitating a transition to proliferative metabolism, following antigenic stimulation. In the absence of Noxa expression, stimulated T cells delay cell death and exhibit longer lifespans. Specifically, effector T cells exhibit significantly longer lifespans following antigen stimulation when Noxa expression has been knocked down compared to control cells. Without wishing to be bound by theory, it is described herein that Noxa may promote cell death in T cells at the end of response following antigen stimulation and, therefore, knocking out Noxa may interrupt cell death. Similarly, as described herein for the first time, Noxa expression is significantly induced in human NK cells following stimulation (FIG. 15). Thus, Noxa may induce cell death in these and other hematopoietic cells. Human Noxa (hNoxa) is encoded by the PMAIP1 gene, located on the positive strand of chromosome 18 from nucleotide 59,899,996 to nucleotide 59,904,305 (UCSD Genome Browser, hg38). The Noxa protein (UniProt A13794-2) includes 54 amino acids and harbors a single BH3 domain that interacts with pro-survival Bcl-2 family members, such as Mcl-1, to promote apoptosis. hNoxa is the only BH3-only Bcl-2 protein that both promotes and prevents cell death in human T cells. Uniquely, hNoxa is also directly involved in the switch to glutaminolysis. Glutaminolysis is a known part of metabolic reprogramming following activation of the T cell immune response. The Noxa / Mcl-1 interaction may be important for apoptosis in T cells at the end of an immune response, and Noxa’s regulation of glutamine metabolism may be independent of its role in apoptosis. Naïve CD8+ T cells lack the Noxa protein, but consistently induce its expression following TCR engagement. The protein remains highly expressed during the T cell expansion phase, but its expression is suppressed in memory T cells. Post-translational regulation of hNoxa by phosphorylation also contributes to its growth- promoting role, as it curtails Noxa’s ability to bind Mcl-1 and promote apoptosis but typically does not affect its metabolic function. In proliferating T leukemia (T-ALL) cells, Noxa is stably phosphorylated on Ser13 and functions as a growth promoter when phosphorylated. In contrast, the dephosphorylated protein triggers apoptosis. In CD8+ T cells, Noxa remains phosphorylated during the expansion phase. CDK5 has been identified as a Noxa kinase in T leukemia cells. However, it is possible that additional kinases can phosphorylate Noxa, potentially in response to different stimuli. Investigating the role of kinases and phosphatases in regulating Noxa’s function, and the role of nutrients and fuel, in turn, in regulating kinase and phosphatase activity in primary CD8+ T cells should reveal new insights into signaling pathways associated with an immune response. As is shown for the first time herein, Noxa is also significantly induced in human NK cells upon stimulation (FIG 15) and could play a role in proliferative metabolism following activation, similar to the effects of Noxa on human T cells. Because Noxa induces NK cells in a manner similar to T cells and because Noxa is expressed in proliferating hematological cancer cells (e.g., ALL, T-ALL, and CLL), the data provided herein may be extrapolated to other hematopoietic cells. Thus, while the engineered cells and methods described herein are described in the context of exemplary embodiments that involve T cells, the engineered cells and methods can alternatively involve the use of other hematopoietic cells. Other hematopoietic cells include lymphoid cells (e.g., T cells, B cells, NK cells, etc.) and / or myeloid cells (e.g., neutrophils, dendritic cells, etc.). Noxa enhances glucose uptake and imparts a Warburg-like proliferative phenotype to T- ALL cells while, concomitantly, enhancing glutamine utilization for mitochondrial OxPhos. Noxa also promotes the growth and expansion of primary, normal human T lymphocytes through metabolic regulation by facilitating glutaminolysis for mitochondrial respiration. Noxa transcripts are upregulated following TCR engagement in human CD8+ T cells, but Noxa translation requires glutamine (FIG. 6). Conversely, glutamine conversion to glutamate through the TCA cycle is decreased in the absence of Noxa (FIG. 9, FIG. 10). However, mitochondrial fitness is not compromised, suggesting that mitochondria switch to an alternative fuel source for oxidative phosphorylation. The targeted tracer analysis in Noxa siRNA silenced human CD3+T cells described herein (FIG. 9, FIG. 10) places Noxa at the point at which glutamine enters the mitochondrial TCA cycle. Noxa is typically required for the first step in glutaminolysis during activation- induced metabolic reprogramming. The conversion of TCA cycle glutamine to glutamate, the first step in glutaminolysis, is catalyzed by glutaminase, either the “kidney-type” (GLS1) or the “liver-type” (GLS2). GLS2 is the active enzyme in T cells and is upregulated in T cells following activation. Although a pilot study suggested that GLS2 expression may not be affected by Noxa, it is possible that Noxa regulates GLS activity, either through direct interaction with GLS1 and / or GLS2, through a metabolic signaling cascade, or through regulation of upstream kinases that phosphorylate GLS1 / 2. For example, Noxa may directly or indirectly block FAO by reducing levels of carnitine palmitoyl-transferase 1a (CPT1a), transporter of acyl carnitine from cytosol to the mitochondria for FAO. Alternatively, Noxa may prevent phosphorylation of AMPK and its downstream targets, such as ACC1, since pAMPK, the active form of AMPK, is required for FAO. It is also possible that Noxa downregulates levels of fatty acid binding and transport membrane proteins. It is still unclear whether mouse Noxa harbors the pro-growth function of hNoxa. With 103 residues and two BH3 domains, murine Noxa (mNoxa) is a significantly different protein; it also lacks a serine at its amino terminus. mNoxa is therefore unlikely to be phosphorylated. Considering the differences between hNoxa and mNoxa, it is likely that hNoxa is a coordinator of activation, proliferation and T cell fate through metabolic regulation and protein-protein interactions, while mNoxa may play a different role. The present disclosure describes the role of Noxa in metabolically switching activated human CD8+ T cells first to proliferation and subsequently to apoptosis following antigenic activation. Although Noxa was initially classified as a death-promoter, the present disclosure describes a novel pro-growth metabolic function for the phosphorylated protein in human hematological malignancies and primary T cells. Methods of modulating a cell As is described herein, Noxa facilitates glutaminolysis and suppresses FAO during activation-induced metabolic reprogramming. Noxa’s dual roles in proliferative metabolism and apoptotic death in normal T cells makes it a potential target for immunotherapy and cancer therapy. In particular, the present disclosure describes that Noxa is required for metabolic reprogramming in CD8+ T cells following stimulation and modulates both the expansion and the apoptotic phases of an immune response. Accordingly, in one or more embodiments, the present disclosure relates to a method of modulating the activity of Noxa in a cell. Modulating the activity of Noxa may include modulating the expression of Noxa and / or modulating the phosphorylation of Noxa. In one or more embodiments, a method includes modulating the expansion of lymphocytes including modulating expression of Noxa. In one or more embodiments, the present disclosure relates to a method of modulating apoptosis in a lymphocyte including modulating expression of Noxa. In particular, a method of modulating apoptosis in a lymphocyte may include extending the lifespan of the lymphocyte by modulating expression of Noxa. Typically, modulating expression of Noxa includes decreasing or knocking out expression of Noxa. In another aspect, this disclosure describes an isolated nucleic acid sequence that encodes any embodiment of Noxa, or any component fragment of Noxa. In one or more embodiments, the isolated nucleic acid sequence encodes hNoxa. In one or more embodiments, the isolated nucleic acid encodes hNoxa(D34A). Given the amino acid sequence of Noxa (e.g., hNoxa 1-54, SEQ ID NO:1), or one or more component fragments of the Noxa, a person of ordinary skill in the art can determine the full scope of polynucleotides that encode that amino acid sequence using conventional, routine methods. As used herein, the term “nucleic acid” or “oligonucleotide” refers to polynucleotides such as deoxyribonucleic acid (DNA) or ribonucleic acid (RNA). Nucleic acids include but are not limited to genomic DNA, cDNA, mRNA, iRNA, miRNA, tRNA, ncRNA, rRNA, and recombinantly produced and chemically synthesized molecules such as aptamers, plasmids, anti- sense DNA strands, shRNA, ribozymes, nucleic acids conjugates, and oligonucleotides. A nucleic acid may be single-stranded, double-stranded, linear, or covalently circularly closed molecule. A nucleic acid can be isolated. The term “isolated nucleic acid” means that the nucleic acid (i) was amplified in vitro, for example via polymerase chain reaction (PCR), (ii) was produced recombinantly by cloning, (iii) was purified, for example, by cleavage and separation by gel electrophoresis, (iv) was synthesized, for example, by chemical synthesis, or (vi) extracted from a sample. A nucleic might be introduced—i.e., transfected—into cells. When RNA is used to transfect cells, the RNA may be modified by stabilizing modifications, capping, or polyadenylation. As used herein “amplified DNA” or “PCR product” refers to an amplified fragment of DNA of defined size. Various techniques are available and well known in the art to detect PCR products. PCR product detection methods include, but are not restricted to, gel electrophoresis using agarose or polyacrylamide gel and adding ethidium bromide staining (a DNA intercalant), labeled probes (radioactive or non-radioactive labels, southern blotting), labeled deoxyribonucleotides (for the direct incorporation of radioactive or non-radioactive labels) or silver staining for the direct visualization of the amplified PCR products; restriction endonuclease digestion, which relies on agarose gel electrophoresis, polyacrylamide gel electrophoresis, or high-performance liquid chromatography (HPLC); dot blots, using the hybridization of the amplified DNA on specific labeled probes (radioactive or non-radioactive labels); high-pressure liquid chromatography using ultraviolet detection; electro- chemiluminescence coupled with voltage-initiated chemical reaction / photon detection; and direct sequencing using radioactive or fluorescently labeled deoxyribonucleotides for the