Gioacchino Robustelli della Cuna

High dose medroxyprogesterone acetate (MPA) treatment in metastatic carcinoma of the breast: a dose-response evaluation.

The results of a controlled clinical trial that used high doses of medroxyprogesterone acetate (MPA) in the treatment of metastatic breast cancer are reported. Two treatment regimens were used: regimen A, 500 mg daily with a total dose of 30 g; regimen B, 1,000 mg daily with a total dose of 60 g. The overall response rates were similar, with no statistically significant difference between the two treated groups. Regimen A (lower dosage group) reached a remission rate of 44%, whereas regimen B (higher dosage group) had a remission rate of 41%. The mean duration of response was 8 months with regimen A and 9 months with regimen B. The advantages of the lower dosage regimen as opposed to the higher dosage regimen of MPA in the treatment of advanced breast cancer are discussed.

Diverse expansion potential and heterogeneous avidity in tumor-associated antigen-specific T lymphocytes from primary melanoma patients.

While tumor‐associated antigen (TAA)‐specific CD8+ T lymphocytes have been detected in metastatic melanoma patients, immune response in early disease phases has not yet been carefully evaluated. We looked for circulating cytotoxic T lymphocytes (CTL) directed against Melan‐A / MART1, tyrosinase, gp100 and MAGE‐3 antigens in patients with a diagnosis of primary cutaneous melanoma by using fluorescent HLA‐A2 tetramers. In five out of six cases high numbers of CD8+ / tetramer+ cells could be detected by flow cytometry, and in four patients lymphocyte populations specific for two different melanoma antigens (Melan‐A / MART1 and tyrosinase) were contemporarily present. The TAA‐specific cells could represent as much as 1 / 220 T lymphocytes in the circulating CD8+ population. When tetramers were used to monitor the in vitro expansion of TAA‐specific CTL precursors upon antigen‐specific stimulation, a diverse expansion potential was evidenced in CTL from the different donors and, more strikingly, in CTL specific for the different TAA. Melan‐A / MART1‐specific CTL clones derived from two patients exhibited a broad range of avidity. Only the highest avidity clones, representing about 50 % of the cases analyzed, were tumor specific. By correlating tetramer staining with clone avidity, we found that tetramer fluorescence intensity could represent a good indicator of TCR affinity, but not of overall clone avidity.

Dominant TCR-α Requirements for a Self Antigen Recognition in Humans

TCR-α and -β chains are composed of somatically rearranged V, D, and J germline-encoded gene segments that confer Ag specificity. Recent crystallographic analyses revealed that TCR-α has more contacts with peptide than TCR-β, suggesting the possibility that peptide recognition predominantly relies on TCR-α. T cells specific for the self Ag Melan-A/MART-1 possess an exceptionally high precursor frequency in human histocompatibility leukocyte Ag-A2 individuals. This provided a unique situation for assessment of the structural relationship between TCR and peptide/MHC ligand at both the pre- and postimmune levels. Molecular and phenotypic analysis of many different Melan-A-specific T cell populations revealed that a structural constraint is imposed on the TCR for engagement with Melan-A peptides presented by HLA-A2, namely the highly preferential use of a particular TCRAV segment, AV2. Examination of CD8 single-positive thymocytes indicated that this preferential use in forming the Melan-A-specific TCR is mainly imposed by intrathymic positive selection. Our data demonstrate a dominant function of TCRAV2 segment in forming the TCR repertoire specific for the human self Ag Melan-A/MART-1 and support the view that Ag recognition is mediated predominantly by TCR-α.

T-cell dynamics after high-dose chemotherapy in adults: elucidation of the elusive CD8+ subset reveals multiple homeostatic T-cell compartments with distinct implications for immune competence.

