Poster · 2022 · ESMO

Tumor Infiltrating Lymphocytes Expressing Membrane-bound IL-2 and IL-12 Exhibit Enhanced Proliferation, Function, and Persistence Without Requiring Exogenous IL-2 Support

Rebecca J. Silver, Murillo Silva, Michael F. Maloney, Emrah Ilker Ozay, Ashley A. Brate, Nader El-Sayes, Benjamin Cossette, Drishti Parwanda, Cara Bartlett, Armon Sharei, Scott M. Loughhead, Jonathan Gilbert, and Devin T. Bridgen

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Available IL-7Rα [HIL-7R-M21] % IL-7Rα availability (norm to Control TILs) Control mbIL-7 Cell Expansion Cell sorting Tumor dissociation Background: Tumor Infiltrating Lymphocyte (TIL) therapies have shown significant solid tumor activity in patients, but many current TIL regimens require lymphodepletion and high-dose IL-2 after cell infusion. Removing these requirements that cause systemic toxicity while maintaining TIL product functionality by ex vivo engineering of the TIL product with transient expression of mRNA could dramatically improve the patient experience, expand the eligible patient population, and allow repeat dosing. Overview Tumor Infiltrating Lymphocytes Expressing Membrane-bound IL-2 and IL-12 Exhibit Enhanced Proliferation, Function, and Persistence Without Requiring Exogenous IL-2 Support Rebecca J. Silver, Murillo Silva, Michael F. Maloney, Emrah Ilker Ozay, Ashley A. Brate, Nader El-Sayes, Benjamin Cossette, Drishti Parwanda, Cara Bartlett, Armon Sharei, Scott M. Loughhead, Jonathan Gilbert, and Devin T. Bridgen SQZ Biotechnologies, Watertown MA, USA mbIL-2 binds IL-2R and drives STAT5 phosphorylation and proliferation without rhIL-2 support R Engineered mbIL-2/12 Tumor Infiltrating Lymphocytes: Summary Kinetics of mRNA expression, viability, and proliferation in squeezed TILs Figure 4. Cryopreserved post-REP TILs from an ovarian tumor were thawed and cultured for 3 days in rhIL-2 and were either not squeeze processed and cultured with (Control+rhIL-2) or without (Control) rhIL-2 support or squeezed with mbIL-2 mRNA (mbIL-2) and/or mbIL-12 mRNA (mbIL-2+mbIL-12) and cultured with donor-matched primary autologous tumor cells. (A) Experimental layout, image partially made with BioRender. (B-E) Representative images from at least N=3 independent experiments after 24 hours of co-culture between 1E4 tumor cells (red) and 5E4 post-REP TILs (undyed) per well. Images were taken on an IncuCyte S3 fluorescent imager with a Caspase-3/7 dye (green) to mark dying/dead cells. (F) Caspase-3/7 Dye brightness when sub-gating for tumor cells (red; CellTrace Far Red; CTFR) via flow cytometry at 24 hr. (G) IFN-γ content from the supernatant of wells imaged in (B-E). Figure 3. Cryopreserved post-REP TILs from a (A) melanoma tumor or (B) ovarian tumor were thawed and cultured for 3 days in rhIL-2 and squeezed with either media only (Control; solid line) and cultured with or without exogenous cytokine support (Control+rhIL-2; dashed line) or squeezed with mbIL-2 mRNA and/or mbIL-12 mRNA and cultured for 6 days. (A) Percent expression of a representative donor from at least N=3 independent experiments for CD62L expression. (B) Representative dot plots of a single donor from at least N=3 independent experiments gated on CD8-positive cells and sub-gated on CD45RO (x-axis) by CD62L (yaxis). (C) Fresh post-REP TILs from lung and melanoma tumors were squeeze processed as above and immediately cryopreserved, then thawed and cultured for 8 days. Percent expression of a representative donor for CD127 expression sub-gated