Thrombocytopenia induced by the histone deacetylase inhibitor abexinostat involves p53-dependent and -independent mechanisms.

Abexinostat is a pan histone deacetylase inhibitor (HDACi) that demonstrates efficacy in malignancy treatment. Like other HDACi, this drug induces a profound thrombocytopenia whose mechanism is only partially understood. We have analyzed its effect at doses reached in patient plasma on in vitro megakaryopoiesis derived from human CD34(+) cells. When added at day 0 in culture, abexinostat inhibited CFU-MK growth, megakaryocyte (MK) proliferation and differentiation. These effects required only a short incubation period. Decreased proliferation was due to induction of apoptosis and was not related to a defect in TPO/MPL/JAK2/STAT signaling. When added later (day 8), the compound induced a dose-dependent decrease (up to 10-fold) in proplatelet (PPT) formation. Gene profiling from MK revealed a silencing in the expression of DNA repair genes with a marked RAD51 decrease at protein level. DNA double-strand breaks were increased as attested by elevated γH2AX phosphorylation level. Moreover, ATM was phosphorylated leading to p53 stabilization and increased BAX and p21 expression. The use of a p53 shRNA rescued apoptosis, and only partially the defect in PPT formation. These results suggest that HDACi induces a thrombocytopenia by a p53-dependent mechanism along MK differentiation and a p53-dependent and -independent mechanism for PPT formation.

Marked antitumor activity of a new potent acronycine derivative in orthotopic models of human solid tumors.

S 23906-1 is a novel acronycine derivative selected on the basis of its potency in vitro. We investigated the antitumor activity of S 23906-1 against several murine transplantable tumors (C38 colon carcinoma, P388 leukemia, B16 melanoma, and Lewis lung carcinoma) and in orthotopic models of human lung (NCI-H460 and A549), ovarian (IGROV1 and NIH:OVCAR-3), and colorectal cancers (HCT116 and HT-29). Against established C38 colon carcinoma, S 23906-1 administered twice i.v. from 1.56-6.25 mg/kg markedly inhibited tumor growth. Treatment at the optimal dose (6.25 mg/kg) induced tumor regression in all of the mice. Acronycine was 16-fold less potent and only moderately active at the maximum tolerated dose, 100 mg/kg. Against other murine tumors of the former National Cancer Institute panel, S 23906-1 was either only moderately active or totally inactive. When evaluated in human orthotopic models, S 23906-1 given p.o. or i.v. demonstrated a marked antitumor activity against human carcinomas. In the two human lung cancer models, S 23906-1 increased the survival of the animals in a dose-dependent manner and induced treated versus control values of 162% (NCI-H460) and 193% (A549). Vinorelbine was less active, with treated versus control values of 119% and 174%, respectively. A significant survival benefit was also observed against the two i.p. ovarian tumors in which S 23906-1 was as active as paclitaxel, inducing 80% long-term survivors in the NIH:OVCAR-3 model. Lastly, S 23906-1 inhibited the growth of primary HT-29 and HCT116 colon tumors grafted onto the cecum as efficiently as irinotecan and eradicated the formation of lymph node, hepatic, and pulmonary metastases in the aggressive HCT116 model. The novel spectrum of activity of S 23906-1 compared with existing anticancer agents warrants further preclinical investigation.

Synthesis and cytotoxic and antitumor activity of benzo[b]pyrano[3, 2-h]acridin-7-one analogues of acronycine.

Benzo¿bacronycine (6-methoxy-3,3,14-trimethyl-3, 14-dihydro-7H-benzo¿bpyrano¿3,2-hacridin-7-one, 4), an acronycine analogue with an additional aromatic ring linearly fused on the natural alkaloid basic skeleton, was synthesized in three steps, starting from 3-amino-2-naphthalenecarboxylic acid (5). Eight 1, 2-dihydroxy-1,2-dihydrobenzo¿bacronycine esters and diesters (17-24) were obtained by catalytic osmic oxidation, followed by acylation. All these compounds were significantly more cytotoxic than acronycine, when tested against L1210 leukemia cells in vitro. The potency of the cyclic carbonate 24 was in the range of the most active drugs currently used in cancer chemotherapy. Two selected diesters (17 and 24) were evaluated in vivo against P388 leukemia and colon 38 adenocarcinoma implanted in mice. Both compounds were markedly active at doses 16-fold lower than the dose of acronycine itself. Against colon 38 adenocarcinoma, compounds 17 and 24 were highly efficient, inhibiting tumor growth by more than 80%. Diacetate 17 was the most active, inhibiting tumor growth by 96% at 6.25 mg/kg, with two of seven mice being tumor-free on day 43.