Recovery of total T cell numbers after in vivo T‐cell depletion in humans is accompanied by complex perturbation within the CD8+ subset. We aimed to elucidate the reconstitution of CD8+ T cells by separate analysis of putative naïve CD95− CD28+, memory CD95+ CD28+ and CD28− T cell compartments after acute maximal depletion by high‐dose chemotherapy (HD‐ChT) in women with high‐risk breast cancer. We found that recovery of putative naïve CD8+ CD95− CD28+ and CD4+ CD95− CD28+ T cells, was compatible with a thymus‐dependent regenerative pathway since their recovery was slow and time‐dependent, their values were tightly related to each other, and their reconstitution patterns were inversely related to age. By analysing non‐naïve T cells, a striking diversion between putative memory T cells and CD28− T cells was found. These latter increased early well beyond normal values, thus playing a pivotal role in total T‐cell homeostasis, and contributed to reduce the CD4 : CD8 ratio. In contrast, putative memory T cells returned to values not significantly different from those seen in patients at diagnosis, indicating that this compartment may recover after HD‐ChT. At 3–5 years after treatment, naïve T cells persisted at low levels, with expansion of CD28− T cells, suggesting that such alterations may extend further. These findings indicate that CD28− T cells were responsible for ‘blind’ T‐cell homeostasis, but support the notion that memory and naïve T cells are regulated separately. Given their distinct dynamics, quantitative evaluation of T‐cell pools in patients undergoing chemotherapy should take into account separate analysis of naïve, memory and CD28− T cells.

Transfusion of platelet concentrates cryopreserved with ThromboSol plus low‐dose dimethylsulphoxide in patients with severe thrombocytopenia: a pilot study

We have recently reported the possibility of supporting the phase of severe thrombocytopenia after high‐dose chemotherapy (HDC) and stem cell transplantation using 5% dimethylsulphoxide (DMSO)‐cryopreserved autologous platelet concentrates (PCs). The aim of the present study was to evaluate the therapeutic potential of ThromboSol (a recently developed platelet storage solution) plus PCs cryopreserved in 2% DMSO in patients undergoing myeloablative chemotherapy and autologous transplantation. PCs were collected from 14 women with breast cancer by a single plateletapheresis and cryopreserved in ThromboSol/2% DMSO by either direct insertion in a −80°C freezer or in liquid nitrogen after computer‐controlled rate (CR) freezing. When required, PCs were thawed, centrifuged to remove the cryoprotectants and transfused. In vitro studies on thawed platelets showed loss of epitopes of surface glycoproteins and a marked reduction of functional activity compared with fresh platelets. Transfusion of CR‐frozen PCs was associated with a mean 1 h corrected count increment (CCI) of 9.2 ± 5.4 × 109/l and only one allogeneic PC was required in this group. In contrast, six out of seven patients required additional allogeneic transfusions in the −80°C group (CCI = 2.7 ± 1.4 × 109/l). ThromboSol‐treated PCs have the ability to overcome thrombocytopenia if processed by a CR freezing protocol, but appear ineffective when frozen by direct placing at −80°C.

Circulating CD33+ large mononuclear cells contain three distinct populations with phenotype of putative antigen‐presenting cells including myeloid dendritic cells and CD14+ monocytes with their CD16+ subset

BACKGROUND In peripheral blood, myeloid markers identify a heterogeneous mixture of cells in transit from the bone marrow to peripheral tissues. Similarly, HLA-class II DR expression usually identifies mononuclear cells with the potential for developing antigen-presenting activity. We gathered putative antigen presenting cells bearing myeloid markers (My-APC) to study their composition by cell surface phenotype. METHODS To gather and dissect My-APC phenotype while excluding lymphocytes and granulocytes, we developed a strategy based on staining red cell-lysed peripheral blood and gating cells bearing myeloid markers and physical parameters of large mononuclear cells. RESULTS Phenotypic analysis within the My-APC gate showed three distinct populations. The largest fraction was constituted by CD14+ monocytes that extended into the other two populations, each expressing gradually lower levels of CD14 surface antigen along with increasing levels of CD16 and CD2, respectively. The CD16 and CD2 expression patterns extended from CD16+CD14+ or CD2+CD14+ double- positive intermediate cells toward each single positive subset, but they were reciprocally exclusive. Interestingly, CD2+CD14- cells within the My-APC gate were equivalent to myeloid dendritic cell precursors (pre-DC) defined previously by the absence of lineage markers and expression of HLA-DR and myeloid markers. Phenotypic analysis of each population revealed differences in the expression of costimulatory molecules and CD62L. CONCLUSIONS This novel analytical approach allowed us to distinguish circulating My-APC in three subsets and to identify relationships between monocytes and other related myeloid populations including DC.

