A model of in vivo purging with Rituximab and high-dose AraC in follicular and mantle cell lymphoma.

We studied a model of in vivo purging with Rituximab and high-dose (HD) cytarabine in 14 patients with relapsed/refractory follicular lymphoma and two with refractory mantle cell lymphoma enrolled in a program of HD chemotherapy and autotransplant. After two courses of debulking immunochemotherapy with Rituximab, Vincristine and Cyclophosphamide, we used a combination of Rituximab, HD cytarabine and granulocyte colony-stimulating factor for peripheral blood stem cells (PBSC) mobilization. The median number of CD34+ cells collected was 14.69 x 10(6)/kg (range 5.74-73.2). Monitoring of peripheral CD19+ and CD20+ B cells prior to and throughout the purging period showed that a treatment with Rituximab, Vincristine and Cyclophosphamide results in a profound depletion of B cells in peripheral blood. B-cell depletion persists during mobilization with Rituximab and HD cytarabine allowing a collection of PBSC free of B cells (median CD19+ and CD20+ cells counts 0%). Of nine patients PCR positive for bcl-2 or bcl-1 in blood and marrow at the start of immunochemotherapy, all showed PCR-negative PBSC. In conclusion, in patients with indolent lymphoma, the concurrent administration of Rituximab and HD cytarabine is a safe and efficient method to obtain in vivo purged PBSC. Immunochemotherapy prior to mobilization produces B-cell depletion and seems to be a useful preparative step.

A sequence of immuno-chemotherapy with Rituximab, mobilization of in vivo purged stem cells, high-dose chemotherapy and autotransplant is an effective and non-toxic treatment for advanced follicular and mantle cell lymphoma.

Options for relapsed/refractory indolent lymphoma include chemotherapy, immunotherapy and high-dose therapy with autologous support. The best combination of these approaches, however, is not defined. We treated 10 patients with relapsed/refractory follicular (n = 7) or mantle cell lymphoma (n = 3) using chemotherapy, immunotherapy, high-dose therapy and autotransplant in a sequence of four phases, each designed to play a specific role in tumour eradication. After the debulking with VACOP-B (doxorubicin, cyclophosphamide, etoposide, vincristine, prednisone, bleomycin) (phase 1), 9/10 patients responded but none achieved a molecular response. After the immuno-chemotherapy phase, which combined Rituximab with vincristine and cyclophosphamide, seven patients were in complete response (CR) and three in good partial response (PR), and all those with a molecular marker of disease showed a disappearance of the signal from marrow and blood. Phase 3, which coupled high-dose cytarabine with Rituximab, was effective in mobilizing an adequate number of progenitor cells that were polymerase chain reaction negative in all informative cases. Phase 4 consisted of high-dose therapy with autologous support followed by two doses of Rituximab. Autograft was performed in nine patients. The haematopoietic recovery was as expected. This sequence of chemotherapy, immuno-chemotherapy, stem cell mobilization with in vivo purging and autotransplant, organized in four blocks of treatment, was simple to administer and devoid of toxic effects. It permits rapid attainment of clinical and molecular response and enables the harvest of lymphoma-free peripheral blood progenitor cells even in heavily pretreated patients with relapsed or refractory disease.

Biological activity of all-trans-retinoic acid with and without tamoxifen and alpha-interferon 2a in breast cancer patients.

In addition to suppressing breast cancer cell growth, retinoids potentiate growth inhibition in human breast cancer when tested in vitro and in vivo with tamoxifen and/or interferon. The purpose of this study was to ascertain the biologic effects of all-trans-retinoic acid (ATRA) administered alone and with tamoxifen +/- interferon and to identify the relationship between ATRA plasma concentrations and optimal biological dose (the lowest dose that produces a biological response). Three consecutive groups of 15 patients with locally advanced operable breast cancer were treated, in accordance with good clinical practice (GCP) requirements, with ATRA at 3 dose levels alone or with tamoxifen +/- alpha-interferon 2a at flat doses. After 3 weeks, the tumors were surgically removed. Biological parameters measured at the beginning (in biopsy tissue) and end (in surgical tissue) of the study were compared. The optimal biological dose for ATRA was 15 mg/m2/day. Treatments influenced tumor grade but not cell cycle kinetics (G0-G1 phase) or proliferation (Ki67 levels). ATRA induced progesterone receptors independent of dose level and co-administered drugs, but did not induce estrogen receptors when administered alone. Retinoic acid receptor (RAR)-alpha was not affected by treatment and RAR-alpha was moderately influenced whereas RAR-beta (concomitantly with transforming growth factor-beta) was induced in 33% of patients by ATRA alone. ATRA pharmacokinetics were dose- and time-dependent. Neither the ATRA + tamoxifen nor the ATRA + tamoxifen + interferon combinations potentiated the ATRA-induced biological changes. Future studies evaluating the role of RAR-beta as a biological marker of retinoid activity are warranted.

