Patent Application
20080063699
Tumor size of L3.6pI pancreatic tumors 19 days after start of treatment. Treatment with 10% trehalose, paclitaxel, MBT-0206, Gemzar (gemcitabine) and the combination of both MBT-0206 and Gemzar started 8 days after tumor cell inoculation. Gemzar was applied i.p. at a dose of 100 mg/kg bw twice a week (Mon, Thu). paclitaxel and MBT-0206 were applied i.v. on a Mon, Wed, Fri schedule at a paclitaxel dose of 5 mg/kg bw. The combination group received both MBT-0206 and Gemzar with the respective schedule. Tumors were measured by palpation with a calliper on day 23 and 27. Mean±SEM; n=9 per group.
Metastases at day 19 after start of treatment. Treatment with 10% trehalose, paclitaxel, MBT-0206, Gemzar (gemcitabine) and the combination of both MBT-0206 and Gemzar started at day 8 after tumor cell inoculation. Geinzar was applied i.p. at a dose of 100 mg/kg bw twice a week (Mon, Thu). paclitaxel and MBT-0206 were applied i.v. on a Mon, Wed, Fri schedule at a paclitaxel dose of 5 mg/kg bw. The combination group received both MBT-0206 and Gemzar with the respective schedule, n=9 per group.
The growth inhibitory assay was performed in 24-well plates with each drug concentration tested in duplicate (n=2 wells). 4×104 cells per well were seeded into a 24-well plate and incubated over night. The following day, 11 concentrations of the respective drug formulation were added for 72 h to cover the range depicted in the respective graph. Finally, the cell viability was determined by a standard MTT-assay measuring the activity of mitochondrial dehydrogenases.
The following examples should be illustrative only but are not meant to be limiting to the scope of the invention. Other generic and specific configurations will be apparent to those skilled in the art.
This example is concerned with human treatment protocols using the formulations disclosed. Treatment will be of use preventing and/or treating various human diseases and disorders associated with enhanced angiogenic activity. It is considered to be particularly useful in anti-tumor therapy, for example, in treating patients with solid tumors and hematological malignancies or in therapy against a variety of chronic inflammatory diseases such as rheumatoid arthritis or psoriasis.
A feature of the invention is that several classes of diseases and/or abnormalities may be treated by directly targeting angiogenic epithelial cells without directly targeting the tissue or cells involved in the abnormality e.g., by inhibiting angiogenesis the blood supply to a tumor is cut off and the tumor is killed without directly targeting the tumor cells in any manner. Other classes of diseases and/or abnormalities may be treated by directly targeting angiogenic endothelial cells and by directly targeting the tissue or cells involved in the abnormality.
In an other application, drug resistant cells such as drug resistant cancer cells or highly proliferative synoviocytes in rheumatoid arthritis can be affected directly.
The various elements of conducting a clinical trial, including patient treatment and monitoring, will be known to those skilled in the art in light of the present disclosure.
For regulatory approval purposes, it is contemplated that patients chosen for a study would have failed to respond to at least one course of conventional therapy and would have objectively measurable disease as determined by physical examination, laboratory techniques, or radiographic procedures. Such patients would also have no history of cardiac or renal disease and any chemotherapy should be stopped at least 2 weeks before entry into the study.
Prior to application, the formulation can be reconstituted in an aqueous solution in the event that the formulation was freeze dried. As outlined above, the required application volume is calculated from the patient's body weight and the dose schedule.
The disclosed formulations may be administered over a short infusion time. The infusion given at any dose level should be dependent upon the toxicity achieved after each. Thus, if Grade II toxicity was reached after any single infusion, or at a particular period of time for a steady rate infusion, further doses should be withheld or the steady rate infusion stopped unless toxicity improved. Increasing doses should be administered to groups of patients until approximately 60% of patients showed unacceptable Grade III or IV toxicity in any category. Doses that are ⅔ of this value would be defined as the safe dose.
Physical examination, tumor measurements and laboratory tests should, of course, be performed before treatment and at intervals of about 3-4 weeks later. Laboratory tests should include complete blood cell counts, serum creatinine, creatine kinase, electrolytes, urea, nitrogen, SGOT, bilirubin, albumin and total serum protein.