on CD8/CD62L/CD45RO triple-positive cells. Figure 5. Cryopreserved post-REP TILs from a single ovarian tumor were thawed and cultured for 3 days in rhIL-2 and squeezed with either media only (Control) or mbIL-2 mRNA (mbIL-2) and/or mbIL-12 mRNA (mbIL-2+mbIL-12) and 5M TILs per mouse injected i.v. into immunodeficient NOD SCID gamma (NSG) mice. A peripheral bleed was performed at day 1 post-transfer and a terminal bleed and spleen harvest were performed at day 5 post-transfer. Cells were isolated and quantified via flow cytometry. (A) Experimental design. (B) Percent human CD45-positive cells when sub-gated on total CD45 cells in the blood. (C) Absolute cell count of human CD45-positive cells in the spleen on day 5. (D) Percent positive for CD62L and/or CD45RO expression when sub-gated on human CD45-positive cells in the blood on day 5. (C-D) Statistical significance was determined via (C) a One-way ANOVA with Dunnett’s post-test, or (D) a Two-way ANOVA with Dunnett’s post-test. Ns = not significant, **** = p<0.0001. Note: This is a representative donor; variability is seen across donors. IL-2 Available to TILs in Patient Proposed mbIL-2-TILs: Cells supported with consistent and localized IL-2 for >1 day Current Systemic IL-2 Support: Proleukin dosing every 8 to 12 hours for up to 6 doses unless toxicity limit reached. Systemic exposure with elimination half life of <1.5hrs. 8-12 hrs Conceptual Kinetics: mbIL-2 vs Systemic IL-2 mbIL-2 TILs Current TILs Figure 1. Cryopreserved post-REP TILs from N=2-3 donors from lung and melanoma tumors were thawed and cultured for 3 days then squeezed with either media only (Control), mbIL-2 mRNA, and/or mbIL-12 mRNA and cultured for 8 days. (A-B) Percent expression of biological replicates in N=2-3 independent experiments for (A) mbIL-2 and (B) mbIL-12 over 8 days or (C-D) representative histograms from a single donor 4 hours post-squeeze. (E-F) Viability and cell counts of TILs from a single donor that were processed as above and cultured for 5 days. mbIL-7 Expression % mbIL-7 (of CD3+) Control mbIL-7 Figure 2. Cryopreserved post-REP TILs from melanoma and ovarian tumors were thawed and cultured for 3 days in rhIL-2 and squeezed with either media only (Control), mbIL-2 mRNA, and/or mbIL-12 mRNA and cultured for 3 days. (A) Experimental layout. (B-C) Percent expression of N=3 donors for (B) Total CD25 [clone M-A251] and (C) Available CD25 [clone BC96] at 4 hours postsqueeze and sub-gated on mbIL-2 positive TILs (mbIL-2 positive). (D) Available CD25 [clone BC96] in a representative donor over time. (E) Phosphorylated STAT5 from an individual donor processed as above and cultured for 3 days. Figure 6. Cryopreserved post-REP TILs from N=1-2 donors from melanoma tumors were thawed and cultured for 3 days in rhIL-2 and squeezed with either media only (Control), mbIL-7 mRNA (mbIL-7), or Bcl-2 mRNA (Bcl-2) and cultured for 1-5 days. (A) Percent mbIL-7 expression and (B) Available IL-7R-alpha (CD127) in a representative donor over 3 days culture post-squeeze processing. (C) Geometric mean fluorescence intensity (gMFI) of Bcl-2 in TILs a single donor measured 24 hours post-squeeze processing. (D) Viability of TILs from a single donor measured 5 days post-squeeze processing and cultured with or without rhIL-7. Future Targets: Pre-conditioning cytokine IL-7 and pro-survival protein Bcl-2 mbIL-12 TILs upregulate CD62L and CD127, markers of central memory T cells (TCM) 0 1 2 3 4 5 1×10 5 2×10 5 3×10 5 4×10 5 5×10 5 in vitro Proliferation Cell Counts