determination of the precise order of nucleotides with a DNA fragment of interest, oligo ligation assay (OLA), PCR, qPCR, DNA sequencing, fluorescence, gel electrophoresis, magnetic beads, allele specific primer extension (ASPE) and / or direct hybridization. Generally, nucleic acid can be extracted, isolated, amplified, or analyzed by a variety of techniques well known to those of ordinary skill in the art. Examples of nucleic acid analysis include, but are not limited to, sequencing and DNA-protein interaction. Sequencing may be by any method known in the art. DNA sequencing techniques include classic dideoxy sequencing reactions (Sanger method) using labeled terminators or primers and gel separation in slab or capillary, and next generation sequencing methods such as sequencing by synthesis using reversibly terminated labeled nucleotides, pyrosequencing, 454 sequencing, Illumina / Solexa sequencing, allele specific hybridization to a library of labeled oligonucleotide probes, sequencing by synthesis using allele specific hybridization to a library of labeled clones that is followed by ligation, real time monitoring of the incorporation of labeled nucleotides during a polymerization step, polony sequencing, and SOLiD sequencing. Separated molecules may be sequenced by sequential or single extension reactions using polymerases or ligases as well as by single or sequential differential hybridizations with libraries of probes. In one or more embodiments, modulating expression of Noxa includes administering an exogenous nucleic acid. In one or more such embodiments, the exogenous nucleic acid comprises a nucleic acid associated with RNA interference, such as siRNA, shRNA or miRNA. In one or more embodiments, the exogenous nucleic acid encodes one or more molecules that decrease expression of the Noxa protein. In one or more embodiments, the exogenous nucleic acid encodes an amino acid sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% identity to SEQ ID NO:1. In one or more embodiments, modulating expression of Noxa includes administering a gene editing composition. In one or more such embodiments, the gene editing composition includes CRISPR / Cas components, such as Cas9, a transcription activator-like effector nuclease, a zinc finger nuclease (ZFN), or an endonuclease. The gene editing composition may include additional components such as guide nucleic acids (e.g., gRNA, sgRNA), repair template nucleic acids, polymerases, or molecules associated with gene editing and repair. In one or more embodiments, a gene editing composition includes a donor vector including a desired sequence, such as the sequence of hNoxa. In one or more embodiments, a donor vector encodes the amino acid sequence of SEQ ID NO:1. In one or more embodiments, the donor vector encodes an amino acid sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% identity to SEQ ID NO:1. In one or more embodiments, the present disclosure relates to a method of modulating a lymphocyte including altering the PMAIP1 gene. Altering the PMAIP1 gene typically includes introducing one or more genetic alterations to the sequence of the gene. The genetic alteration may include, without limitation, a point mutation, a deletion of one or more nucleotides, or an insertion of one or more nucleotides. In one or more embodiments, the alteration includes a genetic alteration that encodes mutation of the aspartate at position 34 of the Noxa protein to alanine (D34A). In one or more embodiments, the alteration includes a genetic alteration that encodes a mutation at an aspartate equivalent to the aspartate at position 34 of hNoxa (SEQ ID NO:1) to alanine (D34A). In one or more embodiments, the alteration includes a genetic alteration that encodes mutation of the serine at position 13 to a phosphomimic residue, such as glutamate (S13E) or aspartate (S13D). In one or more embodiments, the alteration includes a genetic alteration that encodes a mutation at a serine equivalent to the serine at position 13 of hNoxa (SEQ ID NO:1) to a phosphomimic residue, such as glutamate (S13E) or aspartate (S13D). The core promoter region of the Noxa gene (PMAIP) contains several consensus sequences for binding of transcription factors, including a TATA box and a CCAAT box. These have been well-researched, at least because Noxa is typically silent in cells and epithelial cancers, while it is activated in response to stress stimuli. Several transcription factors including p53, c-MYC, ATF3, shown to bind the promoter region of the gene and positively regulate its expression, and the polycomb group protein BM11, a negative regulator, will be characterized in relation to Noxa. In particular, transcription factors relevant to Noxa expression in T cells representing each of the five activation phases identified in FIG. 1B will be characterized. In one or more embodiments, a method of modulating expression of Noxa includes administering one or more transcriptional repressors. In one or more embodiments, a method of modulating expression of Noxa includes administering one or more transcriptional activators. In particular, a method may include administering one or more transcriptional activators or transcriptional repressors associated with expression of the Noxa protein. For example, a genetic alteration may include mutation or deletion of a transcription factor, such as p52, MYC (e.g., c-MYC), ATF2, or BM11. In one or more embodiments, the genetic alteration to the PMAIP1 gene results in reduced or no transcription of the transcript that encodes the Noxa protein. In one or more embodiments, the genetic alteration to the PMAIP1 gene results in reduced or no translation of the Noxa protein. Noxa is induced in activated human CD8+ T cells and contributes to metabolic regulation and to apoptotic cell death in the T cell’s response and is both transcriptionally controlled and post-translationally modulated during this period. Phosphorylation of Noxa inhibits its ability to interact with Mcl-1 and trigger apoptosis, while enabling its role as growth promoter. Noxa’s phospho-motif (QPSPARA, SEQ ID NO:6, typically phosphorylated on the bold serine (S) of SEQ ID NO:1 below) is a strong consensus substrate for CDK5 which was identified as a Noxa kinase in T-ALL cells and in CD3+ T cells. The QPSPARA (SEQ ID NO:6) motif is also a strong substrate for protein phosphatase 2 (PP2C) phosphatases, such as the PH domain and leucine rich repeat protein phosphatase (PHLPP) family phosphatases. PHLPP2 cofractionates with pNoxa by size exclusion chromatography and co-localizes with pNoxa in a cytosolic multiprotein complex, which also harbors Mcl-1, in both Jurkat and pan T cells. In one or more embodiments, the genetic alteration to the PMAIP1 gene results in reduced or no phosphorylation of the Noxa protein. In particular, the genetic alteration may result in reduced or no phosphorylation of the Noxa protein. In another aspect, the present disclosure relates to a method of modulating Noxa phosphorylation. A method of modulating phosphorylation of Noxa may include administering one or more phosphatase inhibitors. Suitable phosphatase inhibitors include, without limitation, NSC45586, an inhibitor specific to the PHLPP2 phosphatase which has been shown to be resistant to most known phosphatase inhibitors. A method of modulating phosphorylation of Noxa may include administering one or more kinase inhibitors. In one or more embodiments, a method of modulating the activity of Noxa includes modulating the expression of one or more proteins associated with Noxa. Unphosphorylated Noxa is known to be associated with the protein myeloid cell leukemia 1 (Mcl-1). In one or more embodiments, a method of modulating activity of Noxa includes modulating the activity of Mcl- 1. In one or more embodiments, a method of modulating activity of Noxa includes modulating the expression of Mcl-1. In one or more embodiments, a method of modulating the activity of Noxa includes modulating the intracellular localization of Noxa. Phosphorylated Noxa (pNoxa) is typically localized to the cytosol in T-ALL cells. However, Noxa is found localized to both mitochondria and cytosol in CD8+ T cells (FIG. 5). Further, it is likely that Noxa is incompletely phosphorylated 48 hours post-stimulation (FIG. 4). Accordingly, it is likely that Noxa’s role in glutaminolysis (FIG. 8) is mitochondrial and may not require a phosphorylated protein. Thus, the activity of Noxa may be modulated by directing the protein to a particular cellular location. In one or more embodiments, a method includes localizing Noxa to the mitochondria. In one or more embodiments, a method includes localizing Noxa to the nucleus of a cell. In one or more embodiments, a method includes localizing Noxa to the cytosol. As it is used herein, “localizing” typically refers to appending to a protein one or more small molecules, such as amino acid tags, that traffics the protein to a desired cellular location. The methods disclosed herein may be compatible with any cells described herein, including engineered cells, described in greater detail. Engineered cells In one or more embodiments, the present disclosure relates to a cell, such as an engineered cell. In one or more embodiments, the cell includes a reduced amount of Noxa protein relative to a comparable non-engineered cell. In one or more embodiments, the cell includes a reduced amount of phosphorylated Noxa protein relative to a comparable non- engineered cell. The amount of Noxa and phosphorylated Noxa protein present in a cell may be measured by any suitable assay, such as by Western blot, ELISA, or flow cytometry. The sequence of hNoxa (1-54, SEQ ID NO: 1) is shown below: MPGKKARKNAQPSPARAPAELEVECATQ(LRRFGDK)LNFRQKLLNLISKLFCSGT Serine 13 is shown in bold. The pro-apoptotic BH3 domain is enclosed in parentheses (LRRFGDK, SEQ ID NO:2). In one or more embodiments, the cell includes one or more mutations to the PMAIP1 gene. The mutation may be an insertion, a deletion, or a point mutation. In one or more embodiments, the mutation may encode a D34A mutation to the Noxa protein. In one or more embodiments, the D34A mutation is within the BH3 domain, and serves to reduce binding to Mcl-1 and, thus, Noxa’s apoptotic function, while leaving its metabolic function unaffected. In one or more embodiments, the mutation encodes mutation of the serine at position 13 of the Noxa protein. As is described herein, mutation of the serine at position 13 may alter phosphorylation of Noxa and resultantly alter its pro-apoptotic activity. The serine at position 13 of the Noxa protein may be mutated to any phosphomimetic residue, such as glutamate or aspartate. In one or more embodiments, the cell includes one or more mutations to a gene associated with Noxa. For example, the cell may include a mutation to a