Role of CAR-T cell therapy in pancreatic cancer: an updated narrative review of current evidence

Rationale and Background: Immunotherapy of cancer has gained much attention in the past decade with the development of immune checkpoint inhibitors and chimeric antigen receptor (CAR) technology that can activate and redirect patient T cells to kill tumors that over-express a specific antigen. CARs are fusion receptors that are comprised of an antibody-derived single-chain variable fragment (scFv) coupled via hinge and transmembrane elements to a T cell signaling and co-stimulatory domain. This technology is in its early stages of development and has not been fully exploited for the treatment of metastatic epithelial cancers. We focus our studies on Pancreatic Ductal Adenocarcinomas (PDAC) and Triple Negative Breast Cancer (TNBC). However, if successful, the studies will be applicable to other epithelial tumors. For CAR-T cells to work, and to avoid off target toxicities, both the target antigen and the antibody recognizing the target have to be highly specific. The challenge is that there are few such antigen-antibody combinations for solid tumors. We have recently developed a novel patented antibody (designated TAB 004) that specifically recognizes ONLY the tumor-associated form of MUC1 (tMUC1) but not the normal form of MUC1 (nMUC1) in several subtypes of breast cancers including TNBC and in PDAC. We show compelling data that TAB004 recognizes tMUC1 in >90% of human TNBCs and 85% of PDAC but spares all normal epithelial tissues. The antigenic isoform that TAB004 recognizes is completely hidden in normal epithelia making it extremely safe for development of CAR-T cells. Hypothesis: TNBC and PDAC can be specifically targeted with the tMUC1-CAR-engineered T cells, whilst sparing normal organs. Methodology and Results: We have engineered several TAB-specific CAR constructs using the scFv fragment of TAB 004. Six constructs are developed, 3 for human T cells and 3 for mouse T cells: 1. TAB- CD28-CD3zeta (2nd generation CAR), 2. TAB-CD28-41BB-CD3zeta (3rd generation CAR), and 3. TAB-CD28-OX40-CD3zeta (3rd generation CAR). Data shows that we can successfully engineer human T cells to express the TAB-CAR on their surface and that these engineered T cells can bind specifically to tumor cells expressing the unique tMUC1 epitope, become activated, and effectively kill the tumor cells. We show that these engineered T cells only minimally bind and kill normal epithelial cell lines. However, some of the cell lines are more resistant than others. We are therefore conducting combination therapy with various drugs that are known to enhance immune based therapies including checkpoint inhibitors, COX-2 inhibitors, cyclophosphamide and others. We may also have data to show that the engineered TAB-CAR T cells kill tMUC1-expressing TNBC and PDAC cells in vivo. We will show data that TAB-CAR-T cell kill human PDAC and TNBC cells in vitro and propose to conduct the same in vivo xenograft model of human PDAC and metastatic TNBC. In the following months, we will test if TAB-CAR-T cell retards tumor growth in an orthotopic syngeneic mouse model of PDAC and metastatic BC in human MUC1.Tg immune competent mice. We further propose to demonstrate that the TAB-CAR-T cell can mediate apoptosis in the immune competent KCM mice (KC X human MUC1.Tg mice) that develop spontaneous PDAC and the MMT mice (PyVMT X human MUC1.Tg mice) that develop spontaneous mammary gland tumors. Both models mimic the human disease progression and express human MUC1 in a tissue specific manner. This is important since all normal epithelia in these mice express the nMUC1 except the tumors that expresses the target, tMUC1. Impact: If successful, this project will have a major impact and accelerate progress toward a clinical trial for PDAC and metastatic TNBC. This will be the first attempt to test the efficacy of a CAR-T cell in an immune competent, human MUC1.Tg mouse model that develops spontaneous tumors within the appropriate stromal and hormonal microenvironment. Citation Format: Pinku Mukherjee, Ru Zhou, Mahboubeh Yazdanifar, Das Roy Lopamudra. Development and future of CAR T cell therapy for pancreatic ductal adenocarcinoma and triple negative breast cancer [abstract]. In: Proceedings of the AACR International Conference: New Frontiers in Cancer Research; 2017 Jan 18-22; Cape Town, South Africa. Philadelphia (PA): AACR; Cancer Res 2017;77(22 Suppl):Abstract nr A37.