Histology and sensitivity to anticancer drugs of two human non-small cell lung carcinomas implanted in the pleural cavity of nude mice.

We have established two metastatic models of human non-small cell lung carcinoma (NSCLC)-the NCI-H460 large-cell carcinoma and the A549 adenocarcinoma-by inoculating tumor cells into the pleural space of nude mice. The objectives of this work were as follows: (a) to study the histological characteristics and growth and dissemination patterns of these tumors in nude mice; (b) to assess their sensitivity to drugs that have demonstrated significant clinical therapeutic effect in the treatment of NSCLC; and (c) to investigate the antitumor activity of S 16020-2, a new olivacine derivative, currently in Phase II clinical evaluation. In each of the two models, all animals developed lung tumors, resulting in 100% mortality. Histopathological study showed that these two tumors spread locally to contiguous structures, including the mediastinal pleura and diaphragm, with histological characteristics consistent with the human pathology. Anticancer drugs used for the treatment of NSCLC, such as cisplatin, doxorubicin, vinblastine, and etoposide, enhanced the life span of treated mice in the two models and were more active in the NCI-H460 than in the A549 model. The increases of survival time as compared to control groups were from 60 (P < or = 0.05) to 83% (P < or = 0.01) and from 21 to 40% for NCI-H460 and A549, respectively. Vinorelbine, paclitaxel, and irinotecan showed similar activities in the two models and increased the survival of treated mice by between 38 and 79% (P < or = 0.001) and between 58 (P < or = 0.01) and 78% in the NCI-H460 and A549 models, respectively. However, none of these drugs was curative, reflecting the resistance of this disease to chemotherapy. S 16020-2 exhibited a remarkable antitumor activity, increasing the survival by 82% (P < or = 0.01) for NCI-H460 and by 126% (P < or = 0.001) for A549. This drug was among the most active compounds in these models, thereby indicating its potential for the chemotherapy of this disease.

Biological and pharmacological characterisation of three models of human ovarian carcinoma established in nude mice: use of the CA125 tumour marker to predict antitumour activity.

In an attempt to improve the relevance of human tumour xenografts to the clinical situation, we have established 3 models of human ovarian carcinoma (IGROV1, A2780 and NIH:OVCAR-3) in nude mice in which progressive peritoneal carcinomatosis resulted in the death of tumour-bearing animals (median survival times: 32, 40 and 64 days, respectively). Histological analyses revealed both common and different characteristics in growth patterns and dissemination profiles. In each case, three stages of the disease were defined (early, intermediate and late). The antitumour activities of adriamycin, cisplatin, cyclophosphamide and paclitaxel were then compared when administered at the early stage where small multifocal tumour nodules were detectable in the peritoneal cavity of the animals. Significant antitumour activities of cisplatin and particularly paclitaxel were noted in terms of increase in survival time of the treated mice (T/C values for IGROV1, A2780 and NIH:OVCAR-3 respectively: 152%, 167%, and 187% for cisplatin and 211%, 179% and >283% for paclitaxel), paclitaxel being curative against the NIH:OVCAR-3 xenograft. These results reflect the high efficacy of these two drugs in the clinic in the treatment of ovarian carcinoma. The clinically used CA125 tumour marker, not detectable in healthy mice, was measured in the serum of mice bearing IGROV1 and NIH:OVCAR-3 tumours. CA125 serum levels increased as a function of time and were well correlated to disease progression. Moreover, treatment with cisplatin and paclitaxel led to significant decreases in these levels of between 58% and 100%. This human serum marker could be used to predict early on the efficacy of chemotherapy in these two models. In conclusion, the three experimental ovarian carcinomas possess several important characteristics of the human disease and may thus be used as a screen to select new antitumour drugs potentially active in this pathology.

Synthesis of 9-O-substituted derivatives of 9-hydroxy-5, 6-dimethyl-6H-pyrido[4,3-b]carbazole-1-carboxylic acid (2-(dimethylamino)ethyl)amide and their 10- and 11-methyl analogues with improved antitumor activity.

