Submitted Manuscripts
*co-first authors; +corresponding authorsSubmitted Manuscripts
CAR-T cell therapy targeting cell-surface MUC17 in gastric tumors
Park S, Ho CE, Birocchi F, Wolff AN, Bouffard AA, Kelly C, Salas-Benito D, Escobar G, Mucci A, Berger TR, Maus MV. Submitted (2025)Submitted (2025)Park S*, Paek JH*, Colville MJ, Huang LT, Struzyk PA, Womack SJ, Neelamegham S, Reesink HL, Paszek MJ. BioRxiv (2024)Ibrutinib and PD1-blockade potentiate mesothelin-targeting CAR-T cell therapy in preclinical models of pancreatic cancer
Armstrong A, Plancke GVD, Nishiguchi S, Salas-Benito D, Bouffard AA, Gonclaves S, Kelly C, Birocchi F, Park S, Leick M, Berger T, Maus MV, Escobar G. Submitted (2025)The microenvironment dictates glycocalyx construction and immune surveillance
Tharp K, Park S, Timblin G, Richards A, Berg J, Twells N, Riley NM, Peltan E, Shon DJ, Stevenson E, Tsui CK, Palomba F, Lefebvre AE, Soens R, Ayad N, Hoeve-Scott JT, Healy K, Digman M, Dilin A, Bertozzi C, Mahal LK, Swaney D, Cantor J, Paszek MJ, Weaver V. BioRxiv (2023)Publications
*co-first authors; +co-corresponding authorsPublications
2025
2025
Tuning CAR-T cells by targeting cancer-associated glycan in pancreatic cancer
Park S, Ho CE, Darnell EP, Wolff AN, Takei H, Birocchi F, Bouffard AA, Salas-Benito D, Escobar G, Leick MB, Mucci A, Berger TR, Maus MV. Nature Communications, In press (2025)"We incorporate an additional binder targeting Tn-MUC1, termed a "glyco-bridge", into mesothelin-targeted CAR-T cells to enhance CAR-T cell-mediated killing. This bridge not only facilitates CAR-T cell infiltration and increases avidity for their target cells but also activates CAR molecules in a manner dependent on the density of the bridge-target antigens and the affinity of the antigen-binding domain. To broaden the glyco-bridge targeting range, we modified the binding domain to be composed of tandem Helix pomatia agglutinin (HPA) lectins, enabling effective recognition of Tn antigens across various cancer types. Our CAR-T cells with the HPA glyco-bridge exhibit superior cytotoxicity in cell-line and patient-derived xenograft (PDX) models of pancreatic cancer. By advancing binding strategies to penetrate the cancer glycocalyx, this approach opens new pathways for enhanced CAR-T cell efficacy in solid tumor immunotherapy."
In vivo CRISPR screens identify key modifiers of CAR T cell function in myeloma
Knudsen NH, Escobar G, Korell F, Kienka T, Nobrega C, Aderson S, Cheng A, Zschummel M, Armstrong A, Bouffard A, Kann MC, Goncalves S, Pope HW, Pezeshki M, Rojas A, Suermondt JS, Philips M, Berger T, Park S, Salas-Benito D, Darnell E, Birocchi F, Leick M, Larson R, Doench JG, Sen D, Yates KB, Mangusso RT, Maus MV. Nature (2025)"We performed in vivo loss-of-function CRISPR screens in BCMA-targeting CAR-T cells to investigate genes influencing CAR-T cell persistence and function in a human multiple myeloma model. We tracked the expansion and persistence of CRISPR-library edited T cells in vitro and at early and late timepoints in vivo to track the performance of gene modified CAR-T cells from manufacturing to survival in tumors. The screens revealed context-specific regulators of CAR-T cell expansion and persistence. Ablation of RASA2 and SOCS1 enhanced T cell expansion in vitro, while loss of PTPN2, ZC3H12A, and RC3H1 conferred early growth advantages to CAR-T cells in vivo. Strikingly, we identified cyclin-dependent kinase inhibitor 1B (CDKN1B), a cell cycle regulator, as the most important factor limiting CAR-T cell fitness at late timepoints in vivo. CDKN1B ablation increased CAR-T cell proliferation and effector function, significantly enhancing tumor clearance and overall survival. Our findings reveal differing effects of gene-perturbation on CAR-T cells over time and in different environments, highlight CDKN1B as a promising target to generate highly effective CAR-T cells for multiple myeloma, and underscore the potential of in vivo screening for identifying genes to enhance CAR-T cell efficacy."Salas-Benito D, Birocchi F, Park S, Ho C, Armstrong A, Parker A, Bouffard AA, Frank JA, Kim E, Kienka T, Graham K, Kelly C, Goncalves S, Leick MB, Escobar G, Rueda B, Berger TR, Maus MV. Journal for ImmunoTherapy of Cancer (2025)
"We show that the scFv arrangement and linker length impacted antigen binding and CAR expression in T cells. Tandem CAR configuration (TanCAR1) (with SS1 scFv located distally and one G4S repeat as the linker between scFvs) had the best binding and activation profile in vitro and outperformed SS1 and 4H11 monospecific CAR-T cells in mixed tumor models in vitro and in vivo, showing an antigen-driven killing of tumor cells based on antigen density. Moreover, acoustic force microscopy, using tumor cells with different levels of antigen expression, revealed that TanCAR1-T cells likely bind to one antigen at a time rather than simultaneously."