Clinical responses may be defined by acceptable measure or changes in laboratory values e.g. tumor markers. For example, a complete response may be defined by the disappearance of all measurable disease for at least a month, whereas a partial response may be defined by a 50% or greater reduction.
All of the compositions and methods disclosed and claimed herein can be made and executed without undue experimentation in light of the present disclosure. While the compositions and methods of this invention have been described in terms of preferred embodiments, it will be apparent to those skilled in the art that variations may be applied to the composition, methods and in the steps or in the sequence of steps of the method described herein without departing from the concept, spirit and scope of the invention. More specifically, it will be apparent that certain agents which are both chemically and physiologically related may be substituted for the agents described herein while the same or similar results would be achieved. All such similar substitutes and modifications apparent to those skilled in the art are deemed to be within the spirit, scope and concept of the invention as defined by the appended claims.
Some variation in dosage will necessarily occur depending on the condition of the subject being treated. The person responsible for administration will, in any event, determine the appropriate dose for the individual subject. Moreover, for human administration, preparations should meet sterility, pyrogenicity, general safety and purity standards as required by the FDA Office of Biologics standards.
The present invention includes a method of delivery of a pharmaceutically effective amount of the inventive formulation of an active agent to a target site such as an angiogenic vascular target site of a subject in need thereof. A “subject in need thereof” refers to a mammal, e.g. a human.
The route of administration preferably comprises peritoneal or parenteral administration.
For use with the present invention the “pharmacologically effective amount” of a compound administered to a subject in need thereof will vary depending on a wide range of factors. The amount of the compound will depend upon the size, age, sex, weight, and condition of the patient, as well as the potency of the substance being administered. Having indicated that there is considerable variability in terms of dosing, it is believed that those skilled in the art can, using the present disclosure, readily determine appropriate dosing by first administering extremely small amounts and incrementally increasing the dose until the desired results are obtained. Although the amount of the dose will vary greatly based on factors as described above, in general, the present invention makes it possible to administer substantially smaller amounts of any substance as compared with delivery systems which only target the pathologic tissue e.g., target the tumor cells themselves.
Standard formulation liposomal paclitaxel 50 mol % DOTAP:47 mol % DOPC:3 mol % paclitaxel (MBT-0206=EndoTag 1)
Response will be evaluated according to the WHO or RECIST criteria.
50 mol % DOTAP:47 mol % DOPC:3 mol % paclitaxel Treatment schedule for liposomal paclitaxel in ongoing studies
A further planned study comprises administration of liposomal paclitaxel, e.g. 0.5 or 1.0 or 1.5 mg/kg and gemcitabine, e.g. 1000 mg/m2 once weekly for three weeks followed by one week without treatment, preferably for an interval of at least one year.
The treatment schedule for liposomal paclitaxel will be as described above for ongoing studies.
Response will be evaluated according to the WHO or RECIST criteria.
Patient:
Dosing Schedule:
Results:
One patient with liver cell carcinoma, who had disease progression after multiple chemotherapies, has been treated with MBT-0206.
Lyophilized MBT-0206 has been reconstituted with water for injection and a total infusion volume of 300400 ml MBT-0206 (equivalent to a dose of 1.0 mg liposomal paclitaxel/kg body weight) has been administered by central or peripheral intravenous infusion over a period of 2- 4 h. The infusion rate has been increased slowly up to a maximum speed of 2,5 ml/min. Premedication depended on the patient's sex, age, condition. In the specific case of the above mentioned patient it has been given dexamethasone and an antihistamine.
MBT-0206 has been administered once weekly with a dose escalation schedule, beginning with 2 times 0.25 mg liposomal paclitaxel/kg bw, 1 times 0.5 mg liposomal paclitaxel/kg bw) and than a consolidation dose of 19 times 1.0 mg liposomal paclitaxel/kg bw. This treatment is after 22 weekly administrations still ongoing and up to now no adverse drug reactions have been reported. Besides the favourable safety profile the last evaluation of tumor size, which has been performed by CT-Scan of the liver, showed stable disease.
