Gail Colbern

Comparative pharmacokinetics, tissue distribution, and therapeutic effectiveness of cisplatin encapsulated in long-circulating, pegylated liposomes (SPI-077) in tumor-bearing mice

Purpose: The pharmacokinetics (PK), biodistribution and therapeutic efficacy of cisplatin encapsulated in long-circulating pegylated (Stealth®) liposomes (SPI-077) were compared with those of nonliposomal cisplatin in two murine (C26 colon carcinoma and Lewis lung) tumor models. Methods: In therapeutic effectiveness studies, mice bearing murine C26 or Lewis lung tumors received multiple intravenous doses of SPI-077 or cisplatin in a variety of treatment schedules and cumulative doses. In the PK and biodistribution study, mice received a single intravenous bolus injection of 3 mg/kg of either SPI-077 or cisplatin 14 days after inoculation with 106 C26 tumor cells. Plasma and tissues were analyzed for total platinum (Pt) content by graphite furnace (flameless) atomic absorption spectrophotometery (GF-AAS). Results: Efficacy studies showed that SPI-077 had superior antitumor activity compared to the same cumulative dose of cisplatin. When lower doses of SPI-077 were compared to cisplatin at its maximally tolerated dose in Lewis lung tumors, equivalent SPI-077 antitumor activity was seen at only half the cisplatin dose. Higher cumulative doses of SPI-077 were well tolerated and had increased antitumor effect. SPI-077 PK were characterized by a one-compartment model with nonlinear (saturable) elimination, whereas cisplatin PK were described by a two-compartment model with linear elimination. SPI-077 had a 55-fold higher volume of distribution, 3-fold higher peak plasma levels, and a 60-fold larger plasma AUC compared with cisplatin. In addition, SPI-077-treated animals displayed a 4-fold reduction in Pt delivered to the kidneys (primary target organ of toxicity) relative to cisplatin, but a 28-fold higher tumor AUC than cisplatin. Conclusions: Based on the results of our studies, encapsulation of cisplatin in long-circulating pegylated liposomes has overcome limitations experienced with other liposomal cisplatin formulations. SPI-077 has a prolonged circulation time and increased tumor Pt disposition, and its antitumor effect is significantly improved compared to cisplatin in murine colon and lung cancer models.

Therapy of a xenografted human colonic carcinoma using cisplatin or doxorubicin encapsulated in long-circulating pegylated stealth liposomes

We compared the therapeutic effects of low doses of cisplatin and doxorubicin hydrochloride encapsulated in long-circulating liposomes composed of cholesterol/hydrogenated soy phosphatidylcholine-polyethylene glycol-distearoyl-phosphatidyl-ethanolamine. The encapsulation of cisplatin and doxorubicin in these liposomes made ineffectively low doses of the free drugs able to inhibit the growth of and affect cures of a human colonic carcinoma growing in nude mice. Liposome-encapsulated cisplatin had minor systemic toxic side effects indicated by an average 9% weight loss which was recovered 3-4 weeks after the last treatment. Toxicity was not observed in mice treated with liposome-encapsulated doxorubicin.

Significant Increase in Antitumor Potency of Doxorubicin Hc1 by its Encapsulation in Pegylated Liposomes

AbstractPegylated liposomal doxorubicin (PL-DOX or Doxi®) is currently being used in the clinic to treat solid tumors in humans (ovarian and breast cancer and Kaposis sarcoma). Previous preclinical studies comparing the antitumor activity of nonliposomal doxorubicin and PL-DOX have shown that PL-DOX has significantly greater antitumor activity at equivalent doses, but these studies have not reported the degree of increase in antitumor potency associated with liposome encapsulation at lower doses of PL-DOX. The studies presented here were designed to determine the dose of PL-DOX that produces the same antitumor activity as the maximum tolerated dose (MTD) of nonliposomal doxorubicin. Conventional mice were inoculated with Lewis lung or C26 colon cells, and nude mice were inoculated with BT474 or MCF7 human breast cancer cells. Tumor-bearing mice were treated with nonliposomal doxorubicin at the MTD or with PL-DOX at the same or lower doses. As in previously published studies, PL-DOX had significantly grea...

