Ras inhibition enhances autophagy,which partially protects cells from death

Autophagy, a process of regulated turnover of cellular constituents, is essential for normal growth control but may be defective under pathological conditions. The Ras/PI3K/mTOR signaling pathway negatively regulates autophagy. Ras signaling has been documented in a large number of human cancers. In this in-vitro study we examined the effect of the Ras inhibitor Salirasib (S-trans, trans-farnesylthiosalicylic acid; FTS) on autophagy induction and cell viability. We show that Ras inhibition by FTS induced autophagy in several cell lines, including mouse embryonic fibroblasts and the human cancer cell lines HeLa, HCT-116 and DLD-1. The autophagy induced by FTS seems to inhibit the cell death induced by FTS, since in the absence of autophagy the death of FTS-treated cells was enhanced. Therefore, inhibition of autophagy may promote the inhibition of tumor cell growth and the cell death mediated by FTS.     

Effects of SCH 59228, an orally bioavailable farnesyl protein transferase inhibitor, on the growth of oncogene-transformed fibroblasts and a human colon carcinoma xenograft in nude mice

The products of the Ha-, Ki-, and N-ras proto-oncogenes comprise a family of 21 kDa guanine nucleotide-binding proteins which play a crucial role in growth factor signal transduction and in the control of cellular proliferation and differentiation. Activating mutations in the ras oncogenes occur in a wide variety of human tumors. Ras proteins undergo a series of posttranslational processing events. The first modification is addition of the 15-carbon isoprene, farnesyl, to a Cys residue near the carboxy-terminus of Ras. Prenylation allows the Ras oncoprotein to localize to the plasma membrane where it can initiate downstream signalling events leading to cellular transformation. Inhibitors of the enzyme which catalyzes this step, farnesyl protein transferase (FPT), are a potential class of novel anticancer drugs which interfere with Ras function. SCH 59228 is a tricyclic FPT inhibitor which inhibits the farnesylation of purified Ha-Ras with an IC₅₀ of 95 nM and blocks the processing of Ha-Ras in Cos cells with an IC₅₀ of 0.6  M. SCH 59228 has favorable pharmacokinetic properties upon oral dosing in nude mice. The in vivo efficacy of SCH 59228 was evaluated using a panel of tumor models grown in nude mice. These included several rodent fibroblast lines expressing mutationally-activated (val¹²) forms of the Ha-Ras oncogene. In some cases, these proteins contain their native C-terminal sequence (CVLS) which directs farnesylation. In one model, the C-terminal sequence was altered to CVLL, making the expressed protein a substrate for a distinct prenyl transferase, geranylgeranyl protein transferase-1. When dosed orally at 10 and 50 mg/kg (four times a day, 7 days a week) SCH 59228 significantly inhibited tumor growth of cells expressing farnesylated Ha-Ras in a dose-dependent manner; over 90% growth inhibition was observed at the 50 mg/kg dose. Tumor growth of cells expressing the geranylgeranylated form of Ha-Ras was less potently inhibited. Growth of tumors derived from a rodent fibroblast line expressing activated Ki-Ras containing its native C-terminal sequence (CVIM), which preferentially directs farnesylation, was also inhibited by SCH 59228. Inhibition in the Ki-Ras model was less than that observed in the Ha-Ras model. In contrast, tumors derived from cells transformed with the mos oncogene were not significantly inhibited even at the highest dose level. SCH 59228 also significantly and dose-dependently inhibited the growth of human colon adenocarcinoma DLD-1 xenografts (which express activated Ki-ras). These results indicate that SCH 59228 possesses in vivo antitumor activity upon oral dosing in tumor models expressing activated ras oncogenes. This is the first report of oral antitumor activity with an FPT inhibitor. These results are discussed in light of recent observations on alternative prenylation of some Ras isoforms. Received: 14 January 1998 / Accepted: 13 May 1998     

Interfering with the interaction between ErbB1, nucleolin and Ras as a potential treatment for glioblastoma

