Biomedical subjects

F Ravagnani

Publications and source records attributed to F Ravagnani.

At least 37 records · Page 2Linked to original sources

PubMed1996

Elimination of bcl-2-IgH-positive follicular lymphoma cells from blood transplants with high recovery of hematopoietic progenitors by the miltenyi CD34+ cell sorting system.

Contamination of autologous blood cell transplants with cells of follicular non-Hodgkin's lymphoma (F-NHL) may contribute to relapse of the malignancy after potentially curative high doses of chemotherapy and radiotherapy. In an attempt to circumvent this limitation, we have evaluated various techniques of selection of CD34+ cells to eliminate malignant cells from blood cell transplants of five patients with F-NHL undergoing high-dose sequential therapy. The contamination of F-NHL cells was evaluated using a nested PCR assay for the detection of bcl-2-IgH rearrangement with a sensitivity of one F-NHL cell in 10(5) normal cells. In two experiments with blood cell transplant fractions of 0.5 x 10(9) nucleated cells, negative selection of CD34+ cells by removal of B cells and other mature cells that naturally adhere to nylon wool fibers decreased the number of CD19+ B cells detectable by flow cytometry but failed to eliminate bcl-2-IgH-positive F-NHL cells detectable by PCR. In contrast, positive selection of CD34+ cells by the Miltenyi MiniMACS high gradient magnetic cell sorting system in five separate experiments resulted in: (1) the elimination of F-NHL cells in four out of five cases as detected both flow cytometry and bclk-2-IgH PCR; (2) a highly purified population of hematopoietic progenitors comprising 90.8% +/- 2.3% CD34+ cells; and (3) the recovery of 77.9% +/- 3.2% CD34+ cells. These favorable results were confirmed on a large-scale with a blood cell transplant comprising 5.8 x 10(9) nucleated cells in which positive selection of CD34+ cells by the Miltenyi SuperMACS system resulted in: (1) the elimination of F-NHL cells as detected both by flow cytometry and bcl-2-IgH PCR; (2) a highly purified population of hematopoietic progenitors comprising 94.6% CD34+ cells; and (3) the recovery of 62.7% CD34+ cells. These results, attained with the newly available Super MACS system, compare favorable with previous techniques because they show the feasibility of eliminating F-NHL cells from blood cell transplants without relevant nonspecific loss of hematopoietic progenitors.

Adsorption↗

PubMed1996

Lack of T-cell-mediated recognition of the fusion region of the pml/RAR-alpha hybrid protein by lymphocytes of acute promyelocytic leukemia patients.

In previous studies, it was shown that the fusion region of the pml/RAR-alpha protein, expressed by acute promyelocytic leukemia (APL) cells, can be specifically recognized in vitro by donor (D. E. ) CD4 T cells in a HLA class II DR11-restricted fashion. We present here the results on the recognition of several pml/RAR-alpha peptides by APL patients expressing HLA DR11. The in vitro immunization of peripheral blood lymphocytes from four patients in remission (S. R., F. R., M. M., P. G.) with BCR1/25, a 25-mer pml/RAR-alpha, did not elicit either a polyclonal or a clonal immune response specific to the peptide. We then generated new donor anti-pml/RAR-alpha CD4(+) T-cell clones. These clones were tested for their recognition of BCR1/25. One clone (C3/5, CD3(+), CD4(+), CD8(-)) was selected for further analysis. Clone C3/5 showed specific proliferation, cytotoxicity, and cytokine (tumor necrosis factor alpha, granulocyte-macrophage colony-stimulating factor) production when challenged with autologous lymphoblastic cell lines pulsed with peptide BCR1/25. C3/5 cells developed specific proliferation and cytotoxicity when challenged with peptide-pulsed lymphoblastic cell lines and peripheral blood lymphocytes from the four DR11(+) APL patients. APL blasts, available only from patients F. R. and P. G., were not lysed by C3/5 and were unable to present peptide BCR1/25. Incubation of APL cells with IFN-gamma failed to induce HLA class II molecules and recognition by the C3/5 clone. Since APL cells do not express HLA class II molecules, we tested in two donors (D. E. and C. H. R.) and in patients S. R. and P. G. whether the use of 9-mer peptides (BCR1/9) would generate a CD8/HLA class I-restricted response. No peptide-specific T-cell line or clone could be generated from both donors and patients. These findings are discussed in relation to possible therapeutic approaches to the immunotherapy of APL.