Analogues of the antitumor drug S 16020-2 modified at the 9, 10, or 11 position were synthesized and evaluated in vitro and in vivo on the P388 leukemia and B16 melanoma models. Starting from 9-methoxy-5, 11-dimethyl-6H-pyrido[4,3-b]carbazole-1-carboxylic acid ethyl ester, the 11-CH3 analogue of 9-hydroxy-5,6-dimethyl-6H-pyrido[4, 3-b]carbazole-1-carboxylic (2-(dimethylamino)ethyl)amide (1), compound 4, was synthesized using a four-step sequence, whereas its 10-CH3 analogue 5 was prepared using a two-step pathway, starting from compound 1. Finally starting from the 9-OH compounds 1, 4, and 5, a series of variously 9-O-substituted derivatives were synthesized. In these series, the most active compounds resulted from esterification of the 9-OH group with various aliphatic diacids, which led to 9-O-CO-( )-COOH derivatives of 1, 4, and 5. For these compounds, the number of long-term surviving mice obtained at the optimal dose were 60-100% in the ip/iv P388 leukemia and 10-35% in the ip/ip B16 melanoma, corresponding to an improved therapeutic index with respect to 1 and 4. This high antitumor activity, with curative examples in both models, was not due to a higher cytotoxicity since these compounds were equally or slightly less potent in vitro than 1 and 4. The most active compounds were thus selected for further in vivo evaluation.

Antitumor activity of S 16020-2 in two orthotopic models of lung cancer.

S 16020-2, a new olivacine derivative selected on the basis of its cytotoxicity in vitro and antitumor activity in vivo, was evaluated against the human A549 and the murine Lewis lung tumor models implanted s.c. and i.v. Against Lewis lung carcinoma implanted s.c., S 16020-2 was found to be curative, with an activity and therapeutic index (Ti = 4) similar to that of cyclophosphamide. S 16020-2 administered weekly demonstrated a high therapeutic efficacy against A549 non-small cell lung carcinoma implanted s.c. in nude mice and induced tumor regression at 80 mg/kg. When A549 tumor cells were injected i.v. in SCID mice, experimental metastases rapidly developed and the progressive invasion of the lung tissue by tumor preceded the death of animals. In this model, S 16020-2 administered at 40 mg/kg i.v. following an early (days 8, 18 and 28) or delayed (days 20, 30 and 40) treatment schedule prolonged the survival of tumor-bearing mice with T/C values of 150 and 145%, respectively. Against the i.v. Lewis lung carcinoma, S 16020-2 was also highly active since when administered at 60 mg/kg on days 5, 9 and 13 it totally inhibited tumor growth and cured up to 89% of mice. When administered on days 11, 15 and 19 to animals with established tumors, S 16020-2 was still active but not curative. In the presented studies, S 16020-2 antitumor activity was superior to that of adriamycin and comparable or superior to cyclophosphamide (used as reference compounds). Our results demonstrate the efficacy of S 16020-2 against these highly aggressive and chemoresistant tumor models.

Experimental antitumour activity of S 16020-2 in a panel of human tumours.

The antitumour activity of S 16020-2, a new topoisomerase II inhibitor, was evaluated in comparison with doxorubicin against 13 human tumours, including colon (HT-29, Colo320DM), breast (MCF7, MDAMB-231), ovary (SK-OV-3, A2780, NIH:OVCAR-3), non-small cell lung (NCI-H460, A549, Calu-6, NCI-H125) and small-cell lung (NCI-H69, SCLC6) cancers. S 16020-2 was administered weekly intravenous within a dose range of 20-90 mg/kg for 3 weeks. Antitumour responses were obtained in all the tumour types tested except in the two colon cancers. S 16020-2 produced significant growth delays in nine tumour models and induced regressions of all A549 lung tumours. The antitumour activity of S 16020-2 was superior to that of doxorubicin against the NCI-H460, A549, NCI-H69, SCLC6 and NIH:OVCAR-3 xenografts. These results demonstrate the broad spectrum of antitumour activity of S 16020-2 in a large panel of in vivo experimental models and confirm its interest as a potential agent in the treatment of malignant disease.

Up-regulated expression of the beta3 integrin and the 92-kDa gelatinase in human HT-144 melanoma cell tumors grown in nude mice.