transcription factor associated with expression of Noxa from the PMAIP1 gene. Additionally or alternatively, the cell may include one or more mutations to molecules associated with the metabolic processes and signaling cascades described herein. In one or more embodiments, the cell includes one or more exogenous nucleic acids. The one or more exogenous nucleic acids may be associated with RNA interference, such as siRNA, shRNA or miRNA. In one or more embodiments, the cell includes one or more nucleic acid molecules that decrease expression of the Noxa protein. In one or more embodiments, the cell treated by a method is a mammalian cell. The cell may be, without limitation, a human cell, a murine cell, an ovine cell, a hircine cell, a feline cell, a canine cell, or a porcine cell. In one or more embodiments, the cell is a mammalian cell including a nucleic acid encoding hNoxa. For example, the cell may be a murine cell encoding wild-type hNoxa or a murine cell encoding hNoxa(D34A). Examples of murine cells consistent with the cells of the present disclosure are described in greater detail in Example 9 and Example 10. In one or more embodiments, the cell is a hematopoietic cell. Typically, hematopoietic cells include lymphoid cells, such as T cells and B cells, and myeloid cells, such as a neutrophil. In one or more embodiments, the cell is an immune cell. Typically, immune cells are a subclass of hematopoietic cells, and include T cells, B cells, dendritic cells, and natural killer cells, in part. In one or more embodiments, the cell is a T cell. The T cell may be a cytotoxic T cell, such as a CD8+ T cell. The cytotoxic T cell may express a T cell receptor (TCR) against a specific antigen, such as an antigen associated with cancer or infection. The TCR may be any suitable TCR, such as an αβ-TCR. In one or more embodiments the T cell expresses a TCR against a known antigen. In one or more embodiments, the T cell is a CD3+ T cell. In one or more embodiments, the T cell expresses one or more of IFNγ, lymphotoxin-α, TFNα, IL-15, IL- 10, IL-17, IL-21, IL-18, IL-12, and IL-2. In one or more embodiments, the T cell may be a naïve T cell, an effector T cell, an effector memory T cell, a central memory T cell, or a regulatory T cell. Based upon this finding and other findings related to T cells described herein, it is believed that Noxa may play a similar role in other hematopoietic cell types, particularly lymphatic cells. Thus, while described herein primarily in the context of T cells and NK cells, the methods and compositions of the present disclosure may be applied to other cell types such as B cells and dendritic cells. In one or more embodiments, the cell is a B cell, such as a primary B cell. In one or more embodiments, the primary B cell is a non-clonal cell. In one or more embodiments, the primary B cell is a proliferating cell. In one or more embodiments, the B cell is cultured in the presence of CD40L. In one or more embodiments, the B cell is a naive B cell. In one or more embodiments, a “naive B cell” is CD19+, lgD+, lgM+, CD27; CD21+, and / or CXCR5+. In one or more embodiments, the B cell is a memory B cell. In one or more embodiments, a “memory B cell” is CD19+, IgD; CD27+ or CD27; CD21+, and / or CXCR5+. In one or more embodiments, the B cell is an activated memory B cell. In one or more embodiments, an “activated memory B cell” is CD19+, IgD; CD27+, CD21; and / or CXCR5+. In one or more embodiments, the B cell is a natural effector B cell. In one or more embodiments, a “natural effector B cell” is CD19+, lgD+, lgM+, and / or CD27+. In one or more embodiments, the B cell is a plasmablast. In one or more embodiments, a “plasmablast” is CD19+, CXCR5; CD38+, CD27; and / or CD20’. In one or more embodiments, the cell is a chimeric antigen receptor (CAR) T cell. A CAR T cell may include an exogenous nucleic acid encoding a chimeric antigen receptor. In one or more embodiments, knocking out Noxa from CAR T cells expands lifespan of the CAR T cells without compromising effector function. Similarly, modulating activity or otherwise modulating expression of Noxa in CAR T cells may expand the lifespan of CAR T cells without compromising effector function. In one or more embodiments, the CAR is specific to an antigen associated with a disease, such as cancer. In one or more embodiments, the CAR includes a stimulatory domain against CD28. In one or more embodiments, the CAR is specific to CD30, CD33, CD123, FLT3, mesothelin, B7H3, CD19, or BCMA. In one or more embodiments, the cell is a CAR T cell specific to mesothelin. Example 13 describes mesothelin-specific CAR T cells. These CAR T cells were evaluated for effector function was evaluated in response to TCR (anti-CD3 and anti-CD28) co-stimulation (FIG. 16, FIG. 17). In one or more embodiments, the cell is a CAR T cell specific to CD19. CD19 is a transmembrane protein typically expressed on B cells. Often, CD19 is targeted for treatment of B cell cancers, such as lymphoma. In addition, CD19 is often targeted for treatment of leukemia, such as acute lymphoblastic leukemia, and chronic lymphocytic leukemia. In one or more embodiments, the cell is a CAR T cell specific to B7H3. B7H3, sometimes referred to as CD276, is often targeted for treatment of lung cancer. Example 14 describes mesothelin, CD19, and B7H3-specific CAR T cells. In one or more embodiments, the cell is a CAR NK cell. CAR NK cells are natural killer (NK) cells engineered to include a chimeric antigen receptor, much like a CAR T cell. A CAR NK cell may be specific to any suitable antigen, such as mesothelin, CD19, B cell maturation antigen (BCMA), or B7H3. A CAR NK cell may be specific to any antigen described herein. Tumor infiltrating lymphocyte (TIL) therapy, an adoptive cellular therapy, involves harvesting infiltrated lymphocytes from tumors, culturing and expanding them in vitro, then infusing them back into patients as treatment. It is being increasingly preferred over other adoptive cellular therapies, such as CAR T cell therapy, for treating solid tumors, for its diverse TCR clonality, superior tumor-homing ability and low off-target toxicity, and has been successful against several cancer types, including lung and colorectal cancer. In one or more embodiments, the cell is a tumor infiltrating lymphocyte (TIL). In one or more embodiments, the TIL may include a T cell, such as a tumor-specific T cell. The TIL may be patient-derived, such as from a cancer patient. Typically, an engineered TIL of the present disclosure includes an exogenous nucleic acid encoding or modulating Noxa, such as hNoxa. In one or more embodiments, an engineered TIL has an altered level of Noxa (e.g., hNoxa) expression or activity. In one or more embodiments, an engineered TIL may be a CAR T cell, such as a CAR T cell with specificity against mesothelin, B7H3, or CD19. The cell may be derived from any suitable source. Typically, the cell is a primary (e.g., a non-immortalized) cell. For example, the cell may be derived from a donor sample, such as a patient sample. In one or more embodiments, the cell may be derived from a patient sample, engineered using methods disclosed herein, and readministered to the patient (e.g., autologously transplanted). In one or more embodiments, the cell may be derived from a donor sample, engineered using methods disclosed herein, and administer to an unrelated patient (e.g., allogenically transplanted). In one or more embodiments, the cell is derived from an immortalized cell. In one or more embodiments, the cell is a leukemia cell. For example, the cell may be derived from an immortalized leukemia cell line. Suitable leukemia cell lines include the KASUMI-1, HL-60, THP-1, K-562, RS4;11, MOLT-4, CCRF-CEM cell lines, available from American Type Culture Collection (ATCC). In one or more embodiments, the cell exhibits distinctive properties relative to a non- engineered cell. For example, the cell may exhibit a longer lifespan than a non-engineered cell. In one or more embodiments, the cell is more resistant to apoptosis following antigenic activation relative to a non-engineered cell. In one or more embodiments, an engineered cell may exhibit a longer lifespan than a non-engineered cell by at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, or at least 35%. Lifespan may be measured using any suitable method such as tissue culture. Characteristics of the cell may be measured using microscopy, such as fluorescent microscopy or phase contrast microscopy. In one or more embodiments, the cell exhibits higher levels of proliferation relative to a non-engineered cell. For example, the cell may yield a greater number of progeny following expansion relative to a comparable non-engineered cell. Proliferation may be measured using any suitable method, such as nuclear staining and cellular counting or flow cytometry. Animal models Although in vitro studies on hNoxa continue to reveal new insights into its unique role in human T cells, no suitable animal models have historically been available. Many studies of hNoxa use donor samples, which lack of a uniform genetic background, compromising rigor and reproducibility. Described herein are two gene replacement mouse models of hNoxa generated to address, the absence of a reliable, reproducible in vivo model. In one or more embodiments, the engineered mouse includes a genetic modification to replace the entire mouse mNoxa gene and promoter with the corresponding promoter / gene region of hNoxa. In one aspect, the present disclosure relates to an animal model. For example, an animal model may be an engineered animal including a nucleic acid encoding Noxa, such as hNoxa. In one or more embodiments, the engineered animal is an engineered mus musculus, or house mouse. In one or more embodiments, an engineered mouse may be from a C57BL / 6 background. Examples 8-10 describe engineered mice expressing hNoxa generated by replacing the murine Noxa gene / promoter region with the corresponding region of the human gene. This model allows the unique biological properties of hNoxa (versus mNoxa) to be investigated in a genetic model for the first time. The sequence of mouse Noxa (1-103, SEQ ID NO:3) is shown below: MPGRKARRNAPVNPTRAELPPEFAAQ(LRKIGDK)VYCTWSAPDITVVLAQMPGKSQKS RMRSPSPTRVPADLKDECAQ(LRRIGDK)VLNRQKLLNLISKLFNLVT The pro-apoptotic BH3 A and BH3 B domains are enclosed in parentheses (LRKIGDK, SEQ ID NO:4) and (LRRIGDK, SEQ ID NO:5). While mouse Noxa (mNoxa) and hNoxa are related, the pro-growth function and phospho-regulation of hNoxa are not shared by the mNoxa protein. Although the mouse protein is a potent death promoter, its gene is a product of duplication, and codes for a protein that is significantly larger and harboring two apoptotic BH3 domains. In contrast, hNoxa only includes one BH3 domain (see