Effects Of Flavone Acetic Acid Research Articles

Vasoacting agents flavone acetic acid and hydralazine given in combination enhance antitumor effects under condition of hyperthermia.

The combined effects of flavone acetic acid (FAA), hydralazine (HYD) and hyperthermia on B16 melanoma cells and solid tumor were examined in vitro and in vivo. In vitro, hyperthermia did not enhance the cytotoxicity of the combined use of FAA and HYD. In vivo, growth inhibition of B16 melanoma solid tumor by FAA (150 mg/kg) combined with HYD (5.0 mg/kg) could not be differentiated from that by FAA or HYD alone. Increased antitumor effect was recognized when FAA combined with HYD was used under conditions of hyperthermia. FAA combined with HYD significantly reduced tumor blood flow compared to FAA alone or HYD alone. We thus conclude that the significant reduction in tumor blood flow may play an important role in the enhanced antitumor effects of the combined treatment with FAA, HYD, and hyperthermia.

The in vivo interaction between flavone acetic acid and hyperthermia.

The in vivo interaction between flavone acetic acid (FAA) and hyperthermia was studied in a C3H mammary carcinoma grown in the feet of female CDF1 mice and in normal foot skin. FAA was intraperitoneally injected prior to local tissue heating in restrained non-anaesthetized animals. Alone, FAA at doses of 100 mg/kg and above, inhibited tumour growth in a dose-dependent fashion. FAA also enhanced the tumour response to heat, the effect being dependent on both the time interval between the two modalities and the FAA dose, the greatest effect occurring when FAA doses of > or = 150 mg/kg preceeded heat by 3-48 h. These effects of FAA correlated with the drug's ability to decrease tumour blood perfusion measured using the RbCl extraction procedure. Injecting 150 mg/kg FAA 3 h before heating (42.7 degrees C) resulted in a 2.2-fold increase in tumour heat damage, but had little effect on the response of normal foot skin in non-tumour-bearing mice. However, this treatment gave a 2.0-fold increase in normal tissue damage when the skin experiments were repeated in tumour-bearing animals. These effects in skin occurred in the absence of any blood perfusion changes, but appeared to be associated with FAA-induced TNF-alpha production.

Combined treatments of heat, radiation, or cytokines with flavone acetic acid on the growth of cultured endothelial cells.

The antitumour effects of flavone acetic acid (FAA) against a broad spectrum of established experimental tumours has been demonstrated. Damage to the vasculature, which rapidly disrupts blood flow and induces haemorrhagic necrosis, is believed to be a major mechanism contributing to the observed antitumour effects. Despite these established observations, FAA has shown little effect against human tumours. However, other applications of FAA, for examples, for an extended period of treatments or in combination with other antitumour modalities, have not been sufficiently explored. In order to test the direct effects of FAA on vasculature, endothelial cells isolated from human umbilical vein (HUVEC) and bovine pulmonary artery (CPAEC) were used in this study. FAA at the concentrations of 50 to 200 micrograms/ml causes reduction in cell number (from 20 to > 30% of the cells) of HUVEC as measured by MTT assay after 1, 3, and 5 h of treatment at 37 degrees C. FAA did not produce significant effects on similarly treated human squamous cell carcinoma, cell line UM-SCC-2. After 1 h treatment of FAA at 300 micrograms/ml, a large number of HUVECs failed to react with an actin stain, NBD-phallacidin. The growth of HUVECs and CPAEC in the presence of FAA for 1-3 days was progressively reduced. The number of HUVEC treated for 3 days at the concentrations of 100, 200, and 300 micrograms/ml were reduced by 75-86% in comparison with the control culture. The experiments with CPAEC showed similar results. The inhibition of the growth of endothelial cells by FAA was enhanced when it combines with tumour necrosis factor-alpha but not with interleukin-1, interferon-gamma, heat, or radiation. We observed that FAA can initiate both immediate effects and growth inhibition on cultured endothelial cells. These results support the notion that FAA rapidly induces vasculature damage. Furthermore, cytokines such as tumour necrosis factor-alpha can enhance the toxicity of FAA on endothelial cells.

Flavone acetic acid increases the antitumor effect of hyperthermia in mice.

The combined effects of flavone acetic acid (FAA), a synthetic flavonoid, and hyperthermia on B16 melanoma cells were investigated. In vitro, FAA alone at concentrations below 100 micrograms/ml was not cytotoxic with a 60-min exposure at 37 degrees C. Hyperthermia at 43 degrees C for 60 min enhanced the cytotoxicity of FAA only at concentrations over 100 micrograms/ml. Inhibition of the growth of B16 melanoma solid tumor by FAA and/or hyperthermia was examined in vivo. FAA (100-200 mg/kg) inhibited tumor growth in a dose-dependent manner. The combined treatment of FAA (200 mg/kg) and hyperthermia (43 degrees C, 15 min) significantly inhibited tumor growth compared to a treatment of FAA or hyperthermia alone. The maximum antitumor effect of FAA combined with hperthermia was obtained when FAA was administered 2 or 4 h before heat. The significantly increased cytotoxicity of FAA combined with hyperthermia seems to relate to specific decreases in tumor blood flow, a reduction in tumor pH, and an increased tumor temperature, without altering pH in the normal tissues. This combined treatment of FAA and hyperthermia warrants further study for treating subjects with solid tumors.

