Daniele Pozzessere

Recruitment of dendritic cells and enhanced antigen-specific immune reactivity in cancer patients treated with hr-GM-CSF (Molgramostim) and hr-IL-2: results from a phase Ib clinical trial

Experimental findings suggest that granulocyte-monocyte-colony stimulating factor (GM-CSF) synergistically interacts with interleukin-2 (IL-2) in generating an efficient antigen-specific immune response. We evaluated the toxicity, antitumour activity and immunobiological effects of human recombinant (hr)-GM-CSF and hr-IL-2 in 25 cancer patients who subcutaneously (s.c.) received hr-GM-CSF 150 microg/day for 5 days, followed by hrIL-2 s.c. for 10 days and 15 days rest. Two of the most common side-effects were bone pain and fever. Of the 24 patients evaluable for response, 3 achieved partial remission, 13 experienced stable disease, and 8 progressed. Cytokine treatment increased the number of monocytes, dendritic cells (DC), and lymphocytes (memory T cells) in the peripheral blood and enhanced the antigen-specific immunoreactivity of these patients. Our results show that the hr-GM-CSF and hr-IL-2 combination is active and well tolerated. Its biological activity may support tumour associated antigen (TAA)-specific anticancer immunotherapy by increasing antigen presenting cell (APC) activity and T cell immune competence in vivo.

High-affinity HLA-A(*)02.01 peptides from parathyroid hormone-related protein generate in vitro and in vivo antitumor CTL response without autoimmune side effects.

Parathyroid hormone-related protein (PTH-rP), a protein produced by prostate carcinoma and other epithelial cancers, is a key agent in the development of bone metastases. We investigated whether the protein follows the self-tolerance paradigm or can be used as a target Ag for anticancer immunotherapy by investigating the immunogenicity of two HLA-A(*)02.01-binding PTH-rP-derived peptides (PTR-2 and -4) with different affinity qualities. PTH-rP peptide-specific CTL lines were generated from the PBMC of two HLA-A(*)02.01+ healthy individuals, stimulated in vitro with PTH-rP peptide-loaded autologous dendritic cells and IL-2. The peptide-specific CTLs were able to kill PTH-rP+HLA-A(*)02.01+ breast and prostate carcinoma cell lines. The two peptides were also able to elicit a strong antitumor PTH-rP-specific CTL response in HLA-A(*)02.01 (HHD) transgenic mice. The vaccinated mice did not show any sign of side effects due to cell-mediated autoimmunity or toxicity. In this study we describe two immunogenic and toxic-free PTH-rP peptides as valid candidates for the design of peptide-based vaccination strategies against prostate cancer and bone metastases from the most common epithelial malignancies.

Tumour-associated antigen (TAA)-specific cytotoxic T cell (CTL) response in vitro and in a mouse model, induced by TAA-plasmids delivered by influenza virosomes

We investigated influenza virosomes as a TAA-gene delivery system for use in TAA-directed anti-cancer vaccine therapy. An engineered plasmid (GC90) expressing the parathyroid hormone-related peptide (PTH-rP), a protein secreted by prostate and lung carcinoma cells, was included in influenza virosomes (GC90V). The ability of GC90V to elicit a PTH-rP-specific cytotoxic T cell (CTL) response was demonstrated in BALB/c mice immunised with intranasal (i.n.) GC90V+/-adjuvant subcutaneous (s.c.) interleukin-2 (IL-2). A PTH-rP-specific CTL response with antitumour activity was also demonstrated in human peripheral blood mononuclear cells (PBMC) stimulated in vitro with GC90V infected autologous dendritic cells (DC). These results provide a rationale for investigating GC90V in clinical trials of anticancer vaccine therapy.

Weekly Low-Dose Docetaxel in Advanced Hormone-Resistant Prostate Cancer Patients Previously Exposed to Chemotherapy

Objective: The aim of this study was to evaluate the activity and tolerability of docetaxel in patients with hormone-resistant prostate cancer previously exposed to chemotherapy. Methods: We enrolled 27 patients with hormone-resistant prostatic cancer that had progressed during first-line chemotherapy. The primary end-point was palliative response defined as a 2-point reduction in the 6-point present pain intensity scale, and an improvement in Karnofsky performance status of one 10-point category. The treatment consisted of weekly docetaxel 25 mg/m² body surface area administered by means of a 1-hour intravenous infusion with corticosteroid premedication. Results: The primary criterion of palliative response was met in 13 patients (48%) after eight treatment cycles; its median duration was 6 months (range 1–8). Mean global quality of life improved in 8 and 10 patients after respectively four and eight treatment cycles. After a median follow-up of 8 months, 21 patients had died: the median survival was 9+ months (range 2–18). Weekly docetaxel was very well tolerated: grade 3 neutropenia occurred in 1 patient and grade 3 anemia in 2. Conclusions: Weekly low-dose docetaxel is an effective and well-tolerated treatment for patients with hormone-resistant prostate cancer previously exposed to chemotherapy.

