Sabrina
Chen
92 Program for Research in Science and Engineering Defining andTargeting Cell Surface Protein Adaptations to Oncogenic KRAS Inhibition in Pancreatic Ductal Adenocarcinoma (PDAC)
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Authors:
Sabrina Chen, Phillip Kong, Joseph Mancias
Date Created:
2025-01-01
Course Title:
Professor:
Not specified
About Paper:
Pancreatic ductal adenocarcinoma (PDAC), the most common We also conducted IC50 assays to compare the sensitivities of and lethal form of pancreatic cancer, lacks early diagnostic tools these lines to MRTX1133 (KRASi), MMAE (drug payload), and and effective treatments. A key driver of PDAC progression ALPP ADC. We then tested the combination of KRASi and is KRAS, a dominant oncogene mutated in ▯90% of tumors, ALPP ADC in synergy assays and live-cell imaging studies. making it an important target for therapy. However, resistance Our experiments confirmed increased ALPP expression in both to KRAS inhibitors (KRASi) emphasizes the need for effective sensitive and resistant cell lines. We also found high sensitivity combination strategies. To address this, the Mancias Lab has to MMAE, confirming its suitability as the ADC drug payload. appliedquantitativemassspectrometry-basedtemporalproteomics The cell lines displayed variable sensitivities to MRTX1133, and to map changes in PDAC cells following KRAS inhibition. Using our synergy assays demonstrated that combining KRASi with this data, my project focuses on identifying and evaluating cell ALPP ADC produced synergistic effects, with live-cell imaging surface proteins upregulated in response to KRASi as targets for confirming ADC internalization. In the coming weeks, we plan antibody-drug conjugate (ADC) delivery. ALPP was selected due to knock out ALPP using CRISPR/Cas9 genome editing to test if to its consistent upregulation following KRAS inhibition across upregulation of ALPP is related to KRASi resistance. Lastly, we multiple PDAC cell lines. To validate ALPP upregulation on the plan to test this combination in immunodeficient mice with PDAC cell surface, we performed flow cytometry on several pancreatic cell lines. If successful, this approach could advance to clinical cancer cell lines (AsPC-1, Panc-1, Panc0203, SUIT2, SW1990) trials as a promising PDAC therapy. to measure percent positivity and mean fluorescence intensity. Engineering a Non-Pathogenic E. coli Surface Display Platform for Cancer Immunotherapy Alice Chen, Stephanie Sendker, Rizwan Romee Harvard College | Eliot House | Chemical and Physical Biology | 2026 Cancerimmunotherapyhasrevolutionizedoncologybyharnessing proteins: Neae, YiaT181, YiaT232, OmpA, and C-IgAP. Protein the body’s immune system to target malignant cells. However, presentation was tracked via FLAG-tag AF647 fluorochrome the broader field still faces several key challenges, including thedetection by flow cytometry, with staining confirmed for both immunosuppressive tumor microenvironments, poor therapeutic murine and human IL-12 through the shared p40 domain. We trafficking, and systemic cytotoxicity. To address this, our lab evaluated IL-12 bioactivity using HEK-Blue reporter assays and has exploited the tumor tropism and genetic tractability of non- assessed therapeutic efficacy through cytotoxicity assays against pathogenic Escherichia coli (E. Coli) as a novel bioengineering OCI-AML3, NOMO1, K562, and AsPC1 cancer cell lines. NK platform to deliver immune-activating cytokines via surface cell activation was assessed through immunophenotyping using display. In our previous work, we have successfully engineered multicolor flow cytometry. In vivo efficacy was tested using these non-pathogenic E. coli to display the immune-activating intratumoral injection in B16-F10 melanoma C57BL/6 mouse decoy-resistant interleukin IL-18, leading to enhanced anti-tumor models. Our results showed successful IL-12 surface display, immunity while avoiding systemic cytotoxicity. Building on this promoting NK cell activation and driving potent anti-leukemia platform, this project aims to enhance E. coli by expressing IL- cytotoxicity. Mouse models confirmed therapeutic efficacy 12, one of the most potent immunostimulatory cytokines that following intratumoral bacterial injection. Future work includes effectively activates NK cells and promotes anti-tumor immunity. exploring combinatorial therapeutic strategies and optimizing We transformed the non-pathogenic E. coli strain DH5alpha to delivery methods for broader cancer treatment applications. display human and murine IL-12 tethered to five different scaffold
