HER-2 pulsed dendritic cell vaccine can eliminate HER-2 expression and impact ductal carcinoma in situ.

BACKGROUND: HER-2/neu overexpression plays a critical role in breast cancer development, and its expression in ductal carcinoma in situ (DCIS) is associated with development of invasive breast cancer. A vaccine targeting HER-2/neu expression in DCIS may initiate immunity against invasive cancer. METHODS: A HER-2/neu dendritic cell vaccine was administered to 27 patients with HER-2/neu-overexpressing DCIS. The HER-2/neu vaccine was administered before surgical resection, and pre- and postvaccination analysis was conducted to assess clinical results. RESULTS: At surgery, 5 of 27 (18.5%) vaccinated subjects had no evidence of remaining disease, whereas among 22 subjects with residual DCIS, HER-2/neu expression was eradicated in 11 (50%). When comparing estrogen receptor (ER)(neg) with ER(pos) DCIS lesions, vaccination was more effective in hormone-independent DCIS. After vaccination, no residual DCIS was found in 40% of ER(neg) subjects compared with 5.9% in ER(pos) subjects. Sustained HER-2/neu expression was found in 10% of ER(neg) subjects compared with 47.1% in ER(pos) subjects (P = .04). Postvaccination phenotypes were significantly different between ER(pos) and ER(neg) subjects (P = .01), with 7 of 16 (43.8%) initially presenting with ER(pos) HER-2/neu(pos) luminal B phenotype finishing with the ER(pos) HER-2/neu(neg) luminal A phenotype, and 3 of 6 (50%) with the ER(neg) HER-2/neu(pos) phenotype changing to the ER(neg) HER-2/neu(neg) phenotype. CONCLUSIONS: Results suggest that vaccination against HER-2/neu is safe and well tolerated and induces decline and/or eradication of HER-2/neu expression. These findings warrant further exploration of HER-2/neu vaccination in estrogen-independent breast cancer and highlight the need to target additional tumor-associated antigens and pathways.

Differential production of IL-23 and IL-12 by myeloid-derived dendritic cells in response to TLR agonists.

The recently delineated role for IL-23 in enhancing Th-17 activity suggests that regulation of its expression is distinct from that of IL-12. We hypothesized that independent TLR-mediated pathways are involved in the regulation of IL-12 and IL-23 production by myeloid-derived dendritic cells (DCs). The TLR 2 ligand, lipoteichoic acid (LTA), the TLR 4 ligand, LPS, and the TLR 7/8 ligand, resimiquod (R848), induced production of IL-23 by DCs. None of these TLR ligands alone induced significant IL-12 production, except when combined with IFN-gamma or other TLR ligands. Notably, IL-23 production in response to single TLR ligands was inhibited by IL-4. DCs treated with single TLR agonists induced IL-17A production by allogeneic and Ag-specific memory CD4(+) T cells, an effect that was abrogated by IL-23 neutralization. Moreover, these DCs stimulated IL-17A production by tumor peptide-specific CD8(+) T cells. In contrast, DCs treated with dual signals induced naive and memory Th1 responses and enhanced the functional avidity of tumor-specific CD8(+) T cells. These results indicate that distinct microbial-derived stimuli are required to drive myeloid DC commitment to IL-12 or IL-23 production, thereby differentially polarizing T cell responses.

Targeting HER-2/neu in early breast cancer development using dendritic cells with staged interleukin-12 burst secretion.

Overexpression of HER-2/neu (c-erbB2) is associated with increased risk of recurrent disease in ductal carcinoma in situ (DCIS) and a poorer prognosis in node-positive breast cancer. We therefore examined the early immunotherapeutic targeting of HER-2/neu in DCIS. Before surgical resection, HER-2/neu(pos) DCIS patients (n = 13) received 4 weekly vaccinations of dendritic cells pulsed with HER-2/neu HLA class I and II peptides. The vaccine dendritic cells were activated in vitro with IFN-gamma and bacterial lipopolysaccharide to become highly polarized DC1-type dendritic cells that secrete high levels of interleukin-12p70 (IL-12p70). Intranodal delivery of dendritic cells supplied both antigenic stimulation and a synchronized preconditioned burst of IL-12p70 production directly to the anatomic site of T-cell sensitization. Before vaccination, many subjects possessed HER-2/neu-HLA-A2 tetramer-staining CD8(pos) T cells that expressed low levels of CD28 and high levels of the inhibitory B7 ligand CTLA-4, but this ratio inverted after vaccination. The vaccinated subjects also showed high rates of peptide-specific sensitization for both IFN-gamma-secreting CD4(pos) (85%) and CD8(pos) (80%) T cells, with recognition of antigenically relevant breast cancer lines, accumulation of T and B lymphocytes in the breast, and induction of complement-dependent, tumor-lytic antibodies. Seven of 11 evaluable patients also showed markedly decreased HER-2/neu expression in surgical tumor specimens, often with measurable decreases in residual DCIS, suggesting an active process of "immunoediting" for HER-2/neu-expressing tumor cells following vaccination. DC1 vaccination strategies may therefore have potential for both the prevention and the treatment of early breast cancer.

High-avidity antitumor T-cell generation by toll receptor 8-primed, myeloid- derived dendritic cells is mediated by IL-12 production.

