WO1994004159A2

WO1994004159A2 - Treatment of cancer with azt and alkylating or intercalating agents

Info

Publication number: WO1994004159A2
Application number: PCT/GB1993/001789
Authority: WO
Inventors: Bangaru Chandrasekaran; David Stanley Duch
Original assignee: The Wellcome Foundation Limited
Priority date: 1992-08-24
Filing date: 1993-08-23
Publication date: 1994-03-03
Also published as: IL106763A0; EP0655918A1; MX9305106A; JPH08500356A; AU4967993A; GB9217998D0; ZA936166B; WO1994004159A3

Abstract

This invention relates to methods of treating cancer in a mammal which comprise administering to said mammal a potentiating effective amount of AZT, or a pharmaceutically acceptable salt or ester thereof, with an effective amount of either an alkylating agent or a DNA intercalating agent. The preferred alkylating agent is melphalan while the preferred DNA intercalating agent is adriamycin.

Description

TREATMENT OF CANCER WITH AZT AND ALKYLATING OR INTERCALATING AGENTS

This invention relates to the field of cancer therapy and particularly to chemotherapeutic methods of treating cancer as well as to pharmaceutical compositions for use in such methods.

Chemotherapy is an important method of treating cancer which comprises the administration of substances, which are usually to a greater or lesser degree cytotoxic, to a patient. Because of the nature of these compounds patients often suffer from side effects which can in many cases be quite severe.

Drugs 39(4) (1990) p 489 is a review of certain well known categories of anti-cancer agents.

3'-azido-3'-deoxythymidine (also known as A T or zidovudine) has been found to have a particularly beneficial clinical effect against a wide range of conditions associated with HTV infections such as Acquired Immune Deficiency Syndrome (AIDS), ALDS- related complex (ARC) and asymptomatic infections. This compound at low doses is generally very well tolerated by patients and is now widely used in the treatment of HTV infections.

In addition, however, AZT has been found to be useful in combination with certain known anti-cancer agents in the treatment of cancer.

US patent No. 5116823 discloses the use of AZT in combination with 5-fluorouracil (5-FU), its precursor 5-fluoro-2'-deoxyuridine (FTJDR) or methotrexate in the treatment of carcinomas, particularly of the colon.

It has now been found that the action of alkylating agents or of DNA intercalating agents as anti-cancer agents is potentiated by using AZT in combination with them. This potentiation results in a surprisingly large synergistic cytotoxic effect. Thus, treatment with the combination is more effective than treatment with either compound alone.

Therefore in a first embodiment the invention provides a method of treating cancer in a mammal such as a human which comprises administering to said mammal a potentiating effective amount of AZT, or a pharmaceutically acceptable salt or ester thereof, and an effective amount of an alkylating agent. Examples of such alkylating agents are chloromethine, cyclophosphamide, ifosfamide, melphalan and chlorambucil, with melphalan being preferred.

Preferably the two agents should be administered separately, such separate administration being carried out within a time period sufficient to allow the desired therapeutic effect to take place.

More preferably the administration of the AZT precedes administration of the alkylating agent.

Most preferably AZT is administered both before and after administration of the alkylating agent.

In addition the present inventors have found that administration of AZT, or a pharmaceutically acceptable salt or ester thereof, potentiates the effect of DNA intercalating agents producing synergistic cytotoxic effects in cancer cells.

Therefore a further embodiment of the invention provides a method of treating cancer in a mammal such as a human which comprises administering to said mammal a potentiating effective amount of AZT, or a pharmaceutically acceptable salt or ester thereof and an effective amount of a DNA intercalating agent. Examples of such intercalating agents are anthracyclines such as adriamycin or daunorubicin, with adriamycin being prefered.

More preferably the administration of the AZT precedes administration of the DNA interclating agent.

Most preferably AZT is administered both before and after administration of the DNA intercalating agent. The methods of the invention described above are particularly useful in treating breast, colon, ovarian, bladder and thyroid carcinomas, as well as both Hodgkin and non- Hodgkin lymphomas, lymphoblastic leukaemia and multiple myelomas.

In the methods discussed, above the AZT, or a pharmaceutically acceptable salt or ester thereof, and the alkylating agent or DNA intercalating agent, may be administered by any suitable route, including oral, rectal, nasal, topical (including transdermal, buccal and sublingual), vaginal and parenteral (including subcutaneous intramuscular, intravenous and intradermal). It will be appreciated that the preferred route or routes will vary with the condition and age of the recipient, the nature of the condition and other clinical factors.

However a preferred route of administration would be via intravenous dosing, either by bolus or infusion.