Binding Sites↗

PubMed1995

Adjuvant therapy with essential fatty acids (EFAs) for primary liver tumors: some hypotheses.

Hepatocarcinoma is responsible for approximately 1 million deaths annually. It is usually discovered at an advanced stage and, if inoperable, has a poor prognosis. New therapies combining chemotherapy, hyperthermia, radiotherapy and immunomodulators have been recently attempted with various levels of success. Once the tumor is detected at an early stage, some possibilities of cure seem to emerge either by intratumoral percutaneous injection (PEI) of alcohol or by chemoembolization and interstitial hyperthermia. When the tumor volume is more than 5 cm, these therapies are less successful and radiotherapy can be used. All the techniques described have some limits; PEI, for instance, does not achieve a complete eradication of lesions > 3 cm and a non-homogenous alcohol distribution within the tumor leads to areas of necrosis. Radiotherapy, even if effective, is limited by dose-related radiation hepatitis. Another important limiting factor is the incomplete response to therapy and tumor recurrence. Essential fatty acids, especially gamma linolenic acid (GLA) and eicosapentaenoic acid (EPA) are discussed here for their ability to control primary tumor proliferation and increase response to chemotherapy, radiotherapy and hyperthermic treatment, thanks to their effects on cellular membranes (increased lipoperoxidation and modification of tumor stroma).

Animals↗

PubMed1995

Massive ex vivo generation of functional dendritic cells from mobilized CD34+ blood progenitors for anticancer therapy.

We report that blood cell autografts, collected by single leukapheresis in cancer patients (n = 11) at the time of mobilization of hematopoietic progenitors into peripheral blood following anticancer therapy with high-dose cyclophosphamide (HD-CTX) plus interleukin-3 (IL-3) and granulocyte colony-stimulating factor (G-CSF/filgrastim), comprise 1.98 +/- 0.39 x 10(5)/kg (mean +/- SE) CD34+ progenitors of dendritic cells (DCs). This number corresponds to 140-fold more progenitors than in a control autograft collected in the steady state. DCs derived from mobilized CD34+ cells, morphologically and immunophenotypically undistinguishable from skin Langerhans cells and DCs from bone marrow and cord blood CD34+ cells, are shown to be powerful stimulators of allogeneic T cell proliferation in primary MLR and of autologous HLA-DR-restricted CD4+ T cell proliferation in response to presentation of xenogenic antigens. We show that the GM-CSF-plus-TNF-alpha-dependent ex vivo generation of DCs from mobilized CD34+ cells is 2.5-fold enhanced by flk-2/flt-3 ligand or c-kit ligand (stem cell factor) and five-fold enhanced by a combination of these growth factors. In addition, the optimal serum for the generation of DCs is autologous HD-CTX recovery-phase serum rather than fetal calf serum (FCS) or steady-state human serum, which are clinically inadequate and ineffective, respectively. In practice, the stimulation of CD34+ cells in a blood cell autograft (15.75 +/- 2.46 x 10(6)/kg) provided by the above four growth factors should permit ex vivo generation of approximately 40 x 10(9) DCs in an adult patient. These new findings provide advantageous tools for the large-scale generation of DCs that are potentially usable for clinical protocols of immunotherapy or vaccination in patients undergoing cancer treatment.

Adult↗

PubMed1994

Large-scale enrichment of mobilized CD34+ peripheral blood hematopoietic progenitors by removal of nylon wool-adherent mature cells.