above). The mouse protein is also unlikely to be post-translationally regulated like its human counterpart as it lacks the N-terminal serine residue which converts hNoxa to a growth promoter when phosphorylated. Thus, studies of mNoxa have limited applicability to humans. An animal model where the promoter / gene region of endogenous Noxa are replace with the promoter / gene region of hNoxa may be generated using a Bacterial Artificial Chromosome or BAC. In both cases, the endogenous mNoxa gene locus was deleted and replaced with (1) WT hNoxa (PMAIP1) or (2) mutant (hNoxa-D34A). A schematic of a strategy consistent with the present disclosure is shown in FIG. 27. Mice generated using this strategy are described in greater detail in Example 8. Briefly, hNoxa mice are healthy, show no overt phenotypic abnormalities, and develop at a normal rate compared to wildtype (C57BL / 6) mice. A pilot in vitro activation experiment with CD8+T cells isolated from spleen of homozygous WT hNoxa mice confirmed that Noxa protein is induced within 2 days of co-stimulation (FIG. 28B), suggesting that the hNoxa promoter is functioning normally. The Noxa antibody used was specific to hNoxa and did not detect protein in WT mouse T cells (FIG.28A). While described herein in the context of an engineered mouse, other animal models are contemplated. In one or more embodiments, the animal model is a rat, a ferret, a non-human primate, a guinea pig, a dog, or a fruit fly. In the preceding description and following claims, the term “and / or” means one or all of the listed elements or a combination of any two or more of the listed elements; the terms “comprises,” “comprising,” and variations thereof are to be construed as open ended—i.e., additional elements or steps are optional and may or may not be present; unless otherwise specified, “a,” “an,” “the,” and “at least one” are used interchangeably and mean one or more than one; and the recitations of numerical ranges by endpoints include all numbers subsumed within that range (e.g., 1 to 5 includes 1, 1.5, 2, 2.75, 3, 3.80, 4, 5, etc.). As used herein, “have,” “has,” “having,” “include,” “includes,” “including,” “comprise,” “comprises,” “comprising” or the like are used in their open-ended inclusive sense, and generally mean “include, but not limited to,” “includes, but not limited to,” or “including, but not limited to.” Further, wherever embodiments are described herein with the language “have,” “has,” “having,” “include,” “includes,” “including,” “comprise,” “comprises,” “comprising” and the like, otherwise analogous embodiments described in terms of “consisting of” and / or “consisting essentially of” are also provided. The term “consisting of” means including, and limited to, that which follows the phrase “consisting of.” That is, “consisting of” indicates that the listed elements are required or mandatory, and that no other elements may be present. The term “consisting essentially of” indicates that any elements listed after the phrase are included, and that other elements than those listed may be included provided that those elements do not interfere with or contribute to the activity or action specified in the disclosure for the listed elements. As used herein, the word “exemplary” means to serve as an illustrative example and should not be construed as preferred or advantageous over other embodiments. As used herein, the terms “preferred” and “preferably” refer to embodiments of the invention that may afford certain benefits under certain circumstances. However, other embodiments may also be preferred under the same or other circumstances. Furthermore, the recitation of one or more preferred embodiments does not imply that other embodiments are not useful and is not intended to exclude other embodiments from the scope of the invention. In the preceding description, particular embodiments may be described in isolation for clarity. Reference throughout this specification to “one embodiment,” “an embodiment,” “certain embodiments,” “one or more embodiments,” or “some embodiments,” etc., means that a particular feature, configuration, composition, or characteristic described in connection with the embodiment is included in at least one embodiment of the disclosure. Thus, the appearances of such phrases in various places throughout this specification are not necessarily referring to the same embodiment of the disclosure. Furthermore, the particular features, configurations, compositions, or characteristics may be combined in any suitable manner in one or more embodiments. Thus, features described in the context of one embodiment may be combined with features described in the context of a different embodiment except where the features are necessarily mutually exclusive. In several places throughout the above description, guidance is provided through lists of examples, which examples can be used in various combinations. In each instance, the recited list serves only as a representative group and should not be interpreted as an exclusive list. For any method disclosed herein that includes discrete steps, the steps may be performed in any feasible order. And, as appropriate, any combination of two or more steps may be performed simultaneously. ILLUSTRATIVE EMBODIMENTS Embodiment 1 is an engineered cell including a decreased amount of a Noxa protein relative to a comparable non-engineered cell. Embodiment 2 is an engineered cell including an engineered Noxa protein including lower pro-apoptotic activity relative to a non-engineered Noxa protein. Embodiment 3 is the engineered cell of embodiment 1 or embodiment 2, wherein the engineered cell includes one or more modifications to the PMAIP1 gene. Embodiment 4 is the engineered cell of embodiment 3, wherein the one or more modifications decreases the activity of the Noxa protein. Embodiment 5 is the engineered cell of embodiment 3, wherein the modification to the PMAIP1 gene encodes mutation of an aspartate at a position equivalent to aspartate 34 in SEQ ID NO:1 to alanine (D34A). Embodiment 6 is the engineered cell of embodiment 3, wherein the modification to the PMAIP1 gene encodes mutation of a serine at a position equivalent to serine 13 in SEQ ID NO:1 to glutamate (S13E) or aspartate (S13D). Embodiment 7 is the engineered cell of any preceding embodiment, wherein the cell is an immune cell. Embodiment 8 is the engineered cell of embodiment 7, wherein the engineered cell includes an engineered T cell. Embodiment 9 is the engineered cell of embodiment 8, wherein the T cell includes a CD8+ T cell. Embodiment 10 is the engineered cell of embodiment 8 or embodiment 9, wherein the T cell includes a human T cell. Embodiment 11 is the engineered cell of any preceding embodiment, wherein the engineered cell includes a chimeric antigen receptor T cell (CAR T cell). Embodiment 12 is the engineered cell of embodiment 11, wherein the CAR includes an anti-B7H3, anti-CD19, or anti-mesothelin CAR T cell. Embodiment 13 is the engineered cell of any preceding embodiment, wherein the engineered cell includes a tumor infiltrating lymphocyte (TIL). Embodiment 14 is the engineered cell of embodiment 7, wherein the engineered cell includes an engineered NK cell. Embodiment 15 is the engineered cell of embodiment 14, wherein the NK cell includes a CAR NK cell. Embodiment 16 is the engineered cell of embodiment 15, wherein the CAR NK cell includes an anti-B7H3, anti-CD19, or anti-mesothelin CAR T cell. Embodiment 17 is the engineered cell of any preceding embodiment wherein the engineered cell is derived from a primary cell. Embodiment 18 is the engineered cell of any preceding embodiment, wherein the engineered cell exhibits a longer lifespan than a comparable non-engineered cell. Embodiment 19 is the engineered cell of embodiment 18, wherein the engineered cell exhibits at least a 30% longer lifespan than a comparable non-engineered cell. Embodiment 20 is a method of regulating Noxa expression in a cell including administering a gene editing composition to the cell. Embodiment 21 is a method of regulating Noxa phosphorylation in a cell including administering a gene editing composition to the cell. Embodiment 22 is the method of embodiment 20 or embodiment 21, wherein the gene editing composition introduces one or more mutations to the PMAIP1 gene. Embodiment 23 is the method of embodiment 22, wherein the one or more mutations to the PMAIP1 gene encodes mutation an aspartate at a position equivalent to aspartate 34 in SEQ ID NO:1 to alanine (D34A). Embodiment 24 is the method of embodiment 22, wherein the one or more mutations to the PMAIP1 gene encodes mutation of the serine at a position equivalent to serine 13 in SEQ ID NO:1 to glutamate (S13E) or aspartate (S13D). Embodiment 25 is a method of extending the lifespan of a cell including decreasing the activity of the Noxa protein. Embodiment 26 is the method of embodiment 25, wherein the activity of the Noxa protein includes pro-apoptotic activity. Embodiment 27 is the method of any preceding embodiment, wherein the cell includes a human cell. Embodiment 28 is the method of any preceding embodiment, wherein the cell includes a CD8+ T cell. Embodiment 29 is the method of any preceding embodiment, wherein the cell includes a chimeric antigen receptor T cell (CAR T cell). Embodiment 30 is the method of any preceding embodiment, wherein the cell includes a tumor infiltrating lymphocyte (TIL). Embodiment 31 is the method of any preceding embodiment, wherein the cell includes an NK cell. Embodiment 32 is the method of any preceding embodiment, wherein the cell includes a CAR NK cell. EXAMPLES The present invention is illustrated by the following examples. It is to be understood that the particular examples, materials, amounts, and procedures are to be interpreted broadly in accordance with the scope and spirit of the invention as set forth herein. Example 1 - Noxa activation throughout T cell engagement in vitro It was previously shown that Noxa was induced in human T cells populations within 24 hours to 48 hours of TCR engagement in vitro. Preliminary studies with human CD8+ T cells included here show that Noxa expression is transcriptionally, post-transcriptionally and post- translationally regulated during an immune response and may contribute to several phases of the response. This Example focuses on a CD8+ T cell model to follow Noxa expression through proliferation, differentiation, and apoptosis, to memory cell formation, in a largely single subset in contrast to a CD4+ T cell model. Induction of Noxa protein expression in CD8+ T cells within 40 hours of TCR (CD3) / CD28 co-stimulation has been consistently observed. Noxa was first identified in Jurkat T cells as a PMA-inducible protein; but human CD8+ T cells also upregulate Noxa protein in response to activators, PMA / ionomycin, as well as the mitogen ConA. An in vitro assay was developed to further investigate the contribution of Noxa to the CD8+ T cell immune response. Human CD8+ T cells (naïve or total CD8+ T cells) were isolated from peripheral blood mononuclear cells (PBMCs) obtained from leukoreduction system chambers and stimulated with anti-CD3 / CD28 antibodies in media supplemented with IL-2. Cells were harvested at specific time points and evaluated for activation, proliferation, metabolic state, and apoptosis, using flow cytometry, Western blotting, and mitochondrial respiration rates. FIG. 1A shows a representative Western blot of Noxa protein in T cells over a 21-day period after activation of a naïve CD45RA+ enriched population. The naïve population is replaced by day 5 with a CD45RO+ effector / memory component (FIG. 1C); other markers shown are CD69 (an early activation marker), CD107a (an effector cell marker) and HLA-DR (a late cytotoxic T lymphocyte (CTL) marker) (FIG. 2); CD62L (naïve cell marker) and cleaved caspase-3 (apoptosis marker), which peaks between 5 days and 8 days depending on the donor, are not shown. A drop in HLA-DR levels signals the end of the immune response and emergence of a viable memory-like population within the apoptotic milieu. This enriched viable cell population of memory cells (Mem-US) can be restimulated (FIG. 1A) and evaluated. Thus, Noxa protein expression in CD8+ T cells correlates with proliferative, effector and apoptotic phases but not with the quiescent, non-dividing, naïve or memory cell state, depicted in the simple model in FIG.1B. Example 2 - Identifying transcription factors associated with the PMAIP1 gene The observed induction of Noxa protein in CD8+ T cells upon TCR engagement is transcriptional (FIG. 3A, B). Thus, the PMAIP1 gene encoding Noxa is silent in naïve cells and in memory populations. Given the pattern of Noxa expression over the course of the immune response, upstream signals and transcription factors that regulate hNoxa gene expression may be used to uniquely manipulate Noxa expression. Nucleic from cell representing each of the 5 phases (naïve, 48 hours, 7 days, Mem-US and Mem-restimulated at 24 hours), shown in FIG. 1B, were isolated, subjected to fragmentation and tagmentation with TN5 Transposase and sequencing adapters, and libraries analyzed by next generation sequencing (NGS) and bioinformatics. FIG. 3C shows are bias-corrected open chromatin profiles from bioinformatics analysis of naïve, 48 hour and 7 days post-stimulation CD8+ T cells in this pilot study. Example 3 - Characterizing Noxa phosphorylation profiles The Noxa protein is typically post-translationally regulated by Ser13 phosphorylation in CD8+ T cells. It has been shown that Noxa is phosphorylated on Ser13 in proliferating T-ALL cells and in stimulated CD3+T cells, and that dephosphorylation, which promotes Mcl-1 / Noxa interactions was required to activate its pro-apoptotic function and trigger apoptosis. It was hypothesized that Noxa is first phosphorylated upon induction in an antigenically stimulated T cell and promotes growth during rapid expansion, and subsequently dephosphorylated during the contraction phase, interacting with its canonical binding partner to promote apoptosis. The role of phosphorylation in regulating Noxa in stimulated T cells was investigated using a pS13-Noxa- specific antibody specific to pS13-Noxa. FIG. 4 shows pS13-Noxa in Western blots of lysates from naïve CD8+ T cells and at several time points post-stimulation. These data suggest that both pNoxa and Noxa are present in the activated cell populations. Example 4 - Characterizing intracellular distribution of Noxa Intracellular distribution of Noxa is regulated during an immune response. hNoxa can be localized to both cytosol and mitochondria, and the protein harbors a mitochondrial targeting domain in its C-terminus. Mitochondrial / cytosolic fractionation of human CD8+ T cells was carried out in accordance with manufacturer’s instruction (ABCAM kit ab65320). T cells (40×106), collected at the times indicated, were resuspended in cytosol extraction buffer, homogenized, and centrifuged to recover the supernatant as the cytosolic fraction and the pellet as the mitochondrial fraction. A Western blot of a representative experiment is shown in FIG. 5. Given that Noxa promotes both survival and death, and that phosphorylation of Ser13 regulates this function, studies correlating its opposing functions and posttranslational modification with its localization should further inform strategies to modulate Noxa activity. Example 5 - Investigating the effect of glutamine on Noxa protein expression Glutamine is typically required for induction of Noxa protein in activated CD8+ T cells. CD8+ T cells failed to induce Noxa protein when co-stimulated in the absence of glutamine (FIG. 6A). This regulation was likely to be at the translational as Noxa mRNA levels were not affected (FIG. 6B). In T-ALL cells, Noxa expression has been associated with increased entry of glutamine carbons into the TCA cycle. As expected, mitochondrial SRC was significantly impaired in the absence of glutamine (FIG.7C, D). To determine whether glutaminolysis was essential for the observed induction of Noxa two inhibitors, CB-839, a glutaminase inhibitor, and DON, a glutamine agonist, were tested. Glutaminase (GLS) inhibitor CB-839 did not affect Noxa protein induction or mitochondrial SRC, but glutamine antagonist and broad-based inactivator of glutamine-utilizing reactions, DON, had the same effect as glutamine deprivation on Noxa induction, proliferation, and effector formation (FIG. 7, FIG. 8A). Additionally, cell-permeable glutamate and α-ketoglutarate (αKG), TCA cycle products of glutaminolysis could not restore Noxa expression (FIG.8B), did not affect early activation (not shown) but reduced differentiation to effector cells (FIG. 8C). Thus, Noxa expression required glutamine, not glutaminolysis. Example 6 - Characterizing the role of Noxa and glutaminolysis on T cell proliferation Noxa is typically required for the metabolic switch to glutaminolysis, but not for mitochondrial respiration following activation. Because induction of the Noxa protein in primary T cells required the presence of glutamine (FIG. 6 to FIG. 8), it has been historically unclear whether Noxa regulates the switch to the proliferative phase in T cells. Stable isotope tracer analysis of siNoxa and control activated panT cells labeled with [U-13C5]-glutamine for 24 hours show glutamate-enrichment was severely impaired in the absence of Noxa (FIG. 9A, B). Additionally, with CB-839 serving as positive control, Noxa siRNA expressing CD8+ T cells showed significant reduction in intracellular glutamate upon activation (FIG. 10A, B). Despite the block in glutamate enrichment, mitochondrial fitness was not impaired, and even improved slightly, in siNoxa cells (FIG. 9C, D, FIG. 10D-G). This data demonstrated that (i) Noxa protein translation during activation requires glutamine, (ii) Noxa is required for glutaminolysis (simple model in FIG. 10C). The high SRC in activated siNoxa T cells suggests that they switch to an alternate fuel source for energy generation in the absence of glutaminolysis. Glutamine regulation of Noxa protein expression is mediated through the mTORC1 pathway. Given that Noxa induction required glutamine itself rather than glutamine catabolism, it was hypothesized that glutamine limitation could indirectly be modulating mTORC1 activity. mTOR regulates protein translation through effector molecules eukaryotic initiation factor binding protein 4EBP-1 and ribosomal protein p70S6K. Data for this set of completed experiments are not shown but results are summarized in the model in FIG. 11A. Inhibition of mTORC1 activity with rapamycin and everolimus in the presence of glutamine completely reversed the induction of Noxa as well as effector formation but not activation (not shown). Glutamine transporters and antiporters, such as CD98, xCT / SLC7A11, ASCT2 / SLC1A5, and SLC3A2 / SLC7A5, facilitate the uptake and / or rapid efflux of glutamine in exchange for essential amino acids (e.g., leucine, cystine) that are important for mTOR function. The glutamine-leucine exchange is a rate-limiting step in mTORC1 activation. To test whether reduced function of the glutamine-leucine antiport affected Noxa translation, cells were incubated with one of two specific antagonists of glutamine transporters (V9302 and JPH203, see model in FIG.11A and legend) and an inhibitor of the less specific cationic amino acid transporter NEM N-Ethylmaleimide CAT-1 transporter inhibitor. Inhibition of glutamine influx as well as glutamine-leucine exchange abrogated induction of Noxa and differentiation to effectors while the CAT-1 inhibitor was only weakly effective. Thus, glutamine may regulate Noxa protein levels in stimulated T cells primarily through mTORC1 via the glutamine-leucine antiporter. To summarize, Noxa gene expression is activated in CD8+ T cells in response TCR engagement. However, translation of mRNA requires glutamine to be available to facilitate intracellular entry of leucine, an activator of the mTOR pathway and, thus, protein translation. CD8+ T cell activation is not inhibited in the absence of glutamine or by mTOR inhibition, but the activated cells do not differentiate to effectors and show reduced mitochondrial fitness. This Example further suggests that Noxa regulates the conversion of glutamine to glutamate to control the entry of glutamine carbons into the TCA cycle at the time cells switch to glucose (and aerobic glycolysis) for growth and to glutamine (and glutaminolysis) for energy production. Example 7 - Characterizing the role of Noxa during apoptosis following an immune response To further test the hypothesis that Noxa is required both for the initial metabolic switch to proliferation and for apoptotic cell death at the end of the immune response, the effects of knocking out Noxa on CD8+ T cell activation and expansion, effector function, viability, and apoptosis were investigated. Two major differences that emerged are the lower dependence on glutamine for mitochondrial respiration (not shown) and delayed apoptosis. FIG. 11C shows representative Western blots of early and late passage Noxa (CRISPR) KO CD8+ T cells. The data on the right (FIG. 11C, average of 3 independent donors) show that NoxaKO cells are significantly more viable than control cells during the contraction phase, indicating that Noxa contributes to apoptosis at the end of the immune response. Memory cells generated in vitro display increased mitochondrial fitness and show dependence on fatty acids for respiration. Western blots of cell lysates from in vitro stimulation experiments (e.g., FIG. 1A) do not detect Noxa in memory