# PBMC-based cancer vaccines generated with microfluidics squeezing demonstrate synergistic and durable tumor reduction in combination with PD-1 checkpoint and FAP targeted IL-2 variants 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: PBMC-based cancer vaccines generated with microfluidics squeezing demonstrate synergistic and durable tumor reduction in combination with PD-1 checkpoint and FAP targeted IL-2 variants - Document type: Poster - Year: 2020 - Outlet: SITC - Cells: T cells, PBMCs, Monocytes, B cells, Dendritic Cells - Cargos: Polymers, Protein, Peptide - Page URL: https://portal.bio/publications/2020-pbmc-based-cancer-vaccines-generated-with-microfluidics-squeezing-demonstrate-synergi--ajkHChEAACsAc-Bg - Markdown URL: https://portal.bio/publications/2020-pbmc-based-cancer-vaccines-generated-with-microfluidics-squeezing-demonstrate-synergi--ajkHChEAACsAc-Bg.md - Original file: https://portal-bio.cdn.prismic.io/portal-bio/65ccf1639be9a5b998b5d264_SITC-2020-SQZ-Roche-Combo-Poster-S.-Loughhead-FINAL.pdf ## Summary Matthew G. Booty, Adam Stockmann, Olivia Pryor, Melissa Myint, Christine Trumpfheller, Valeria Nicolini, Christian Klein, Laura Codarri Deak, Pablo Umana, Armon Sharei, Howard Bernstein, Katherine J. Seidl, and Scott M. Loughhead ## Publication text PBMC-based cancer vaccines generated with microfluidics squeezing demonstrate synergistic and durable tumor reduction in combination with PD-1 checkpoint and FAP targeted IL-2 variants Matthew G. Booty1, Adam Stockmann1, Olivia Pryor1, Melissa Myint1, Christine Trumpfheller2, Valeria Nicolini2, Christian Klein2, Laura Codarri Deak2, Pablo Umana2, Armon Sharei1, Howard Bernstein1, Katherine J. Seidl1, and Scott M. Loughhead1 1SQZ Biotechnologies, Watertown, MA, USA 2Roche Innovation Center Zurich, Roche Pharma Research and Early Development (pRED), Wagistrasse 10, 8952 Schlieren, Switzerland ### Conclusions Monotherapy with SQZ-PBMC-based cancer vaccines can drive anti-tumor responses in murine systems. These responses are enhanced by combined administration of targeted immunocytokines. SQZ-PBMC-HPV is currently under clinical evaluation for HPV16+ tumors as both a monotherapy and in combination with other therapies. Preclinical data support the combination of SQZ-PBMC with FAP-IL2v or PD1-IL2v targeted immunocytokine as promising cancer immunotherapies. ### Background Microfluidic squeezing of peripheral blood mononuclear cells (PBMCs) enables direct delivery of antigen into PBMCs and bypasses the need for cross-presentation and directs the delivered antigen into the MHC-I pathway. This allows for efficient MHC-I presentation of a variety of antigens to CD8+ T cells by PBMCs (MF Maloney et. al, SITC 2020, #169). We have also shown that squeezed mouse cells elicit anti-tumor immunity in an HPV tumor model (EI Ozay et. al., SITC 2020, #170) and is the basis for an ongoing Phase 1 trial (C Eng et. al., SITC 2020, #418). Here, we investigated the combined efficacy of our PBMC-based cancer vaccine with targeted interleukin 2 variants (IL2v); anti-Programmed Cell Death Protein 1 (muPD1- IL2v) and anti-Fibroblast Activation Protein (muFAP-IL2v) using the TC-1 mouse tumor model. Day 0 Day 14 Day 24Day 21 TC-1 Implant ### M-sqz-pbmc-hpv Immunization Treatment with PD1-IL2v TIL Analysis0 20 40 60 80 Time (days) Tumor Volume (mm3) Untreated 1e6 M-SQZ-PBMC-HPV alone 1e6 M-SQZ-PBMC-HPV + FAP-IL-2v FAP-IL2v alone 1e6 M-SQZ-PBMC-HPV + PD1-IL2v PD1-IL2v alone Imm. Treatment with Targeted IL-2v Group % Untreated 0 ### Sqz 17 SQZ+FAP-IL2v 75 FAP-IL2v 0 SQZ+PD1-IL2v 100 PD1-IL2v 0 % Tumor-Free Mice on Day 8480 100 120 140 SQZ alone (n=2)80 100 120 140 SQZ + FAP-IL2v (n=8)80 100 120 140 SQZ + PD1-IL2v (n=10) SQZ Technology and APC Platform Engineering Cells (All Types) to Function as Antigen Presenting Cells: Cytosolic Antigen Delivery: Traditionally, cross-presentation is