Anna Karolina Palucka

Induction of ICOS+CXCR3+CXCR5+ TH Cells Correlates with Antibody Responses to Influenza Vaccination

A T cell subset that emerges in blood after seasonal influenza vaccinations correlates with the development of protective antibody responses. What Lies Beneath Although the seasonal flu vaccine, which can protect 60 to 90% of young healthy adults, has been in use for decades, we still know surprisingly little about how it actually induces protective antibody responses. This information is especially important to improve vaccination efficacy in populations that are more susceptible to infection such as the very young and the elderly. Now, Bentebibel et al. take us a step further into understanding what is required for protective antibody responses in humans. The authors identified a subset of CD4+ T cells that were associated with protective antibody responses after seasonal flu vaccination in humans. These cells expressed the costimulatory molecules ICOS as well as two chemokine receptors, CXCR3 and CXCR5, which identify these cells as circulating memory T follicular helper (TFH) cells. TFH cells traditionally are thought to reside in the B cell follicles and be instrumental for germinal center formation and subsequent memory antibody response. Indeed, these circulating cells were influenza antigen–specific, could induce memory B cells to differentiate into plasma cells, and correlated with specific antibody titer. Further studies that find ways to harness these cells could thus improve vaccine design. Seasonal influenza vaccine protects 60 to 90% of healthy young adults from influenza infection. The immunological events that lead to the induction of protective antibody responses remain poorly understood in humans. We identified the type of CD4+ T cells associated with protective antibody responses after seasonal influenza vaccinations. The administration of trivalent split-virus influenza vaccines induced a temporary increase of CD4+ T cells expressing ICOS, which peaked at day 7, as did plasmablasts. The induction of ICOS was largely restricted to CD4+ T cells coexpressing the chemokine receptors CXCR3 and CXCR5, a subpopulation of circulating memory T follicular helper cells. Up to 60% of these ICOS+CXCR3+CXCR5+CD4+ T cells were specific for influenza antigens and expressed interleukin-2 (IL-2), IL-10, IL-21, and interferon-γ upon antigen stimulation. The increase of ICOS+CXCR3+CXCR5+CD4+ T cells in blood correlated with the increase of preexisting antibody titers, but not with the induction of primary antibody responses. Consistently, purified ICOS+CXCR3+CXCR5+CD4+ T cells efficiently induced memory B cells, but not naïve B cells, to differentiate into plasma cells that produce influenza-specific antibodies ex vivo. Thus, the emergence of blood ICOS+CXCR3+CXCR5+CD4+ T cells correlates with the development of protective antibody responses generated by memory B cells upon seasonal influenza vaccination.

Dendritic cells loaded with killed allogeneic melanoma cells can induce objective clinical responses and MART-1 specific CD8+ T-cell immunity.

Dendritic cells (DCs) loaded with killed allogeneic tumors can cross-prime tumor-specific naive CD8+ T cells in vitro, thereby providing an option to overcome human leukocyte antigen restriction inherent to loading DC vaccines with peptides. We have vaccinated 20 patients with stage IV melanoma with autologous monocyte-derived DCs loaded with killed allogeneic Colo829 melanoma cell line. DCs were generated by culturing monocytes with granulocyte macrophage-colony stimulating factor (granulocyte macrophage-colony stimulating factor) and interleukin (IL-4) and activated by additional culture with tumor necrosis factor and CD40 ligand. A total of 8 vaccines were administered at monthly intervals. The first patient was accrued December 2002 and the last November 2003. Fourteen patients were alive at 12 months, 9 patients were alive at 24 months, and 8 patients are alive as of January 2006. The estimated median overall survival is 22.5 months with a range of 2 to 35.5 months. Vaccinations were safe and tolerable. They induced, in 2 patients who failed previous therapy, durable objective clinical responses, 1 complete regression (CR) and 1 partial regression (PR) lasting 18 and 23 months, respectively. Three out of 13 analyzed patients showed T-cell immunity to melanoma antigen recognized by autologous T cells (MART-1) tissue differentiation antigen. Two of 3 patients showed improved immune function after vaccinations demonstrated by improved secretion of interferon (IFN)-γ or T-cell proliferation in response to MART-1 derived peptides. In one of these patients, vaccination led to elicitation of CD8+ T-cell immunity specific to a novel peptide-derived from MART-1 antigen, suggesting that cross-priming/presentation of melanoma antigens by DC vaccine had occurred. Thus, the present results justify the design of larger follow-up studies to assess the clinical response to DC vaccines loaded with killed allogeneic tumor cells in patients with metastatic melanoma.

