Andrea Piga

Rapid down-regulation of protein kinase C and membrane association in phorbol ester-treated leukemia cells

Peripheral blood lymphocytes from patients with chronic lymphocytic leukemia (CLL) acquire after several days of exposure to 12‐O‐tetradecanoylphobol‐13‐acetate (TPA) several morphological, immunological and histochemical features of hairy cell leukemia. We have investigated the short term effects of TPA treatment on protein kinase C and its subcellular distribution. Within minutes of addition of TPA to CLL cells 20% of the cytosolic protein kinase C had associated with the particulate fraction. The remaining 80% of protein kinase C activity was down‐regulated. The association with the membrane dramatically increased the resistance of the enzyme to inhibition by the non‐ionic detergent, Triton X‐100. These results suggest that activation of protein kinase C causes multiple biological changes in CLL cells.

Cytotoxic effects of the lipophilic iron chelator omadine

Cytotoxic effects were observed following 4 h incubation of human leukaemic cells with the iron chelator 1‐hydroxypyridine‐2‐thione (omadine). Its Cytotoxic activity was comparable to that of the cytotoxic drug doxorubicin. At the same concentration two other effective iron chelators, desferrioxamine and 1,2‐dimethyl‐3‐hydroxypyrid‐4‐one, were not cytotoxic. Addition of iron augmented the effect of omadine. It is suggested that the lipophilic properties of omadine and of its iron complex cause their intracellular accumulation and potent cytotoxic activity.

Predictors of short-term survival and progression to chemotherapy in patients with advanced colorectal cancer treated with 5-fluorouracil-based regimens.

The aim of this study was to assess in patients with advanced colorectal cancer which factors were associated with short-term survival (6 months or less) and progression to first-line 5-fluorouracil (5-FU) chemotherapy. Three hundred twenty-one consecutive nonselected patients with advanced colorectal cancer were treated with conventional 5-FU-based regimens as first-line treatment from 1988 to 1999. Factors related to patient, tumor, or treatment were analyzed by univariate and multivariate logistic regression analysis by comparing short survivors (SS, those who survived ≤6 months) with those who survived longer than 6 months. The same statistical methods were used to analyze 200 patients, all treated with bolus 5-FU regimens, by comparing who progressed to treatment with those who did not. Sixty-two patients (19.3%) were SS, the remaining 259 patients survived more than 6 months. First-line chemotherapy included 5-FU in all patients; 112 (35%) and 27 (8.4%) patients were offered, after disease progression, second and third-line chemotherapy, respectively. The overall response rate to first-line chemotherapy was 12.9%. No SS patient achieved an objective response. To investigate factors associated with progression to first-line chemotherapy, we considered only those patients treated with bolus 5-FU regimens, to eliminate the variable of regimen used. Ninety-six of them progressed to treatment and 104 did not. At multivariate analysis, SS patients were characterized by the following: right and transverse colon primary (p = 0.006), younger age (p = 0.043), poor performance status (Eastern Cooperative Oncology Group ≥ 2) (p = 0.015), elevated (≥5 &mgr;g/l) serum carcinoembryonic antigen (CEA) (p = 0.015), and more than one site of metastatic disease (p < 0.001). Progression to first-line chemotherapy (p < 0.001) was also a strong factor associated with short survival in multivariate analysis; factors predictive of progression were elevated CEA (p = 0.027) and diffuse metastatic disease (p = 0.029). Our data indicate the relevance of some clinical prognostic factors (younger age, poor performance status, elevated CEA, site of primary, number of metastatic sites, resistance to chemotherapy) as independent factors associated with poor survival and progression to first-line chemotherapy in patients with metastatic colorectal cancer treated with conventional 5-FU regimens. Patients identified by these factors as having a poor prognosis and low probability of response to treatment should be considered either for more aggressive regimens or supportive care only: conventional 5-FU treatments do not impact on response or survival.

Assessing quality of life in patients with cancer: a comparison of a visual-analogue and a categorical model.

