Open access Original research J Immunother Cancer: first published as 10.1136/jitc-2019-000474 on 8 April 2020. Downloaded from Vaccine efficacy against primary and metastatic cancer with in vitro- generated CD103+ conventional dendritic cells Yifan Zhou, Natalie Slone, Taylor T Chrisikos, Oleksandr Kyrysyuk, Rachel L Babcock, Yusra B Medik, Haiyan S Li, Eugenie S Kleinerman, Stephanie S Watowich To cite: Zhou Y, Slone N, ABSTRACT BACKGROUND Chrisikos TT, et al. Vaccine Background Type 1 conventional dendritic cells (cDC1s) T cell-based immunotherapy and antibody- efficacy against primary and possess efficient antigen presentation and cross- mediated immune checkpoint blockade are metastatic cancer with in vitro- presentation activity, as well as potent T cell priming generated CD103+ conventional among the most exciting advances in cancer ability. Tissue- resident cDC1s (CD103+ cDC1s in mice, dendritic cells. Journal for + therapy over the past decade, eliciting durable ImmunoTherapy of Cancer CD141 cDC1s in humans) are linked with improved tumor control, yet the efficacy of immunotherapy using this control of several cancers and prolonging 2020;8:e000474. doi:10.1136/ survival rates.1 2 Nonetheless, limitations exist jitc-2019-000474 population is understudied. Methods We generated murine CD103+ cDC1s in vitro with current immunotherapies including non- responsiveness or adverse events.3 Thus, ► Additional material is and examined their expression of cDC1- related factors, published online only. To view antigen cross- presentation activity, and accumulation approaches to improve the specificity, effec- please visit the journal online in tumor- draining lymph nodes (TdLNs). The antitumor tiveness, and safety of cancer immunotherapy + (http:// dx. doi. org/ 10. 1136/ jitc- efficacy of the in vitro- generated CD103 cDC1s was across patient populations and cancer types 2019- 000474). studied in murine melanoma and osteosarcoma models. are needed. We evaluated tumor responses on vaccination with Dendritic cells (DCs) are the principal Accepted 21 February 2020 CD103+ cDC1s, compared these to vaccination with + antigen- presenting cells of the immune monocyte- derived DCs (MoDCs), tested CD103 cDC1 system and therefore shape adaptive, anti- vaccination with checkpoint blockade, and examined the 4 antimetastatic activity of CD103+ cDC1s. tumor immunity. These features indicate Results In vitro- generated CD103+ cDC1s produced DCs as a promising tool for anticancer treat- 5–7 cDC1- associated factors such as interleukin- 12p70 ment. The majority of DCs used in clin- http://jitc.bmj.com/ and CXCL10, and demonstrated antigen cross- ical trials have been generated from human presentation activity on stimulation with the toll- like CD14+ monocytes (MoDCs) or CD34+ progen- receptor 3 agonist polyinosinic:polycytidylic acid 8 + itors in culture. While these DCs can be (poly I:C). In vitro- generated CD103 cDC1s also produced in abundance and are capable of migrated to TdLNs following poly I:C treatment and inducing tumor- specific T cells with minimal intratumoral delivery. Vaccination with poly I:C- 7–9 activated and tumor antigen- loaded CD103+ cDC1s side effects, their efficacy remains limited. on September 28, 2021 by guest. Protected copyright. enhanced tumor infiltration of tumor antigen- specific More recently, specific DC populations and interferon-γ+ CD8+ T cells, and suppressed including plasmacytoid DCs (pDCs) and type melanoma and osteosarcoma growth. CD103+ cDC1s 2 conventional DCs (cDC2s) have yielded showed superior antitumor efficacy compared with clinical responses,10 11 yet these subsets are MoDC vaccination, and led to complete regression of relatively sparse in vivo. The efficacy or feasi- 100% of osteosarcoma tumors in combination with bility of current DC vaccines, therefore, may CTLA-4 antibody- mediated checkpoint blockade. In be limited by issues such as use of suboptimal vitro- generated CD103+ cDC1s effectively protected or rare DC subsets. © Author(s) (or their mice from pulmonary melanoma and osteosarcoma employer(s)) 2020. Re- use metastases. Type 1 cDCs (cDC1s) exhibit several permitted under CC BY-NC. No features that predict important roles in acti- commercial re- use. See rights Conclusions Our data indicate an in vitro- generated + and permissions. Published by CD103 cDC1 vaccine elicits systemic and long- vating antitumor immunity, and abundance BMJ. lasting tumor- specificT cell- mediated cytotoxicity, of cDC1s within tumors correlates with Immunology, University of Texas which restrains primary and metastatic tumor growth. improved patient outcomes and response to + MD Anderson Cancer Center, The CD103 cDC1 vaccine was superior to MoDCs immune checkpoint blockade.12 13 The cDC1 