Rituximab (IDEC-C2B8): validation of a sensitive enzyme-linked immunoassay applied to a clinical pharmacokinetic study.

Rituximab is a chimeric monoclonal antibody (MAb) directed against the B-cell CD20 antigen that has been approved for therapy of relapsed and resistant follicular non-Hodgkin's lymphoma (NHL). This study describes the development and validation of a highly sensitive, rapid, accurate, precise enzyme-linked immunosorbent assay (ELISA) to measure Rituximab serum concentrations. This study also describes the application of the ELISA method to a pharmacokinetic study in a homogeneous group of patients with follicular lymphoma who received 4 weekly doses of MAb at the standard dose of 375 mg/m2 as consolidation of chemotherapy. In the patients in this study, the median Rituximab serum concentrations increased during therapy, and showed a slow decline during the posttreatment period. The Rituximab elimination half-life of approximately 20 days accounts for the demonstrated accumulation of MAb in serum samples. Because previous pharmacokinetic studies showed a correlation between Rituximab serum levels and tumor response, the ELISA method used in this study, which allows a precise control of serum concentrations, could be useful for predicting the final response to the MAb and for selecting patients able to benefit from higher dosage or repeated drug administration.

All-trans retinoic acid and interferon-alpha in the treatment of a patient with resistant metastatic osteosarcoma.

BACKGROUND: A boy age 14 years who was in complete remission from Stage IIB small cell osteosarcoma, which was misdiagnosed as Ewing sarcoma and consequently was treated, developed inoperable lung metastases when he was off therapy. He received second-line treatment for recurrent Ewing sarcoma, including chemotherapy and radiotherapy, and obtained only a temporary response. A compassionate treatment with all-trans retinoic acid (ATRA) and interferon-alpha (IFNalpha) was then undertaken. METHODS: The patient initially was treated according to the national SE91 protocol for nonmetastatic Ewing sarcoma. After a bilateral pulmonary recurrence, he received second-line chemotherapy and irradiation of the largest metastasis, with a temporary partial response. The patient was then treated with a combination of oral ATRA (90 mg/m(2) for 3 days per week) and subcutaneous IFNalpha (3 x 10(6) U/m(2) 5 days per week) for 4 months. The same therapy also was administered for the control of residual disease after surgery for a total duration of 1 year of ATRA/IFN treatment. During the first 3 weeks of therapy, ATRA pharmacokinetics were studied. RESULTS: After progression of the patient's disease, despite the administration of first-line and second-line chemotherapy, combined treatment with ATRA/IFNalpha yielded a partial remission, which allowed surgical resection of the largest metastasis. The same therapy was effective in preventing tumor recurrence after incomplete removal of the remaining metastases. Treatment was well tolerated, and the patient is in stable complete remission 14 months after the end of therapy. The pharmacokinetics results confirmed the indication of an intermittent schedule for oral ATRA therapy. CONCLUSIONS: ATRA/IFNalpha treatment may be considered as an alternative approach in the treatment of patients with metastatic osteosarcoma who have disease that is resistant to conventional chemotherapy and in the treatment of patients with minimal tumor residue.

[The stability of all-trans retinoic acid in "retinoid solutions" of "Di Bella's Cure"]. / Stabilità dell'acido all-trans retinoico nella "soluzione ai retinoidi" della "cura Di Bella".

Recently all-trans retinoic acid, an oxidation product of retinol, has become famous, since it is a component of "retinoids solution" of "Di Bella's therapy". Since all-trans retinoic acid is rapidly destroyed in the presence of light and oxidants, we verified its stability in samples of "retinoids solution" stored in conditions believed optimal (under stream of nitrogen) and we compare the obtained results with those observed in other "retinoids solution" samples daily open and close again, simulating the intake from the patient. All samples showed a decrease of all-trans retinoic acid recovery at the end of the study, with a more rapid decline in samples daily open. From our observations, we decided to indicate an expiration date of one months from the date of "solution" preparation. The patient's manipulation brings out a change in the final composition of "retinoids solution".

All-trans retinoic acid (ATRA) in patients with chronic myeloid leukemia in the chronic phase.