The three oncogenes, ErbB receptors, Ras proteins and nucleolin may contribute to malignant transformation. Previously, we demonstrated that nucleolin could bind both Ras protein and ErbB receptors. We also showed that the crosstalk between the three proteins facilitates anchorage independent growth and tumor growth in nude mice, and that inhibition of this interaction in prostate and colon cancer cells reduces tumorigenicity. In the present study, we show that treatment with Ras and nucleolin inhibitors reduces the oncogenic effect induced by ErbB1 receptor in U87-MG cells. This combined treatment enhances cell death, reduces cell proliferation and cell migration. Moreover, we demonstrate a pivotal role of nucleolin in ErbB1 activation by its ligand. Nucleolin inhibitor prevents EGF-induced receptor activation and its downstream signaling followed by reduced proliferation. Furthermore, inhibition of Ras by Salirasib (FTS), mainly reduces cell viability and motility. The combined treatment, which targets both Ras and nucleolin, additively reduces tumorigenicity both in vitro and in vivo. These results suggest that targeting both nucleolin and Ras may represent an additional opportunity for inhibiting cancers, including glioblastoma, that are driven by these oncogenes.     

Ras inhibits endoplasmic reticulum stress in human cancer cells with amplified Myc

In neuroblastoma LAN‐1 cells harboring an amplified MycN gene, disruption of cooperation between Ras and MycN proteins by the Ras inhibitor farnesylthiosalicylic acid (FTS, Salirasib) reportedly arrests cell growth. Our aim was to establish whether this is a general phenomenon. We examined the effects of FTS on gene‐expression profiles, growth and death of NCIH929 myeloma cells and K562 leukemia cells, which—like LAN‐1 cells—exhibit Myc gene amplification and harbor active Ras. Under specified conditions, FTS reduced Ras and Myc and induced cell growth arrest and death in all Myc‐amplified cell lines but not in SHEP, a neuroblastoma cell line without Myc gene amplification. Gene‐expression analysis revealed a common pattern of FTS‐induced endoplasmic reticulum (ER) stress, known as the unfolded protein response (UPR), in Myc‐amplified cells, but not in SHEP. Thus, Ras negatively regulates ER stress in cells with amplified Myc. ER stress was also inducible by dominant‐negative (DN)‐Ras or shRNA to Ras isoforms, all of which induced an increase in BIP (the master regulator of ER stress) and its downstream targets Nrf2 and eIF2α, both regulated by active p‐PERK. FTS also induced an increase in p‐PERK, while small interfering RNA to PERK reduced Nrf2 and ATF4 and rescued cells from FTS‐induced death. BIP and its downstream targets were also increased by inhibitors of MAPK p38 and MEK. Ras, acting through MAPK p38 and MEK, negatively regulates the ER stress cascades BIP/PERK/Nrf2 and eIF2α/ATF4/ATF3. These findings can explain the Ras‐dependent protection of Myc‐amplified cells from ER stress‐associated death.     

A new functional Ras antagonist inhibits human pancreatic tumor growth in nude mice.

Constitutively active Ras proteins, their regulatory components, and overexpressed tyrosine kinase receptors that activate Ras, are frequently associated with cell transformation in human tumors. This suggests that functional Ras antagonists may have anti-tumor activity. Studies in rodent fibroblasts have shown that S-trans, transfarnesylthiosalicylic acid (FTS) acts as a rather specific nontoxic Ras antagonist, dislodging Ras from its membrane anchorage domains and accelerating its degradation. FTS is not a farnesyltransferase inhibitor, and does not affect Ras maturation. Here we demonstrate that FTS also acts as a functional Ras antagonist in human pancreatic cell lines that express activated K-Ras (Panc-1 and MiaPaCa-2). In Panc-1 cells, FTS at a concentration of 25-100 microM reduced the amount of Ras in a dose-dependent manner and interfered with serum-dependent and epidermal growth factor-stimulated ERK activation, thus inhibiting both anchorage-dependent and anchorage-independent growth of Panc-1 cells in vitro. FTS also inhibited tumor growth in Panc-1 xenografted nude mice, apparently without systemic toxicity. Daily FTS treatment (5 mg/kg intraperitoneally) in mice with tumors (mean volume 0.07 cm3) markedly decreased tumor growth (after treatment for 18 days, tumor volume had increased by only 23+/-30-fold in the FTS-treated group and by 127+/-66-fold in controls). These findings suggest that FTS represents a new class of functional Ras antagonists with potential therapeutic value.     

