McKenna, David H.

Abstract Organoids, which are tiny, lab-grown 3D structures that mimic some organizational and functional properties of human organs, are slowly transforming the face of systems and developmental biology, biomedical research, pharmaceutical testing, environmental toxin testing, and healthcare. Significant investments are essential for the mass production, preservation, and distribution of organoids, with the aim to accelerate innovation and progress across multiple fields—much like the investments made in cell and biologics manufacturing over the past 2 decades.

Abstract Background: Given the dramatic rise in incidence of colorectal cancer in young adults, there remain limited treatment options effective in metastatic disease. Although T cell-based therapies have been successful in treating highly antigenic cancers, like melanoma and lung, they have not yet been shown to induce consistent and durable tumor regression in common epithelial malignancies, including metastatic colorectal cancer (mCRC). CISH (Cytokine inducible SH2 containing protein) is a novel intracellular checkpoint target demonstrating PD-L1 ligand independent mechanisms of enhanced anti-cancer activity. Given that CISH is not currently tractable via antibody or small molecule drug modalities, we employed CRISPR for precise inhibition of CISH in tumor infiltrating lymphocytes (TILs). Here, we provide the first clinical report of safety and anti-tumor activity of CISH edited TILs in 12 patients with metastatic gastrointestinal cancers (NCT04426669), including a clinical complete response (CR) in a young-adult patient with immunotherapy-refractory mCRC. Methods: Tumors were surgically resected for TIL harvest, followed by a rapid expansion protocol and CRISPR/Cas9 knockout (KO) CISH. CISH KO, neoantigen-reactive TILs were expanded, then infused following non-myeloablative lymphocyte depleting (LD) chemotherapy (cyclophosphamide & fludarabine) followed by high-dose IL-2. To establish mechanistic attribution of the CR, we used iR-RepSeq+ for serial analysis of the TCR immune repertoire. Results: Our CRISPR engineering led to KO of CISH in T cells with high-efficiency (>90%) without detectable off-target editing. We safely dosed patients at all five dose levels (range: 1.91e8 to 9.93e10 cells). No dose-limiting toxicities attributable to the CISH-KO TIL product were observed. Adverse events were consistent with established risks of LD chemotherapy, IL-2, or disease progression. A durable ongoing complete response (>21 months) was achieved in a patient with microsatellite instability-high (MSI-H) mCRC refractory to anti-PD1/CTLA-4 combination therapy. Conclusions: We provide the first clinical report of CISH checkpoint targeting via genetically modified T cell therapy, with complete response in a patient with mCRC. Persistence and expansion of unique TCR clonotypes detected in neoantigen responsive TIL was temporally consistent with spikes in CISH edited alleles detected by NGS assay; in the patient with CR, four of the clonotypes exhibiting prolonged persistence greater than one-year post-infusion exhibited significantly reduced or undetectable expression of CISH compared to the total infused TIL population. Given these findings, further investigation of CISH checkpoint inhibition, via gene and cell therapy, and next-generation small molecule drugging modalities, is underway. Citation Format: Emil Lou, Modassir S. Choudhry, Timothy K. Starr, Timothy Folsom, Jason Bell, Blaine Rathmann, Anthony P. DeFeo, Ji Hyun Kim, Nicholas Slipek, Zhaohui Jin, Darin Sumstad, Christopher A. Klebanoff, Katherine Ladner, Akshat Sarkari, R Scott McIvor, Thomas A. Murray, Jeffrey Miller, Madhuri Rao, Eric Jensen, Jacob Ankeny, Mahmoud A. Khalifa, Anil Chauhan, Benjamin Spilseth, Ajay Dixit, Paolo P. Provenzano, Wenjing Pan, Daniel Weber, Miranda Byrne-Steele, Tom Henley, David H. McKenna, Matthew J. Johnson, Beau R. Webber, Branden S. Moriarity. First-in-human trial in patients with metastatic colorectal cancer using CRISPR-engineered tumor infiltrating lymphocytes in which the intracellular immune checkpoint CISH is inhibited [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_2):Abstract nr CT269.

2546 Background: We recently reported that tumor cell vaccines cultured in 5% oxygen (O2) had enhanced the immunogenicity relative to those grown in standard atmospheric O2. In order to develop an “off-the-shelf” source of whole cell antigen we screened a panel of primary glioblastoma (GBM) cell lines, leading to identification of (GBM6) as an ideal candidate. GBM6 was extensively characterized and shown to express glioma-associated antigens (IL13Ra2, Sox2, Epha2, etc.). A Phase I clinical trial was initiated to evaluate the safety and feasibility of a vaccine consisting of dendritic cells (DC) pulsed with apoptotic bodies from GBM6 grown in 5% O2. Methods: Patients ranging from 3 to 71 years with recurrent GBM (n=6) or ependymoma (n=1) were enrolled. Monocytes were collected via apheresis, matured into DC and pulsed with apoptotic bodies derived from GBM6. The first three patients received escalating doses of DC (5x106, 10x106, and 15x106), the remainder received 15 x 106 DC. Pulsed dendritic cellswere injectedsubcutaneously into the supra-scapular region. Imiquimod cream was applied at the injection site just prior to vaccination and 24 hours later. The vaccine schedule dictated administration every 2 weeks for 8 weeks (total of 5 doses) then monthly to progression or a total of 52 weeks. Prior to each vaccination, patients met eligibility parameters and had no progression on MRI. Patients were imaged monthly and blood was drawn to evaluate toxicity and immune response. Results: No vaccine-related toxicity has been reported. Time to progression ranges from 6.5 weeks for the patient treated at the first dose level to 35 weeks for one patient receiving 15 x 106 DC. The latter patient experienced a partial response at 20 weeks. Two patients have stable disease at 18.5 and 28 weeks, respectively. One patient was not evaluable. Flow cytometric analysis demonstrated expansion of central memory T cells amidst declining effector memory cells following vaccination in three patients at a dose of 15x106 DC. Conclusions: Apoptotic body-pulsed DC vaccination was well tolerated and preliminarily demonstrated clinical activity but a minimum of 15x106 DC was required for a modulation of central memory T cells.