Now in its third year, the Selective Targeting of Pancreatic Cancer (SToP) Specialized Program of Research Excellence (SPORE) grant has teamed up with UNC Lineberger’s Career Enhancement Program (CEP) to support a diverse group of young researchers seeking to gain key insight into critical areas of pancreatic cancer research. During each year of this five-year grant, we have the unique opportunity to fund the work of early-career or mid-career investigators who are transitioning to focus on pancreatic cancer and whose research is directed toward elucidating mechanisms of this particular cancer’s development, progression or therapy.
Below is an overview of the innovative research we’ve been able to support through the SToP Cancer SPORE CEP in its first two years, including each of the project’s goals and the investigators’ significant findings and conclusions.
Justin Milner, PhD: “Molecular regulation of T cell activity in pancreatic ductal adenocarcinoma”
Although cancer immunotherapy has revolutionized the field of oncology and rapidly emerged as a first-line treatment for diverse malignancies, pancreatic ductal adenocarcinoma (PDAC) tumors are especially immunosuppressive and broadly refractory to current immunotherapy approaches. Milner’s project sought to define key molecular regulators of the tumor immune microenvironment (TIME) in PDAC.
Through high-throughput genetic screens, new roles have been uncovered for numerous chromatin regulatory factors in controlling immune cell fate and function in vivo, including unappreciated roles for Bromodomain and extraterminal domain (BET) family proteins. BET proteins are chromatin “readers” broadly expressed in cancer cells and immune cells. Inhibition of BET proteins typically results in profound anti-tumorigenic effects in vitro or in immunocompromised mouse models for many cancer types, including PDAC; however, results from BETi clinical trials using BET inhibitors (BETi) have not shown clinical activity for BETi in cancer patients. Critically, how BET proteins control the tumor immune microenvironment (TIME) remains ill-defined. Milner posited that complex roles of BET proteins in the immune system may contribute to mixed results of BETi therapies in cancer, and he anticipated that careful, cell-intrinsic studies of BET proteins in the PDAC TIME may provide new insights for therapeutically modifying the composition of the TIME.
Milner first extensively optimized and validated a spectral flow cytometry approach permitting comprehensive analysis of the TIME across 13 different syngeneic C57BL6 tumor models, including orthotopically implanted KrasG12DTrp53R172H (KPC) tumors. He and his team are in the process of leveraging this TIME profiling approach to investigate how depletion of BRD2, BRD3, BRD4 modulate the TIME in KPC tumors. They started by profiling the role of BRD4 in regulating the composition of the TIME through use of Brd4 floxed mice. As CD8 T cells are crucial regulators of PDAC progression, Milner assessed how depletion of Brd4 in CD8 T cells impacted their differentiation in the TIME and found that induced depletion of Brd4 in CD8 T cells blunted exhausted differentiation but also impaired effector differentiation. Lastly, they generated complementary in vitro data with a small molecule inhibitor of BET proteins, JQ1, also demonstrating that BET proteins are crucial in the differentiation of CD8 T cell exhaustion.
Milner’s team then looked to determine how BET protein BRD4 restrains anti-tumor CD8 T cell responses in PDAC. As BRD4 critically regulates CD8 T cell responses in PDAC, they sought to investigate mechanisms by which BRD4 controls gene expression programs and CD8 T cell fate. Preliminary findings indicate that the transcription factor (TF) BATF exhibits overlapping binding patterns with BRD4 at key loci that regulate T cell differentiation. Therefore, they are in the process of investigating whether BRD4 interacts with BATF to regulate expression of effector- and exhaustion-specifying genetic programs in CD8 T cells in the PDAC TIME. To further discern the molecular signals regulating the differentiation and function of CD8 T cells in the PDAC TIME and complement their BRD4 studies, they have performed single-cell omics in PDAC-specific CD8 T cells in mice. They then generated a KPC cell line expressing the model CD8 T cell epitope GP33-41 derived from the LCMV pathogen.
