UbiVac (United States)

Abstract Description Background NCPs derived from non-coding regions of the genome are a recently discovered source of non-mutated HLA-presented cancer antigens. Here we studied the immune response of a patient with NSCLC that received adjuvant treatment with DPV-001 immunotherapy that contained canonical and non-canonical antigens. The patient is disease free at 10 years. Materials and Methods This patient received 7 doses of DPV-001 at 3-week intervals. PBMCs and serum were collected regularly to assess antibodies (PhIPseq), peripheral lymphocytes populations (flow cytometry), and TCR repertoires. A metastasis resected 5 months into treatment generated a tumor cell line and tumor-infiltrating lymphocyte (TIL) culture. The HLA-presented immunopeptidome of the patient’s NSCLC cell line was characterized by mass spectrometry and identified more than 300 NCP from 5’UTR. Antibody responses to the canonical protein downstream of the 5’UTR were used to prioritize peptides for detection of a T cell response in TIL. The TCR of a responding T cell was identified, expressed, and evaluated. Results The identified TCR recognized a NCP and autologous NSCLC cell line, it was not detected pretreatment. Current studies are evaluating whether this TCR recognizes other cancer cell lines. Conclusions The detection of a T cell response to a NCP presented by the patient’s cancer expands the range of possible cancer antigens that can be targeted with the potential to improve outcomes for patients with cancer. Funding Sources Support: Murdock Charitable Trust, Providence Portland Medical Foundation, Nancy Lematta, Lynn Loacker, Cindy and Steve Harder, The Chiles Foundation, Robert W. Franz, Elsie Franz Finley and by NCI SBIR grant #R44 CA121612. Topic Categories Tumor Immunology: Cellular Responses and Tumor Microevironment (TIME)

Abstract Description Background Single cell RNA-seq combined with CITE-seq and TCR-seq has allowed for a deep characterization of anticancer immunity. Here we report on a single patient with head and neck squamous cell cancer (HNSCC) who underwent initial resection in 2020. Twenty-one months later, when the patient recurred and went on a combination immunotherapy trial, they had three serial biopsies performed. Methods Samples of blood and tumor from 2020 were sorted into 6 CD4 subsets: Blood Mem CD4, Blood Treg, PD1/ICOS(-/-) TIL, PD1/ICOS(+/+) TIL (putative tumor-specific CD4), PD1/ICOS(+/-) TIL, and TIL Treg. On progression the patient received a biologic containing >300 proteins for genes that are overexpressed by HNSCC followed by anti-PD-1 at wk 2, with biopsies taken at baseline, wk 2 and 8. Phenotypic and molecular studies were performed. Results Over 20% of the total peripheral T cell repertoire at the start of immunotherapy were CD4 clonotypes present in 2020; this shrank to less than 15% by week 8. Within the biopsy samples PD1+ TIL expanded while Treg contracted during immunotherapy. Blood Mem, Blood Treg and PD1/ICOS(-/-) TIL were all mainly found in the periphery, while PD-1 TIL were found in both blood and tumor. Interestingly, the majority of TIL Treg were later found in either the blood or tumor, not both. Conclusions These results provide insights into how Immunotherapy may change the clonal and phenotypic distribution of CD4 T cells. Ethics Approval: PSJH IRB# 2020000480 Funding Sources Support: Incyte, Murdock Charitable Trust, Providence Portland Medical Foundation, Nancy Lematta, Lynn Loacker, Cindy and Steve Harder, The Chiles Foundation, Robert W. Franz and Elsie Franz Finley Topic Categories Tumor Immunology: Cellular Responses and Tumor Microevironment (TIME)

