This study focuses on the specific CD4+ T cell requirement for optimal induction of cytotoxicity against MHC class II negative autologous tumors (AuTu) collected from patients with various types of cancer at advanced stages. CD4+ T cells were induced in cultures of cancer patients’ malignant effusion-associated mononuclear cells with irradiated AuTu (mixed lymphocyte tumor cultures (MLTC)) in the presence of recombinant IL-2 and recombinant IL-7. Tumor-specific CD4+ T cells did not directly recognize the AuTu cells, but there was an MHC class II-restricted cross-priming by autologous dendritic cells (DCs), used as APC. CD8+ CTL, also induced during the MLTC, lysed specifically AuTu cells or DCs pulsed with AuTu peptide extracts (acid wash extracts (AWE)) in an MHC class I-restricted manner. Removal of CD4+ T cells or DCs from the MLTC drastically reduced the CD8+ CTL-mediated cytotoxic response against the AuTu. AWE-pulsed DCs preincubated with autologous CD4+ T cells were able, in the absence of CD4+ T cells, to stimulate CD8+ T cells to lyse autologous tumor targets. Such activated CD8+ T cells produced IL-2, IFN-γ, TNF-α, and GM-CSF. The process of the activation of AWE-pulsed DCs by CD4+ T cells could be inhibited with anti-CD40 ligand mAb. Moreover, the role of CD4+ T cells in activating AWE-pulsed DCs was undertaken by anti-CD40 mAb. Our data demonstrate for the first time in patients with metastatic cancer the essential role of CD4+ Th cell-activated DCs for optimal CD8+ T cell-mediated killing of autologous tumors and provide the basis for the design of novel protocols in cellular adoptive immunotherapy of cancer, utilizing synthetic peptides capable of inducing T cell help in vivo.

Nine labdane type diterpenes isolated from the plant Cistus creticus subsp. creticus and from the resin "Ladano" which is excreted on the surface of the leaves and stems of this plant, were examined for their in vitro cytotoxic activity against 14 human leukemic cell lines. Compound 1, (13E)-labd-13-ene-8 alpha,15-diol, exhibited cytotoxic activity against 13 of the cell lines tested, while compound 2, (13E)-labd-7,13-dienol, was active only against HL60 cells. Further compound 1 was examined for its effect on the uptake of [3H]-thymidine as a marker of DNA synthesis.

HER2/neu-derived peptides inducing MHC class II-restricted CD4+ T helper lymphocyte (Th) responses, although critical for tumour rejection, are not thoroughly characterized. Here, we report the generation and characterization of CD4+ T cell clones specifically recognizing a HER-2/neu-derived peptide (776–788) [designated HER2(776–788)]. Such clones yielded specific proliferative and cytokine [gamma-interferon(IFN)-γ] responses when challenged with autologous dendritic cells (DCs) loaded with HER2(776–788). By performing blocking studies with monoclonal antibodies (MAbs) and by using DCs from allogeneic donors sharing certain HLA-DR alleles, we found that HER2(776–788) is a promiscuous peptide presented, at least, by DRB5*0101, DRB1*0701 and DRB1*0405 alleles. One TCRVbeta6.7+ clone recognized the HLA-DRB5*0101+ FM3 melanoma cell line transfected with a full length HER-2/neu cDNA. Moreover, this clone recognized the HER-2/neu+ SKBR3 breast cancer cell line induced to express HLA-DR, thus demonstrating that HER2(776–788) represents a naturally processed and presented epitope. Our data demonstrate that helper peptide HER2(776–788) represents a promiscuous epitope binding to at least three HLA-DR alleles, thus offering a broad population coverage. The use of antigenic peptides presented by major histocompatibility complex (MHC) class II in addition to those presented by class I may improve the therapeutic efficacy of active immunization.© 2001 Cancer Research Campaign  http://www.bjcancer.com

Incubation of kaempferol-3-O-beta-D-(6"-E-p-coumaroyl)-glucopyranoside (tiliroside) (1) with Aspergillus nidulans gives the 7-methyl ether of tiliroside (2) which is a new compound. Its structure is determined by spectroscopic methods. Cytotoxic studies of 2 and of its acetylated derivative 2a were carried out in vitro against fourteen human leukemic cell lines. Results clearly show that compound 2 is ineffective against all leukemic cell lines tested. On the contrary, compound 2a exhibited cytotoxic activity against four of the cell lines (HL60, DAUDI, HUT78 and MOLT3) and additionally, a dose- and time-dependent effect on DNA synthesis.

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