A simple instrument for self-assessment of quality of life (QL) in patients with cancer was elaborated using a linear analogue scale (LAS). The instrument was based on five questions, exploring different functional areas; the same questions were also addressed in a parallel format, where problems were seen from an opposite point of view (positive/negative). The LAS was given to 222 patients, for a total of 372 tests collected. Internal consistency was satisfactory (Cronbachs alpha = 0.75); QL score was significantly correlated to parameters of disease. Concordance between scales, as judged by comparison of parallel formats, was statistically significant but poor. A questionnaire was then elaborated with similar items, based on a categorical scale. A direct comparison between LAS and our questionnaire was made on a group of 41 patients. Internal consistency was poor for the LAS (alpha = 0.58) and good for the questionnaire (alpha = 0.93); Spearmans rank correlation coefficients were disappointing for the LAS and good for the questionnaire; the questionnaire was judged reliable in 82.9% of cases, the LAS in 29.3% only; the questionnaire score, and not the LAS score, was significantly correlated with PS and disease status. In conclusion, many patients appeared unable to correctly interpret the visual-analogue scale; the categorical scale was more immediate and correctly understood by the large majority of patients; the correlation between score and important parameters of QL was maintained, and internal consistency was excellent, indicating a satisfactory reliability of this instrument.

Higher tyrosine protein kinase activity in resting lymphocytes than in proliferating normal or leukaemic blood cells

We have studied tyrosine phosphorylation in particulate fractions from 11 leukaemic cell lines by using as substrate either a synthetic tyrosine containing peptide or the endogenous proteins. The results were compared with those obtained using similar fractions from normal lymphocytes and bone marrow cells. Particulate fractions from all the leukaemic cell lines and normal bone marrow cells exhibited lower levels of tyrosine protein kinase activity compared to normal lymphocytes. When the phosphorylation of endogenous substrates was assayed, proteins were phosphorylated on tyrosine residues (rather than serine or threonine residues) to a larger extent in normal lymphocytes than in leukaemic cell lines. Separation of labelled endogenous substrates on sodium dodecyl sulfate-polyacrylamide gels showed a number of phosphorylated alkali-resistant bands in the range 14-175kd in the lymphoid cell lines; normal lymphocytes exhibited a smaller number of strongly phosphorylated bands. Normal lymphocytes from different individuals showed reproducible patterns of phosphorylated substrates. Normal bone marrow cells and myeloid leukaemia lines showed weak, if any, phosphorylation. Among the leukaemic cell lines no particular pattern of phosphorylated substrates common to cells of similar phenotype could be detected. We suggest that the level of overall tyrosine protein kinase activity in these fractions reflects their position in the cell cycle rather than their normal or malignant status.

ATTITUDES OF NON-ONCOLOGY PHYSICIANS DEALING WITH CANCER PATIENTS. A SURVEY BASED ON CLINICAL SCENARIOS IN ANCONA PROVINCE, CENTRAL ITALY

Aims and Background With this study we attempted to determine to what extent recent acquisitions in clinical oncology had reached categories of physicians involved in the management of patients with cancer, namely general surgeons, internists and family doctors. Methods A questionnaire was prepared with scenarios based on the following clinical situations: Scenario A, Adjuvant therapy in colon cancer; Scenario B, Treatment of small-cell lung cancer; Scenario C, Adjuvant therapy in high-risk, node-negative breast cancer; Scenario D, Treatment of early stage breast cancer; Scenario E, Asymptomatic transient myelosuppression during chemotherapy. Questionnaires were mailed to 365 family doctors, 54 general surgeons and 61 internists of the Province of Ancona in central Italy. Results A total of 198 completed questionnaires were returned (41%). Respondents were 36.7% of family doctors, 54.1% of internists and 57.4% of surgeons. Less than half of respondents selected an adequate approach such as adjuvant chemotherapy for colon cancer and high-risk, node-negative breast cancer or chemotherapy as first-line treatment for small-cell lung cancer. Conservative surgery plus radiotherapy (QUART) for early stage breast cancer was indicated by 69% of respondents. Over three quarters of physicians would give treatment for asymptomatic transient chemotherapy-induced leukopenia. In most of the scenarios, significant differences were detected in the distribution of preferences according to category of physicians. Family doctors and young physicians (<40 years) generally performed worse than hospital-based physicians (general surgeons and internists) and older physicians. Conclusions Non-oncology physicians showed insufficient awareness of currently available knowledge in cancer treatment. Basic concepts in cancer management should be part of the professional knowledge of all medical doctors, and key advances in clinical oncology should spread outside the oncologic environment more promptly, with a wide circulation among all physicians who care for cancer patients.