BackgroundOur group developed a strategy for generating an ‘off-the-shelf’ multivalent proteasome-blocked autophagosome vaccine that contains proteins for many genes commonly overexpressed in adenocarcinoma and squamous cell cancers. This strategy exploits in vitro manipulation of the antigen presentation pathway to concentrate the dominant epitopes presented by MHC, including short-lived proteins (SLiPs), defective ribosomal products (DRiPs), and Dark Matter, the short-lived non-canonical peptides that are not expressed in the thymus and represent potential shared alternative cancer neoantigens.1 In preclinical models this vaccine strategy provides significant protection as a single agent,2 and significantly increased therapeutic efficacy when combined with anti-GITR and anti-PD-1.3 Based on these studies we hypothesize that addition of anti-GITR to the human vaccine, DPV-001, and anti-PD-1, will augment expansion and limit contraction of the anti-cancer immune response. A phase I clinical trial was initiated, and preliminary immunological monitoring data will be presented.MethodsPatients received DPV-001, with sequenced checkpoint inhibition (aPD-1 mAb; retifanlimab), with or without aGITR agonist mAb (INCAGN1876), in recurrent or metastatic HNSCC (NCT04470024). Tumor biopsies were taken pre-treatment, week 2 and 8. Blood samples were taken pre-treatment and at multiple timepoints and analyzed by flow cytometry and seromics. Tumor biopsies and blood were assayed by CITE-seq, scRNA-seq, BCR-seq, and TCR-seq. Multiplex immunofluorescence (mIF) was performed on biopsies.ResultsIn the first 4 patients evaluated, tumor-infiltrating T cells at week 8 increased from pre-treatment levels by an average of 4.3 fold (range 2.9 – 6.7, p=0.032). The density of CD39/CD103 double positive cells, previously shown to identify tumor-reactive T cells,4 also increased in all week 8 biopsies (mean 14.7 fold, range 5–40). All patients showed increased numbers of cells expressing IFN-γ and GZMB and increased numbers of T cells expressing LAG3+ in week 8 biopsies. Preliminary TCR evaluation of the tumor identified proliferation of clones previously undetected in PBL, including αβ T cells, iNKT and MAIT cells. Expansion of clones that predated treatment was also identified.ConclusionsAn increase in intra-tumoral T cells expressing activation and effector molecules is encouraging and studies are underway to expand the number of patients analyzed. Increased expression of LAG3 by T cells that infiltrate and have expanded in the tumor, provide a rationale for including anti-LAG-3 in this treatment strategy. Future plans include evaluating whether immune responses target shared non-canonical alternative neoantigens, or Dark Matter, contained in DPV-001 and whether antibody responses identify targets of cellular immunity.AcknowledgementsSupport: Incyte, UbiVac, Providence Medical Foundation, The Chiles Foundation, The Harder Family, Nancy Lematta, Robert W. Franz and Elsie Franz Finley, The Murdoch Memorial TrustTrial RegistrationNCT04470024ReferencesFox BA, Urba WJ, Jensen SM, Page DB, Curti BD, Sanborn RE, Leidner RS. Cancer’s Dark Matter: Lighting the Abyss Unveils Universe of New Therapies. Clinical Cancer Research 2023;29:2173–2175. 10.1158/1078–0432.CCR-23–0422.Twitty CG, Jensen SM, Hu H-M, Fox BA. Tumor-Derived Autophagosome Vaccine: Induction of Cross-Protective Immune Responses against Short-lived Proteins through a p62-Dependent Mechanism. Clinical Cancer Research 2011;17:6467–6481. 10.1158/1078–0432.CCR-11–0812.Patel JM, Cui Z, Wen Z-F, Dinh CT, Hu H-M. Peritumoral administration of DRibbles-pulsed antigen-presenting cells enhances the antitumor efficacy of anti-GITR and anti-PD-1 antibodies via an antigen presenting independent mechanism. J Immunother Cancer 2019;7:311. 10.1186/s40425–019-0786–7.Duhen T, Duhen R, Montler R, Moses J, Moudgil T, de Miranda NF, Goodall CP, Blair TC, Fox BA, McDermott JE, et al. Co-expression of CD39 and CD103 identifies tumor-reactive CD8 T cells in human solid tumors. Nat Commun 2018;9:2724. 10.1038/s41467–018-05072–0.Ethics ApprovalPSJH IRB# 2020000480