Anti-HER2 immunoliposomes for targeted drug delivery.

The pharmaceutical application of liposomes has been greatly advanced over the last decade by the introduction of long-circulating liposomes,1,2 refinement of liposome preparation techniques,3 and efficient loading of drug into liposomes.4 The development of stable, long circulating liposomes has led to a new era in liposome drug delivery. For example, sterically stabilized liposomes (Doxil, Sequus Pharmaceuticals, Inc.) were generated by grafting polyethyleneglycol (PEG) onto the liposome surface. These liposomes display prolonged drug circulation and extravasate in solid tumors. However, these liposomes do not interact directly with tumor cells in vitro or in vivo, and instead they release drug for eventual diffusion into tumor cells. Similarly, progress in monoclonal antibody (mAb)-based therapy of cancer has led to clinical validation after two decades of research (reviewed in ref. 5). A leading example has been the development of mAb directed against the p185HER2 (HER2) receptor tyrosine kinase, the product of the HER2 (erbB2, neu). HER2 is highly overexpressed in a significant proportion of cancers and HER2 overexpression is clearly associated with poor prognosis in breast cancer.6 Anti–HER2 immunoliposomes were developed to combine the tumor-targeting properties of mAbs such as rhuMAbHER2 (recombinant humanized anti-HER2 monoclonal antibody) with the drug delivery properties of sterically stabilized liposomes. We previously showed that anti-HER2 immunoliposomes efficiently bind to and internalize in HER2-overexpressing cells in vitro, resulting in intracellular drug delivery.7,8 Here we show the therapeutic properties of anti-HER2 immunoliposomes containing doxorubicin (dox) in animal models (FIG.1) and the novel mechanism of intracellular drug delivery of anti-HER2 immunoliposomes in these models (FIG. 2). The antitumor efficacy of anti-HER2 immunoliposome–dox against human breast cancer xenografts was compared with control groups: saline and nontargeted liposome-dox + free rhuMAbHER2 (FIG. 1). Anti-HER2 immunoliposome-dox tested produced marked antitumor effects, including tumor growth inhibition, tumor regressions, and cures of mice and showed superior activity to the other treatment conditions.

Dual promoter-controlled oncolytic adenovirus CG5757 has strong tumor selectivity and significant antitumor efficacy in preclinical models.

Purpose: Transcriptionally controlled oncolytic adenovirus CG5757 is engineered with two tumor-specific promoters from E2F-1 and human telomerase reverse transcriptase genes. This virus has broad anticancer spectrum and higher specificity. The objective of the current study is to show its antitumor selectivity and therapeutic potential. Experimental Design: The antitumor specificity of E2F-1 and human telomerase reverse transcriptase promoters was evaluated in a panel of tumor and normal cells. Under the control of these promoters, the tumor-selective expression of E1a and E1b genes was evaluated. Further in vitro antitumor specificity and potency of this virus were characterized by viral replication and cytotoxicity assays followed by a newly developed ex vivo tumor culture assay. Subsequently, in vivo antitumor efficacy and toxicology studies were carried out to assess the therapeutic potential of this oncolytic agent. Results: In a broad panel of cells, E2F-1 and human telomerase reverse transcriptase promoters were activated in a tumor-selective manner. Under the control of these promoters, expression of E1a and E1b genes appears only in tumor cells. This specificity is extended to viral replication and hence the cytotoxicity in a broad range of cancer cells. Furthermore, CG5757 only replicates in cancer tissues but not in normal tissues that are derived from clinical biopsies. The safety profile was further confirmed in in vivo toxicology studies, and strong efficacy was documented in several tumor xenograft models after CG5757 was given via different routes and regimens. Conclusions: CG5757 has strong antitumor selectivity and potency. It has low toxicity and has great potential as a therapeutic agent for different types of cancers.