With the aim of facilitating the ex vivo manipulation of peripheral blood hematopoietic progenitors (CPCs = circulating progenitor cells) collected by leukapheresis, we removed polymorphonuclear cells and monocytes that naturally adhere to nylon wool fibers. Leukapheresed cells harvested at the time of hematopoietic recovery after cancer therapy with high-dose cyclophosphamide plus hematopoietic growth factors were incubated with nylon wool fibers for 1 h at 37 degrees C. Evaluation of the cells non-adherent to the nylon wool in all experiments (n = 14) showed that the median recovery of nucleated cells and CPCs detected as CD34+ cells, CFU-GM and BFU-E was 16.4% (range 4.8%-34.0%), 60.0% (range 30.8-80.8%), 60.9% (range 33.4-74.5%) and 65.5% (range 30.8-69.2%), respectively. Therefore exposure to the nylon wool determined a selective removal of mature cells and a complementary enrichment of CPCs. The wide range of results depended on the significantly different cell compositions of the unmanipulated leukaphereses. The latter from patients receiving rhG-CSF (n = 10) comprised a median of 88.5% (range 77.8-93.8%) and 11.5% (range 6.2-22.2%) polymorphonuclear and mononuclear cells, respectively. In contrast, leukaphereses from patients receiving rhGM-CSF or PIXY321 (n = 4) comprised a median of 71.1% (range 55.4-85.0%) and 28.9% (range 15.0-44.6%) polymorphonuclear and mononuclear cells, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Antigens, CD↗

PubMed1993

Curability of advanced Burkitt's lymphoma in children by intensive short-term chemotherapy.

The treatment programme (regimen I) we designed in 1982 for advanced Burkitt's lymphoma was modified in 1986 as regimen IIA and IIB for patients presenting without or with bone marrow (BM) and/or nervous system involvement, respectively. Following a 5-week course of cytoreductive chemotherapy, including vincristine (VCR), cyclophosphamide (CPM), doxorubicin (DXR), high-dose methotrexate (HDMTX) and intrathecal methotrexate and cytarabine (ARAC), high-dose ARAC and cisplatin were given as a 4-day continuous infusion. Regimen I continued with an additional 3-week course including VCR, CPM, DXR and HDMTX, which was omitted in regimen IIA. In regimen IIB the initial cytoreductive chemotherapy was complemented by adding etoposide and increasing HDMTX doses, and by modifying the high-dose ARAC administration modality and was followed, once the bone marrow had recovered, by ifosfamide that concluded the programme. A total of 44 children (22 in regimen I and 22 in regimens IIA and IIB) were treated, with an overall response rate of 98%. 4 patients died as a result of treatment related complications. Survival, progression-free and event-free survival rates were 73, 70 and 63%, respectively, for regimen I, and 82, 90 and 82%, respectively, for regimen IIA and IIB. A short chemotherapeutic regimen, using alternating phase-specific and non-specific agents, is able to cure the majority of patients with advanced Burkitt's lymphoma.

Adolescent↗

PubMed1993

Recombinant human interleukin-3 hastens trilineage hematopoietic recovery following high-dose (7 g/m2) cyclophosphamide cancer therapy.

BACKGROUND: Interleukin-3, a recombinant cytokine with multilineage stimulatory effect on hematopoietic cells, was administered to 22 previously untreated breast cancer patients following high-dose therapy with cyclophosphamide (7 g/m2). PATIENTS AND METHODS: The growth factor, administered through continuous intravenous infusion at 1 (3 patients), 2.5 (3 patients), 5 (10 patients) and 10 micrograms/kg/day (6 patients), was well tolerated up to 5 micrograms/kg/day. RESULTS: Nausea, vomiting, fever and headache prevented administration of the intended dose to all 6 patients in the 10 micrograms/kg/day cohort. At the maximal tolerable dose (5 micrograms/kg/day) the growth factor significantly accelerated granulocyte, platelet and reticulocyte recovery as compared to matched historical controls who received high-dose cyclophosphamide without cytokine infusion. Moreover, no platelet transfusions and fewer erythrocyte transfusions were required in interleukin 3-treated patients. In contrast to GM-CSF and G-CSF, interleukin 3 showed no effect on the mobilization of hematopoietic progenitor cells in the peripheral blood. CONCLUSIONS: Interleukin-3 represents a well-tolerated cytokine, clinically useful for accelerating trilineage hematopoietic recovery following severely myelotoxic treatments such as high-dose cyclophosphamide.