Targeting self- and foreign antigens to dendritic cells via DC-ASGPR generates IL-10–producing suppressive CD4+ T cells

Targeting antigens to the lectinlike DC-ASGPR receptor on human DCs and in nonhuman primates results in the induction of antigen-specific IL-10–producing CD4+ T cells.

Dendritic cells: a critical player in cancer therapy?

Cancer immunotherapy seeks to mobilize a patients immune system for therapeutic benefit. It can be passive, that is, transfer of immune effector cells (T cells) or proteins (antibodies), or active, that is, vaccination. Early clinical trials testing vaccination with ex vivo generated dendritic cells (DCs) pulsed with tumor antigens provide a proof-of-principle that therapeutic immunity can be elicited. Yet, the clinical benefit measured by regression of established tumors in patients with stage IV cancer has been observed in a fraction of patients only. The next generation of DC vaccines is expected to generate large numbers of high avidity effector CD8+ T cells and to overcome regulatory T cells and suppressive environment established by tumors, a major obstacle in metastatic disease. Therapeutic vaccination protocols will combine improved DC vaccines with chemotherapy to exploit immunogenic chemotherapy regimens. We foresee adjuvant vaccination in patients with resected tumors but at high risk of relapse to be based on in vivo targeting of DCs with fusion proteins containing anti-DCs antibodies, antigens from tumor stem/propagating cells, and DC activators.

Concomitant Activation and Antigen Uptake via Human Dectin-1 Results in Potent Antigen-Specific CD8+ T Cell Responses

Dectin-1, a C-type lectin recognizing fungal and mycobacterial pathogens, can deliver intracellular signals that activate dendritic cells (DCs), resulting in initiation of immune responses and expansion of Th17 CD4+ T cell responses. In this paper, we studied the roles of human Dectin-1 (hDectin-1) expressed on DCs in the induction and activation of Ag-specific CD8+ T cell responses. We first generated an agonistic anti–hDectin-1 mAb, which recognizes the hDectin-1 Glu143-Ile162 region. It bound to in vitro monocyte-derived DCs and to in vivo CD1c+CD1a+ dermal DCs but not to epidermal Langerhans cells. Anti–hDectin-1–mediated DC activation resulted in upregulation of costimulatory molecules and secretion of multiple cytokines and chemokines in a Syk-dependent manner. DCs activated with the anti–hDectin-1 mAb could significantly enhance both neo and foreign Ag-specific CD8+ T cell responses by promoting both the expansion of CD8+ T cells and their functional activities. We further demonstrated that delivering Ags to DCs via hDectin-1 using anti–hDectin-1-Ag conjugates resulted in potent Ag-specific CD8+ T cell responses. Thus, hDectin-1 expressed on DCs can contribute to the induction and activation of cellular immunity against intracellular pathogens, such as mycobacteria, that are recognized by DCs via Dectin-1. Vaccines based on delivering Ags to DCs with an agonistic anti–hDectin-1 mAb could elicit CD8+ T cell-mediated immunity.

Dengue virus and dendritic cells.

Little is known about the pathogenesis of the Dengue virus, a mosquito-borne flavivirus that causes about 50 million human infections annually, some of which may be fatal. Recent studies have demonstrated two subsets of dendritic cells, monocyte-derived dendritic cells and human skin Langerhans cells, that are targets for Dengue virus infection (pages 816–820).