Next, Milner’s team sort purified GP33-41-specific CD8 T cells from KPC-GP tumors and performed combined scRNAseq+ scATACseq. Intriguingly, the BATF motif is enriched in both
effector and exhausted CD8 T cell clusters in PDAC, and they are currently investigating putative gene targets of BRD4-BATF through mining this scATACseq data and in-hand BRD4 binding data. They have recently completed CUT&RUN assays to profile genome wide binding patterns of BRD4 in exhausted CD8 T cells, which will be used to clarify gene targets and regulatory activity of BRD4 in CD8 T cells. Lastly, they have performed scRNAseq+scVDJ profiling to investigate PDAC-specific CD8 T cell responses of expanded T cell clones that presumably recognize native KPC tumor antigens. They find extensive overlap in the clusters and genetic programs between our KPC-GP single-cell omics data and the scVDJ experiment, indicating their KPC-GP model represents a robust approach to investigating how BRD4 modulates KPC-specific CD8 T cells responses in PDAC.
Through this development of new tumor models, new single-cell omics data, and breeding of uncharacterized mouse strains in PDAC, Milner has begun to uncover important signals that regulate the differentiation and accumulation of CD8 T cells in the PDAC TIME. In contrast to current publications on the role of BRD4 in leukemia models, BRD4 is simultaneously important for both exhaustion and effector function in PDAC. This may explain why BETi monotherapy is not especially effective in treating PDAC, as CD8 T cell antitumor responses are likely constrained. He and his team have also generated preliminary evidence that BRD4 interacts with BATF to regulate important effector programs in CD8 T cells. Through generation of new single-cell omics datasets, they have begun to clarify the extensive heterogeneity of CD8 T cells in PDAC and have uncovered putative regulatory mechanisms governing T cell fate and function in PDAC.
Ashwin Somasundaram, MD: “A Phase Ib study of patients with advanced basal PANcreatic adenocarcinoma treated with Gemcitabine, Erlotinib, and nab-paclitaxel”
This project supported the opening of a two-arm clinical trial to determine the optimal combination treatment for patients with advanced unresectable or metastatic basal-like subtype PDAC. Investigators seek to examine the safety, efficacy, tolerability, and anti-tumor effects of low-dose epidermal growth factor receptor (EGFR) inhibitors in combination with bi-weekly gemcitabine/nab-paclitaxel (GnP).
The standard of care chemotherapy for first-line advanced pancreatic adenocarcinoma is FOLFIRINOX, NALIRIFOX, or gemcitabine/nab-paclitaxel. However, multiple studies suggest that basal-like subtypes of pancreatic adenocarcinoma do not respond to FOLFIRINOX. Based upon existing retrospective analyses, the addition of epidermal growth factor receptor (EGFR) inhibitors such as erlotinib to gemcitabine and nab-paclitaxel suggest improved rates of response in subjects with basal-like pancreatic adenocarcinoma compared to FOLFIRINOX.
The Purity Independent Subtyping of Tumors (PurIST), previously developed by researchers in the UNC Lineberger Pancreatic Cancer Center of Excellence, is used to classify a patient’s type of cancer as either “basal type” or “classical.” Subjects with a classical subtype are being treated on standard-of-care oxaliplatin-based triplet chemotherapy. Subjects with basal type are receiving the erlotinib combination treatment. The erlotinib combination treatment is not FDA-approved, however, the combinations of erlotinib, gemcitabine, and nab-paclitaxel are both approved by the FDA for the treatment of pancreatic cancer. All three drugs have been used in combination before in other clinical trials and a certain amount of safety data exists.
Since receiving support for this project, Somasundaram and his team worked aggressively to open the clinical study, which began accruing patients in February 2025. Currently, they have enrolled 12 patients within 2 months including 3 basal subtype patients. This concept has also led to the expansion into a two-arm, randomized, basal study. Through this development, Somasundaram has submitted five proposals for further funding given the strong feasibility and impact of this project, which has allowed for expansion to additional local and national sites and has garnered significant interest as well as expansion into the neoadjuvant setting and with novel combination therapies. He has hypothesized that the tumors from patients responsive to anti-EGFR and gemcitabine-based therapy have a highly activated stromal signature, and they also posit that the tumors with the least response to combination therapy have decreased molecular signature of the basal subtype but increased expression of metagenes associated with the classical subtype.
To test these hypotheses, Somasundaram will evaluate the formalin-fixed paraffin-embedded (FFPE) tumor samples from patients both before and after treatment with E+GnP utilizing the GeoMx platform by NanoString. They will analyze these samples in the same methodology used to validate the PurIST signature. They will also evaluate the molecular signature of their new treatment regimens and retrospectively identify any novel markers of resistance for future consideration.