Ras inhibition by FTS attenuates brain tumor growth in mice by direct antitumor activity and enhanced reactivity of cytotoxic lymphocytes

A major concern in targeted drug therapy is that the inhibition of receptors and signaling molecules in tumor cells may also affect similar components in the tumor microenvironment or in the immune system, with undefined consequences for inhibition of tumor growth. One example is given by the Ras inhibitor salirasib (Farnesythiosalycilic acid, FTS), which in addition to its antitumor activity in mice and humans also exhibits anti-inflammatory activity. Here we show three major effects through which Ras inhibition by FTS provides a favorable antitumor environment in immune-competent mice with subcutaneous or intracranial tumors. First, FTS exhibited antitumor activity in intracranial immune-competent tumor-bearing mice and increased their survival relative to tumor-bearing immune-compromised mice. Second, FTS induced an increase in regulatory T cells in mouse splenocytes, in which Foxp3+ T cells did not interfere with the tumor growth inhibitory effects of FTS. Third, FTS induced an increase in antitumor cytotoxic T-cell reactivity in glioma cells by downregulating their own expression of Foxp3. This downregulation induced a TGF-β-associated mechanism in glioma cells altering the tumor microenvironment and causing reduced resistance of the tumor to the immune system. These results are important as they might explain some of the major beneficial effects of Ras inhibitors. They may provide an experimental framework for examination of the impact of other anticancer drugs on cancer and the immune system.     

The Ras inhibitor farnesylthiosalicylic acid as a potential therapy for neurofibromatosis type 1.

PURPOSE: Farnesylthiosalicylic acid (FTS) is a Ras inhibitor that dislodges all active Ras isoforms from the membrane. We assessed the ability of FTS to reverse the transformed phenotype of neurofibromatosis type 1 (NF1)-associated tumor cell lines of malignant peripheral nerve sheath tumor (MPNST). EXPERIMENTAL DESIGN: nf1 mutations were genotyped, allelic losses were analyzed, and neurofibromin expression levels were determined in MPNST cell lines ST88-14, S265P21, and 90-8. The effects of FTS on GTP-bound Ras (Ras-GTP) and its prominent downstream targets, as well as on cell morphology, anchorage-dependent and anchorage-independent growth, and tumor growth in mice, were assessed. RESULTS: The MPNST cell lines were biallelic, NF1 inactive, and neurofibromin deficient. We show that FTS treatment shortened the relatively long duration of Ras activation and signaling to extracellular signal-regulated kinase, Akt, and RalA in all NF1-deficient MPNST cell lines (NF1 cells) to that observed in a non-NF1, normally expressing neurofibromin MPNST cell line. These effects of FTS led to lower steady-state levels of Ras-GTP and its activated targets. Both anchorage-dependent and anchorage-independent growth of NF1 cells were dose dependently inhibited by FTS, and the inhibition correlated positively with Ras-GTP levels. NF1 cells were found to possess strong actin stress fibers, and this phenotype was also corrected by FTS. NF1 tumor growth in a nude mouse model was inhibited by oral FTS. CONCLUSIONS: FTS treatment of NF1 cells normalized Ras-GTP levels, resulting in reversal of the transformed phenotype and inhibition of tumor growth. FTS may therefore be considered as a potential drug for the treatment of NF1.     

Salirasib inhibits the growth of hepatocarcinoma cell lines in vitro and tumor growth in vivo through ras and mTOR inhibition

Background  

Dysregulation of epidermal growth factor and insulin-like growth factor signaling play important roles in human hepatocellular carcinoma (HCC), leading to frequent activation of their downstream targets, the ras/raf/extracellular signal-regulated kinase (ERK) and the phosphoinositide 3-kinase (PI3K)/Akt/mammalian Target of Rapamycin (mTOR) pathways. Salirasib is an S-prenyl-cysteine analog that has been shown to block ras and/or mTOR activation in several non hepatic tumor cell lines. We investigated in vitro the effect of salirasib on cell growth as well as its mechanism of action in human hepatoma cell lines (HepG2, Huh7, and Hep3B) and its in vivo effect in a subcutaneous xenograft model with HepG2 cells.     