In situ changes in the repertoire of integrins and proteolytic enzymes have been demonstrated during melanoma metastasis. To investigate whether established human melanoma cell lines, injected into nude mice, could undergo phenotypic changes similar to those observed in in situ lesions, we studied 3 melanoma cell lines of distinct metastatic origin, adherent HT-144 and SK-MEL-2 cells, and non-adherent SK-MEL-1 cells for integrin expression, proteolytic enzyme repertoire and invasive potential after in vitro culture. Heterogeneity in integrin expression, such as elevated levels in alpha(v)beta3 in SK-MEL-1 and SK-MEL-2 cells and low expression in HT-144 cells, correlated with their in vitro invasiveness, since only the adherent HT-144 and SK-MEL-2 cells were able to invade Matrigel, and in addition, secreted a 72-kDa gelatinase. In contrast, no similar correlation could be established in nude mice, as all 3 cell lines, including the non-adherent SK-MEL-1 cells, were tumorigenic when injected s.c., while only HT-144 consistently produced experimental lung metastasis. Immunochemical analysis of the integrin profile in s.c. xenografts revealed over-expression of alpha(v), beta1 and beta3 integrins exclusively in HT-144 cells, as well as increased expression of beta3 in HT-144 cell lung metastases, as confirmed by PCR analysis using species-specific primers, while zymography and Western-blot analysis demonstrated de novo expression of the 92-kDa gelatinase MMP-9 in HT-144 xenografts. Our results highlight a positive correlation between up-regulated beta3 integrin and MMP-9 expression in human HT-144 melanoma cell tumors grown in nude mice.

Synthesis and cytotoxic and antitumor activity of esters in the 1,2-dihydroxy-1,2-dihydroacronycine series.

Seven 1,2-dihydroxy-1,2-dihydroacronycine and 1,2-dihydroxy-1,2-dihydro-6-demethoxyacronycine esters and diesters were synthesized via osmic oxidation of acronycine or 6-demethoxyacronycine followed by acylation. The 6-demethoxyacronycine derivatives were found to be inactive, whereas in contrast, all of the acronycine derivatives were more potent than acronycine itself when tested against L1210 cells in vitro. Four selected acronycine derivatives (17,19, 21, and 22) were evaluated in vivo against murine P388 leukemia and colon 38 adenocarcinoma implanted in mice. All compounds were markedly active against P388 at doses 4-16-fold lower than acronycine itself. Against the colon 38 adenocarcinoma, the three compounds 17, 21, and 22 were highly efficient. 1,2-Diacetoxy-1,2-dihydroacronycine (17) was the most active, all the treated mice being tumor-free on day 23.

Synthesis and biological evaluation of 6-(9-hydroxy-5-methyl (and 5,6-dimethyl)-6H-pyrido[4,3-b]carbazol-1-yl)picolinic amides as new olivacine derivatives.

Starting from 2-(6-methoxy-1-methylcarbazol-2-yl)ethylamine and diethyl-2,6-pyridine dicarboxylate, the title compounds were obtained through five or six steps. The new compounds retained significant cytotoxicity towards various tumor cell lines, but in vivo studies on murine P388 leukemia, B16 melanoma and Lewis lung carcinoma showed a lowered antitumor activity with respect to that of the related olivacine lead compound 1.

New purines and purine analogs as modulators of multidrug resistance.

A series of 36 purine and purine analog derivatives have been synthesized and tested for their ability to modulate multidrug resistance in vitro (P388/VCR-20 and KB-A1 cells) and in vivo (P388/VCR leukemia). Compounds were compared to S9788, a triazine derivative which has already shown some activity during phase 1 clinical trials and also a limiting cardiovascular side effect possibly linked to its calcium channel affinity. The fact that active compounds increase adriamycin accumulation in the resistant KB-A1 cells, and not in the sensitive KB-3-1 cells, suggests they act predominantly by inhibiting the P-glycoprotein-catalyzed efflux of cytotoxic agents. No direct relation was found between the affinity for the phenylalkylamine binding site of the calcium channel and in vitro sensitization of resistant cells. In vivo, when administered po in association with vincristine (0.25 mg/kg), five compounds (3, 4, 9, 25, and 26), of very differing calcium channel affinities (Ki from 5 to 560 nM), fully restored (T/V > or = 1.4) the sensitivity of P388/VCR leukemia to vincristine.

In vivo antitumor activity of S 16020-2, a new olivacine derivative.