Adolescent↗

PubMed1993

Clinical application of growth factors for collection of circulating hematopoietic progenitors in breast cancer patients treated with high-dose cyclophosphamide.

Seventy-seven (68 operable breast cancer with > 9 metastatic axillary nodes and 9 inflammatory breast cancer) entered this study. During hematopoietic recovery after cancer therapy with high-dose cyclophosphamide (7 g/m2; HD-CTX) circulating hematopoietic progenitors were collected by leukapheresis (LK) in all patients and then cryopreserved for autologous transplantation. Following HD-CTX, 70 patients were treated with hematopoietic growth factor(s) for 14 days: 38 with rhGM-CSF (group a), 16 with rhIL-3 (group b), 11 with sequential rhIL-3 and rhGM-CSF (group c), 5 with sequential rhIL-3 and rhG-CSF (group d). Seven control patients (group e) did not receive any growth factor. Leukaphereses, carried out over 2-4 consecutive days per patient, were started earlier in group c and in group d patients (mean day: +12 after HD-CTX). The sequential administration of rhIL-3 and rhG-CSF (group d) resulted in clearly higher yield of CFU-GM and CD34+ cells per leukapheresis (65.9 x 10(4)/Kg versus 20.9 x 10(6)/Kg, respectively) if compared with other groups of treatment.

Adult↗

PubMed1993

Quantization of CD34+ peripheral blood hematopoietic progenitors for autografting in cancer patients.

After myeloablative regimens, combined reinfusion of peripheral blood hematopoietic circulating progenitor cells (CPC) and bone marrow, yields a very rapid hematopoietic recovery. Therefore, based on the knowledge that CPC express the CD34 and CD33 differentiation antigen, we have developed a direct immunofluorescence flow cytometry assay to detect the peak of CPC in the peripheral blood of patients treated with high dose chemotherapy and growth factors. This assay, compared to CFU-GM assay, has the following advantages: 1) easy to do 2) standardized method 3) real time information on CPC number. This work illustrates the practical aspects of this assay and substantiate the widespread use of the CD34/33 flow cytometry assay to guide harvesting of circulating hematopoietic progenitors for autologous transplantation.

Antigens, CD↗

PubMed1991

Flow cytometry for clinical estimation of circulating hematopoietic progenitors for autologous transplantation in cancer patients.

Optimum methods of harvesting circulating hematopoietic progenitors for autologous transplantation to support myeloablative cancer therapy are still uncertain, mostly because of the lack of an assay for marrow-repopulating stem cells. The CFU-GM assay, the commonly used indirect indicator of the quality of the graft, is poorly standardized and provides results evaluable only retrospectively. Based on the knowledge that hematopoietic progenitors express CD34 and CD33 differentiation antigens, we developed a dual-color direct immunofluorescence flow cytometry assay with the aim of replacing the CFU-GM assay advantageously. For this purpose, we applied both assays to 157 blood samples obtained daily throughout 20 different recoveries from pancytopenia induced by high-dose cyclophosphamide or etoposide cancer therapy with or without recombinant human GM colony-stimulating factor (rhGM-CSF). The appearance of CD34+ cells in the circulation indicated that hematopoietic progenitors had increased to more than 500 CFU-GM/mL, a level clinically adequate for large-scale harvest by leukapheresis. Total CD34+ cells correlated well with CFU-GM (r = .89), and data could be fitted by a linear regression line described by the equation y = 388.3 + 64.0x, where y = CFU-GM/mL and x = CD34+ cells per microliter. Moreover, in a series of six patients treated with myeloablative chemoradiotherapy, early hematopoietic recovery of marrow functions was predicted more accurately by the number of transplanted CD34+/CD33+ cells than by either total nucleated cells, CFU-GM, CD34+/CD33- cells, or CD34-/CD33+ cells. Data presented in this article favor clinical use of the CD34/CD33 flow cytometry assay to guide harvesting of circulating hematopoietic progenitors for autologous transplantation and contribute to better understanding of the role played by circulating hematopoietic progenitor cell subsets in marrow recovery after myeloablative cancer therapy.