Abstract:
Pancreatic ductal adenocarcinoma (PDAC), the most common We also conducted IC50 assays to compare the sensitivities of and lethal form of pancreatic cancer, lacks early diagnostic tools these lines to MRTX1133 (KRASi), MMAE (drug payload), and and effective treatments. A key driver of PDAC progression ALPP ADC. We then tested the combination of KRASi and is KRAS, a dominant oncogene mutated in ▯90% of tumors, ALPP ADC in synergy assays and live-cell imaging studies. making it an important target for therapy. However, resistance Our experiments confirmed increased ALPP expression in both to KRAS inhibitors (KRASi) emphasizes the need for effective sensitive and resistant cell lines. We also found high sensitivity combination strategies. To address this, the Mancias Lab has to MMAE, confirming its suitability as the ADC drug payload. appliedquantitativemassspectrometry-basedtemporalproteomics The cell lines displayed variable sensitivities to MRTX1133, and to map changes in PDAC cells following KRAS inhibition. Using our synergy assays demonstrated that combining KRASi with this data, my project focuses on identifying and evaluating cell ALPP ADC produced synergistic effects, with live-cell imaging surface proteins upregulated in response to KRASi as targets for confirming ADC internalization. In the coming weeks, we plan antibody-drug conjugate (ADC) delivery. ALPP was selected due to knock out ALPP using CRISPR/Cas9 genome editing to test if to its consistent upregulation following KRAS inhibition across upregulation of ALPP is related to KRASi resistance. Lastly, we multiple PDAC cell lines. To validate ALPP upregulation on the plan to test this combination in immunodeficient mice with PDAC cell surface, we performed flow cytometry on several pancreatic cell lines. If successful, this approach could advance to clinical cancer cell lines (AsPC-1, Panc-1, Panc0203, SUIT2, SW1990) trials as a promising PDAC therapy. to measure percent positivity and mean fluorescence intensity. Engineering a Non-Pathogenic E. coli Surface Display Platform for Cancer Immunotherapy Alice Chen, Stephanie Sendker, Rizwan Romee Harvard College | Eliot House | Chemical and Physical Biology | 2026 Cancerimmunotherapyhasrevolutionizedoncologybyharnessing proteins: Neae, YiaT181, YiaT232, OmpA, and C-IgAP. Protein the body’s immune system to target malignant cells. However, presentation was tracked via FLAG-tag AF647 fluorochrome the broader field still faces several key challenges, including thedetection by flow cytometry, with staining confirmed for both immunosuppressive tumor microenvironments, poor therapeutic murine and human IL-12 through the shared p40 domain. We trafficking, and systemic cytotoxicity. To address this, our lab evaluated IL-12 bioactivity using HEK-Blue reporter assays and has exploited the tumor tropism and genetic tractability of non- assessed therapeutic efficacy through cytotoxicity assays against pathogenic Escherichia coli (E. Coli) as a novel bioengineering OCI-AML3, NOMO1, K562, and AsPC1 cancer cell lines. NK platform to deliver immune-activating cytokines via surface cell activation was assessed through immunophenotyping using display. In our previous work, we have successfully engineered multicolor flow cytometry. In vivo efficacy was tested using these non-pathogenic E. coli to display the immune-activating intratumoral injection in B16-F10 melanoma C57BL/6 mouse decoy-resistant interleukin IL-18, leading to enhanced anti-tumor models. Our results showed successful IL-12 surface display, immunity while avoiding systemic cytotoxicity. Building on this promoting NK cell activation and driving potent anti-leukemia platform, this project aims to enhance E. coli by expressing IL- cytotoxicity. Mouse models confirmed therapeutic efficacy 12, one of the most potent immunostimulatory cytokines that following intratumoral bacterial injection. Future work includes effectively activates NK cells and promotes anti-tumor immunity. exploring combinatorial therapeutic strategies and optimizing We transformed the non-pathogenic E. coli strain DH5alpha to delivery methods for broader cancer treatment applications. display human and murine IL-12 tethered to five different scaffold
Source:
Harvard / Gissell Chapa, Felicia Wranitz, Caroline Burns / 2025
Topics:
cell, alpp, pdac, line, surface, krasi, adc, coli, kras, assay, cancer, using