The antitumor activity of S 16020-2, a new olivacine derivative, was investigated in vivo and compared with that of Adriamycin and elliptinium acetate in a panel of murine (P388 leukemia, M5076 sarcoma, Lewis lung carcinoma, and B16 melanoma) and human (NCI-H460 non-small-cell lung and MCF7 breast carcinomas) tumor models. S 16020-2 given i.v. was active against P388 leukemia implanted i.p., s.c., or intracerebrally. The therapeutic effect of an intermittent schedule (administration on days 1, 5, 9) was superior to that of single-dose treatment, allowing the i.v. administration of high total doses of S 16020-2 and resulting in the cure of 60% of mice in the i.p. P388 model. In this model, S 16020-2 was more active than elliptinium acetate and showed a better therapeutic index than Adriamycin: > or = 8 versus 2. A good therapeutic effect of S 16020-2 was also observed in three P388 leukemia sublines displaying the classic multidrug-resistance phenotype, namely, P388/VCR, P388/VCR-20, and P388/MDRC.04, the latter being totally insensitive to vincristine and Adriamycin. However, S 16020-2 was not active against the P388/ADR leukemia, a model highly resistant to adriamycin in vivo. S 16020-2 was both more active than Adriamycin and curative in the M5076 sarcoma and Lewis lung carcinoma implanted s.c. In the B16 melanoma implanted i.p. or s.c., S 16020-2 was less active than Adriamycin. Against the NCI-H460 human tumor xenograft, S 16020-2 demonstrated activity superior to that of Adriamycin (T/C = 20% versus 43% on day 21). Against the MCF7 breast cancer xenograft, S 16020-2 was active, but less so than Adriamycin (T/C = 23% versus 9% on day 21), whereas elliptinium acetate was marginally active (T/C = 49% on day 24). The hematological toxicity of S 16020-2 given to B6D2F1 mice at pharmacological dose appeared to be less severe than that of Adriamycin, particularly in bone-marrow stem cells. These results demonstrate that S 16020-2 is a highly active antitumor drug in various experimental tumor models and is markedly more efficient than elliptinium acetate. Because of its pharmacological profile, which is globally different from that of Adriamycin, S 16020-2 is considered an interesting candidate for clinical trials.

Synthesis and evaluation of 9-hydroxy-5-methyl-(and 5,6-dimethyl)-6H-pyrido[4,3-b]carbazole-1-N- [(dialkylamino)alkyl]carboxamides, a new promising series of antitumor olivacine derivatives.

Starting from 2-(2-aminoethyl)-6-methoxy-1-methylcarbazole, ethyl 9-methoxy-5-methyl-6H-pyrido[4,3-b]carbazole-1-carboxylate was obtained through a three-step sequence. This compound and its 6-methyl derivative react with (dialkylamino)alkylamines to provide various 9-methoxy-5-methyl-6H-pyrido[4,3-b]carbazole-1-(N-substituted carboxamides) whose boron tribromide demethylation afforded corresponding 9-hydroxy-1-(N-substituted carbamoyl)-olivacines. The same pathway but starting from 2-(2-aminoethyl)-6-methoxy-1,4-dimethylcarbazole led to ethyl 9-methoxy-5,11-dimethyl-6H-pyrido[4,3-b]carbazole-1-carboxylate which did not normally react with amines. It provided either the recovered starting material at 120 degrees C or 9-methoxyellipticine resulting from an unexpected decarboethylation in a steel vessel at 180 degrees C. Biological testing of the newly obtained 1-carbamoylolivacine derivatives showed that 9-hydroxylated compounds displayed high cytotoxicity for cultured L1210 and colon 38 cells (IC50 range 5-10 nM) and good antitumor activity in vivo in the P388 leukemia and colon 38 models when administered by the iv route. The most active compound in these series is 9-hydroxy-5,6-dimethyl-1-[N-[2-(dimethylamino)ethyl]carbamoyl]-6H- pyrido[4,3-b]carbazole which was selected for further evaluation on murine solid tumors and for toxicological studies.

In vivo reversal of multidrug resistance by two new dihydropyridine derivatives, S16317 and S16324.

Two new dihydropyridine derivatives with low calcium channel affinity, S16317 and S16324, were found to fully overcome multidrug resistance in vitro. These two compounds increased doxorubicin cytotoxicity on the human COLO 320DM cell line and completely reversed the vincristine resistance of murine P388/VCR cells. In vivo, S16324 administered p.o. (200 mg/kg on days 1 to 4) or i.p. (50 mg/kg on days 1, 5, 9) in combination with vincristine (i.p.) restored the antitumor activity of vincristine in P388/VCR-bearing mice. S16317 showed a reversing activity when administered p.o., i.v. (days 1 to 4) or i.p. (days 1, 5, 9) at the same dose (25 mg/kg), suggesting a remarkable bioavailability. Moreover, these two compounds potentiated the antitumor activity of vincristine in the sensitive P388 leukemia, increasing the number of long-term survivors. These results suggest that combination chemotherapy using S16317 or S16324 would be effective not only in circumventing multidrug resistance but also in preventing the emergency of a population of resistant tumor cells in sensitive tumors.