Secretion of a VEGF-blocking scFv enhances CAR-T cell potency
Supper VM*, Donner H*, Birocchi F, Bratt A, Escobar G, Kann MC, Park S, Martin G, Korell F, Takei H, Parker A, Salas-Benito D, Darnell EP, Bailey SR, Kienka T, Bouffard A, Goncalves S, Choi BD, Haradhvala NJ, Maus MV, Leick MB. Cancer Immunology Research (2025)"We show that CAR T cells from patients treated with FDA-approved CAR T-cell products express members of the VEGF signaling pathway and this expression is correlated with patient non-response. To overcome putative VEGF-induced CAR T-cell dysfunction and deliver local VEGF blockade, we generated CAR T cells that secrete a VEGF-targeting scFv to block T-cell and tumor-derived VEGF within the TME. These CAR T cells potently inhibited VEGF signaling and angiogenesis in vitro, and exhibited enhanced activation, cytotoxicity, proliferation, and effector function across different antigen and solid tumor contexts. VEGF scFv-secreting CAR T cells showed improved tumor control in immunocompromised murine metastatic and orthotopic models of ovarian and lung cancer."Jani P, Colville MJ, Park S, Ha Y, Paszek MJ+, and Abbott NL+. Soft Matter (2025)
"We report single-vesicle-level characterization of giant plasma membrane vesicles (GPMVs) derived from cells overexpressing Muc1, revealing a 40x variation in Muc1 density between GPMVs from a single preparation and a strong correlation between GPMV size and Muc1 density. By dispersing GPMVs in aqueous liquid crystals (LCs), we show that the elasticity of the LC can be used to strain individual GPMVs into spindle-like shapes, consistent with the straining of fluid-like membranes. Overall, our study advances the understanding of heterogeneity in size and Muc1 density in GPMVs, and establishes single-vesicle-level methods for characterization of mechanical properties within a heterogeneous population of GPMVs."
2024
2024
CAR-T cell therapy for the treatment of adult high-grade gliomas [review]
Park S, Maus MV, Choi BD. NPJ Precision Oncology (2024)"Treatment for malignant primary brain tumors, including glioblastoma, remains a significant challenge despite advances in therapy. CAR-T cell immunotherapy represents a promising alternative to conventional treatments. This review discusses the landscape of clinical trials for CAR-T cell therapy targeting brain tumors, highlighting key advancements like novel target antigens and combinatorial strategies designed to address tumor heterogeneity and immunosuppression, with the goal of improving outcomes for patients with these aggressive cancers."Dimerization activates the Inversin complex in C. elegans
Beyrent E, Wei DT, Beacham GM, Park S, Zheng J, Paszek MJ, and Hollopeter G. Molecular Biology of the Cell (2024)"Genetic, colocalization, and biochemical studies suggest that the ankyrin repeat-containing proteins Inversin (INVS) and ANKS6 function with the NEK8 kinase to control tissue patterning and maintain organ physiology. It is unknown whether these three proteins assemble into a static “Inversin complex” or one that adopts multiple bioactive forms. Through the characterization of hyperactive alleles in C. elegans, we discovered that the Inversin complex is activated by dimerization. We propose that dynamic switching between at least two functionally distinct states – an active dimer and an inactive monomer – gates the output of the Inversin complex."Park S*, Choi S*, Shimpi AA, Estroff LA, Fischbach C+, Paszek MJ+. Advanced Materials (2024)
"Skeletal metastasis is common in patients with advanced breast cancer and often caused by immune evasion of disseminated tumor cells (DTCs). Here, a combination of synthetic bone matrix models with controlled mineral content, nanoscale optical imaging, and flow cytometry are utilized to evaluate how collagen type I mineralization affects the biochemical and biophysical properties of the tumor cell glycocalyx, a dense layer of glycosylated proteins and lipids decorating their cell surface. These results suggest that collagen mineralization upregulates mucin-type O-glycosylation and sialylation by tumor cells, which increases their glycocalyx thickness while enhancing resistance to attack by natural killer (NK) cells."
Immunoengineering can overcome the glycocalyx armor of cancer cells
Park S, Colville MJ, Paek JH, Shurer CR, Singh A, Secor EJ, Sailer CJ, Huang L, Kuo JCH, Goudge MC, Su J, Kim M, DeLisa MP, Neelamegham S, Lammerding J, Zipfel WR, Fischbach C, Reesink HL, Paszek MJ. Nature Materials (2024)*Highlight [Nature Materials] [Technology Networks]
"Cancer cell glycocalyx is a major line of defence against immune surveillance. However, how specific physical properties of the glycocalyx are regulated on a molecular level, contribute to immune evasion and may be overcome through immunoengineering must be resolved. Here we report how cancer-associated mucins and their glycosylation contribute to the nanoscale material thickness of the glycocalyx and consequently modulate the functional interactions with cytotoxic immune cells. Natural-killer-cell-mediated cytotoxicity is inversely correlated with the glycocalyx thickness of the target cells. Together, our results motivate the development of immunoengineering strategies that overcome the glycocalyx armour of cancer cells."