Divergent effects of flavone acetic acid on established versus developing tumour blood flow

Flavone Acetic Acid (FAA) exerts much of its effect by reducing tumour blood flow. Previous studies on FAA-induced changes in blood flow have used established tumours with a functional microvasculature. Using radioactive Xenon(133Xe) clearance to monitor local blood flow we show that the effects of FAA are dependent on the presence of this functional microvasculature with no evidence that FAA inhibits the actual development of tumour microcirculation. Thus, administration of multiple doses of FAA around the time of tumour cell injection failed to diminish t1/2 values of 133Xe (e.g. t1/2 16 min for FAA vs 14 min for saline controls at 10 days) or to affect tumour volumes (5.55 +/- 0.06 cm3 in FAA-treated animals vs 5.7 +/- 1.3 cm3 in controls at 25 days). In marked contrast a single dose of FAA (200 mg kg-1 body weight) 2 weeks after tumour cell injection dramatically extended t1/2 times (47 min for FAA vs 7 min for controls; P less than 0.001) and significantly reduced tumour burden. This effect is specific for tumour microvasculature and is not directed simply at new vessels since a similar treatment of animals with implanted-sponge-induced granulation tissue had no effect on t1/2 times (6.8 +/- 1.1 min for FAA at 200 mg kg-1 vs 7.2 +/- 1.0 min for saline-treated controls.

Interaction between flavone acetic acid (LM-975, NSC 349512) and radiation in Glasgow's osteogenic sarcoma in vivo

Flavone acetic acid (FAA) is a new anticancer agent in Phase II trials in Europe. In preclinical testing FAA showed broad activity against murine solid tumors and minimal activity against murine leukemias. Our interest in studying the combination of FAA and radiation was based on two of its biological effects which might modify radiation damage. First, FAA depletes ATP and inhibits macromolecular synthesis which are needed to repair radiation-induced DNA strand breaks; and second, inhibition of tumor blood flow by FAA could lead to radiobiological hypoxia. Various schedules of FAA (170 mg/kg I.V.) (n = 9, SF = 0.44) and radiation (10 Gy) (n = 9, SF = 0.37) were investigated against s.c. implanted Glasgow osteogenic sarcoma. In the same model we studied both the kinetics of ATP depletion by 31P-Nuclear Magnetic Resonance Spectroscopy and the repair of radiation induced single and double strand breaks by alkaline elution. The combined response was not significantly different from log-additive when radiation was given 24, 5 or 1 hr before FAA. When FAA was given immediately before radiation an increase in tumor response, significantly different from log-additive ( p = 0.03) was observed. This enhancement disappeared when radiation was delayed for between 1 and 48 hr after FAA. While decreased ATP levels and increased response to radiation occurred within minutes after FAA administration, no effect of FAA at either 180 or 200 mg/kg was observed on the repair of radiation induced single or double strand breaks (10 and 50 Gy, respectively; 5 hr after FAA) in spite of significant ATP depletion in the tumors.

Flavone acetic acid induces a coagulopathy in mice.

The effects of flavone acetic acid (FAA) on the coagulation properties of plasma from tumour-bearing and non-tumour-bearing mice have been investigated. The study was carried out primarily on CBA mice and the CaNT tumour, although substantiating data are included for two other tumours grown in the WH strain. FAA was injected at a range of single doses up to a maximum of 300 mg kg-1, and clotting properties of the plasma were measured in vitro at various times after FAA administration. Platelet numbers and the concentration of fibrin degradation products (FDP) in the plasma were also determined. Following a dose of 300 mg kg-1, the clotting times were significantly reduced at 15-30 min in both tumour-bearing and non-tumour-bearing mice of both strains. Detailed studies on coagulation in the CBA strain (+/- CaNT tumour) indicate that in tumour-bearing animals the initial decrease in clotting time is followed 4-6 h later by an increase in clotting time, thrombin time and FDP levels. Platelet counts of tumour-bearing mice also decreased significantly over this period. Similar experiments in non-tumour-bearing mice did not show these late effects. All the data from the coagulation tests on mice with CaNT tumours are consistent with the hypothesis that intravascular coagulation occurs following treatment with FAA, and that vascular occlusion in tumours, as a results of FAA-induced coagulopathy, may contribute to tumour regression.