needed for efficient presentation on MHC-I. SQZ delivers materials directly to cytosol, bypassing need for cross-presentation. ### Pd1 Fap PD-1/Tumor Targeted IL-2v Cytokines - Binds to fibroblast activation protein (FAP) in tumor stroma to deliver IL-2 (IL-2v) - Lacks preferential induction of regulatory T cells due to abolished IL-2Rα (CD25) binding - Binds to PD1+ T cells to deliver IL-2 (IL-2v) to antigen-experienced TILs - Lacks preferential induction of regulatory T cells due to abolished IL-2Rα receptor (CD25) binding Combination of M-SQZ-PBMC-HPV with FAP-IL2v or PD1-IL2v Elicits Protective Immunological Memory0 10 20 30 40 50 60 E7-Specific (% among CD45+) Untreated SQZ E7-Specific SQZ Imm. E7 Tetramer ### Pd-1 CD8+ TILS from TC-1 Tumors Rationale for Combination Tumor-infiltrating E7-specific CD8+ T cells express high levels of PD-1 TC-1 tumor stroma expresses FAP - In the preclinical model using the HPV16 E6/E7 expressing tumor, TC-1, therapeutic SQZ immunization elicits influx of E7-specific CD8+ T cells that initially control tumor growth - Over time, the fraction of E7-specific CD8+ T cells in the tumor falls and correlates with timing of tumor relapse - Therefore, this is a useful model with which to test combinations like PD1-IL2v and FAP-IL2v, which may augment waning activity of infiltrating CD8+ T cells and help them maintain tumor control FAP Staining of TC-1 Tumor Weekly administration of PD1-IL2v or FAP-IL2v in combination with M-SQZ- PBMC-HPV results in enhanced anti-tumor activity. 14 days after TC-1 implantation, mice were immunized with M-SQZ-PBMC-HPV. Starting on day 21, mice were dosed intravenously with PD1-IL2v or FAP-IL2v on days 21, 28, and 35. Rechallenge of tumor-free mice demonstrates immunological memory with durable ant-tumor activity. Remaining tumor-free mice from the study above were rechallenged by s.c. implantation of TC-1 on the opposite flank 84 days after the initial challenge with TC-1. Naïve mice were also challenged by s.c. implantation of TC-1 as a control. M-SQZ-PBMC-HPV Vaccination has Synergistic Anti-Tumor Activity when Combined with FAP-IL2v or PD1-IL2vUntr 50000 100000 150000 CD45+ Cells ### # Cd45+ (per mg tumor) 5.4x 2.9x 1.1xUntr E7-Specific CD8+ T Cells # E7 Tetramer + (per mg tumor) 1770x 480xUntr CD8+ T Cells ### # Cd8+ (per mg tumor) 75.1x 21.1x 1.5xUntr 1000 Tumor Mass Tumor Mass (mg)Untr Tregs ### # Cd4+ Foxp3+ Cd25+ (per mg tumor) 2.1x 0.7x 1.3xUntr CD8/Treg Ratio Ratio (CD8+/ FOXP3+CD25+) 0.8x 34x 61x M-SQZ-PBMC-HPV Immunization Drives 480-Fold Increase in Tumor Infiltration of E7-Specific CD8+ T Cells that is Augmented by PD1-IL2v Administration PD1-IL2v augments tumor infiltration of E7-specific CD8+ T cells elicited by M-SQZ- PBMC-HPV. 14 days after TC-1 implantation, mice were immunized with M-SQZ-PBMC- HPV. On day 21, indicated mice were treated intravenously with PD1-IL2v. On day 24, mice were sacrificed and tumors harvested for analysis of immune infiltrates by flowcytometry. Recovered tumor masses were weighed prior to cell dissociation. Numbers of indicated cell populations were measured by flowcytometry and normalized per mg of tumor harvested. Mean fold increase for each group is shown relative to the untreated group. PD1-IL2v Preferentially Expands CD8+ T Cells Over Tregs Treg numbers were assessed from the study outlined above. Treg numbers increased moderately over untreated (~2-fold) but were not significantly affected with M-SQZ-PBMC- HPV immunization or in combination with PD1-IL2v. Change in CD8/Treg ratio was 34-fold above untreated for M-SQZ-PBMC-HPV vaccination and 61-fold when combined with PD1- IL2v, highlighting preferential expansion of CD8 T cells over Tregs within the tumor.0 30 60 90 ### M-sqz-pbmc-hpv SQZ + FAP-IL-2v FAP-IL2v alone0 30 60 90 SQZ + PD1-IL2v PD1-IL2v alone #140 ## 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.