Desmoplasia of Pancreatic Ductal Adenocarcinoma

Therapeutic Advances in Pancreatic Cancer

B7-H3 as a Novel CAR-T Therapeutic Target for Glioblastoma

Advancing together and moving forward: Combination gene and cellular immunotherapies

Pancreatic ductal adenocarcinoma (PDAC) is projected to become the second leading cause of cancer death by 2030. PDAC lacks effective treatment options and innovative strategies for therapeutic intervention are urgently needed. Chimeric Antigen Receptor (CAR)-T cell therapy has shown remarkable success in treating hematologic malignancies, yet achieving efficacy in solid tumors remains a significant challenge. Early results from clinical trials show limited efficacy of CAR T-cell therapy in PDAC. To uncover tumor-intrinsic mechanisms of resistance to CAR T-cell therapy in vivo, we performed iterative CRISPR/Cas9-based pooled screens in a fully immunocompetent, orthotopic model of PDAC. We identified a variety of genes influencing CAR T-cell treatment efficacy. In particular, loss of genes involved in redox biology and oxidative stress sensitized PDAC tumors to CAR T-cell therapy in vivo. Additionally, single-cell RNA sequencing of PDAC tumors subjected to CAR T-cell therapy unveiled a distinct subset of tumor cells resilient to the treatment, characterized by changes in oxidative stress. Our findings indicate a potential avenue for therapeutic synergy by targeting pathways involved in oxidative stress to increase the efficacy of CAR T cell therapy in PDAC. Citation Format: Julia Fröse, Charlie Whittaker, Paul Leclerc, Adam Langenbucher, Riley Hellinger, Daniel R. Goulet, Michael T. Hemann. Unveiling resistance mechanisms to CAR T-cell therapy in pancreatic cancer through in vivo CRISPR/Cas9 knockout screening [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr B060.

Cell surface GRP78-directed CAR-T cells are effective at treating human pancreatic cancer in preclinical models

The engineering of immune cells with chimeric antigen receptors (CARs) has enabled the development of CAR T cell therapies, which eliminate tumor cells in an antigen-specific, HLA-independent manner without prior education. While CAR T cells have shown remarkable success in CD19+ B cell malignancies, their efficacy in solid tumors remains limited due to heterogeneous antigen expression, complex tumor trafficking, and an immunosuppressive tumor microenvironment (TME). To overcome these challenges, this study explores a CAR-based natural killer (NK) cell therapy targeting CD70, an emerging tumor-associated antigen, for metastatic colorectal cancer (mCRC) and pancreatic ductal adenocarcinoma (PDAC). CD70 is part of the CD70-CD27 co-stimulatory axis, which is normally tightly regulated but becomes dysregulated in various cancers, contributing to tumor progression and immune evasion. While CD70 is primarily expressed on tumor cells, recent findings, including those from this study, have identified CD70+ cancer-associated fibroblasts (CAFs) in the TME of CRC and PDAC, where they promote tumor migration and immune suppression. Given their pro-tumorigenic role, we hypothesized that CD70+ CAFs represent a viable therapeutic target. CAR NK cells offer several advantages over CAR T cells, including off-the-shelf availability, lower toxicity, and multiple tumor-killing mechanisms. In this study, CD70-directed CAR NK cells were developed and evaluated for their efficacy against CD70+ tumor cells and CAFs in CRC and PDAC. Notably, interleukin-15 (IL-15) stimulation was found to be essential for optimal CAR NK cell activation and cytotoxicity. IL-15 significantly enhanced CAR expression and NK cell activation, reducing dependence on high CD70 target expression. To optimize patient selection, we investigated soluble CD27 (sCD27) as a potential predictive biomarker for CD70+ CAFs. However, in vitro analyses revealed no correlation between sCD27 levels and CD70+ CAF presence, limiting its utility as a stratification marker for CD70-CAR NK therapy. Given the central role of CAFs in therapy resistance and tumor progression, targeting CD70+ CAFs may improve treatment responses. However, to achieve comprehensive tumor eradication, this strategy may need to be combined with direct tumor-targeting therapies. Chemotherapy, a standard-of-care treatment for mCRC, was found to enhance CD70 expression and induce tumor cell stress responses that could synergize with CAR NK therapy. In conclusion, this study provides a strong preclinical rationale for translating CD70-CAR-IL-15 NK cells into clinical applications for mCRC and PDAC. Furthermore, the findings support investigating combination strategies with chemotherapy to enhance therapeutic efficacy.