In another case a prostate cancer patient who became refractory to hormone therapy, has been treated with 1.0 mg liposomal paclitaxel/kg bw, 3 times weekly every third day under the same conditions of preparation and administration as described above. The premedication contained dexamethasone and antihistamines. The accumulated dose of liposomal paclitaxel for this patient in 7 days was 3.0 mg liposomal paclitaxel/kg bw.
Immortalised endothelial cells (EA.hy926) are seeded into 24-well plates (4×104 cells per well) and grown over night. The following day, 9 wells are treated for 1 h with the low dose of 51.2 ng/ml liposomal paclitaxel (60 nM) formulated as MBT-0206. In addition, 3 wells per formulation are treated with the high dose of 153.7 ng/ml (180 nM) liposomal paclitaxel formulated as MBT-0206 for 1 h and 3 wells remain untreated. Approximately 24 h later, 6 of the 9 low dose-treated wells are again treated with the same low doses of MBT-0206 for 1 h (i.e. 2× treatment groups). Again 24 h later, 3 of these 6 two times-treated wells are treated for the third time with 51.2 ng/ml paclitaxel formulated as MBT-0206 for 1 h (3× treatment groups). Approximately 96 h after this third treatment, the cell viability of all wells is quantitated. For this purpose, an assay which measures the activity of mitochopdrial dehydrogenases using the tetrazolium salt 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) is applied according to standard protocols (e.g. {Lindl, 1994 #28} with slight modifications).
The results demonstrate that the viability of cells treated for 3 times with a low dose of MBT-0206 is at least as strongly reduced as the viability of cells treated only one time with a high dose. Cells treated one time or two times with the low dose of MBT-0206 exhibit a somewhat increased viability which is, however, reduced in comparison to untreated cells.
Treatments with high doses of MBT-0206 can be replaced by using low doses at a higher frequency. There is a correlation between treatment density (no. of treatments per week) and treatment efficacy. Three weekly treatments with low doses were superior to 1 or 2 weekly treatments. This optimised dosing regimen potentially reduces toxic side effects caused by high dose treatments.
The mortality was low before day 21: only 1 control and 1 MBT-0206 animal died. At day 24 the mortality increased for unknown reason: 3 MBT-0206, 1 Gemzar and 4 control animals were dead at day 24.
The body weight was not effected by either treatment, only the weight of control tumors decreased by 18% during the last week.
Tumor volume palpated at day 10, 12, 14, 17, 19, 21, 24 after inoculation and at day 26, 28 after harvesting
Necrospy after harvest at day 26 and 28
No effect of any treatment could be observed by palpation three days after begin of treatment. Strong anti-tumor effect was observed after one week and at day 24 by palpation, with the following ranking in efficacy: Gemzar-50≈paclitaxel <MBT-0206<MBT-0206+Gemzar. However, after harvest at day 26, the measured tumor volumes of all groups were clearly lower compared to day 24. This difference in size is most likely due to imprecise palpation before harvest. At day 24 tumor size is reduced to ˜30% by the mono treatments compared to the control group (n=2). The combination of MBT-0206+Gemzar resulted in the strongest reduction of the tumor size to 13%, which was significantly (p<0.05) more effective compared to paclitaxel, Gemzar and MBT-0206 alone. At day 28 the control group (n=2) is not shown because one of the two tumors was extremely small compared all other tumors (day 24, 26, 28) and thus considered as not representative. The tumors after the mono treatments showed a weak increase in tumor size compared to day 26 (paclitaxel: 536 mm3; MBT-0206: 392 mm3; Gemzar: 398 mm3), whereas the combination therapy led to a slight tumor regression between day 26 and 28 to 88 mm3.
These data show a strong anti-tumor efficacy of paclitaxel, Gemzar and MBT-0206 in this model. The anti-tumor action of MBT-0206 is slightly stronger than paclitaxel but similar to Gemzar. The combination of MBT-0206 and Gemzar shows an impressive anti-tumor efficacy.
Tumor volume palpated at day 23, 26 after inoculation and after harvesting
Body weight from day 1, 7,12, 16, 19, 21, 23, 27
Necrospy after harvest at day 27
Clear anti-tumor effect of all therapeutic treatments was observed at day 23 after tumor inoculation, with an prominent efficacy of the combination therapy (
The mortality was slightly increased in the treatment groups, particularly during combinatory treatment (4 mice died). No control animal died before harvest.