Growth inhibition of ras-dependent tumors in nude mice by a potent ras-dislodging antagonist

A lipophilic farnesyl moiety attached to the carboxyl terminal cysteine of ras proteins structurally supports their membrane anchorage, required for ras-dependent growth-factor signaling and for transforming activity of ras oncoproteins. It has been shown that inhibition of ras farnesylation can block tumor growth in nude mice but that some ras-dependent tumors escape such blockage as a result of prenylation of ras. S-trans-transfarnesylthiosalicylic acid (FTS) is a potent ras-dislodging antagonist that does not affect ras prenylation but rather acts on the mature, membrane-bound ras and facilitates its degradation. Here we demonstrate that FTS induces reappearance of stress fibers in H-ras-transformed rat-1 cells (EJ cells) in vitro, inhibits their anchorage-independent growth in vitro, and blocks EJ-tumor growth in nude mice. The anchorage-independent growth of cells expressing ErbB2 (B104), but not that of v-raf-transformed cells, is also inhibited by FTS, suggesting specificity towards activated ras. FTS treatment (5 mg/kg i.p. daily) caused inhibition (75-80%) of tumor growth in nude mice implanted with EJ, but not in mice implanted with v-raf-transformed cells, with no evidence of systemic toxicity. Moreover, FTS treatment increased the survival rate of EJ-tumor-bearing mice from 48 to 68 days. Here we demonstrate anti-tumor potency in a synthetic, non-toxic, ras-dislodging antagonist acting independently of farnesyltransferases.     

Plasma pharmacokinetics of butyrate after intravenous administration of sodium butyrate or oral administration of tributyrin or sodium butyrate to mice and rats

Purpose: To define the plasma concentrations of butyrate achieved and the profile of plasma butyrate concentrations versus time in mice and rats treated with tributyrin or sodium butyrate. Methods: Female CD₂F₁ mice were treated with tributyrin by oral gavage or with sodium butyrate by i.v. bolus or oral gavage. Oral tributyrin doses delivered to mice were 3.1, 5.2, 7.8, and 10.3 g/kg. Intravenous sodium butyrate doses were 0.31, 0.62, 0.94, and 1.25 g/kg. Oral sodium butyrate was given to mice at 5 g/kg. Subsequently, similar studies were performed in female Sprague-Dawley rats. Rats were given tributyrin by oral gavage at doses of 3.6, 5.2, or 10.3 g/kg or sodium butyrate i.v. at a dose of 500 mg/kg. Plasma butyrate concentrations were determined by gas chromatography. Results: In mice, oral dosing with tributyrin resulted in detectable plasma butyrate concentrations as early as at 5 min after treatment and produced peak plasma butyrate concentrations at between 15 and 60 min after dosing. Peak plasma butyrate concentrations increased proportionally with increasing tributyrin dose, but as the oral tributyrin dose increased there was a greater than proportional increase in the area under the curve of plasma butyrate concentrations versus time (AUC). At a tributyrin dose of 10.3 g/kg, plasma butyrate concentrations peaked at approximately 1.75 mM and remained ≥1 mM for between 10 and 60 min after dosing. However, approximately 10% of mice treated with this dose died acutely. At a tributyrin dose of 7.8 g/kg, plasma butyrate concentrations reached approximately 1 mM by 15 min after dosing and remained between 0.8 and 1 mM until 60 min after dosing. No mouse treated with this dose died acutely. Mice given tributyrin doses of 5.2 and 3.1 g/kg achieved peak plasma butyrate concentrations of approximately 0.9 and 0.5 mM, respectively, by 45 min after dosing. Plasma butyrate concentrations in these mice remained above 0.1 mM until 120 and 90 min after dosing, respectively. The four i.v. doses of sodium butyrate resulted in plasma concentration-time profiles that also indicated nonlinear pharmacokinetics and were well described by a one-compartment model with saturable elimination. Values recorded for the Michaelis-Menten constant (K _m) and the maximal velocity of the process (V_max) ranged between 1.02 and 5.65 mM and 0.60 and 1.82 mmol/min, respectively. Values noted for the volume of the central compartment (V_c) varied between 0.48 and 0.72 l/kg. At 1.25 g/kg, i.v. sodium butyrate produced peak plasma butyrate concentrations of 10.5–17.7 mM, and plasma butyrate concentrations remained above 1 mM for 20–30 min. Sodium butyrate delivered orally to mice at 5 g/kg produced peak plasma butyrate concentrations of approximately 9 mM at 15 min after dosing and plasma butyrate concentrations exceeding 1 mM for 90 min after dosing. In rats the 10.3-g/kg oral dose of tributyrin produced peak plasma butyrate concentrations of approximately 3 mM by 75 min after dosing and butyrate concentrations excedding 1 mM from 30 to 90 min after dosing. The plasma butyrate concentrations produced in rats by 5.2- and 3.6-g/kg doses were appropriately lower than those produced by the 10.3-g/kg dose, and there was no evidence of nonlinearity. The 500-mg/kg i.v. dose of sodium butyrate produced peak plasma butyrate concentrations in rats of approximately 11 mM, and the decline in plasma butyrate concentrations with time after dosing was consistent with saturable clearance. Conclusion: These studies document the ability to use oral administration of tributyrin to achieve pharmacologically relevant concentrations of butyrate in rodent plasma. They also document the nonlinear nature of butyrate clearance. These data are being used in the design of clinical trials of oral tributyrin in patients with malignancies and hemoglobinopathies. Received: 27 July 1998 / Accepted: 3 November 1998     