BackgroundToll-like receptors (TLRs) are commonly expressed on innate immune cells such as macrophages and dendritic cells (DC), and they play crucial roles in 1) mediating the first-line innate immune response against a wide variety of pathogens and 2) promoting adaptive immunity involving T and B cells. The TLR family consists of 10 subtypes (TLR1-TLR10) in humans and 12 (TLR1-TLR9, TLR11-TLR13) in mice. CAN1012 was developed as a selective TLR7 agonist uniquely designed for intratumoral (IT) administration.MethodsCAN1012 was evaluted by in vitro cell-based assays and in vivo animal models. Its DMPK, safety and toxicitology properties were also studied.ResultsBased on the receptor screening study, CAN1012 has been proved to have a significant stimulatory effect (NF-κB activation) via human and mouse TLR7. In addition, CAN1012 stimulated significant IFN-α release from human PBMC at low concentrations (nM), whereas TNF-α secretion required a relatively high concentration.Most importantly, CAN1012 induced a robust production of IFN-α exclusively by plasmacytoid DCs in human PBMC. When given intratumorally, CAN1012 exposure in tumor tissues was over 1,000-fold higher than in blood, resulting in much less systemic toxicity. The antitumoral effects of IT-administered CAN1012 monotherapy or combined with other therapeutical agents were investigated in mouse SCC7, CT26, and MC38 syngeneic tumor models. The results showed that CAN1012 possessed significant anti-tumor growth effects in a dose-dependent and schedule-dependent manner, and antitumoral effects could be augmented when combined with other agents. Pharmacodynamic studies revealed that IT administration of CAN1012 increased CD4+ and CD8+ T cell infiltration but decreased the number of myeloid-derived suppressor cells (MDSCs) and tumor-associated macrophages (TAMs) in the tumor microenvironment (TME). CAN1012 has a favorable ADME/PK profile when administered subcutaneously in mice, rats and monkeys, and its pharmaceutical properties have also been optimized for IT administration.ConclusionsIn conclusion, CAN1012 is a potent, safe, and selective TLR7 agonist and could be a best-in-class agent for cancer immunotherapy. A first-in-human Phase I clinical study in advanced cancer patients is ongoing in the US (NCT04987112).Trial RegistrationNCT04987112Ethics ApprovalAniaml studies had been approved by the ethics review committee at the institution at which the studies were conducted

Meeting abstracts Analysis of tumor-infiltrating immune cells using quantitative immunohistochemistry (IHC) has proved to be a powerful prognostic biomarker in colon cancer [[1][1], [2][2]]. Similar observations have been made in patients with oral, head and neck squamous cell carcinoma (OHNSCC),

Meeting abstracts We hypothesize that immune responses that develop following immunotherapy, particularly in patients with objective responses, are directed against a wide range of antigenic determinants. However, in the absence of autologous tumor cells, monitoring is frequently limited to a small

Abstract CD80 expression by APC provides signal 2, a critical element in priming T cell immune responses to antigen. As limited CD80 expression on APC can result in reduced immune responses, strategies that increase CD80 may have benefit. We report here that the autophagosome-enriched vaccine named DRibbles (DRiPs and SLiPs in Blebs), derived from human tumor cells transfected with a plasmid encoding huCD80, are highly efficient at transferring membrane proteins onto human PBMCs. When PBMCs were incubated with a DRibble vaccine derived from a CD80 expressing breast-tumor cell line, a significant increase in surface expression of CD80 was observed on PBMCs within 5 minutes (p<0.001). This effect was time and concentration-dependent and repeatable across multiple donors (N=5) in multiple experiments (N≥16). DRibble vaccine could transfer CD80 onto as high as 60-86.4% of monocytes and 41-86.1% of CD4+ T cells. Transfer of CD80 to CD8 T cells was substantially less effective. Transfer of membrane-associated proteins between leukocyte populations is a well-documented phenomenon, however, this is the first report of transfer of membrane-associated proteins from cell-free autophagosomes. Current efforts are investigating the functional properties associated with increased expression of CD80. Given the critical role of CD80 in augmenting immune responses this provides a novel strategy to further augment the response to DRibble vaccines, which are being explored in clinical trials.