BACKGROUND: High-level production of heterodimeric p70 interleukin (IL)-12 by myeloid-derived dendritic cells (DCs) requires 2 signals: interferon gamma (IFN-gamma) and a maturation signal provided by CD40 ligation (CD40L) or lipopolysaccharide (LPS). METHODS: In the current study we demonstrate that signaling through toll-like receptor (TLR) 8, but not TLR3, TLR2, or TLR4, provides a priming signal to myeloid-derived DC for high IL-12 p70 heterodimer production. RESULTS: All the TLR agonists induced maturation of DC as evidenced by increased expression of CD83, CD80, and CD86. Both IFN-gamma and TLR7/8 agonist R848 increased expression of TLR8 in immature monocyte-derived DCs. The combination of TLR7/8 agonist R848 and maturation signals LPS or CD40L induced high-level expression of IL-12p35 and p40 similar to that induced by IFN-gamma plus LPS. In contrast, receptor agonists specific for TLR7 did not prime for IL-12 production. The p70 IL-12 produced by the TLR8-primed DC polarized CD4+ T for Th1 cytokine production and induced CD8+ T cells, displaying high functional avidity with enhanced tumor cell recognition. CONCLUSIONS: The data suggest that toll 8 receptor agonists are useful for inducing type-1 polarized DCs for vaccine design in treating cancer and infectious disease.

A novel dendritic cell-based immunization approach for the induction of durable Th1-polarized anti-HER-2/neu responses in women with early breast cancer.

Twenty-seven patients with HER-2/neu overexpressing ductal carcinoma in situ of the breast were enrolled in a neoadjuvant immunization trial for safety and immunogenicity of DC1-polarized dendritic cells (DC1) pulsed with 6 HER-2/neu promiscuous major histocompatibility complex class II-binding peptides and 2 additional human leukocyte antigen (HLA)-A2.1 class I-binding peptides. DC1 were generated with interferon-γ and a special clinical-grade bacterial endotoxin (lipopolysaccharide) and administered directly into groin lymph nodes 4 times at weekly intervals before scheduled surgical resection of ductal carcinoma in situ. Patients were monitored for the induction of new or enhanced antipeptide reactivity by interferon-γ ELISPOT and enzyme-linked immunosorbentassays performed on Th cells obtained from peripheral blood or excised sentinel lymph nodes. Responses by cytotoxic T lymphocyte against HLA-A2.1-binding peptides were measured using peptide-pulsed T2 target cells or HER-2/neu-expressing or nonexpressing tumor cell lines. DC1 showed surface phenotype indistinct from "gold standard" inflammatory cocktail-activated DC, but displayed a number of distinguishing functional characteristics including the secretion of soluble factors and enhanced "killer DC" capacity against tumor cells in vitro. Postimmunization, we observed sensitization of Th cells to at least 1 class II peptide in 22 of 25 (88%; 95% exact confidence interval, 68.8%-97.5%) evaluable patients, whereas 11 of 13 (84.6%; 95% exact confidence interval, 64%-99.8%) HLA-A2.1 patients were successfully sensitized to class I peptides. Perhaps most importantly, anti-HER-2/neu peptide responses were observed up to 52-month postimmunization. These data show that even in the presence of early breast cancer such DC1 are potent inducers of durable type I-polarized immunity, suggesting potential clinical value for development of cancer immunotherapy.

Is there a role for the Fas-/Fas-Ligand pathway in chemoresistance of human squamous cell carcinomas of the head and neck (SCCHN)?

The aim of the present investigation was to determine the expression of the Fas-receptor/ligand system in established cell lines of squamous cell carcinomas of the head and neck (SCCHN), and to study it's functional impact on chemotherapy-induced apoptosis in these SCCHN cell lines. We observed constitutive expression of Fas and FasL in 13 SCCHN cell lines by RT-PCR, Southern-blotting and immunocytochemistry, respectively. Administration of the agonistic Fas-antibody CH-11 led to a significant reduction of viable cells in the colorimetric MTT-assay in 5 out of 13 (38%) cell lines tested and preincubation with Interferon-gamma (IFN-gamma) rendered 3 (23%) primarily resistant cell lines sensitive. Cisplatin (cDDP) and bleomycin (BLM) caused dose-dependent cytotoxicity in all cell lines as determined by the 50% inhibitory concentration (IC(50)) and induction of apoptosis. Furthermore, both antineoplastic agents led to an enhanced surface expression of Fas and FasL in all cell lines, and this effect was independent of the respective p53-status. This upregulation of Fas/FasL surface expression increased preexisting Fas-sensitivity only, but failed to make primarily resistant cell lines undergo Fas-mediated growth reduction or apoptosis. Vice versa, blockade of Fas-receptor-ligand-interactions by monoclonal antibodies directed against FasL was able to attenuate the cytotoxic effect of cDDP and BLM in 2 out of 5 (40%) cell lines tested only. In conclusion, in contrast to many other solid tumors, the Fas/FasL-system does not seem to play an exclusive role in anticancer drug mediated apoptosis in SCCHN.

Asymptomatic changes in cardiac function can occur in ductal carcinoma-in-situ patients following treatment with HER-2/neu-pulsed dendritic cell vaccines.

BACKGROUND: Targeting HER-2/neu with Trastuzumab has been associated with development of cardiac toxicity. METHODS: Twenty-seven patients with ductal carcinoma in situ (DCIS) of the breast completed an IRB approved clinical trial of a HER-2/neu targeted dendritic cell based vaccine. Four weekly vaccinations were administered prior to surgical resection. All subjects underwent pre- and post-vaccine cardiac monitoring by MUGA/ECHO scanning allowing for a comparison of cardiac function. RESULTS: In 3 of 27 vaccinated patients (11%) transient asymptomatic decrements in ejection fraction of greater than 15% were noted after vaccination. Notably, evidence of circulating anti-HER-2/neu antibody was found prior to vaccination in all three patients, but cardiac toxicity was not noted until induction of cellular mediated immune responses. CONCLUSIONS: This is the first description of HER-2/neu targeted vaccination associated with an incidence of cardiac changes, and the induction of cellular immune responses combined with antibody may contribute to changes in cardiac function.