cells but show it is rapidly induced upon their restimulation. However, it has been observed that a significant decrease in HLA-DR levels signals the emergence of a viable memory-like population (FIG. 2) This reliable pattern has allowed the consistent enrichment of large, relatively pure, memory cell populations, primed for rapid response to recall stimulation, and displaying the robust mitochondrial respiratory capacity associated with the memory phenotype in a mitochondrial stress test (FIG. 12A, B). The many-fold higher spare respiratory capacity (SRC) and max OCR in an unstimulated memory population (Mem-US) compared to activated naïve (40h) and reactivated CD8+ T cells (40h-re) from the same donor is evident. While SRC values varied between donors, unstimulated memory cells consistently exhibited higher SRC before re-stimulation and dependence on fatty acids for respiration in the form of reduced OCR and SRC in response to (5 μM) etomoxir, FAO inhibitor, (FIG. 12C, D). Example 8 - Characterization of hNoxa and hNoxa-D34A mice A pilot in vitro activation experiment with CD8+ T cells isolated from spleen of hemizygous WT hNoxa mice showed that Noxa protein is induced within 40 hours of co- stimulation and the induction is glutamine dependent (FIG. 13A), suggesting that the hNoxa promoter is functioning normally. In addition, splenic CD8+ T cells from a WT hNoxa mouse showed a greater dependence on OxPhos for the unstimulated population and a switch to aerobic glycolysis following 16 hours of co-stimulation (FIG. 13B, C), indicating that the cells exhibited a normal metabolic profile. To further investigate this finding, novel gene-replacement mouse models for in vivo validation of hNoxa regulation were generated. Previous conditional knockout (KO) Noxa models have failed to produce viable animals, indicating that properly timing NoxaKO is important. To create a sustainable model for hNoxa that allowed us to investigate its post- translational regulation and role in T cell expansion and apoptosis in vivo, without interference from the mNoxa protein, two gene replacement (GR) models were generated. Each GR model replaced the entire mouse (m)Noxa gene / regulatory region with the corresponding promoter / gene region of hNoxa using a Bacterial Artificial Chromosome or BAC. In both cases, the endogenous mNoxa gene locus was deleted and replaced with WT hNoxa (PMAIP1) or mutant (hNoxa-D34A), on a C57BL / 6 background. The D34A point mutation to the BH3 domain, abrogates the interaction with Mcl-1L preventing apoptosis. Mice were generated using a donor vector including 5′ and 3′ arms homologous to the genomic regions flanking the mNoxa allele. CRISPR / Cas9 was used to induce breaks within the genome, and the donor vector was inserted using endogenous repair machinery (FIG. 27). hNoxa mice and hNoxa-D34A mice were viable and produce viable offspring when self- crossed. The hNoxa mice were healthy, showed no overt phenotypic abnormalities, and developed at a normal rate compared to wildtype (C57BL / 6) mice. A pilot in vitro activation experiment with CD8+T cells isolated from spleen of homozygous WT hNoxa mice confirmed that Noxa protein is induced within two days of co-stimulation (FIG. 28B), suggesting that the hNoxa promoter is functioning normally. The Noxa antibody is specific to hNoxa, so no protein was detected in the WT mouse T cells (FIG.28B). Example 9 - Response of hNoxa and hNoxa(D34A) mice to viral challenge hNoxa GR and hNoxa(D34A, “hNoxaDA” in FIG. 29) GR mice, and WT C57BL / 6 controls (6 to12 weeks old), were infected with Lymphocytic choriomeningitis virus (LCMV:Armstrong strain), which induces a robust and well characterized CD8+ T cell response. Spleens were harvested 8 days, 15 days, or 30 days post-infection (corresponding to peak effector, contraction, and memory immune response phases, respectively), LCMV specific CD8+ T cells were identified using a set of peptide / MHC tetramers (from the NIH Tetramer Facility) and assayed for proliferation, markers of effector / memory differentiation and survival. Effector cells were assayed for in vitro function (IFN-γ production, cytolysis). Noxa expression was detected by flow cytometry (once detection parameters are optimized) and by Western blots. Robust induction of Noxa protein levels at the peak response time point and significant reduction in levels at the 30-day time point suggested the hNoxa promoter is functioning similarly in human and mouse T cells (FIG.29). Example 10 - Characterizing the role of Noxa in TIL therapy Expression and regulation of Noxa in expanded TILs was investigated. To investigate regulation of Noxa in expanded TILs, large numbers of cytotoxic CD8+ T cells were isolated from outgrown TIL and the effects of manipulating Noxa expression on TIL endurance, memory cell formation, and viability were studied. TIL were outgrown from tumor slurries. Briefly, dissected tumor fragments (~2 mm) were seeded into individual wells of a 24-well GRex plates and cultured in the presence of 6000 IU / mL for 4-6 weeks (FIG. 30A). Expanded CD8+ TIL populations were genetically engineered to knock out Noxa or to express an apoptosis deficient mutant using a Rapid Expansion Protocol (REP), following CRISPR / Cas9-mediated gene disruption in TIL (FIG. 14). NoxaKO and apoptosis-deficient populations were tested for proliferative capacity, reduced dependency on glutamine, and extended viability and survival compared to controls following antigenic stimulation. Flow cytometry was used to quantify the percentage of CD8+, CD69+, and GZMB+ cells, as well as the percentage of viable cells. It was found that NoxaKO TILs exhibited extended viability and survival compared to controls (FIG. 30F). In addition, NoxaKO TILs exhibited peak activation around day 4 or day 6 (FIG. 30D), and peak cytotoxic potential around day 4 or day 6 (FIG. 30E). A Western blot was used to measure Noxa expression in Cas9 (control) and NKO cells, and it was found that Noxa was not expressed in NKO cells (FIG. 30C). Example 11 – Noxa regulation in human NK cells NK cell isolation and stimulation Natural killer (NK) cells were isolated from peripheral blood mononuclear cells (PBMCs) using the EASYSEP Human NK Cell Isolation Kit (StemCell Technologies, Cat# 17955). Isolated NK cells were cultured in CTS AIM V SFM medium supplemented with 5% CTS Immune Cell SR (Thermo Fisher Scientific, Inc., Waltham, MA), penicillin / streptomycin, and IL-2 (100 IU / mL). NK cells were activated through co-culture with X-irradiated (100 Gy) feeder cells (C2, K562 expressing membrane-bound IL-21 and 41BB-L). Western blot analysis For Western blot analysis, unstimulated and 5 day-stimulated NK cells were harvested and lysed using RIPA Lysis and Extraction Buffer (Thermo Scientific, Cat# 89900) supplemented with Protease Inhibitor Cocktail (Thermo Scientific, Cat# 78410) and HALT Phosphatase Inhibitor Cocktail (Thermo Scientific, Cat# 78420). Protein lysates (40 μg) were denatured with 2× Laemmli Buffer (Bio-Rad, Cat# 1610737) at 85 °C for three minutes, resolved by SDS-PAGE, and transferred onto nitrocellulose membranes. The membranes were blocked in TBS-TM buffer (20 mM Tris-HCl, pH 7.6, 137 mM NaCl, 0.1% Tween-20, and 5% skim milk) and incubated overnight at 4 °C with primary antibodies diluted in TBS-T buffer. Primary antibodies used were β-Actin (Santa Cruz Biotechnology, Cat# sc-69879, 1:7000) and Noxa 114C307 (Santa Cruz Biotechnology, Cat# sc-56169, 1:250). Following incubation with HRP- conjugated secondary antibodies (Invitrogen) in TBS-TM buffer for one hour at room temperature, bands were detected using the Pierce ECL Western Blotting Substrate (Thermo Scientific, PI80196). NK cells were counted following isolation using the Stemcell NK isolation kit. 20×106clone 2 cells in ~20 mL are irradiated with 100 Gy, and clone 2 cells were counted after irradiation. NK cells were cocultured with irradiated clone 2 cells at a 1:1 ratio. For stimulation experiments, 1.25×105NK cells / mL and 1.25×105clone 2 cells / mL were used. IL-2 was added to cultured cells at a concentration of 50 IU / mL. Media was refreshed every 2-3 days. Cells were harvested for lysate preparation on days 5, 6, 7, 8, and 9 to detect Noxa expression by Western blot. Western blots detecting markers such as IFNγ production, expression of degranulation marker CD107a (for activation), perforin, granzymes, and TNFα (for cytotoxic potency) and cleaved caspase-3 (for apoptosis) were used to evaluate activation status, cytotoxic potency and apoptotic cell death in the NK cells (not shown). Noxa was induced upon activation and as observed in T cells and remains highly expressed through the activation and functional phase until the onset of apoptosis (FIG. 15). Example 12 –Noxa engineering and knockout in NK cells NK cells are cultured in CTS AIM V SFM containing 5% CTS Immune cell SR (ThermoFisher, Waltham, MA), Penicillin / Streptomycin, and IL-2 (100 IU / mL). NK cells are activated by coculture with X-irradiated (100 Gy) feeder cells at indicated feeder:NK ratios (2:1 prior to electroporation, 5:148 hours after electroporation, or 1:1 for all subsequent expansions). The engineering protocol is based on the method described in Skeate et. al., Evolution of the clinical-stage hyperactive TcBuster transposase as a platform for robust non-viral production of adoptive cellular therapies. Molecular Therapy, 2024. 