The preferred daily dose range for AZT is l-25g/m2 as a constant i.v. infusion or 1- lOg/m^ as a bolus dose.. For the alkylating agent it is 5-50mg/m2, preferably 15- 25mg/m2, if given i.v. or 0.1-0.5mg/kg, preferably 0.25mg/kg if given orally. For the DNA intercalating agent the preferred dose range is 5-100mg/m2, preferably 20-

The doses of AZT and the alkylating or DNA intercalating agent can be administered as single doses or as a series of sub-doses administered at appropriate intervals.

AZT, or a pharmaceutically acceptable salt or ester thereof, and the other agent are employed in an appropriate ratio whereby the above-mentioned therapeutic effects are obtained.

The AZT and the alkylating or DNA intercalating agent used in the method of the invention can be presented in any conventional pharmaceutically effective form, e.g. pharmaceutical formulations comprising the active ingredient together with one or more pharmaceutically acceptable carriers or excipients.

Further embodiments of the invention are; a) AZT for use in the potentiation of the anticancer activity of an alkyiating agent or DNA intercalating agent; and

b) The use of AZT in the manufacture of a medicament for the potentiation of the anticancer activity of an alkylating agent or DNA intercalating agent.

As specified above the carrier, excipient, or diluent must be "pharmaceutically acceptable"; in the sense of being compatible with the other components of the formulation and not being injurious to the patient. Suitable formulations include those adapted for oral, rectal, nasal, topical (including buccal and sublingual), vaginal and parenteral (including subcutaneous, intramuscular, intravenous and intradermal) administration. The formulations may conveniently be presented in unit dosage form and may be prepared by any methods well known in the art of pharmacy. Such methods include the step of bringing into association the active ingredients with the other components. In general the formulations are prepared by uniformly and intimately bringing into association the active ingredients with liquid carriers or finely divided solid carriers or both and then if necessary shaping the product.

Formulations of the present invention adapted for oral administration may be presented as discrete units such as capsules, sachets or tablets each containing a predetermined amount of the active ingredients, as a powder or granules; as a solution or a suspension in an aqueous or non-aqueous liquid; or as an oil-in-water liquid emulsion or a water- in-oil liquid emulsion. The active ingredients may also be presented as a bolus, electuary or paste.

A tablet may be made by compression or moulding, optionally with one or more accessory ingredients. Compressed tablets may be prepared by compressing in a suitable machine the active ingredients in a free flowing form such as a powder or granules, optionally mixed with a binder (eg. povidone. gelatin, hydroxypropylmethylcellulose), lubricant, inert diluent, preservative, disintegrant (eg. sodium starch glycollate, cross-linked povidone, cross-linked sodium carboxymethyl cellulose) surface-active or dispersing agent. Moulded tablets may be made by moulding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent. The tablets may optionally be coated or scored and may be formulated so as to provide controlled release of the active ingredients therein using, for example, hydroxypropylmethyl cellulose in varying proportions to provide the desired release profile.

Formulations for parenteral administration include aqueous and non-aqueous isotonic sterile injection solutions which may contain anti-oxidants, buffers, bacteriostats and solutes which render the formulation isotonic with the blood of the intended recipient; and aqueous and non-aqueous sterile suspensions which may include suspending agents and thickening agents. The formulations may be presented in unit-dose or multi-dose sealed containers, for example, ampoules and vials, and may be stored in a freeze-dried (lyophilized) condition requiring only the addition of the sterile liquid carrier, for example water for injections, immediately prior to use. Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets of the kind previously described.

Preferred unit dosage formulations are those containing a daily dose or unit, daily sub- dose, as herein above recited, or an appropriate fraction thereof, of active ingredients.

The two active compounds can be presented together, for instance in a unitary pack or the like, together with suitable instructions regarding their administration. Therefore, in a further aspect the present invention provides a pack which consists essentially of AZT, or a pharmaceutically acceptable salt or ester thereof, and an alkylating agent, provided separately therein and presented for co-administration to a patient. Similarly the invention also provides a pack which consists essentially of AZT, or a pharmaceutically acceptable salt or ester thereof, and a DNA intercalating agent, provided separately therein and presented for co-administration to a patient.

Such packs can be presented as single dose or multi-dose depending on the treatment regime and other clinical factors.

The following examples serve only to illustrate the present invention: Example 1

In Vitro Potentiation of Antitumour Drug Activity bv AZT

Interaction between AZT and clinically used anti-cancer agents melphalan. navelbine, 5-fluorouracil (5-FTJ), methotrexate, cytosine arabinoside (ara-C) and adriamycin was studied in the human colon carcinoma cell lines HCT-8 and WiDr. The cells were grown in the presence of folate free RPMI-1640 containing 10% fetal bovine serum and lOnM Leucovorin. IC50 value for AZT, melphalan, navelbine, ara-C, 5-FU and methotrexate were determined as 40, 1, 0.05, 0.4, 3 and 0.004μM respectively against HCT-8 cells and 100, 3, 0.0015, 0.03, 1 and 0.002μM respectively against WiDr cells. The IC50 value for adriamycin was O.OOlμM against WiDr cells.