Biochemical mechanisms of deoxycoformycin toxicity in chronic leukemias

The in-vitro effects of deoxycoformycin (dCF) on dATP, NAD, ATP and DNA strand breaks have been evaluated in the cells from 42 patients with various types of chronic lymphoid leukemia. These included 18 with B-cell chronic lymphoid leukemias of different types (BCL); 10 with hairy cell leukemia (HCL) and 14 with T-cell chronic lymphoid leukemias of different types (TCL). The dATP concentrations in HCL, BCL and TCL increased from means of 2.9, 1.8 and 3.0 to 100.3, 68.2 and 51.3 pmol/10(6) cells respectively after 2 h with 10(-5) M dCF and 10(-4)M deoxyadenosine. After 18-24 h, the NAD levels and total double-stranded DNA decreased to 37 and 12.5% (HCL) to 36 and 21.6% (BCL) and 40 and 20.5% (TCL) of control values respectively. Similar decreases were observed in ATP levels. The results do not suggest that these measurements in vitro would predict which patients with these disorders will respond to dCF therapy. Although HCL responds particularly well to dCF in vivo, no difference in the in-vitro effects of dCF studied here could be detected between cases of HCL and the other types of chronic leukemia.

Synchronous primary hepatic lymphoma and epidermoid lung carcinoma treated with chemotherapy and surgery.

While involvement of the liver by non-Hodgkins lymphoma is a relatively frequent event, primary liver lymphoma is an uncommon disease. We describe a case of synchronous primary hepatic lymphoma and epidermoid lung carcinoma occurring in a 61-year-old male patient. Complete remission of both diseases was achieved with a radical approach, which included combination chemotherapy and surgery. The patient has now been in persisting complete remission for 40 months after surgery.

Continuing medical education through the videotex system in Italy

Continuing education for medical doctors is not compulsory in Italy. The link with the university is lost shortly after the final medical examination, and there is no other teaching institution for structured continuing medical education (CME). Distance learning gives physicians the opportunity to use updating programs at home at their convenience. Videotel, the Italian videotex system, is the first telematic tool that, at low cost, can reach every home nationwide. Through this system information can be exchanged 24 hours a day, using the Videotel database as central memory, the telephone network as connecting system, and low-cost devices as peripherals. The authors evaluated the technical capacity and didactic efficacy of the Videotel system as a vehicle for CME (in both oncology and general medicine). In an exploratory phase they surveyed physicians of the Italian Province designated for the study, with the objective of promoting the initiative and enrolling physicians interested in this innovative approach to CME. Teachers at Italian universities provided the educational material: interactive lessons, clinical case discussions and problem solving, and multiple-choice questions. Twenty-nine physicians agreed to participate. Despite the interest shown by these physicians, they made very little use of the didactic database. The main reasons for failure to connect with the educational database were the lack of time and unfamiliarity with the instrument. Although the results of the study were discouraging, the authors believe that the resolution of technical problems linked with the system and an increasing familiarity of physicians with telematic and informatic tools in general, together with appropriate incentives, will make the videotex system a feasible, low-cost, efficient vehicle for CME.

Tyrosine protein kinases and their substrates in human leukemia cells.

We have quantitated tyrosine protein kinase (TPK) activity in particulate and cytosolic fractions from human leukemic cells. Slowly proliferating cells from patients with chronic lymphocytic leukemia (CLL) had levels of TPK similar to those of quiescent normal lymphocytes. Cells from patients with acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML) and chronic granulocytic leukemia (CGL) contained markedly lower levels of TPK activity, similar to the levels in phytohaemagglutinin-stimulated (proliferating) normal lymphocytes and in bone marrow cells. This suggested that TPK is part of a mechanism for transducing growth signals and is down-regulated following signal transmission. We also identified endogenous substrates for TPK in leukemic cells. Particulate fractions from ALL, CLL and AML cells contained substrates identical to those previously detected in normal lymphocytes. In particular, a 38kD substrate thought to be involved in early stages of growth signal transduction in normal lymphocytes was found in all samples of these groups examined. Cytosolic fractions from these groups of leukemia cells contained higher molecular weight substrates not found in resting or proliferating normal lymphocytes or bone marrow cells. In contrast, TPK substrates in both particulate and cytosolic fractions from CGL cells resembled those of normal bone marrow cells in that only proteins with molecular weight below 40kD were labelled on tyrosine. We conclude that leukemic cells do not contain higher levels of TPK than do normal hemopoietic cells. Qualitative differences in TPK species or in their substrates may result in aberrant regulation of proliferation in leukemic cells. However, we cannot exclude the possibility that additional TPK substrates detected in leukemic cells were a feature of the normal equivalent hematopoietic cells from which the leukemia cells were derived.