Antigens, CD↗

PubMed1991

Methodologies to estimate circulating hematopoietic progenitors for autologous transplantation in cancer patients.

Optimal criteria for harvesting circulating hematopoietic progenitors (CHP) for autologous transplantation to support myeloablative cancer therapy are still uncertain mostly because the CFU-GM assay, the commonly used indirect indicator of the hematopoietic recovery of the graft, is poorly standardized and provides information evaluable only retrospectively. Based on the knowledge that CHP express CD34 and CD33 differentiation antigens and facilitated by the availability of a very efficient fluorescein-conjugated CD34 antibody (8G12), we developed a direct immunofluorescence flow cytometry assay with the aim of replacing the CFU-GM assay advantageously. Recently, in a comparative study, both assays were applied to 157 blood samples obtained daily throughout 20 different recoveries from pancytopenia induced by high-dose cyclophosphamide (7 g/m2) cancer therapy w/ or w/o rhGM-CSF. Results showed that: a) detectability of CD34+ CHP indicated an increase to greater than 500 CFU-GM/mL, a level clinically adequate for harvesting CHP; b) CD34+ cells correlated well with CFU-GM (R=0.89) and data fitted a linear regression line (y=388.3 + 64.0x; y=CFU-GM/mL and x=CD34+/uL); c) in a series of 8 patients treated with myeloablative chemoradiotherapy, early recovery of marrow functions was predicted more accurately by the number of transplanted blood CD34+/CD33+ cells than by nucleated cells, CFU-GM, CD34+/CD33-cells, or CD34-/CD33+ cells. As a guideline, provided platelets are greater than 70,000/uL, harvest of CHP by leukapheresis during recovery from chemotherapy induced pancytopenia should be started as soon as CD34+ cells appear in the circulation and continued until the threshold dose of 7.8x10(6) CD34+ cells/kg, equivalent to 50 x 10(4) CFU-GM/kg, is achieved.(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies, Monoclonal↗

PubMed1990

Local adoptive immunotherapy of advanced head and neck tumors with LAK cells and interleukin-2.

Since October 1987 a pilot phase I-II study on the effect of loco-regional injections of recombinant interleukin 2 (rIL-2) in association with LAK cells has been performed in advanced, recurrent head and neck cancer patients. Fourteen patients were treated with autologous LAKs and rIL-2 (Glaxo) given peritumorally and in the mastoid region (rIL-2 only in the latter site). LAKs (2-70 x 10(7)) + rIL-2 were injected on the first day of therapy, followed by 9 daily injections of rIL-2 only. The total daily dose of rIL-2 was escalated from 2,400 to 1.8 x 10(6) IU. Clinical evaluation was performed 30 days from the onset of therapy; 3 partial (95%, 66% and 50% reduction) and 3 minor responses were seen in the evaluated patients. All the other patients with a progressive disease after the first cycle were shifted to palliative chemotherapy. The partial responses were found in patients with a tumor burden less than 20 cm2. Cervical node metastasis did not respond to treatment. No relevant side effects occurred. These results indicate that loco-regional immunotherapy with rIL-2 and LAK cells can produce clinical responses in advanced head and neck cancer patients.

Aged↗

PubMed1990

Large-scale collection of circulating haematopoietic progenitors in cancer patients treated with high-dose cyclophosphamide and recombinant human GM-CSF.