32(6): p. 1817-1834. Feeder cell-activated NK cells are washed once with PBS and resuspended at 3×107cells / mL in electroporation buffer. Protector RNase inhibitor (Sigma-Aldrich, St. Louis, MO) is added to the mixture at a concentration of 0.8 U / μL and incubated for five minutes at room temperature. The cell mixture is added to 1.25 μg Cas9 mRNA + 1 μg Noxa guide RNA on ice. Cas9 mRNA alone is used as a control for all experiments. This mixture is electroporated in a 10-μL tip using the Neon Transfection System (ThermoFisher, Waltham, MA) under the following conditions: 1,850 V, pulse width of 10 ms, two pulses. NK cells are allowed to recover at a density of 1.5×106cells / mL in antibiotic-free medium containing 1 μg / mL DNase I solution (STEMCELL Technologies, Cambridge, MA) for 30 minutes at 37 °C, and then cultured in complete NK cell medium at a density of 6×105cells / mL. Forty-eight hours after electroporation, NK cells are expanded with feeder cells at a 5:1 feeder:NK ratio. Knockout of Noxa gene expression is confirmed by DNA analysis using both Surveyor and TIDE algorithms, but primarily by western blot. Cells are stimulated and analyzed for activations, cytotoxicity and viability as described in Example 12. Knocking out Noxa significantly extends the lifespan of NK cells without affecting their cytotoxic potency as previously observed in CD8+T cells. Also, CAR NK cells engineered for Noxa knockout are able to retain their cytotoxic function for significantly longer than CAR NK cells without Noxa knockout. Example 13- Mesothelin CAR T cells with Noxa Knockout Isolation and Activation of Unstimulated Human CD8+ T Cells Peripheral blood mononuclear cells (PBMCs) from the Memorial Blood Center (St. Paul, MN, USA) were incubated in RBC Lysis Buffer (BioLegend, Cat# 420301) to eliminate red blood cells, and allowed to rest for one hour in CTS OPTMIZER T Cell Expansion SFM (ThermoFisher, Cat# A1048501) supplemented with 2% CTS Immune Cell Serum, 2 mM glutamine, 100 U / mL penicillin-streptomycin, and 300 IU / mL IL-2 (PeproTech). Naïve CD8+ T cells were isolated using the EASYSEP Human Naïve CD8+ T Cell Isolation Kit II (STEMCELL Technologies, Cat# 17968) following the manufacturer’s instructions. Isolated naïve CD8+T cells were cryopreserved using CryStor 10 cryopreservation solution (Biolife Solutions) following the manufacturer’s instructions. Cells were thawed the day before the activation experiment and allowed to rest overnight in complete media, before stimulation in IMMUNOCULT Human CD3 / CD28 T Cell Activator (STEMCELL Technologies, Cat# 10791). CAR T Transduction and CRISPR / Cas9 Genomic Editing At 24 hours post-stimulation, CD8+ T cells were transduced by adding medium containing lentiviral particles encoding the mesothelin (MSLN) for generating CAR Ts. The final multiplicity of infection (MOI) was set at 20, with Synperonic F108 (Sigma-Aldrich, 1 μg / μL) included as a transduction enhancer. Electroporation was carried out using an Amaxa P3 Primary Cell kit and 4D-Nucleofecter (Lonza) the following day. Briefly, 100 pmol sgRNAs were incubated with 1ug of Cas9 Nuclease for 10-15 minutes at room temperature to generate Cas9-gRNA ribonucleoprotein (RNP) complexes and a total of 1×106stimulated T cells resuspended in P3 buffer were added to the preincubated Cas9-gRNA RNP complexes. Cells were nucleofected using program EO-115. sgRNAs targeting PMAIP1 (Noxa) sequence: CACCGGCGGAGAUGCCU (SEQ ID NO:7)– Modified were obtained from Synthego (catalog: PMAIP1+59900181) and Clean Cap 3XNLS Cas9 mRNAs were obtained from TriLink (no. L- 606-100). Following electroporation, the cells were transferred to CTS OPTMIZER medium without antibiotics or cytokines and allowed to rest for 30–45 minutes at 37 °C. The cells were then cultured in complete CTS OPTMIZER medium supplemented with an appropriate number of beads for re-stimulation at a bead-to-T cell ratio of 1:2 and expanded for 10 days in G-Rex plates (Wilson Wolf, Saint Paul, MN, USA), with fresh medium supplemented as needed to maintain optimal cell growth and viability. CAR T Cas9 / NoxaKO cell Stimulation The expression of the inserted CAR gene was analyzed by flow cytometry on day 5 and day10. On day11, the CAR T Cas9 / NoxaKO CD8+ T cells were activated by adding IMMUNOCULT Human CD3 / CD28 T Cell Activator (STEMCELL Technologies) or adding to target cell lines with E:T ratio of 1:1 for further Noxa expression and flow cytometry analysis. Western blot analysis For Western blot analysis, 5×10⁶ cells were harvested 48 hours post-stimulation and lysed using RIPA Lysis and Extraction Buffer (Thermo Scientific, Cat# 89900) supplemented with Protease Inhibitor Cocktail (Thermo Scientific, Cat# 78410) and HALT Phosphatase Inhibitor Cocktail (Thermo Scientific, Cat# 78420). Protein lysates (40 μg) were heat-denatured with 2× Laemmli Buffer (Bio-Rad, Cat# 1610737) for three minutes, resolved by SDS-PAGE, and transferred onto nitrocellulose membranes. The membranes were blocked in TBS-TM buffer (20 mM Tris-HCl, pH 7.6, 137 mM NaCl, 0.1% TWEEN-20, and 5% skim milk) and incubated overnight at 4 °C with primary antibodies diluted in TBS-T buffer. Primary antibodies used were β-Actin (Santa Cruz Biotechnology, Cat# sc-69879, 1:7000) and Noxa (Cell Signaling Technologies, Cat# 14766S, 1:500–1:1000). Following incubation with HRP-conjugated secondary antibodies (Invitrogen) in TBS-TM buffer for one hour at room temperature, chemiluminescence detection was performed using the Pierce ECL Western Blotting Substrate (Thermo Scientific, PI80196). 500,000 CAR T Cas9 / NoxaKO cells as single-cell suspensions were analyzed by flow cytometry at different time points after stimulation. Surface staining and staining for proliferation followed the manufacture’s protocol. For intracellular staining, cells were first fixed with 1X FoxP3 / Transcription Factor Fixation / Permeabilization Buffer (Cell Signaling Technology, no. 43481). Flow cytometry was performed on a FACSYMPHONY A3 flow cytometer and the data were analyzed using Flowjo version 10.10.0. The antibodies and sources used for Flow Cytometry are listed in Table 1. Marker expression and percentage of positive populations are shown in FIG. 17 and FIG. 18. These results (Figures 16-18) clearly indicate that knocking out Noxa, in either primary T cells or in CAR-Ts, does not compromise effector function and toxicity but consistently imparts increased lifespan to the T cells. Table 1. Marker panel for cell viability, activation, differentiation and apoptosis Antibody Fluorophore Company Cat# Zombie NIRTMBioLegend 423106 Table 2. Marker panel for Intracellular cytokines: Antibody Fluorophore Company Cat # 5 In Vitro Serial Killing Assay CAR T Cas9 / NoxaKO cytotoxicity was evaluated using a serial killing assay (FIG. 16D). A1847 cells were used as target cells. Target luciferase reporter cell lines were seeded in triplicate at a density of 5×10³ cells per well in 96-well plates containing 100 μL of complete RPMI medium. To initiate co-culture, 100 μL of CAR T cells were added to each well to achieve an effector-to-target (E:T) ratio of 3:1. Co-cultures were incubated for 24 hours, and luminescence was measured as an indicator of target cell viability. After each 24-hour incubation, the cell mixture was centrifuged, and the supernatant was discarded. Fresh complete medium was added, and new target cells were introduced to the wells. This process was repeated daily over a four-day period to evaluate the serial killing activity of the CAR T cells. Efficacy of killing is depicted in FIG. 16E. These data show that MSLN CAR NoxaKO cells were functionally as effective as MSLN CAR WT controls. Example 14- MSLN, CD19, and B7H3 CAR T cells with NoxaKO Isolation and Activation of Unstimulated Human CD8+ T Cells Isolation of naïve CD8+T cells from human donor PBMCs was carried out as described in Example 13 using the STEMCELL EASYSEP Human Naïve CD8+ T Cell Isolation Kit. Naive CD8+ T cells were resuspended in the media at a density of 1×10⁶ cells / mL and activated by adding IMMUNOCULT Human CD3 / CD28 T Cell Activator (STEMCELL Technologies, Cat# 10791). CAR T Transduction and CRISPR / Cas9 Genomic Editing At 24 hours post-stimulation, CD8+ T cells were transduced by adding medium containing lentiviral particles encoding lentiviral-MSLN (anti-mesothelin) CAR, -CD19 CAR, or -B7H3 CAR constructs. The final multiplicity of infection (MOI) was set at 20, with Synperonic F108 (Sigma-Aldrich, 1 μg / μL) included as a transduction enhancer. NoxaKO and Cas9 control CAR Ts were generated and expansion following electroporation as described in Example 13. CAR T Cas9 / NoxaKO cells Stimulation The expression of the inserted CAR gene was analyzed by flow cytometry on day 5 and day 10. On day 11, the CAR T Cas9 / NoxaKO CD8+ T cells were activated by adding IMMUNOCULT Human CD3 / CD28 T Cell Activator (STEMCELL Technologies) or adding to target cell lines with E:T ratio of 1:1 for further Noxa expression and flow cytometry analysis (FIG. 23A-D). Table 3. Target cells used for killing assays with anti-MESO, anti-CD19, and anti-B7H3 CAR T cells. earlier (FIG. 19A, FIG. 20A, FIG. 21A, FIG. 22A). Flow cytometry was performed as described in Example 14 (FIG.19 to FIG. 26). The data in FIG. 19 to FIG.23 show a comprehensive experiment with naïve CD8T cells isolated from an independent donor and utilized to generate Noxa KO CAR-Ts with three different target antigens, in addition to non-CAR-T control knockouts. Here, all three CAR-T populations were activated by TCR engagement as well as by co-culture with target cells expressing the specific CAR-T antigen. Each CAR-T cell group tested was found to effectively kill target cells (FIG. 19 to FIG. 26), and Noxa knockout did not decrease either activation or cytotoxic efficacy of any CAR T cell group, validating our earlier findings that knocking out Noxa does not compromise T effector function, but serves instead to increase the lifespan of in CAR T cells. The complete disclosure of all patents, patent applications, and publications, and electronically available material (including, for instance, nucleotide sequence submissions in, e.g., GenBank and RefSeq, and amino acid sequence submissions in, e.g., SwissProt, PIR, PRF, PDB, and translations from annotated coding regions in GenBank and RefSeq) cited herein are incorporated by reference in their entirety. In the event that any inconsistency exists between the disclosure of the present application and the disclosure(s) of any document incorporated herein by reference, the disclosure of the present application shall govern. The foregoing detailed description and examples have been given for clarity of understanding only. No unnecessary limitations are to be understood therefrom. The invention is not limited to the exact details shown and described, for variations obvious to one skilled in the art will be included within the invention defined by the claims. Unless otherwise indicated, all numbers expressing quantities of components, molecular weights, and so forth used in the specification and claims are to be understood as being modified in all instances by the term “about.” Accordingly, unless otherwise indicated to the contrary, the numerical parameters set forth in the specification and claims are approximations that may vary depending upon the desired properties sought to be obtained by the present invention. At the very least, and not as an attempt to limit the doctrine of equivalents to the scope of the claims, each numerical parameter should at least be construed in light of the number of reported significant digits and by applying ordinary rounding techniques. Notwithstanding that the numerical ranges and parameters setting forth the broad scope of the invention are approximations, the numerical values set forth in the specific examples are reported as precisely as possible. All numerical values, however, inherently contain a range necessarily resulting from the standard deviation found in their respective testing measurements. All headings are for the convenience of the reader and should not be used to limit the meaning of the text that follows the heading, unless so specified.