A) B)

STAT5 Phosphorylation % pSTAT5 Positive Control+rhIL-2+rhIL-12 0 1 2 3 4 5 in vitro Survival Viability (%) Control Control+rhIL-2 mbIL-2 mbIL-2+mbIL-12 A) 0 0.01 0.1 1 10 Day 5 in vitro Survival rhIL-7 (ng/mL) Viability (%squeezed cells) Control mbIL-7 Bcl-2 Control Bcl-2 50000 100000 150000 Bcl-2 Expression gMFI Bcl-2 (of CD3+)

Through microfluidic Cell Squeeze® delivery of mRNAs, we engineered

primary human TILs with transient expression of membrane-bound cytokines IL-2 and IL-12

mbIL-2 signals through endogenous IL-2R to phosphorylate STAT5 and

drive survival and proliferation in the absence of exogenous IL-2 support

mbIL-12 induces upregulation of CD62L, a marker of central memory T

cells which are a T cell subtype known to confer superior antitumor immunity

mbIL-2/12 TILs upregulate CD62L in vivo for at least 5 days in the

blood and spleen of a humanized NSG mouse

mbIL-2/12 TILs release more IFN- γ than Control TILs + rhIL-2

(positive control; current clinical standard) when co-cultured with patientmatched tumor cells Overall, enhanced SQZ® TILs can potentially alleviate current requirements for toxic high-dose IL-2 support and lymphodepleting preconditioning regimens. Methods: mRNAs encoding for membrane-bound (mb) IL-2 or IL-12 were delivered directly to the cytosol of TILs expanded from either melanoma, lung, or ovarian solid tumors using the microfluidic Cell Squeeze® technology. Vision: SQZ® TILs eliminate need for toxic preconditioning and systemic IL-2 support

A) B) C)

mbIL-2/12 TILs release increased IFN-γ during autologous tumor co-culture IFN-γ ELISA IFN-γ (pg/mL)

A) B) C)

G) mbIL-2/12 TILs exhibit memory-like phenotype change in mouse adoptive transfer TIL Phenotype (Blood, Day 5)

Cd62l+cd45ro+

TCM-like

Cd62l-cd45ro+

TEM-like

Cd62l-cd45ro-

TEFF-like % total (gated on hCD45+ cells) ns Proportion Human Cells (Peripheral Blood) mbIL-2+12 20000 40000 60000 80000 Absolute cell count Human Cell Count (Spleen, Day 5)

B) C) D)

Engineering TILs with SQZ® Technology Cells and target mRNA together in suspension Cells are squeezed through chip at high speeds Cell membranes are temporarily disrupted Target mRNA enters the cytosol of the cells4 Membranes reseal 5 mbIL-2 Expression of target mRNA 0 hour 4 hours 24 hours 48 hours 8 days mbIL-2 Expression Time post squeeze mbIL-2+ 0 hour 4 hours 24 hours 48 hours 8 days mbIL-12 Expression Time post squeeze mbIL-12+

E) F)

Til-1 Til-2 Til-3

Total CD25 [M-A251] CD25 positive cells Control mbIL-2 positve

Til-1 Til-2 Til-3

Available CD25 [BC96] CD25 positive cells Control mbIL-2 positive mbIL-12 mRNA 24 hr Media only 24 hr Flow Cytometry

Cd25

pSTAT5 Traditional TIL patient experience SQZ® TIL patient experience

C) D)

TILs: Tumor: IncuCyte S3 Tumor + Control TILs Tumor + mbIL-2/12 TILs (no exogenous cytokines)

D) E)

mbIL-2/12 Tumor Cell Death gMFI Caspase-3/7 Green Dye (gated on all->CTFR+) F)Tumor + Control TILs + rhIL-2 Tumor + mbIL-2 TILs (no exogenous cytokines) mbIL-2+mbIL-12 mRNA CD127+ TILs on Day 8

Cd127+

(% of CD62L+ CD45RO+ CD8+) 0 1 2 3 4 5 6 CD62L Expression % CD62L Positive (gated on CD3+) Available CD25 [BC96] % CD25 availability (norm. to Control TILs) mbIL-2Control +

Cd45ro

Cd62l

t=4 hr t=24 hr t=3 days t=6 days A) mRNA +rhIL-2 5 days Flow Cytometry Persistence

Cd62l

NSG mouse 0 1 2 3 4 5 6 Proportion of human CD45+ cells (gated on total CD45+ cells) Control mbIL-2 mbIL-2+12 Total CD25 [M-A251] Available CD25 [BC96] Available CD25 [BC96] STAT5 Phosphorylation

B) C) D) F)

mbIL-7 Expression Available IL-7Rα [HIL-7R-M21] Bcl-2 Expression Day 5 in vitro SurvivalA) B) C) D) Tumor Cell Death IFN-γ ELISA

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Some publications reference SQZ Biotechnologies, a company Dr. Armon Sharei and colleagues previously founded. Portal is the next generation company leveraging Dr. Sharei's platform technology.