Targeting of Drugs to Solid Tumors Using Anti-Her2 Immunoliposomes

AbstractCancer therapy would clearly benefit from a carrier system capable of intracellular delivery of systemically administered drugs to cancer cells in solid tumors. Sterically stabilized immunoliposomes specific to the cells expressing HER2 protooncogene (anti-HER2 SIL), were designed by conjugating Fab’ fragments of a recombinant humanized anti-HER2 MAb to the distal termini of poly(ethylene glycol) chains on the surface of unilamellar liposomes (size 90–100 nm) of phosphatidylcholine, cholesterol, and poly (ethylene glycol)—derivatized phosphatidylethanolamine. Anti-HER2 SIL avidly and specifically bound to cultured HER2-overexpressing cancer cells (8,000–23,000 vesicles per cell) and became endocytosed (ke = 0.022–0.033 min.−1) via the coated pit pathway. Anti-HER2 SIL showed prolonged circulation lifetime in rats (blood MRT approx. 24 hours) and significantly increased antitumor activity of encapsulated doxorubicin against HER2-overexpressing human breast cancer xenografts in nude mice. Although t...

Antitumor activity of Herceptin in combination with STEALTH liposomal cisplatin or nonliposomal cisplatin in a HER2 positive human breast cancer model.

Single agent antitumor activity of Herceptin, a humanized monoclonal antibody directed against HER2, has been demonstrated in numerous preclinical and clinical studies. Additionally, combination therapy with Herceptin and chemotherapy (CRx) has demonstrated additive antitumor activity in both preclinical models and early clinical trials. STEALTH (pegylated) liposomal (PL) cisplatin, also known as SPI-077, is currently in clinical trials for a variety of solid tumors. The three studies reported here discuss the antitumor activity of the combination of Herceptin and nonliposomal cisplatin or PL-cisplatin in two xenograft tumor models, initiated from the cell lines, BT474 and MDA453, that overexpress the oncogene, HER2. Herceptin alone had significant antitumor activity in all three experiments (p < 0.0001). Nonliposomal cisplatin and PL-cisplatin were both effective antitumor agents but, at tolerable dose levels, PL-cisplatin was superior to nonliposomal cisplatin (p < 0.0003). The effect of combining Herceptin with the chemotherapeutic cisplatin or PL-cisplatin, was most significant at moderate doses of H (0.5 mg/kg, p < 0.0001), but tended to be greater than either agent alone in all experiments. The combination of PL-cisplatin with Herceptin had statistically similar antitumor activity to that of nonliposomal cisplatin with Herceptin in all experiments. We conclude that combination therapy with PL-cisplatin and Herceptin results in significant antitumor activity with the potential for reducing toxicity in metastatic breast cancer patients.

CHAPTER 4.7 – Targeting of sterically stabilized liposomes to cancers overexpressing HER2/neu proto-oncogene

This chapter describes HER2 (c-erbB-2, neu) oncoprotein as a target recognition molecule in cancer. In contrast to a number of previous liposomal targeting systems for anticancer agents, sterically stabilized anti-HER2 immunoliposomes were not only capable of target-specific binding and internalization by cancer cells in culture but were also able to cross the vascular barrier. Malignant phenotype is often associated with the expression of proto-oncogene. The HER2 proto-oncogene encodes a 185 kDa receptor tyrosine kinase, which belongs to the family of receptor tyrosine kinases, including also the products of epidermal growth factor HER3 and HER4 genes. Overexpression of HER2 was first observed in 20–30% of breast carcinomas and was associated with aggressive tumor growth, high recurrence rate, and poor prognosis for the patients. Further studies showed ubiquitous overexpression of HER2 in a variety of malignancies, including cancers of the ovary, endometrium, lung, stomach, pancreas, bladder, and prostate. Over the past decade, the introduction and refinement of “long-circulating” liposomes preparation techniques, and “remote loading” methods for drug loading into liposomes greatly advanced liposomal pharmacology. The “rational design” of cancer cell-targeted sterically stabilized liposomes leads to a re-evaluation of tumor targeting paradigms and opens new avenues for better a treatment of cancer.