Since in vitro observations indicated that all-trans retinoic acid (ATRA), especially in combination with IFNalpha, can exert significant suppressive effects on Ph+ cells, we investigated the effects and the pharmacokinetic profile of ATRA in a selected cohort of patients with Ph+ chronic myeloid leukemia (CML) in chronic phase. Eighteen patients were treated with ATRA at a dose of 80 mg/m2/day (p.o.), divided into two equal doses after meals, for 7 consecutive days every other week for a maximum of 12 courses (1 course = 1 week on and 1 week off). Pharmacokinetic profiles of ATRA were evaluated during intermittent therapy on days 1 and 7 of course 1; on day 1 of course 2; on day 1 of course 6. Out of the 18 patients treated with ATRA, 11 (61%) went off study before the sixth course of treatment because of progressive hyperleukocytosis (seven cases), or thrombocytosis (one case), or refusal (three cases). Seven (39%) patients completed the first six courses (12 weeks) of treatment with ATRA and two of them (11%) maintained a white blood cell (WBC) <10 x 10[9]/l which was induced by the pretreatment with hydroxyurea. One patient completed the 12th course of ATRA maintaining WBC <10 x 10(9)/l, platelets <500 x 10(9)/l and spleen not palpable. The treatment with ATRA was well tolerated and only one patient discontinued the therapy because of non-hematological side-effects. The area under the concentration-time curve (AUC) decreased significantly (P< 0.001) during the first week of therapy. By adopting an intermittent dosing regimen, 1 week on/ 1 week off (1 course), at the start of courses 2 and 6, we obtained the ATRA AUCs equivalent to the ones achieved on day 1 of course 1. In conclusion, our results showed that ATRA alone appeared to be unable to control the WBC expansion in the CML patients in chronic phase. Moreover, it did not induce any remarkable cytoreductive effects on the platelet count and on the hemoglobin level. The major interest of ATRA would be in combination with other therapies. If ATRA was given in combination with IFNalpha or other agents, dose reduction of these would not be planned. On the basis of the pharmacokinetic profile, ATRA should be administered intermittently rather than continuously.

Time-Dependent Kinetics of Tretinoin in Chronic Myelogenous Leukaemia during Intermittent Dose Scheduling: 1 Week On/1 Week Off.

OBJECTIVE: This study investigated the pharmacokinetics of tretinoin during alternating cycles of 1 week of tretinoin treatment and 1 week drug-free in patients with Ph1+ chronic myelogenous leukaemia (CML) in the chronic phase. PATIENTS: Eighteen patients with CML were treated with tretinoin 80 mg/m(2)/day (in two divided doses) for 7 consecutive days every other week (one cycle = 1 week on/1 week off). RESULTS: Body systemic exposure to tretinoin as determined by the area under the plasma concentration-time curve (AUC) decreased significantly during the first week of drug administration, from (mean +/- SD) 678.3 +/- 498.1 to 258.7 +/- 272.4 microg/L.h. In about 40% of the patients the decline in plasma concentrations was >/=80%, while 17% of the population did not experience any decline. On day 7 of cycle 1, the mean apparent oral clearance (CL/F) was 2.6 times the corresponding value on day 1. After 1 week without tretinoin, the mean AUC on day 1 of cycle 2 was lower (down 15%) but not statistically different from the corresponding value observed on day 1 of cycle 1; 62% of patients showed an increase in the AUC, which was 40% higher than the corresponding value on day 7 of cycle 1. On day 1 of cycle 6, the AUC and CL/F of tretinoin during a dosage interval were not statistically different from those observed on day 1 of cycle 1 and cycle 2. On all occasions the peak plasma concentration (C(max)) was strongly correlated to the corresponding AUC. No significant change in the time to observed C(max) (t(max)) and in the elimination half-life (t((1/2))) was observed during the whole study. These results confirmed that the metabolism of tretinoin is rapidly up-regulated in CML patients, with significant declines in plasma drug exposure during the first week of drug administration. After tretinoin was discontinued, a return to the noninduced state followed a mean time-cycle similar to the induction. The strong decrease in the apparent oral drug clearance and the absence of significant variations in the drug half-life demonstrated that the presystemic extraction of tretinoin is the main cause of the marked decline in plasma drug exposure. CONCLUSION: The favourable pharmacokinetic profile of tretinoin obtained by an intermittent regimen, 1 week on/1 week off therapy (vs continuous administration), suggests that such a therapeutic schedule is the most appropriate for the assessment of clinical efficacy in those pathologies in which its use is suitable.

Clinical pharmacokinetics of tretinoin.