In vivo immunopharmacological properties of tuftsin and four analogs.

Four analogs of the natural macrophage-activator peptide tuftsin (T-K-P-R) were synthesized with the aim of obtaining compounds more effective in the stimulation of the immune system than tuftsin. Modifications to the parent tuftsin molecule were (i) substitution of the proline (P) residue, and/or (ii) replacement of the N-terminal residue threonine (T). The study presented here shows that the integrity of the NH2 terminus is not mandatory for a full biological tuftsin-like activity. Our data also suggest that the analogue F-(psi)-K-ABO-R, where ABO is a non-natural amino acid, is a promising agent for immunotherapy of infectious and neoplasic diseases for which tuftsin has already demonstrated some efficacy.

In vitro and in vivo circumvention of multidrug resistance by Servier 9788, a novel triazinoaminopiperidine derivative.

S 9788 is a novel triazinoaminopiperidine derivative which does not belong to any of the classes of compounds known to reverse multidrug resistance (MDR). S 9788 was far more potent than verapamil (VRP) in reversing resistance to adriamycin (ADR) in the ADR-selected murine leukaemia cell lines P388/ADR-1 and P388/ADR-10, and the human chronic myelogenous leukaemia K562/R. Fold reversion with S 9788 (5 microM) was, respectively, 3.5, 5.4 and 11.3 times greater than that with VRP (5 microM). S 9788 was also a more potent reversant of ADR resistance in the intrinsically resistant human colon adenocarcinoma COLO 320DM (2.3 fold), and of vincristine (VCR) resistance in the human MDR1 gene-transfected squamous lung carcinoma line S1/tMDR1 (5.6 fold). The activity of S 9788 depended on both the MDR cell line and the cytotoxic agent. S 9788 (50-100 mg/kg/d) administered IP once a day on days 1-4 resulted in a dose-dependent increase in the chemotherapeutic effect of VCR (0.25 mg/kg/d) in P388/VCR - bearing mice and ADR (4 mg/kg/d) in P388/ADR - bearing mice. Increases in antitumor activity were (% T/C) of +20-34% in the P388/ADR model and + 50-78% in the P388/VCR model with respect to cytotoxic agent treatment alone. S 9788 appeared to be devoid of toxicity at its effective doses. The mechanism of action of S 9788 is unknown but S 9788 (0.5-10 microM) induced a dose-dependent increase in ADR accumulation in KB-Al cells and compared to verapamil its effect was twice as active and approximately seven times more potent. We conclude that S 9788 is a novel agent capable of reversing MDR in vitro and in vivo, and whose pharmacological profile warrants its selection as a candidate drug for eventual assessment in the clinic.

New triazine derivatives as potent modulators of multidrug resistance.

A series of 70 triazine derivatives have been synthesized and tested for their capacity to modulate multidrug resistance (MDR) in DC-3F/AD and KB-A1 tumor cells in vitro, in comparison with verapamil (VRP), a calcium channel antagonist currently used in therapy as an antihypertensive drug, which also shows MDR modulating activity. Among the 12 selected compounds, 16 (S9788) showed high MDR reversing properties in vitro (300- and 6-fold VRP at 5 microM in DC-3F/AD and KB-A1 cells, respectively) and induced a strong accumulation of adriamycin. The relationship between the increase of ADR accumulation and the fold reversal induced by these compounds and their lack of effects on the sensitive DC-3F cells suggest that they act mainly by inhibiting the P-glycoprotein (Pgp) catalyzed efflux of cytotoxic agents, as already described for a majority of MDR modulators. In vivo, in association with the antitumor drug vincristine (0.25 mg/kg), 16 (100 mg/kg) increased the T/C by 39% in mice bearing the resistant tumor cell line P388/VCR. According to these interesting properties, 16 was selected for a clinical development because it was more bioavailable than 34, even though it was less active.

Approaches to some biochemical mechanisms of action of tuftsin and analogues.

Tuftsin, T-K-P-R, is a phagocytosis-stimulating peptide described as a natural immunostimulant. Four analogues of this peptide were synthesized. These compounds were assayed for their ability to compete with [3H]tuftsin for its specific receptor from thioglycollate-elicited mouse peritoneal macrophages. They were also tested for their ability to change level in intracellular cGMP and to stimulate phagocytosis through the nitroblue tetrazolium reduction measurement. Surprisingly, all the analogues were poor competitors of [3H]tuftsin binding but possess potent tuftsin-like activities.