Tumor Uptake and Therapeutic Effects of Drugs Encapsulated in Long-Circulating Pegylated Stealth® Liposomes

Abstract In this study, tumor uptake and clearance of doxorubicin were determined for two formulations of the drug: the free form in aqueous solution and the encapsulated form in polyethylene glycol-coated (pegylated, STEALTH®) liposomes composed of cholesterol/hydrogenated soy phosphatidylcholine/ polyethylene glycol-distearoyl-phosphatidyl-ethanolamine (Doxil®). The determinations used confocal laser scanning microscopy in a pancreatic carcinoma model in nude mice. The movement of pegylated liposomes containing doxorubicin from blood vessels into tumors was studied using confocal microscopy combined with autoradiography of liposomes containing a tritium-labeled phospholipid. Laser microscopy measurements showed that the lipo-some-encapsulated doxorubicin remained in the tumor longer than the free drug and produced a six-fold increase in the area under the concentration-time curve (AUC). Autoradiography showed that the extravasated tritium-labeled lipid had entered the nuclei as well as the cytoplasm of tumor cells. The authors also compared the therapeutic effects of intravenous cisplatin, doxorubicin hydrochloride, vincristine sulfate, and vinorelbine tartrate, each in the aqueous free form or encapsulated in pegylated liposomes. In this pancreatic carcinoma model, the liposome-encapsulated drugs were all more effective than the free drugs in inhibiting tumor growth and in producing cures. Except for cisplatin, all of the free drugs had toxic systemic side effects indicated by an average weight loss of 3 to 5%, which was recovered by 2 to 4 weeks after the last treatment. The liposome-encapsulated drugs did not cause weight loss.

311. Oncolytic Adenovirus CG5757 Preferentially Infects Primary Human Tumor Tissues and Shows Strong Anti-Tumor Activity in Tumor Models Following Systemic Administration

Top of pageAbstract A human adenovirus serotype 5 based oncolytic virus CG5757 was designed to preferentially replicate in and kill cancer cells that have a defective retinoblastoma (Rb) pathway and overexpress telomerase. Tumor selectivity of CG5757 was examined using a newly developed ex vivo primary tumor tissue culture model. Primary human colorectal tumors were obtained after surgical resection and placed in Millicell insert with media containing hormones, which can keep tissue viable for about 5 days. The paired tumor or normal tissues from same tissue sample were used in a viral infection experiment to determine the tumor selectivity of CG5757. Cultured tissues were infected with CG5757 or wild type (wt) Ad5 for five days, and progeny viral production was determined by TCID50 assay. Results of the paired tissue cultures from five colon cancer patients indicate that wt Ad5 has similar viral productivity in tumor and normal tissues, whereas CG5757 has higher productivity in tumor tissues. In general, CG5757 produces 100- to 1,000-fold more virus in tumor tissue compared to normal tissue. Immunohistochemical staining for adenoviral E1A shows that CG5757 has positive infection only in the cancer tissue (colon), whereas no signal is detected in normal tissues of colon, pancreas or spleen. Similarly in vitro viral specificity characterization using a panel of tumor cell lines also suggest a strong tumor selectivity of CG5757. Anti-tumor efficacy of CG5757 following intravenous injection was examined in a subcutaneous tumor model. In this study, pre-established prostate cancer LNCaP xenografts in nude mice were treated with two intravenous injections of 4|[times]|1010 viral particles of CG5757 given at a 3-day interval. Significant anti-tumor efficacy was observed in CG5757-treated animals. Eight weeks after treatment, CG5757-treated animals exhibited approximately 72% inhibition of tumor growth. These studies demonstrate the potential therapeutic efficacy of such dual promoter-controlled oncolytic adenoviruses in cancers that are Rb-defective and telomerase-positive.