Farnesylthiosalicylic acid (salirasib) inhibits Rheb in TSC2-null ELT3 cells: a potential treatment for lymphangioleiomyomatosis

The small GTPase proteins, Ras and Rheb, serve as molecular switches regulating cell proliferation, differentiation and apoptosis. Ras also regulates Rheb by inactivating the tuberous sclerosis complex (TSC), which includes products of the TSC1 and TSC2 genes encoding hamartin (TSC1) and tuberin (TSC2), respectively, and acts as a Rheb-specific GTPase-activating protein. Loss of function of TSC1 or TSC2 results in an increase in active Rheb.GTP with the consequent translational abnormalities and excessive cell proliferation characteristic of the genetic disorders, tuberous sclerosis and lymphangioleiomyomatosis (LAM). To determine whether inactivation of Rheb, Ras or both might be a potential treatment for LAM, we used TSC2-null ELT3 cells as a LAM model. The cells were treated with the Ras inhibitor S-trans,trans-farnesylthiosalicylic acid (FTS; salirasib), which mimics the C-terminal S-farnesyl cysteine common to Ras and Rheb. This C-terminus is critical for their attachment to cellular membranes and for their biological activities. Untreated, the ELT3 cells expressed significant amounts of Rheb but little Ras.GTP, and this phenotype was reversed by TSC2 reexpression. Treatment with FTS decreased Ras.GTP only slightly in the TSC2-null cells, but reduced their overactive Rheb as well as their proliferation, migration and tumor growth. Notably, TSC2 reexpression in these ELT3 cells rescued them from the inhibitory effect of FTS. Evidently, therefore, FTS blocks active Rheb in TSC2-null ELT3 cells and may have therapeutic potential for LAM.     