IFN Priming Is Necessary but Not Sufficient To Turn on a Migratory Dendritic Cell Program in Lupus Monocytes

Blood monocytes from children with systemic lupus erythematosus (SLE) behave similar to dendritic cells (DCs), and SLE serum induces healthy monocytes to differentiate into DCs in a type I IFN–dependent manner. In this study, we found that these monocytes display significant transcriptional changes, including a prominent IFN signature, compared with healthy controls. Few of those changes, however, explain DC function. Exposure to allogeneic T cells in vitro reprograms SLE monocytes to acquire DC phenotype and function, and this correlates with both IFN-inducible (IP10) and proinflammatory cytokine (IL-1β and IL6) expression. Furthermore, we found that both IFN and SLE serum induce the upregulation of CCR7 transcription in these cells. CCR7 protein expression, however, requires a second signal provided by TLR agonists such as LPS. Thus, SLE serum “primes” a subset of monocytes to readily (<24 h) respond to TLR agonists and acquire migratory DC properties. Our findings might explain how microbial infections exacerbate lupus.

Abstract IA17: TSLP-driven inflammation fosters development of epithelial tumors

Solid tumors are often associated with aseptic inflammation. There are two types of inflammation that have opposing effects on tumors, chronic inflammation that promotes cancer cell survival, and metastasis, and acute inflammation which triggers cancer cell destruction. Chronic inflammation is often linked with the presence of type 2-polarized macrophages (M2), which are induced by Th2 cytokines, IL-4 and IL-13. Our recent studies have demonstrated the presence in breast cancer tumors of inflammatory Th2 cells, which produce IL-13, IL-4, and TNF. These CD4+ T cells appear to play a key role in the disease as they accelerate breast tumor development in a xenograft model through the production of IL-13. Breast tumors appear to play a critical role in conditioning the infiltrating myeloid DCs (mDCs) to induce such inflammatory Th2 cells. Our most recent results suggest that thymic stromal lymphopoetin (TSLP) secreted by cancer cells plays a role in mDCs conditioning. Breast cancer cell lines and primary tumors from patients show TSLP protein expression. TSLP-neutralizing antibodies block the upregulation of OX40L by mDCs exposed to tumor supernatant and consequently block mDCs capacity to generate inflammatory Th2 cells in vitro. The TSLP production is mediated by PAR2-signaling in cancer cells. PAR2 is expressed by all 4 breast cancer cell lines, and TSLP production can be induced upon PAR2-agonist peptide treatment. Similar observations were made by us and others in pancreatic cancer as well as lung cancer. Thus, TSLP could serve as therapeutic target in tumors of epithelial origin.

Breast cancer instructs dendritic cells to express OX40 ligand and to prime pro-inflammatory type 2 CD4+ T cells that facilitate tumor development.

Abstract #1050 We found that breast cancer tumors are infiltrated with mature dendritic cells (DCs), which cluster with CD4+ T cells. CD4+ T cells infiltrating breast cancer tumors secrete type 1 (IFN-g), pro-inflammatory (TNF) as well as high levels of type 2 (IL-4 and IL-13) cytokines. Immunofluorescence staining of tissue sections revealed intense IL-13 staining on breast cancer cells. The expression of phosphorylated STAT6 in breast cancer cells suggests that IL-13 actually delivers signals to cancer cells.
 To determine the link between breast cancer, DCs and CD4+ T cells, we implanted human breast cancer cell lines in NOD/SCID b2m-/- mice engrafted with human CD34+ hematopoietic progenitor cells (HPCs) and autologous T cells. There, CD4+ T cells promote early tumor development. This can be prevented by administration of IL-13 antagonists and is dependent on DCs. Furthermore, these humanized mice demonstrate a signature of IL-13.
 Similarly to patient samples, in humanized mice breast cancer but not melanoma tumors are infiltrated with large numbers of human myeloid DCs. DC isolated from breast tumor and its draining lymph nodes induce the production of high level of IL4, IL13 and TNF by naive CD4 T cells further demonstrating that breast tumor modulates DC to induce an inflammatory Th2 response.
 Accordingly, immunofluorescence staining of breast cancer tissue sections from patients shows high expression of OX40 ligand on DCs. This can be ascribed to activity of breast cancer cell-derived soluble factors. Finally, blocking OX40 ligand prevents generation of IL-13 secreting CD4+ T cells.
 Thus, breast cancer targets DCs to generate pro-inflammatory type 2 immunity that promotes its development.
 Supported by U19 AIO57234, RO-1 CA89440, CA78846, and CA85540. Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 1050.