# Tumor Infiltrating Lymphocytes Expressing Membrane-bound IL-2 and IL-12 Exhibit Enhanced Proliferation, Function, and Persistence Without Requiring Exogenous IL-2 Support This page is a text version of a Portal Biotechnologies publication. The bibliographic record is labeled in the text so it can be read without the surrounding website. ## Bibliographic record - Title: Tumor Infiltrating Lymphocytes Expressing Membrane-bound IL-2 and IL-12 Exhibit Enhanced Proliferation, Function, and Persistence Without Requiring Exogenous IL-2 Support - Document type: Poster - Year: 2022 - Outlet: ESMO - Page URL: https://portal.bio/publications/2022-tumor-infiltrating-lymphocytes-expressing-membrane-bound-il-2-and-il-12-exhibit-enhan--ajkGjREAACsAc96_ - Markdown URL: https://portal.bio/publications/2022-tumor-infiltrating-lymphocytes-expressing-membrane-bound-il-2-and-il-12-exhibit-enhan--ajkGjREAACsAc96_.md - Original file: https://portal-bio.cdn.prismic.io/portal-bio/65ccda6b9be9a5b998b5d10a_2022-ESMO-TIL_vf.pdf ## Summary Rebecca J. Silver, Murillo Silva, Michael F. Maloney, Emrah Ilker Ozay, Ashley A. Brate, Nader El-Sayes, Benjamin Cossette, Drishti Parwanda, Cara Bartlett, Armon Sharei, Scott M. Loughhead, Jonathan Gilbert, and Devin T. Bridgen ## Publication text Available IL-7Rα [HIL-7R-M21] % IL-7Rα availability (norm to Control TILs) Control mbIL-7 Cell Expansion Cell sorting Tumor dissociation Background: Tumor Infiltrating Lymphocyte (TIL) therapies have shown significant solid tumor activity in patients, but many current TIL regimens require lymphodepletion and high-dose IL-2 after cell infusion. Removing these requirements that cause systemic toxicity while maintaining TIL product functionality by ex vivo engineering of the TIL product with transient expression of mRNA could dramatically improve the patient experience, expand the eligible patient population, and allow repeat dosing. Overview Tumor Infiltrating Lymphocytes Expressing Membrane-bound IL-2 and IL-12 Exhibit Enhanced Proliferation, Function, and Persistence Without Requiring Exogenous IL-2 Support Rebecca J. Silver, Murillo Silva, Michael F. Maloney, Emrah Ilker Ozay, Ashley A. Brate, Nader El-Sayes, Benjamin Cossette, Drishti Parwanda, Cara Bartlett, Armon Sharei, Scott M. Loughhead, Jonathan Gilbert, and Devin T. Bridgen SQZ Biotechnologies, Watertown MA, USA mbIL-2 binds IL-2R and drives STAT5 phosphorylation and proliferation without rhIL-2 support R Engineered mbIL-2/12 Tumor Infiltrating Lymphocytes: Summary Kinetics of mRNA expression, viability, and proliferation in squeezed TILs Figure 4. Cryopreserved post-REP TILs from an ovarian tumor were thawed and cultured for 3 days in rhIL-2 and were either not squeeze processed and cultured with (Control+rhIL-2) or without (Control) rhIL-2 support or squeezed with mbIL-2 mRNA (mbIL-2) and/or mbIL-12 mRNA (mbIL-2+mbIL-12) and cultured with donor-matched primary autologous tumor cells. (A) Experimental layout, image partially made with BioRender. (B-E) Representative images from at least N=3 independent experiments after 24 hours of co-culture between 1E4 tumor cells (red) and 5E4 post-REP TILs (undyed) per well. Images were taken on an IncuCyte S3 fluorescent imager with a Caspase-3/7 dye (green) to mark dying/dead cells. (F) Caspase-3/7 Dye brightness when sub-gating for tumor cells (red; CellTrace Far Red; CTFR) via flow cytometry at 24 hr. (G) IFN-γ content from the supernatant of wells imaged in (B-E). Figure 3. Cryopreserved post-REP TILs from a (A) melanoma tumor or (B) ovarian tumor were thawed and cultured for 3 days in rhIL-2 and squeezed with either media only (Control; solid line) and cultured with or without exogenous cytokine support (Control+rhIL-2; dashed line) or squeezed with mbIL-2 mRNA and/or mbIL-12 mRNA and cultured for 6 days. (A) Percent expression of a representative donor from at least N=3 independent experiments for CD62L