血清胸腺因子9肽对人结肠癌HT-29的抑瘤作用

目的：观察血清胸腺因子9肽对人结肠癌HT-29移植瘤的抑制作用。方法：裸鼠皮下接种人结肠癌HT-29细胞,瘤块生长至体积约100mm~3后,分为血清胸腺因子9肽高、中、低剂量组（分别为1.25、0.625、0312mg/kg）、环磷酰胺阳性对照组（30mg/kg）和生理盐水空白对照组,每组10只,血清胸腺因子9肽各剂量组和空白对照组裸鼠每天皮下给受试物1次,连续28d。阳性对照组腹腔注射给药,每周2次,连续4周。每周称量体质量、测量瘤块长径和短径2次。于末次给药后24h处死动物,称取体质量、测量瘤块长径和短径,计算体积后,剥离肿瘤称瘤质量,计算相对肿瘤增殖率和抑瘤率。实验重复3次。结果：血清胸腺因子9肽高、中、低剂量组相对肿瘤体积与空白对照组比较均显著减小（P〈0.01或P〈0.05）,其相对肿瘤增殖率分别为50%±20%、62%±20%和77%±35%;血清胸腺因子9肽高、中、低剂量组瘤质量与空白对照组比较均明显减轻（P均〈0.01）,抑瘤率分别为45%±3%、35%±1%、27%±3%;给药前后血清胸腺因子9肽各剂量组裸鼠体质量与空白对照组比较变化不显著（P〉0.05）。结论：血清胸腺因子9肽对人结肠癌HT-29移植瘤具有明显抑制作用。     

Phase 1 first-in-human clinical study of S-trans, trans-farnesylthiosalicylic acid (salirasib) in patients with solid tumors

Purpose

This phase I first-in-human trial evaluated salirasib, an S-prenyl derivative of thiosalicylic acid that competitively blocks RAS signaling.

Methods

Patients with advanced cancers received salirasib twice daily for 21 days every 4 weeks. Doses were escalated from 100 to 200, 400, 600, and 800 mg.

Results

The most common toxicity was dose-related diarrhea (Grade 1–2, 79% of 24 patients). Other toxicities included abdominal pain, nausea, and vomiting. No Grade 3–4 toxicity was noted. Nineteen (79%) patients had no drug-related toxicity >Grade 1. Dose-limiting toxicity (DLT) was not reached, but all three patients treated with 800 mg experienced Grade 1–2 diarrhea, abrogating dose escalation. Six patients were treated at a dose of 600 mg with no DLTs. Seven (29%) patients had stable disease on salirasib for ≥4 months (range 4–23+). The salirasib pharmacokinetic profile was characterized by slow absorption and a rapid elimination phase following oral administration. Salirasib exposure (C _max; day 1 AUC_inf vs. day 15 AUC_0–12 h) was similar between days 1 and 15 (P > 0.05). The T _1/2 (mean ± SD) was 3.6 ± 2.2 h on day 1.

Conclusions

Salirasib therapy was well tolerated. The recommended dose for phase II studies is 600 mg twice daily.     

Ras inhibition boosts galectin-7 at the expense of galectin-1 to sensitize cells to apoptosis

Galectins are a family of β-galactoside-binding lectins that exert diverse extracellular and intracellular effects. Galectin-7 and galectin-1 show opposing effects on proliferation and survival in different cell types. Galectin-7 is a p53-induced gene and an enhancer of apoptosis, whereas galectin-1 induces tumorigenicity and resistance to apoptosis in several types of cancers. We show here that in cells derived from neurofibromin-deficient (Nf1^−/−) malignant peripheral nerve sheath tumors (MPNSTs), Ras inhibition by S-trans,trans-farnesylthiosalicylic-acid (FTS; Salirasib) shifts the pattern of galectin expression. Whereas FTS decreased levels of both active Ras and galectin-1 expression, it dramatically increased both the mRNA and protein expression levels of galectin-7. Galectin-7 accumulation was mediated through JNK inhibition presumably resulting from the observed induction of p53, and was negatively regulated by the AP-1 inhibitor JDP2. Expression of galectin-7 by itself decreased Ras activation in ST88-14 cells and rendered them sensitive to apoptosis. This observed shift in galectin expression pattern together with the accompanying shift from cell proliferation to apoptosis represents a novel pattern of Ras inhibition by FTS. This seems likely to be an important phenomenon in view of the fact that both enhanced cell proliferation and defects of apoptosis constitute major hallmarks of human cancers and play a central role in the resistance of MPNSTs to anti-cancer treatments. These findings suggest that FTS, alone or in combination with chemotherapy agents, may be worth developing as a possible treatment for MPNSTs.     