A novel biweekly pancreatic cancer treatment schedule with gemcitabine, 5-fluorouracil and folinic acid

Pancreatic adenocarcinoma is a common disease considered to be poorly responsive to antiblastic treatment. Recent clinical and preclinical results suggest that a combined treatment of gemcitabine (GEM), 5-flurouracil (5-FU) and folinic acid (FA) offers a clinical benefit in patients with advanced pancreas adenocarcinoma. The aim of this phase II clinical trial was to evaluate the antitumour activity and toxicity of a novel biweekly schedule of this combination in patients with pancreatic adenocarcinoma. A total of 42 patients received a 30 min infusion of FA (100 mg m−2) and 5-FU (400 mg m−2) (FUFA) on days 1–3, and GEM 1000 mg m−2 on day 1 every 15 days. We observed 13 objective responses (two complete, 11 partial) and 23 stable diseases. The median time to progression was 9.75 months (95% Confidence Interval (CI), 6.88–12.62) and the median overall survival was 13.10 months (95% CI 9.64–16.56). There were seven cases of each grade III gastroenteric and haematological toxicity. The GEM plus FUFA combination appears to be well tolerated and very active in patients with pancreatic carcinoma.

Weekly epirubicin in patients with hormone-resistant prostate cancer

The aim of this study was to investigate the benefit of weekly epirubicin in the treatment of metastatic hormone-resistant prostate cancer. One hundred and forty-eight patients with metastatic hormone-resistant prostate cancer received weekly 30-min intravenous infusions of epirubicin 30 mg m2 of body surface area. The primary end-point was palliative response, defined as a reduction in pain intensity and an improvement in performance status. The secondary end-points were the duration of the palliative response, quality of life and survival. Fifty-seven (44%) of the 131 evaluable patients met the primary criterion of palliative response after six treatment cycles and 73 (56%) after 12 cycles; the median duration of the response was 9 months (range 1–11). The median global quality of life improved in 52% of the patients after six cycles and in 68% after 12 cycles. The 12- and 18-month survival rates were respectively 56 and 31%, with a median survival of 13+ months (range 1–36). The treatment was well tolerated: grade 3 neutropenia was observed in 8% of the patients, grade 3 anaemia in 7%, and grade 3 thrombocytopenia in 3%. None of the patients developed grade 4 toxicity or congestive heart failure. Weekly epirubicin chemotherapy can lead to a rapid and lasting palliative result in patients with metastatic HRPC, and have a positive effect on the quality of life and survival.

Gemcitabine increases systemic 5-fluorouracil exposure in advanced cancer patients.

A number of recent clinical trials testing the combination of 5-fluorouracil (5-FU) and gemcitabine in patients with advanced pancreatic adenocarcinoma have shown a significant clinical response rate, but also significant toxicity. As the two antimetabolites may interact at several biochemical levels along their pathways of activation, we investigated whether gemcitabine (GEM) affects 5-FU pharmacokinetics in cancer patients. Thus, we compared 5-FU pharmacokinetics in two groups of patients with various cancers who received the same schedule of 5-FU and folinic acid (FUFA), with or without GEM. There was a significant increase in systemic (5-FU) exposure and toxicity in the FUFA plus GEM group. Our finding may be useful in designing future studies of the combination in order to reduce the occurrence of side-effects and to maximise the antitumour activity.

Weekly low-dose docetaxel in advanced non-small cell lung cancer previously treated with two chemotherapy regimens

BACKGROUND The role of salvage chemotherapy in advanced non-small cell lung cancer (NSCLC) is still controversial, but the recent development of a number of active antineoplastic agents has created new possibilities for disease management. The aim of this study was to evaluate the activity and safety of weekly docetaxel treatment in patients with advanced NSCLC previously treated with two chemotherapy regimens. PATIENTS AND METHODS The study involved 26 patients with histologically documented stage IIIB, IV or recurrent metastatic NSCLC previously treated with two non-taxane based-chemotherapy regimens. They all received docetaxel 25 mg/m(2)/week administered as a 1-h infusion in an outpatient setting with corticosteroid premedication. RESULTS Fourteen of the 26 patients (54%) had distant metastases and 12 (46%) chest-localised disease. Six patients (23%; confidence interval: 9.8-44.1%) showed a partial response, 8 (31%) stable disease, and 12 (46%) progressive disease. The median time to progression was 4 months (range 2-8), and the median survival was 7+ months (range 3-13+). There were no statistically significant differences between the global quality of life scores recorded at baseline and those recorded after subsequent cycles. The treatment was very well tolerated. CONCLUSION The results of this study suggest that weekly low-dose docetaxel is effective, well tolerated and maintains a relatively good quality of life in patients with advanced NSCLC previously exposed to two chemotherapy regimens.

In vitro generation of cytotoxic T lymphocytes against HLA-A2.1-restricted peptides derived from human thymidylate synthase.

Abstract 5-Fluorouracil (5-FU) is a pyrimidine antimetabolite active against colorectal carcinoma and other malignancies of the digestive tract. Over-expression or mutation of thymidylate synthase (TS), the target enzyme of the 5-FU metabolite, 5-fluorodeoxyuridine monophosphate, is strictly correlated with cancer cell resistance to 5-FU. On this basis we investigated whether TS is a potential target for active specific immunotherapy of human colon carcinoma, which acquires resistance to 5-FU. Three TS-derived epitope peptides which fit defined amino acid consensus motifs for HLA-A2.1 binding were synthesized and investigated for their ability to induce human TS-specific cytotoxic T cell (CTL) responses In Vitro. CTL lines specific for each peptide were established by stimulating peripheral blood mononuclear cells (PBMC) from an HLA-A2.1 + healthy donor with autologous dendritic cells loaded with TS peptide. Specific CTL lines showed HLA-A2.1-restricted cytotoxicity In Vitro to HLA-A2.1+ target cells pulsed with the specific TS peptide and to HLA-class I matching colon carcinoma target cells over-expressing TS enzyme after exposure to 5-FU. Recognition by CTL lines suggests that these TS peptides may be potential candidates for use in a peptide-based vaccine against 5-FU resistant colon carcinoma.