expression. (B) Representative dot plots of a single donor from at least N=3 independent experiments gated on CD8-positive cells and sub-gated on CD45RO (x-axis) by CD62L (yaxis). (C) Fresh post-REP TILs from lung and melanoma tumors were squeeze processed as above and immediately cryopreserved, then thawed and cultured for 8 days. Percent expression of a representative donor for CD127 expression sub-gated on CD8/CD62L/CD45RO triple-positive cells. Figure 5. Cryopreserved post-REP TILs from a single ovarian tumor were thawed and cultured for 3 days in rhIL-2 and squeezed with either media only (Control) or mbIL-2 mRNA (mbIL-2) and/or mbIL-12 mRNA (mbIL-2+mbIL-12) and 5M TILs per mouse injected i.v. into immunodeficient NOD SCID gamma (NSG) mice. A peripheral bleed was performed at day 1 post-transfer and a terminal bleed and spleen harvest were performed at day 5 post-transfer. Cells were isolated and quantified via flow cytometry. (A) Experimental design. (B) Percent human CD45-positive cells when sub-gated on total CD45 cells in the blood. (C) Absolute cell count of human CD45-positive cells in the spleen on day 5. (D) Percent positive for CD62L and/or CD45RO expression when sub-gated on human CD45-positive cells in the blood on day 5. (C-D) Statistical significance was determined via (C) a One-way ANOVA with Dunnett’s post-test, or (D) a Two-way ANOVA with Dunnett’s post-test. Ns = not significant, **** = p<0.0001. Note: This is a representative donor; variability is seen across donors. IL-2 Available to TILs in Patient Proposed mbIL-2-TILs: Cells supported with consistent and localized IL-2 for >1 day Current Systemic IL-2 Support: Proleukin dosing every 8 to 12 hours for up to 6 doses unless toxicity limit reached. Systemic exposure with elimination half life of <1.5hrs. 8-12 hrs Conceptual Kinetics: mbIL-2 vs Systemic IL-2 mbIL-2 TILs Current TILs Figure 1. Cryopreserved post-REP TILs from N=2-3 donors from lung and melanoma tumors were thawed and cultured for 3 days then squeezed with either media only (Control), mbIL-2 mRNA, and/or mbIL-12 mRNA and cultured for 8 days. (A-B) Percent expression of biological replicates in N=2-3 independent experiments for (A) mbIL-2 and (B) mbIL-12 over 8 days or (C-D) representative histograms from a single donor 4 hours post-squeeze. (E-F) Viability and cell counts of TILs from a single donor that were processed as above and cultured for 5 days. mbIL-7 Expression % mbIL-7 (of CD3+) Control mbIL-7 Figure 2. Cryopreserved post-REP TILs from melanoma and ovarian tumors were thawed and cultured for 3 days in rhIL-2 and squeezed with either media only (Control), mbIL-2 mRNA, and/or mbIL-12 mRNA and cultured for 3 days. (A) Experimental layout. (B-C) Percent expression of N=3 donors for (B) Total CD25 [clone M-A251] and (C) Available CD25 [clone BC96] at 4 hours postsqueeze and sub-gated on mbIL-2 positive TILs (mbIL-2 positive). (D) Available CD25 [clone BC96] in a representative donor over time. (E) Phosphorylated STAT5 from an individual donor processed as above and cultured for 3 days. Figure 6. Cryopreserved post-REP TILs from N=1-2 donors from melanoma tumors were thawed and cultured for 3 days in rhIL-2 and squeezed with either media only (Control), mbIL-7 mRNA (mbIL-7), or Bcl-2 mRNA (Bcl-2) and cultured for 1-5 days. (A) Percent mbIL-7 expression and (B) Available IL-7R-alpha (CD127) in a representative donor over 3 days culture post-squeeze processing. (C) Geometric mean fluorescence intensity (gMFI) of Bcl-2 in TILs a single donor measured 24 hours post-squeeze processing. (D) Viability of TILs from a single donor measured 5 days post-squeeze processing and cultured with or without rhIL-7. Future Targets: Pre-conditioning cytokine IL-7 and pro-survival protein Bcl-2 mbIL-12 TILs upregulate CD62L and CD127, markers of central memory T cells (TCM) 0 1 2 3 4 5 1×10 5 2×10 5 3×10 5 4×10 5 5×10 5 in vitro Proliferation