The Ras inhibitor S-trans,trans-farnesylthiosalicylic acid chemosensitizes human tumor cells without causing resistance.

Ras transformation requires Ras membrane anchorage, which is promoted by a farnesylcysteine carboxymethyl ester and by additional sequences specific to each Ras isoform. We showed previously that S-trans,trans-farnesylthiosalicylic acid (FTS) disrupts Ras membrane anchorage and that this disturbance contributes to inhibition of cell transformation and tumor growth. Most tumor cells develop resistance to anticancer agents. Here we examined whether tumor cells develop resistance to FTS and evaluated the therapeutic potential of FTS combined with cytotoxic drugs, because oncogenic Ras promotes antiapoptotic signals in tumors of epithelial origin. We showed that Panc-1 pancreatic cancer cells, SW480 colon cancer cells, and H-ras (EJ)-transformed Rat-1 fibroblasts exposed to FTS for prolonged periods (>6 months) do not escape FTS-induced growth inhibition and do not develop drug resistance. These cells continued to express reduced amounts of Ras, exhibit a reversed phenotype, and show an altered response to the cytotoxic drugs doxorubicin and gemcitabine. FTS-treated Panc-1 or SW480 cells acquired sensitivity to the cytotoxic drugs, whereas FTS-treated EJ cells lost sensitivity to doxorubicin, reflecting the opposite effects of oncogenic Ras on the survival of epithelial cells and fibroblasts. Treatment with FTS led to a marked increase in sensitivity to gemcitabine of the formerly resistant SW480 cells and a 100-fold increase in sensitivity to gemcitabine of Panc-1 cells. Such treatment in mice with preexisting Panc-1 tumors provided a synergistic effect of FTS and gemcitabine, leading to enhanced inhibition of tumor growth and a 65% increase in survival rate.     

Inhibitory Effects of Ginsenoside Rh2 on Tumor Growth in Nude Mice Bearing Human Ovarian Cancer Cells

Ginsenoside Rh₂ (Rh₂), isolated from an ethanol extract of the processed root of Panax ginseng CA Meyer, inhibits the growth of B16 melanoma cells. This study was designed to evaluate the ability of Rh₂ to inhibit growth of human ovarian cancer cells (HRA) in vitro and in nude mouse. Rh₂ inhibited proliferations of various established human ovarian cancer cell lines in a dose-dependent manner between 10 and 60 μM in vitro and induced apoptosis at around the IC₅₀ dose. When HRA cells were inoculated s.c. into the right flank of nude mice, all mice formed a palpable tumor within 14 days. Although i.p. administration of Rh₂ alone hardly inhibited the tumor growth, when Rh₂ was combined with cis-diamminedichloroplatinum(II) (CDDP) the tumor growth was significantly inhibited, compared to treatment with CDDP alone. When mice were treated p.o. with Rh₂ daily (but not weekly), the tumor growth was significantly (P<0.01) inhibited, compared to CDDP treatment alone. When Rh₂ was combined with CDDP, the degree of tumor growth retardation was not potentiated. The survival time was significantly (P<, we examined whether p.o. administration of Rh₂ has a dose-dependent inhibitory effect on the tumor growth. I.p. and weekly administration of CDDP had more potent antitumor activity in the order of 1 mg/kg, 2 mg/kg and 4 mg/kg, whereas p.o. and daily administration of Rh₂ (0.4 to 1.6 mg/kg) not only had antitumor activity comparable to that of 4 mg/kg CDDP, but also resulted in a significant increase of the survival. Doses of Rh₂ used in this study did not result in any adverse side-effects as confirmed by monitoring hematocrit values and body weight, unlike 4 mg/kg CDDP, which had severe side-effects. It is noteworthy that p.o. but not i.p. treatment with Rh₂ resulted in induction of apoptotic cells in the tumor in addition to augmentation of the natural killer activity in spleen cells from tumor-bearing nude mice. Thus, particularly in view of the toxicity of CDDP, Rh₂ alone would seem to warrant further evaluation for treatment of recurrent or refractory ovarian tumor.