Promoter usage regulating the surface density of CAR molecules may modulate the kinetics of CAR-T cells in vivo

ASGE guideline on screening for pancreatic cancer in individuals with genetic susceptibility: summary and recommendations

B-cell maturation antigen chimeric antigen receptor T-cell re-expansion in a patient with myeloma following salvage programmed cell death protein 1 inhibitor-based combination therapy.

<div>Abstract<p>Chimeric antigen receptor (CAR) T-cell therapy can lead to dramatic clinical responses in B-cell malignancies. However, early clinical trials with CAR T-cell therapy in non–B-cell malignancies have been disappointing to date, suggesting that tumor-intrinsic features contribute to resistance. To investigate tumor-intrinsic modes of resistance, we performed genome scale CRISPR-Cas9 screens in mesothelin (MSLN)-expressing pancreatic cancer cells. Co-culture with MSLN-targeting CAR T cells identified both antigen-dependent and antigen-independent modes of resistance. In particular, loss of the majority of the genes involved in the pathway responsible for GPI-anchor biosynthesis and attachment abrogated the ability of CAR T cells to target pancreatic cancer cells, suggesting that disruption of this pathway may permit MSLN CAR T-cell evasion in the clinic. Antigen-independent mediators of CAR T-cell response included members of the death receptor pathway as well as genes that regulate tumor transcriptional responses, including <i>TFAP4</i> and <i>INTS12</i>. TFAP4-mediated CAR T resistance depended on the NFκB transcription factor p65, indicating that tumor resistance to CAR T-cell therapy likely involves alterations in tumor-intrinsic states. Overall, this study uncovers multiple antigen-dependent and -independent mechanisms of CAR T-cell evasion by pancreatic cancer, paving the way for overcoming resistance in this disease that is notoriously refractory to immunotherapy.</p>Significance:<p>The identification and validation of key determinants of CAR T-cell response in pancreatic cancer provide insights into the landscape of tumor cell intrinsic resistance mechanisms and into approaches to improve therapeutic efficacy.</p></div>

<div>Abstract<p>Chimeric antigen receptor (CAR) T-cell therapy can lead to dramatic clinical responses in B-cell malignancies. However, early clinical trials with CAR T-cell therapy in non–B-cell malignancies have been disappointing to date, suggesting that tumor-intrinsic features contribute to resistance. To investigate tumor-intrinsic modes of resistance, we performed genome scale CRISPR-Cas9 screens in mesothelin (MSLN)-expressing pancreatic cancer cells. Co-culture with MSLN-targeting CAR T cells identified both antigen-dependent and antigen-independent modes of resistance. In particular, loss of the majority of the genes involved in the pathway responsible for GPI-anchor biosynthesis and attachment abrogated the ability of CAR T cells to target pancreatic cancer cells, suggesting that disruption of this pathway may permit MSLN CAR T-cell evasion in the clinic. Antigen-independent mediators of CAR T-cell response included members of the death receptor pathway as well as genes that regulate tumor transcriptional responses, including <i>TFAP4</i> and <i>INTS12</i>. TFAP4-mediated CAR T resistance depended on the NFκB transcription factor p65, indicating that tumor resistance to CAR T-cell therapy likely involves alterations in tumor-intrinsic states. Overall, this study uncovers multiple antigen-dependent and -independent mechanisms of CAR T-cell evasion by pancreatic cancer, paving the way for overcoming resistance in this disease that is notoriously refractory to immunotherapy.</p>Significance:<p>The identification and validation of key determinants of CAR T-cell response in pancreatic cancer provide insights into the landscape of tumor cell intrinsic resistance mechanisms and into approaches to improve therapeutic efficacy.</p></div>

A Tumor Sorting Protocol that Enables Enrichment of Pancreatic Adenocarcinoma Cells and Facilitation of Genetic Analyses

Recommendations for a More Organized and Effective Approach to the Early Detection of Pancreatic Cancer From the PRECEDE (Pancreatic Cancer Early Detection) Consortium