Figure 1 shows an isobologram analysis of WiDr cells exposed to AZT and melphalan. This shows synergy which is both time and schedule dependent. Data points on the dotted line in the isobologram indicate an additive effect for the drug combination. Data points below this line represent synergy and above the line represent antagonism.

Table 1 shows the interaction of AZT with anti-cancer agents defined in terms of additive, synergistic, or antagonistic effects.

Clonogenic assays were also carried out which showed a maximum synergy of 6-fold with melphalan against HCT-8 cells. This synergy was also time and schedule dependent, as can be seen from figures 2 and 3.

Example 2

In Vivo Potentiation of Antitumour Drug Activity bv AZT

In these experiments the action of AZT in potentiating melphalan activity against A2780 human ovarian Carcinoma and COLO 320 human colon carcinoma cell lines, in vivo, was evaluated. Tumours were implanted subcutaneously in nude mice and antitumor therapy initiated when tumours reached 100-200mg. Animals which received AZT were dosed three times, at 0, 3, and 6 hours with 400mg/kg AZT Animals which received melphalan received a single dose of either lOmg/kg or Table 1 Table 1

INTERACTION OF AZT WITH ANTICANCER AGENTS

DRUG SCHEDULE

15mg/kg at the times indicated. Figures 4 and 5 show the number of days in each case for two tumour doublings to occur. In both tumours AZT significantly potentiated the activity of melphalan.

Example 3

AZT Tablet Formulation

The following formulations A, B and C were prepared by wet granulation of the ingredients with a solution of povidone, followed by addition of magnesium stearate and compression.

Formulation A

Formulation C mg tablet

AZT 100

Lactose 200

Starch 50

Povidone 5

Magnesium Stearate _4 359

The following formulations, D and E were prepared by direct copmpression of the admixed ingredients. The lactose used in formulation E was of the direct compression type (Dairy Crest - "Zeparox")

The formulation was prepared by wet granulation of the ingredients (below) with a solution of povidone followed by the addition of magnesium stearate and compression. mg/tablet

Drug release takes place over a period of about 6-8 hours and was complete after 12 hours.

Example 4

Capsule formulations

Formulation A

A capsule formulation was prepared by admixing the ingredients of Formulation D in Example 2 above and filling into a two-pan hard gelatin capsule. Formulation B (infra) was prepared in a similar manner.

Formulation B

Formulation C me/capsule

(a) AZT 250

(b) Macrogel 4000 B.P. 350

600

Capsules were prepared by melting the Macrogel 4000 B.P., dispersing the AZT in the melt and filling the melt into a two-part hard gelatin capsule.

Formulation D mg/capsule

AZT 250

Lecithin 100

Arachis Oil 100

450

Capsules were prepared by dispersing the AZT in the lecithin and arachis oil and filling the dispersion into soft, elastic gelatin.

Formulation E (controlled release capsule)

The following controlled release capsule formulation was prepared by extruding ingredients a, b and c using an extruder, followed by spheronisation of the extrudate and drying. The dried pellets were than coated with release-controlling membrane (d) and filled into a two-piece, hard gelatin capsule.

m /ca sule

Exam le 5

The AZT was dissolved in most of the water (35°-40°C) and the pH adjusted to between 4.0 and 7.0 with the hydrochloric acid or the sodium hydroxide as appropriate - the batch was then made up to volume with the water and filtered through a sterile micropore filter into a sterile 10ml amber glass vial (type 1) and sealed with sterile closures and overseals. Formulation B

AZT 0.125g

Sterile, pyrogen - free, pH7 phosphate buffer, q.s. to 25ml

Example 6

Intramuscular injection

AZT 0.20g

Benzyl Alcohol 0.1 Og

Glycofurol 75 1.45g

Water for injection q.s. to 3.00ml

The AZT was dissolved in the glycoflirol. The benzyl alcohol was then added and dissolved, and water added to 3 ml. The mixture was then filtered through a sterile micropore filter and sealed in 3ml amber glass vials (type 1).

Example 7

Suppository mg/suppositorv

AZT (631m) * 250

Starch Fat, B.P./Witepsol HI 5-Dynariot NoBel) 1770

2020

* The AZT was used as a powder wherein at least 90% of the particles were of 631m diameter or less. o

One fifth of the Witepsol HI 5 was melted in a steam-jacketed pan at 45 C maximum.