Reduction of cytotoxic effector cell activity in colon 38 tumours following treatment with flavone acetic acid

Host cells have been implicated as being involved in the antitumour effects of flavone acetic acid (FAA), an agent with selectivity towards solid tumours which is currently undergoing clinical trial. To determine whether tumour-associated host cells are affected by FAA treatment, tumour-infiltrating leukocytes (TIL) were isolated from subcutaneous Colon 38 tumours, which are known to be sensitive to FAA. 1–2 × 10 5 TIL were isolated per gram of tumour, comprising mainly small lymphocytes and macrophages. Spontaneous activity against YAC- 1 and P 815 tumour targets was tested in a 4 h 51Cr -release assay for lymphoid cytotoxic effector cells. High levels of activity were exhibited by TIL against both P 815, which is resistant to natural killer (NK) cells, and to NK-sensitive YAC- 1 cells. In contrast, splenic cell populations contained only NK cell activity. Within 1 h of intraperitoneal administration of FAA ( 330 mg/kg) the cytotoxic effector cell activity of the TIL population was dramatically depressed, remaining low during the time in which extensive tumour necrosis became evident. In contrast, splenic NK activity was unchanged at 1 h and elevated at 4 h. The decrease in lymphoid killer activity of the TIL population following treatment argues against the primary involvement of these effector cells in mediating the antitumour action of FAA.

Effect of flavone acetic acid (NSC 347 512) on splenic cytotoxic effector cells and their role in tumour necrosis

Flavone acetic acid (FAA), an antitumour agent currently undergoing clinical trial, has immune-modulatory effects on various cytotoxic cells in mice. Natural killer (NK) cell activity in the spleen was augmented 4 h after FAA treatment, and when spleen cells were cultured with interleukin- 2 to induce the production of lymphokine-activated-killer (LAK) cells, higher levels of LAK cell activity were generated by spleen cells from FAA-treated animals than by spleen cells from untreated, control mice. The response to FAA by spleen cells from mice bearing the Colon 38 tumour was compared to that of non-tumour bearers. Activity against NK-sensitive YAC- 1 tumour targets was augmented to a similar degree, and no activity against NK-resistant P 815 targets was detected. FAA was shown to induce haemorrhagic necrosis in the P 815 tumour grown as a subcutaneous solid tumour. Furthermore, haemorrhagic necrosis was induced by FAA on Colon 38 tumours growing in mice which had been depleted of NK activity by treatment with anti-asialo GM- 1 antibody. Thus, although NK activity could be involved in the long-term host response to the tumour, it does not appear to be a major determinant of FAA-induced haemorrhagic tumour necrosis.

Comparison of the effects of flavone acetic acid, fostriecin, homoharringtonine and tumour necrosis factor α on Colon 38 tumours in mice

Advanced subcutaneous Colon 38 tumours in mice were used for the assessment of activity of a number of anticancer drugs. Activity was measured by histological examination of tumours 24 h after a single dose of the drug and in some cases by tumour growth delay. Agents thought to exert their cytotoxic effect by damaging DNA, including Adriamycin, amsacrine and its analogue CI-921, cyclophosphamide, 5-fluorouracil and methotrexate produced no gross histological changes after 24 h, even though some delayed the growth of subcutaneous tumours. In contrast, flavone acetic acid, fostriecin and homoharringtonine caused extensive necrosis of tumours after 24 h, and each delayed the growth of advanced subcutaneous tumours by at least 10 days when administered as a single dose. The histological effects of flavone acetic acid and fostriecin were indistinguishable from those of recombinant human tumour necrosis factor α. It is proposed that histological assay of advanced tumours may provide a useful adjunct to existing methods in screening for antitumour agents with novel mechanisms of action.

In vitro chemosensitivity testing of flavone acetic acid (LM975; NSC 347512) and its diethylaminoethyl ester derivative (LM985; NSC 293015)

The antitumor effect of flavone acetic acid, LM 975, and its diethylaminoethyl ester derivative, LM 985, was studied in four human malignant cell lines [WiDr, a colon carcinoma; LICR (LON) HN- 3, a tongue carcinoma; MCF 7, a breast carcinoma; K- 562, a leukemia] using a colorimetric assay based on the reduction of dimethylthiazol- 2-yl-diphenyltetrazolium. The cell lines were exposed continuously for 4–6 days to drug concentrations ranging between 0.1 and 500 μg/ml. For LM 975, the concentrations inhibiting the growth of the various cell lines by 50% were 200 ± 10, 97 ± 7, 171 ± 16 and >500 μ g/ml for LICR (LON) HN- 3, WiDr, MCF- 7, and K- 562, respectively. The corresponding concentrations for LM 985 were 151 ± 3, 36 ± 4, 86 ± 3 and 140 ± 18 μg/ml, respectively. The difference between LM 985 and LM 975 was statistically significant for the WiDr and LICR (LON) HN- 3 lines. We also evaluated the cytotoxic activity of the two agents on normal human marrow myeloid progenitor cells in a colony-forming assay. Continuous exposure to the drugs gave a dose-dependent inhibition. The concentrations inhibiting the growth by 50% were 76 ± 31 μg/ml for LM 975 and 134 ± 41 μg/ml for LM 985. One hour incubation with either compound had no toxic effect on the myeloid progenitor cells. In conclusion, LM 975 and LM 985 do not appear to have a specific cytotoxicity for tumor cells. Our results indicate that, in vitro, toxicity on bone marrow myeloid progenitor cells is concentration dependent. Considering the low plasma concentration found in man after i.v. administration of LM 985, our observations correlate well with the absence of drug-induced myelosuppression in patients.