Circulating haematopoietic progenitors from 36 cancer patients were collected by continuous-flow leukapheresis during the phase of rapid haematopoietic recovery after pancytopenia induced by high-dose cyclophosphamide and then cryopreserved for autologous transplantation. 20 of the patients also received intravenous infusion of recombinant human granulocyte macrophage-colony stimulating factor (rhGM-CSF) for 7, 10 or 14 days after cyclophosphamide. 106 leukapheresis procedures were done for 2-5 consecutive days. Leukapheresis was started significantly earlier in patients receiving rhGM-CSF. In these patients, yields of peripheral blood elements (leucocytes, mononuclear cells, haematopoietic progenitors and platelets) were significantly higher than in controls treated with cyclophosphamide only. In particular, the mean number of granulocyte-monocyte colony-forming cells was 43.88 X 10(4) vs. 6.16 X 10(4) per kg patient body weight per leukapheresis. Side-effects of leukapheresis were limited to central venous catheter occlusion and fever in 4% and 2% of all procedures, respectively.

Adult↗

PubMed1989

Circulation of CD34+ hematopoietic stem cells in the peripheral blood of high-dose cyclophosphamide-treated patients: enhancement by intravenous recombinant human granulocyte-macrophage colony-stimulating factor.

We report that hematopoietic progenitor cells expressing the CD34 antigen (CD34+ cells) transiently circulate in the peripheral blood (PB) of cancer patients treated with 7 g/m2 cyclophosphamide (HD-CTX) with or without recombinant human granulocyte macrophage-colony stimulating factor (rHuGM-CSF). In adult humans, CD34+ cells represent a minor fraction (1% to 4%) of bone marrow (BM) cells, comprising virtually all hematopoietic colony-forming progenitors in vitro and probably also stem cells capable of restoring hematopoiesis of lethally irradiated hosts. We show that CD34+ cell circulation is fivefold enhanced by rHuGM-CSF 5.5 protein micrograms/kg/day by continuous intravenous infusion for 14 days after HD-CTX. During the third week after HD-CTX (ie, when CD34+ cells peak in the circulation), large-scale collection of PB leukocytes by three to four continuous-flow leukaphereses allows the yield of 2.19 to 2.73 x 10(9) or 0.45 to 0.56 x 10(9) CD34+ cells depending on whether or not patients receive rHuGM-CSF. The number of CD34+ cells retrieved from the circulation by leukaphereses exceeds the number that can be harvested by multiple BM aspirations under general anesthesia. Thus, after therapy with HD-CTX and rHuGM-CSF, PB represents a rich source of hematopoietic progenitors possibly usable for restoring hematopoiesis after myeloablative chemoradiotherapy. To determine whether CD34+ cells found in the PB are equivalent to their marrow counterpart, we evaluated their in vitro growth characteristics and immunological phenotype by colony assays and dual-color immunofluorescence, respectively. We show that PB CD34+ cells possess qualitatively normal hematopoietic colony growth and high cloning efficiency comparable to that observed with BM CD34+ cells. In addition, PB CD34+ cells display heterogeneous surface membrane differentiation antigens analogous to BM CD34+ cells. The availability of large quantities of CD34+ cells by leukapheresis is relevant to the field of stem cell transplantation and possibly to genetic manipulations of the hematopoietic system in humans.

Antigens, CD34↗

PubMed1989

Differences between in vivo and in vitro activation of cancer patient lymphocytes by recombinant interleukin 2: possible role for lymphokine-activated killer cell infusion in the in vivo-induced activation.