2023
2023
The Lamin A/C Ig-fold undergoes cell density-dependent changes that alter epitope binding
Wallace M, Fedorchak GR, Agrawal R, Gilbert RM, Patel J, Park S, Paszek MJ, Lammerding J. Nucleus (2023)"Lamins A/C are nuclear intermediate filament proteins that are involved in diverse cellular mechanical and biochemical functions. Here, we report that recognition of Lamins A/C by a commonly used antibody (JOL-2) that binds the Lamin A/C Ig-fold and other antibodies targeting similar epitopes is highly dependent on cell density, even though Lamin A/Clevels do not change. We propose that the effect is caused by partial unfolding or masking of the C’E and/or EF loops of the Ig-fold in response to cell spreading. These findings are important for the interpretation of immunofluorescence data for Lamin A/C and also raise the intriguing prospect that the conformational changes may play a role in Lamin A/C mediated cellular function."
Recombinant mucin biotechnology and engineering [review]Park S, Kuo JCH, Reesink HL, Paszek MJ. Advanced Drug Delivery Reviews (2023)
"Mucins represent a largely untapped class of polymeric building block for biomaterials, therapeutics, and other biotechnology. Because the mucin polymer backbone is genetically encoded, sequence-specific mucins with defined physical and biochemical properties can be fabricated using recombinant technologies. The pendent O-glycans of mucins are increasingly implicated in immunomodulation, suppression of pathogen virulence, and other biochemical activities. In this review, we discuss these advances, and the opportunities for engineered mucins in biomedical applications ranging from in vitro models to therapeutics."
Before 2022
Colville MJ, Park S, Singh A, Paszek MJ, Zipfel WR. Biomedical Engineering Technologies (2022)Before 2022
"Azimuthal beam scanning, also referred to as circle scanning, is an effective way of eliminating coherence artifacts with laser illumination in widefield microscopy. With a static excitation spot, dirt on the optics and internal reflections can produce an uneven excitation field due to interference fringes. These artifacts become more pronounced in TIRF microscopy, where the excitation is confined to an evanescent field that extends a few hundred nanometers above the coverslip. In this chapter, we detail the design and construction of the SAIM instrument, including the optical configuration, required peripheral devices, and system calibration."
Litmus-Body: A Molecularly Targeted Sensor for Cell-Surface pH Measurements
Kuo JCH, Goudge MC, Metzloff AE, Huang L, Colville MJ, Park S, Zipfel WR, Paszek MJ. ACS Sensors (2020)"Precise pH measurements in the immediate environment of receptors is essential for elucidating the mechanisms through which local pH changes associated with diseased phenotypes manifest into aberrant receptor function. However, current pH sensors lack the ability to localize and target specific receptor molecules required to make these measurements. Herein we present the Litmus-body, our recombinant protein-based pH sensor, which through fusion to an anti-IgG nanobody is capable of piggybacking on IgG antibodies for molecular targeting to specific proteins on the cell surface. In complex with our Litmus-body, cetuximab therapeutic antibody retained its functionality in binding and inhibiting ligand interaction of its target epidermal growth factor receptor (EGFR), triggering receptor-mediated endocytosis that allowed tracking of local pH from the cell surface through the endocytic pathway."Colville MJ, Park S, Zipfel WR, Paszek MJ. Scientific Reports (2019)
"Azimuthal beam scanning eliminates the uneven excitation field arising from laser interference in through-objective total internal reflection fluorescence (TIRF) microscopy. The same principle can be applied to scanning angle interference microscopy (SAIM), where precision control of the scanned laser beam presents unique technical challenges for the builders of custom azimuthal scanning microscopes. We show how the integration of waveform generation, multiplexed analog outputs, and native hardware triggers into a single central hub provides a versatile platform for performing fast circle-scanning acquisitions, including azimuthal scanning SAIM and multiangle TIRF. Our complete platform, including hardware design, firmware, API, and software, is available online for community-based development and collaboration."Lee H., Choi B, Kang HN, Kim H, Min A, Cha M, Ryu JY, Park S, Sohn J, Shin K, Yun M, Han JY, Shon MJ, Jeong C, Chung J, Lee S, Im S, Cho BC, Yoon T. Nature Biomedical Engineering (2018)
"The accumulation of genetic and epigenetic alterations in cancer cells rewires cellular signalling pathways through changes in the patterns of protein–protein interactions (PPIs). Understanding these patterns may facilitate the design of tailored cancer therapies. Here, we show that single-molecule pull-down and co-immunoprecipitation techniques can be used to characterize signalling complexes of the human epidermal growth-factor receptor (HER) family in specific cancers. Our approach might help predict responses to targeted cancer therapies, particularly for cancers that lack actionable genomic mutations."