Cell Counts ### A) B) STAT5 Phosphorylation % pSTAT5 Positive Control+rhIL-2+rhIL-12 0 1 2 3 4 5 in vitro Survival Viability (%) Control Control+rhIL-2 mbIL-2 mbIL-2+mbIL-12 A) 0 0.01 0.1 1 10 Day 5 in vitro Survival rhIL-7 (ng/mL) Viability (%squeezed cells) Control mbIL-7 Bcl-2 Control Bcl-2 50000 100000 150000 Bcl-2 Expression gMFI Bcl-2 (of CD3+) - Through microfluidic Cell Squeeze® delivery of mRNAs, we engineered primary human TILs with transient expression of membrane-bound cytokines IL-2 and IL-12 - mbIL-2 signals through endogenous IL-2R to phosphorylate STAT5 and drive survival and proliferation in the absence of exogenous IL-2 support - mbIL-12 induces upregulation of CD62L, a marker of central memory T cells which are a T cell subtype known to confer superior antitumor immunity - mbIL-2/12 TILs upregulate CD62L in vivo for at least 5 days in the blood and spleen of a humanized NSG mouse - mbIL-2/12 TILs release more IFN- γ than Control TILs + rhIL-2 (positive control; current clinical standard) when co-cultured with patientmatched tumor cells Overall, enhanced SQZ® TILs can potentially alleviate current requirements for toxic high-dose IL-2 support and lymphodepleting preconditioning regimens. Methods: mRNAs encoding for membrane-bound (mb) IL-2 or IL-12 were delivered directly to the cytosol of TILs expanded from either melanoma, lung, or ovarian solid tumors using the microfluidic Cell Squeeze® technology. Vision: SQZ® TILs eliminate need for toxic preconditioning and systemic IL-2 support ### A) B) C) mbIL-2/12 TILs release increased IFN-γ during autologous tumor co-culture IFN-γ ELISA IFN-γ (pg/mL) ### A) B) C) G) mbIL-2/12 TILs exhibit memory-like phenotype change in mouse adoptive transfer TIL Phenotype (Blood, Day 5) ### Cd62l+cd45ro+ TCM-like ### Cd62l-cd45ro+ TEM-like ### Cd62l-cd45ro- TEFF-like % total (gated on hCD45+ cells) ns Proportion Human Cells (Peripheral Blood) mbIL-2+12 20000 40000 60000 80000 Absolute cell count Human Cell Count (Spleen, Day 5) ### B) C) D) Engineering TILs with SQZ® Technology Cells and target mRNA together in suspension Cells are squeezed through chip at high speeds Cell membranes are temporarily disrupted Target mRNA enters the cytosol of the cells4 Membranes reseal 5 mbIL-2 Expression of target mRNA 0 hour 4 hours 24 hours 48 hours 8 days mbIL-2 Expression Time post squeeze mbIL-2+ 0 hour 4 hours 24 hours 48 hours 8 days mbIL-12 Expression Time post squeeze mbIL-12+ ### E) F) ### Til-1 Til-2 Til-3 Total CD25 [M-A251] CD25 positive cells Control mbIL-2 positve ### Til-1 Til-2 Til-3 Available CD25 [BC96] CD25 positive cells Control mbIL-2 positive mbIL-12 mRNA 24 hr Media only 24 hr Flow Cytometry ### Cd25 pSTAT5 Traditional TIL patient experience SQZ® TIL patient experience ### C) D) TILs: Tumor: IncuCyte S3 Tumor + Control TILs Tumor + mbIL-2/12 TILs (no exogenous cytokines) ### D) E) mbIL-2/12 Tumor Cell Death gMFI Caspase-3/7 Green Dye (gated on all->CTFR+) F)Tumor + Control TILs + rhIL-2 Tumor + mbIL-2 TILs (no exogenous cytokines) mbIL-2+mbIL-12 mRNA CD127+ TILs on Day 8 ### Cd127+ (% of CD62L+ CD45RO+ CD8+) 0 1 2 3 4 5 6 CD62L Expression % CD62L Positive (gated on CD3+) Available CD25 [BC96] % CD25 availability (norm. to Control TILs) mbIL-2Control + ### Cd45ro ### Cd62l t=4 hr t=24 hr t=3 days t=6 days A) mRNA +rhIL-2 5 days Flow Cytometry Persistence ### Cd62l NSG mouse 0 1 2 3 4 5 6 Proportion of human CD45+ cells (gated on total CD45+ cells) Control mbIL-2 mbIL-2+12 Total CD25 [M-A251] Available CD25 [BC96] Available CD25 [BC96] STAT5 Phosphorylation ### B) C) D) F) mbIL-7 Expression Available IL-7Rα [HIL-7R-M21] Bcl-2 Expression Day 5 in vitro SurvivalA) B) C) D) Tumor Cell Death IFN-γ ELISA ## Provenance Some publications reference SQZ Biotechnologies, a company Dr. Armon Sharei and colleagues previously founded. Portal is the next generation company leveraging Dr. Sharei's platform technology.