Recent reports of the dramatic antitumour effect of tretinoin (all-trans retinoic acid) in patients with acute promyelocytic leukaemia (APL) have generated a great deal of interest in the use of this drug as a chemopreventive and therapeutic agent. However, the biological efficacy of tretinoin is greatly impaired by (presumably) an induced hypercatabolism of the drug leading to reduced tretinoin sensitivity and resistance. Several pharmacokinetic studies have shown that plasma drug exposure [as measured by the plasma area under the concentration-time curve (AUC infinity)] declines substantially and rapidly when the drug is administered in a long term daily tretinoin regimen. These observations led to the hypothesis that the rapid development of acquired clinical resistance to tretinoin may have a pharmacological basis and result from an inability to present an effective drug concentration to the leukaemic cells during continuous treatment. The principal mechanisms proposed to explain the increased disappearance of tretinoin from plasma include: (i) decreased intestinal absorption; (ii) enhanced enzymatic catabolism; and (iii) the induction of cytoplasmic retinoic acid binding proteins (CRABP), which leads to increased drug sequestration. The most favoured explanation is that continuous tretinoin treatment acts to induce drug catabolism by cytochrome P450 (CYP) enzymes. Several strategies aimed at preventing or overcoming induced tretinoin resistance have been, and are being, planned. These strategies include intermittent dose administration, administration of pharmacological inhibitors of CYP oxidative enzymes, combination with interferon-alpha and intravenous administration of liposome-encapsulated tretinoin. As these strategies are now under investigation and the number of patients enrolled is small, further studies are needed to determine the efficacy and toxicity of these new schedules of drug administration. In this article we provide an overview of the relevant aspects of tretinoin physiology and pharmacokinetics, and summarise the current status of knowledge to help in the better optimisation of tretinoin administration.

All-trans retinoic acid in hematological malignancies, an update. GER (Gruppo Ematologico Retinoidi).

BACKGROUND AND OBJECTIVE: During the past ten years, the study of retinoids has undergone a total transformation. The Italian Society of Experimental Hematology decided to discuss these advances at a meeting in Florence on April 18, 1996. INFORMATION SOURCES: The material examined in the present review includes articles and abstracts published in journals covered by the Science Citation Index and Medline. In addition, all the authors of the present article have been actively working in the field of retinoids and have contributed several papers. Summaries of their oral presentations at the Florence meeting are reported in the Appendix to this review article. STATE OF ART AND PERSPECTIVES: One of the most important advances has been the elucidation of new molecular mechanisms of control of gene expression by retinoids. A number of new retinoids have been synthesized by chemists, some of which are being screened for potential clinical use, and a few have already had a tremendous impact on clinical practice. The most important achievements have been obtained in acute promyelocytic leukemia. In 1988 a Chinese group working in Shanghai showed that using all-trans retinoic acid (ATRA) alone 94% of acute promyelocytic leukemic patients obtained complete remission through differentiation of the leukemic clone. This result transformed a dream into reality and allowed researchers to move from laboratory experience to clinical applications of this differentiating therapy. Expanding the spectrum of hematological malignancies that may respond to ATRA remains a challenge; however, several results show some activity of retinoids alone or in combination with other drugs in juvenile chronic myeloid leukemia (CML), myelodysplastic syndrome, cutaneous T-cell lymphoma and CML. Particularly interesting are the studies that explored the potential clinical synergism of ATRA-based combination therapies with growth factors, other differentiating agents such as vitamin D3, immunomodulators like interferons, or chemotherapeutic agents, in particular Ara-C, all of which show promising in vitro effects when used in combination with retinoids.

[Flexibility of the surgical model of intestinal transplantation in swine]. / Flessibilità del modello chirurgico di trapianto intestinale sperimentale nel suino.

AIM: To verify the feasibility to introduce variations in the technique of intestinal transplantation, we developed three different intestinal transplant models in pigs. EXPERIMENTAL DESIGN: Feasibility and comparative study. ENVIRONMENT: Pre-clinical organ transplant surgery. MATERIALS AND METHODS: Sixty outbread piglets (mean weight 27.1 +/- 4.4 kg) received a total orthotopic intestinal allograft from equivalent donors perfused through the aorta with UW solution at 4 degrees C. Intraluminal flushing of the graft was always avoided. The animals were divided in 3 groups according to the transplantation procedure adopted. Group 1 (n = 9): excision of small and large bowel and replacement with small bowel only; group 2 (n = 39): excision of small bowel and its replacement; group 3 (n = 12): excision of small and large bowel and their "en-bloc" replacement. The superior mesenteric artery and vein were anastomosed end-to-end in all groups. RESULTS: The lowest perioperative mortality occurred in group 2 (28%), followed by group 3 (58%) and group 1 (78%). However, in group 1 the incidence of perioperative deaths was influenced by our learning curve in surgical and anesthesiologic management. No significant differences were noted in terms of cold and warm ischaemia time of the grafts, length of operative time, histopathologic analysis of preservation injury. The addition of the colon in the transplanted graft resulted in a more critical hemodynamic profile at reperfusion. CONCLUSION: Three different experimental models of intestinal transplantation are feasible in pigs. The choice can be made based on the type of study needed.