In this study 15 consecutive melanoma patients were treated with two courses of bolus recombinant interleukin 2 (rIL2) and rIL2 plus in vitro-generated lymphokine-activated killers (LAK), respectively. The immunological monitoring performed after 4 days of rIL2 or rIL2 plus LAK, indicate that the in vivo peripheral blood lymphocyte (PBL), activation (spontaneous proliferation, tumor cytotoxicity, number of DR+ PBL, obtained after the second cycle of rIL2 plus LAK is significantly higher than after the first cycle of rIL2 alone. During the 5-day interval between the two courses, PBL activation returns to baseline levels and no evidence for increased sensitivity of PBL to rIL2 is present. To further confirm this, two additional patients were studied, in whom rIL2 was administered by continuous i.v. infusion. In these two patients the in vitro versus in vivo PBL activation could be directly and simultaneously compared by using in vitro the same concentration of rIL2 reached and maintained in the patients' sera. The PBL activation induced in vivo by a cycle of rIL2 alone was significantly less (about 10 times) than that obtained in vitro with a comparable rIL2 concentration. Thus, the infusion of in vitro highly activated PBL could explain the increased in vivo lymphocyte activation of the second cycle of rIL2 plus LAK over the first cycle of rIL2 alone.

Cell Line↗

PubMed1988

A new procedure for large scale production and freezing of lymphokine activated killer (LAK) cells to be used in adoptive immunotherapy of cancer.

A new procedure for activation of peripheral blood lymphocytes (PBL) with recombinant interleukin 2 (rIL2) is described. PBL obtained by leukapheresis were subjected to NH4Cl (ACK) treatment to clear erythrocyte contamination; Ficoll separation was not performed. PBL were subsequently seeded in 10-floor multitrays (Cell FactoryTM, CF), gasified and incubated at 37 degrees C for 3-4 days in a humidified 5% CO2 atmosphere. This procedure achieved an activation (evaluated as cytotoxicity and proliferation) comparable with that obtained by culturing PBL in small flasks. Optimal activation of PBL was achieved in CF even in the presence of granulocyte contamination of up to 40%. It was also possible to freeze, thaw and recover most of the frozen cells and their cytotoxic activity. With this procedure therefore large quantities of lymphokine activated killer cells (LAK) can be easily produced to be used in adoptive immunotherapy trials.

Blood Preservation↗

PubMed1988

Diversity of effects of two antitumor anthracycline analogs on the pathway of activation of PKC in intact human platelets.

Two antitumor antibiotics doxorubicin and daunorubicin were tested for their ability to influence the activation of protein kinase C in human platelets. Daunorubicin was found to inhibit the phosphorylation of the 40 K PKC substrate induced by thrombin and 12-O-tetradecanoyl-phorbol-13-acetate as well as the phosphorylation of the 20 K protein induced by thrombin. The serotonin release associated to these phosphorylative events was also inhibited by daunorubicin. In contrast the effects of doxorubicin, though inhibitory on the release reaction, were always stimulatory of the phosphorylations. Doxorubicin alone was able to induce the phosphorylation of both 40 K and 20 K phosphoproteins in a concentration-dependent manner. Whereas the stimulation by doxorubicin was not influenced by pretreatment with dibutyryl-cyclic-AMP which inhibits the effects of thrombin, this effect was inhibited by daunorubicin, neomycin and stimulated by the diacylglycerol-kinase inhibitor R 59 022. It is proposed that doxorubicin activates the protein kinase C by causing the breakdown of phosphoinositides.

Blood Platelets↗

PubMed1984

Inhibition of human melanoma growth in nude mice by autologous, alloactivated peripheral blood lymphocytes.

Peripheral blood lymphocytes of melanoma patients were stimulated in vitro by a pool of allogeneic lymphocytes and shown to be cytotoxic against autologous melanoma cells. To evaluate the in vivo antitumor activity of the cytotoxic alloactivated autologous peripheral blood lymphocytes, tumor neutralization (Winn) assay was carried out by injecting such lymphocytes admixed with autologous melanoma cells in athymic BALB/c nude mice. In 3 of 6 cases, complete inhibition of tumor growth was obtained at lymphocytes to tumor cells ratio of 10:1 and in one case also of 5:1. In all cases the appearance of tumors was delayed and the growth rate was significantly reduced in a dose-dependent fashion as compared to control mice injected with tumor cells alone. We conclude that in vitro alloactivated peripheral blood lymphocytes can inhibit and/or impair the growth of autologous melanoma cells in nude mice.

Animals↗