Combination Chemotherapy with Cyclophosphamide, Fluorouracil, and Either Epirubicin or Mitoxantrone: a Comparative Randomized Multicenter Study in Metastatic Breast Carcinoma

From February 1987 to January 1989, 60 patients with advanced breast cancer and no prior chemotherapy for advanced disease were randomized and studied, with 31 treated with fluorouracil, epirubicin, and cyclophosphamide (FEC) and 29 patients with fluorouracil, mitoxantrone, and cyclophosphamide (FNC). Doses were 500 mg/m2 fluorouracil, 500 mg/m2 cyclophosphamide, and 50 mg/m2 epirubicin2 or 10 mg/m mitoxantrone, i.v. Day 1 every 3 weeks. There were no statistically significant differences in pretreatment patient characteristics between the groups. Fifty-six patients were evaluable for response (29 in the FEC arm and 27 in the FNC arm). The response rates were 48.2% for the FEC group (complete response (CR) 10.3% and partial response (PR) 37.9%) and 40.7% for the FNC group (CR 3.7% and PR 37%) (not significantly different, NS). The median response duration was 247 and 267 days, respectively (NS), the median time to progression and time to treatment failure was 244 and 155.5 days for the FEC group and 86 and 98 days for the FNC group, respectively (NS). The incidence of nausea/vomiting was 87.1% in the FEC group and 79.3% in the FNC group, with comparable severity. Alopecia occurred in 80.6% of FEC patients and 44.8% of FNC patients (p less than 0.05). The incidences and degrees of severity of leukopenia, anemia, and cardiotoxicity were comparable in the two treatment groups. Efficacy and toxicity of the two regimens were quite similar. FNC can improve the quality of life of patients by providing significantly less alopecia.

Steroid receptor enhancement by natural interferon-β in advanced breast cancer

Abstract In the current study we investigated the effect of two different doses of natural interferon-beta (IFN-β) on steroid hormone receptors in 45 patients with advanced breast cancer. IFN-β seems to regulate the receptor mechanisms, inducing in cutaneous metastases an increase of oestrogen and progesterone receptors. Moreover, using IFN-β and tamoxifen as a combined therapy in 23 receptor-positive patients, no negative interference of the two drugs was observed and no relevant side-effects due to the treatment were noticed. The moduation of steroid receptor content by IFN-β in advanced breast cancer might represent an interesting way to ameliorate the clinical responsiveness to anti-oestrogens.

Clinical evaluation of 4′-epi-doxorubicin in advanced solid tumors

SummaryA Phase II clinical evaluation of 4′-epi-doxorubicin has been carried out in 100 patients with various types of solid tumors. Hematopoietic toxicity was dose-limiting but reversible and of mild to moderate degree. Other acute toxic manifestations such as vomiting and alopecia were qualitatively similar to those usually reported for doxorubicin, but lower in frequency and less severe. A number of responding patients received cumulative doses of 4′-epi-doxorubicin in excess of 500 mg/m2. One patient manifested reversible clinical congestive heart failure at cumulative dose of 1,080 mg/m2. Therapeutic activity has been observed in breast carcinoma, in rectal carcinoma and in melanoma. In chemoresistant tumors as rectal cancer and melanoma 4′-epi-doxorubicin deserves further study.

Effects of natural beta-interferon and recombinant alpha-2B-interferon on proliferation, glucocorticoid receptor content, and antigen expression in cultured HL-60 cells

In the current study, we investigated the effects of natural beta‐interferon (β‐IFN) and recombinant alpha‐2b‐interferon (α‐IFN) on the growth of the HL‐60 cell line. Cells cultured in a medium that contains various concentrations (from 10 to 1000 IU/ml) of interferons showed a growth inhibition, which reaches the maximum after a 6‐day treatment, at the highest dose used. Furthermore, we studied the effect of both β‐IFN and α‐IFN on the level of glucocorticoid receptors. This was enhanced more than 30% with respect to control in HL‐60 cells exposed for 24 hours to concentrations of β‐IFN that ranged from 100 to 1000 IU/ml. The increase of the receptor amount was seen even if cells were treated for 5 days, and was not accompanied by a modification of antigen expression of HL‐60 cells. α‐IFN did not modify the glucocorticoid receptor level substantially in our experimental conditions. Our data indicate that both β‐IFN and α‐IFN regulate HL‐60 cell proliferation. Additional studies are required to clarify if modifications of the receptor level induced by β‐IFN could be related to the modulation of hormone‐sensitivity in this model.