The body weight of control mice decreased by 18% during the last 11 days. During this period a transient decrease was also observed for the treated mice leading to a weight loss of 2% for paclitaxel, 12% for MBT-0206, 19% for the combination and 22% for Gemzar.
10. Killing of Paclitaxel Resistant Cells (e.g. Tumor Cell Lines)
To demonstrate the potential of MBT-0206 to directly kill tumors expressing (multi) drug resistance, two highly paclitaxel resistant mammalian tumor cell lines were investigated in vitro. These cell lines were selected by stepwise increasing the concentration of paclitaxel in the culture medium. Both cell lines have developed a high resistance level which is reflected by concentrations for 50% growth inhibition (IC50 value) for paclitaxel around 1 or 5 μM (867 or 5000 ng/ml). In both instances, MBT-0206 is clearly superior to paclitaxel in killing drug resistant tumor cells. In contrast, in drug-sensitve or low-resistant cell lines, MBT-0206 has a more or less identical killing potential to paclitaxel.
The highly paclitaxel resistant derivative cell line Mes-SA/Dx-5MBT was selected with increasing paclitaxel concentrations from the commercially available line Mes-SA/Dx-5 (ATCC, {Harker, 1986 #29}). As shown in
In a similar way to Mes-SA/Dx-5MBT, a highly paclitaxel resistant derivative line of the murine colon carcinoma line Colon-26 (Cell lines Service, Heidelberg) was established and called Colon-26MBT. The IC50 value for paclitaxel is approximately 5 μg/ml (
In highly paclitaxel-resistant cell lines, MBT-0206 has a significantly higher killing potential as paclitaxel. In paclitaxel-sensitive lines, both paclitaxel formulations have a comparable efficacy. MBT-0206 may therefore be able to kill also (multi) drug resistant tumors directly in vitro and in vivo. It may, therefore, be a new approach to treat human tumors (or other diseases) which become unresponsive for paclitaxel.
A safety evaluation of paclitaxel loaded cationic liposomes (MBT-0206) in patients with recurrent, therapy refractory head and neck squamous-cell carcinoma was carried out.
Study Design
Drug Administration
During and after injection no signs of acute or chronic toxicity were observed: Vital and lab safety parameters remained nearly constant.
The results indicate that the dose and schedule suggested from preclinical toxicology studies was well tolerated by the patients.
Cationic liposomes selectively targeted endothelium of human head and neck squamous cell carcinoma. Laser Doppler flowmetry confirmed the antivascular mechanism of action of the therapy.
Two tumor cell lines which show high survival rates upon 72 h treatments with paclitaxel in vitro were used for mouse tumor models to investigate the potential of MBT-0206 in vivo. In these tumor models, it can be demonstrated that MBT-0206 is inhibiting tumor growth significantly more effective than paclitaxel.
1. A highly paclitaxel resistant derivative (Mes-SA/Dx-5MBT, IC50 0.87 μg/ml) was selected from the commercially available human uterus sarcoma derived cell Mes-SA/Dx-5 (ECACC). This moderately resistant line originates from the highly sensitive parental line Mes-SA. As shown in
2. Concerning the human dermal melanoma line Sk-Mel28, already the commercially available line exhibits high paclitaxel resistance (IC50 5.8 μg/ml,
Mes-SA/Dx-5MBT tumor model: tumor volumes on day 23 after start of treatment (9 treatments, i.e. 3 per week, from day 12 to day 31 after tumor cell injection)
Sk-Mel tumor model: tumor volumes on day 7 after start of treatment (6 treatments very other day, i.e. from day 17 to day 28 after tumor cell injection).
| Number | Date | Country | Kind |
|---|---|---|---|
| 03023539.4 | Oct 2003 | EP | regional |
| 04000361.8 | Jan 2004 | EP | regional |
| Filing Document | Filing Date | Country | Kind | 371c Date |
|---|---|---|---|---|
| PCT/EP04/11655 | 10/15/2004 | WO | 00 | 9/28/2007 |