Claims

What is claimed is:

1. An engineered cell comprising a decreased amount of a Noxa protein relative to a comparable non-engineered cell.

2. An engineered cell comprising an engineered Noxa protein comprising lower pro- apoptotic activity relative to a non-engineered Noxa protein.

3. The engineered cell of claim 1 or claim 2, wherein the engineered cell comprises one or more modifications to the PMAIP1 gene.

4. The engineered cell of claim 3, wherein the one or more modifications decreases the activity of the Noxa protein.

5. The engineered cell of claim 3, wherein the modification to the PMAIP1 gene encodes mutation of an aspartate at a position equivalent to aspartate 34 in SEQ ID NO:1 to alanine (D34A).

6. The engineered cell of claim 3, wherein the modification to the PMAIP1 gene encodes mutation of a serine at a position equivalent to serine 13 in SEQ ID NO:1 to glutamate (S13E) or aspartate (S13D).

7. The engineered cell of any preceding claim, wherein the cell is an immune cell.

8. The engineered cell of claim 7, wherein the engineered cell comprises an engineered T cell.

9. The engineered cell of claim 8, wherein the T cell comprises a CD8+ T cell.

10. The engineered cell of claim 8 or claim 9, wherein the T cell comprises a human T cell.

11. The engineered cell of any preceding claim, wherein the engineered cell comprises a chimeric antigen receptor T cell (CAR T cell).

12. The engineered cell of claim 11, wherein the CAR comprises an anti-B7H3, anti-CD19, or anti-mesothelin CAR T cell.

13. The engineered cell of any preceding claim, wherein the engineered cell comprises a tumor infiltrating lymphocyte (TIL).

14. The engineered cell of claim 7, wherein the engineered cell comprises an engineered NK cell.

15. The engineered cell of claim 14, wherein the NK cell comprises a CAR NK cell.

16. The engineered cell of claim 15, wherein the CAR NK cell comprises an anti-B7H3, anti- CD19, or anti-mesothelin CAR T cell.

17. The engineered cell of any preceding claim wherein the engineered cell is derived from a primary cell.

18. The engineered cell of any preceding claim, wherein the engineered cell exhibits a longer lifespan than a comparable non-engineered cell.

19. The engineered cell of claim 18, wherein the engineered cell exhibits at least a 30% longer lifespan than a comparable non-engineered cell.

20. A method of regulating Noxa expression in a cell comprising administering a gene editing composition to the cell.

21. A method of regulating Noxa phosphorylation in a cell comprising administering a gene editing composition to the cell.

22. The method of claim 20 or claim 21, wherein the gene editing composition introduces one or more mutations to the PMAIP1 gene.

23. The method of claim 22, wherein the one or more mutations to the PMAIP1 gene encodes mutation an aspartate at a position equivalent to aspartate 34 in SEQ ID NO:1 to alanine (D34A).

24. The method of claim 22, wherein the one or more mutations to the PMAIP1 gene encodes mutation of the serine at a position equivalent to serine 13 in SEQ ID NO:1 to glutamate (S13E) or aspartate (S13D).

25. A method of extending the lifespan of a cell comprising decreasing the activity of the Noxa protein.

26. The method of claim 25, wherein the activity of the Noxa protein comprises pro- apoptotic activity.

27. The method of any preceding claim, wherein the cell comprises a human cell.

28. The method of any preceding claim, wherein the cell comprises a CD8+ T cell.

29. The method of any preceding claim, wherein the cell comprises a chimeric antigen receptor T cell (CAR T cell).

30. The method of any preceding claim, wherein the cell comprises a tumor infiltrating lymphocyte (TIL).

31. The method of any preceding claim, wherein the cell comprises an NK cell.

32. The method of any preceding claim, wherein the cell comprises a CAR NK cell.

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