The AZT was sifted through a 2001m sieve and added to the molten base with mixing, using a silverson fitted with a cutting head, until a smooth dispersion was achieved. o Maintaining the mixture at 45 C, the remaining Witepsol HI 5 was added to the suspension and stirred to ensure homogenous mix. The entire suspension was passed through a 2501m stainless steel screen and, with continuous stirring, was allowed to o o o cool to 40 C. At a temperature of 38 C to 40 C 2.02g of the mixture was filled into suitable plastic moulds. The suppositories were allowed to cool to room temperature.

Example 8

Pessaries

AZT 250

Anhydrate Dextrose 380 Potato Starch 363 Magnesium Stearate 7

1000

The above ingredients were mixed directly and pessaries prepared by direct compression of the resulting mixture.

Example 9

Melphalan i.v. formulations

(a) Component (i)

Ingredients content per vial

Melphalan BP 50.0 mg

Hydrochloric Acid, BP Ph Eur 34.5 11

Povidone BP* 20.0 mg

Water for Injections, Bp/Ph Eur to 2.0 g * polyvinylpyrrolidone Method

Prepare a suitable dilution of the Hydrochloric Acid in Water for Injeαions.

Add the Melphalan to part of the Water for Injections.

Add the Hydrochloric Acid solution and stir until solution is complete.

Add and dissolve the Povidone.

Make up to weight with Water for Injections.

Sterilise the solution by aseptic filtration.

Fill into sterile vials.

Freeze dry.

Close and seal the vials

Component (ii)

Dissolve the Sodium Citrate in part of the Water for Injections.

Add a mixture of the Propylene Glycol and Ethanol.

Make up to volume with Water for Injections.

Sterilise the solution by aseptic filtration.

Fill into sterile ampoules or vials.

Stopper with sterile closures and secure with alLiminium collars.

Addition of component (ii) to component (i) rapidly reconstitutes the freeze dried product to yield a solution for injection in the pH range 6 to 7.5. (b) Component (i)

The freeze-dried melphalan hydrochloride is prepared as described in Example 1.

Component (ii)

The solvent-diluent is prepared as described in (a) above.

Claims

1. A method of treating cancer in a mammal which comprises administering to said mammal a potentiating effective amount of AZT, or a pharmaceutically acceptable salt or ester thereof, and an effective amount of an alkylating agent.

2. A method as claimed in claim 1, wherein the AZT, or a pharmaceutically acceptable salt or ester thereof, is administered prior to administration of said alkylating agent.

3. A method as claimed in claim 1 or claim 2, wherein the AZT, or a pharmaceutically acceptable salt or ester thereof, is administered as a plurality of separate doses.

4. A method as claimed in claim 3 wherein a further dose of AZT is administered together with the alkylating agent.

5. A method as claimed in any one of claims 1 to 4 wherein after administration of the alkylating agent a further dose of AZT is administered.

6. A method as claimed in anyone of claims 1 to 5 wherein said alkylating agent is melphalan.

7. A method as claimed in claim 6 wherein the AZT, or a pharmaceutically acceptable salt or ester thereof, and/or said alkylating agent are administered by bolus or infusion.

8. A method of treating cancer in a mammal which comprises administering to said mammal a potentiating effective amount of AZT, or a pharmaceutically acceptable salt or ester therof, and an effective amount of a DNA intercalating agent.

9. A method as claimed in claim 8 wherein the AZT, or a pharmaceutically acceptable salt or ester thereof is administered prior to administration of said DNA intercalating agent.

10. A method as claimed in claim 9 wherein after administration of said DNA intercalating agent a second dose of AZT is administered to the mammal.

11. A method as claimed in any one claims 8 to 10 wherein said DNA intercalating agent is adriamycin.

12. A method as claimed in any one of claims 1 to 11 wherein said mammal is a human.

13. A pack consisting essentially of AZT, or a pharmaceutically acceptable salt or ester thereof, and an alkylating agent, provided separately therein and presented for co-administration to a mammal, for the treatment of cancer.

14. A pack consisting essentially of AZT, or a pharmaceutically acceptable salt or ester thereof, and a DNA intercalating agent, provided separately therein and presented for co-administration to a mammal, for the treatment of cancer.

15. A pack as claimed in claim 13 or claim 14 wherein said mammal is man.

16. A pack as claimed in claim 13 wherein said alkylating agent is melphalan.

17. AZT for use in the potentiation of the anticancer activity of an alkylating agent or DNA intercalating agent.

18. The use of AZT in the manufacture of a medicament for the potentiation of the anticancer activity of an alkylating agent or DNA intercalating agent.