Houston, Texas, USA and enhanced response to immune checkpoint subset possesses antigen uptake, antigen blockade. These results indicate the potential for new Correspondence to presentation, and antigen cross-presentation immunotherapies based on use of cDC1s alone or in abilities. Moreover, migratory CD103+ cDC1s Dr Stephanie S Watowich; combination with checkpoint blockade. swatowic@ mdanderson. org transport tissue or tumor antigens to lymph Zhou Y, et al. J Immunother Cancer 2020;8:e000474. doi:10.1136/jitc-2019-000474 1 Open access J Immunother Cancer: first published as 10.1136/jitc-2019-000474 on 8 April 2020. Downloaded from nodes (LNs) and elicit antigen- specific CD8+ T cell METHODS responses.14–18 CD103+ cDC1s can be recruited to tumors CD103+ cDC1 vaccination by T cell- expressed chemokines including XCL1, where In vitro- generated CD103+ cDC1s29 were cultured they participate in further T cell recruitment through (2.5–4.5×106/mL) in Roswell Park Memorial Institute expression of chemoattractants such as CXCL10.12 19 (RPMI) 1640 medium containing 10% heat-inactivated Consistent with these functions, lymphoid organ-resident fetal bovine serum (FBS) (Atlanta Biologicals, Atlanta, CD8α+ cDC1s induced CD8+ T cell responses and protected Georgia, USA), 1% penicillin-streptomycin, 1 mM mice against melanoma engraftment, while treatments sodium pyruvate, and 50 µM β-mercaptoethanol (RPMI to expand and activate locally recruited CD103+ cDC1s 1640 complete medium) ± 20 µg/mL poly I:C (Sigma, St increased the efficacy of B- raf kinase (BRAF) inhibition Louis, Missouri, USA), 5% XG-3 supernatant or 20 ng/ and PD-1 blockade in controlling melanoma.18 20 Collec- mL granulocyte- macrophage colony- stimulating factor tively, these features suggest cDC1-based vaccines will (GM- CSF) and the surrogate tumor antigens ovalbumin elicit antitumor activity, yet this concept requires further (OVA; 100–400 µg/mL) (Sigma) or K7M3 tumor lysate validation. Moreover, whether cDC1- based vaccines for 4 hours. Cells were washed twice with phosphate- 6 protect from metastatic disease is important to examine, buffered saline (PBS) and injected (0.5–3×10 ) into as metastasis is a primary cause of mortality in patients melanoma or osteosarcoma tumors (intratumoral (i.t.)), with cancer. one or two times 4–7 days following tumor implantation, 2 Melanoma and melanoma metastatic disease are respon- when tumor sizes reached 10–20 mm . In bilateral tumor + 2 7 assays, poly I:C- treated and tumor antigen- loaded CD103 sive to immunotherapies such as checkpoint blockade. 6 A number of other tumor types, however, remain poorly cDC1s (2–3×10 cells) were injected into left-side tumors; responsive or refractory. In particular, pediatric solid right- side tumors remained untreated. For metastasis assays, poly I:C- treated and tumor antigen- loaded CD103+ tumors are frequently non-responsive to immunotherapy. 6 Additionally, these tumors often develop resistance to cDC1s (1–2×10 cells) were injected intravenously (i.v.) standard treatments, leaving few clinical options and a approximately 30 days prior to metastasis challenge. In some metastasis assays, mice were injected i.v. with need to identify novel approaches for young patients with + + + 6 cancer. CD45.1 CD45.2 OT- I CD8 T cells (1×10 cells) 1 day prior to CD103+ cDC1 delivery. For combination treat- Osteosarcoma is the most common primary malig- ments with checkpoint inhibitors, DC-vaccinated animals nancy of bone affecting pediatric and adult patients. were injected intraperitoneally (i.p.) with PD-1 antibody Chemotherapy and surgery are standard treatments, yet (RMP1-14; 200 µg/mouse) or CTLA-4 antibody (9H10; the 5- year survival rate is <20% for osteosarcoma patients 200 µg/mouse for the first treatment, 100 µg/mouse for who present with metastases or relapse following treat- subsequent treatments) 4 days following tumor implanta- ment. Negligible improvements have occurred in osteo- 21 22 tion, with two antibody treatments per week for 3 weeks. sarcoma therapeutic options over the past 25 years. http://jitc.bmj.com/ PD-1 and CTLA-4 antibodies were purchased from BioX- Mifamurtide, a liposome-encapsulated immunotherapy Cell company (West Lebanon, New Hampshire, USA). that activates pulmonary macrophages, improved the disease-free and overall survival of patients with osteo- MoDC vaccination sarcoma lung metastases.23 By contrast, MoDC vaccines 24–26 Murine bone marrow (BM) cultures were initiated have yielded little to no clinical responsiveness, while at 0.5×106 cells/mL in RPMI 1640 complete medium checkpoint blockade with PD-1 or CTLA-4 antibodies led containing 20 ng/mL GM-CSF . Cells were collected on September 28, 2021 by guest. Protected copyright. to objective clinical response rates