RU2195937C1 - Combined antituberculosis preparation (rizobutol) - Google Patents

Abstract

FIELD: medicine, phthisiology, pharmacology. SUBSTANCE: invention relates to the combined antituberculosis preparation. The combined antituberculosis preparation comprises rifampicin, ethambutol, isoniazide, pyrazinamide and pyridoxine taken in the definite ratio of components. Invention provides the enhancement of effectiveness of therapy due to simplification of curative process, resistance overcoming in many patients and improvement of the preparation properties. EFFECT: enhanced effectiveness of preparation and treatment. 6 tbl

Description

 The invention relates to the field of pharmacology and medicine, specifically to a new type of anti-TB drugs containing a set of drugs in one tablet (capsule), designed primarily to overcome the possible resistance of the causative agent of tuberculosis to drugs in a patient, to simplify the treatment procedure and, thus, contribute to the full course of treatment and, ultimately, increase the effectiveness of treatment.

 The usual course of tuberculosis monotherapy is to prescribe rifampicin or isoniazid. The preferred course of combination therapy is to prescribe isoniazid, rifampicin and pyrazimide. In the event that resistance to isoniazid is possible, a fourth drug, ethambutol, is usually added. Medicines are prescribed by the attending physician in doses and proportions depending on age, physiological characteristics, the nature of the course of the disease, and, paradoxically, the availability of drugs in pharmacies, which is typical even for developed countries such as the USA or Italy.

 Until recently, research in the field of treatment of tuberculosis went in the direction of searching for new active principles and their combinations with well-known medicines (see, for example, Heb. Pat. 0650728, Pat. US 5399558, Heb. Pat. 0398165, Pat. PCT WO 95 / 13807), or the creation of new dosage forms for such well-known drugs as isoniazid, rifampicin, ethambutol (see, for example, WO 88/06038, US Pat. USA 5811088, Pat. Russia 93015116, 2143900, 94041109, 98100213).

 However, for the most part they did not concern combined medicines, since they do not presuppose the mandatory presence of all active ingredients in one tablet or other dosage form. Thus, they are intended, as a rule, for mono- or combination therapy.

Currently, the World Health Organization (WHO) recommends the abandonment of mono- and combination therapy and indicates the need to create new combination anti-TB drugs containing a specially selected set of fixed-dose drugs in one tablet that provide an optimal therapeutic effect (Consilium Medicum 1999, v.2, p. 170-171. Remedium, 2000, 7.8)
The need for such a strategy is dictated by the fact that at the present time due to an increase in the percentage of patients with resistant strains it includes three or more drugs, the course of treatment is on average 6-12 months. The diagnosis is clinically established within 3-6 weeks, 3-6 weeks are also required to identify primary resistance (i.e., inefficiency of the initially established course of treatment).

 As a consequence of the above, for HIV patients taking combination therapy, it is often not possible to timely carry out tests to select an effective course of treatment, since they die from tuberculosis 4-16 weeks after the clinical manifestations of this disease.

 As a rule, patients with tuberculosis, due to the difficulties and the length of the selection of effective drugs, as well as the course of therapy themselves, either do it not regularly or do not finish it, which causes a return of the disease and the emergence of secondary drug resistance.

 The creation of combined drugs is associated with the resolution of specific problems (except for the selection of an effective composition of drugs, their ratios), such as the compatibility of various drugs in the manufacturing and storage process, the development of technological manufacturing modes acceptable for all active principles, the selection of common carriers that carry out for example, targeted delivery of drugs that contribute to overcoming drug resistance, providing prolonged action, or guides other tasks.

 In the case of the creation of combined anti-tuberculosis drugs based on rifampicin on the recommendation of WHO, special attention should be paid to ensuring its adequate bioavailability, which can be significantly reduced when new combined drugs are created and even in the case of non-compliance with the manufacturing technologies for known combined drugs. (Ibid.).

 Thus, a combined anti-tuberculosis drug is known containing, as an active principle, a mixture of rifampicin, ethambutol and isoniazid in a ratio of 150: 75: 300 (mg) and 30-50 wt.% Inert filler, which is used as chalk, soda, magnesium stearate ( Reference Vidal, 2001, p. B-336), which is closest to the proposed invention and adopted as a prototype.

 Due to the selection of certain doses (and therefore the ratios between medicinal substances) of rifampicin, ethambutol and isoniazid, the drug exhibits high therapeutic activity in a total dose of 140 mg / kg of body weight.

 However, with prolonged use, a pronounced toxic effect is observed.

 An inventive task is to expand the arsenal of combined anti-TB drugs, increase their therapeutic effectiveness, and reduce toxic effects.

The goal is achieved by the fact that a combined anti-tuberculosis drug is proposed containing rifampicin, ethambutol, isoniazid and additionally pyrazimide and pyridoxine as an active ingredient, and as a filler, 2-15 wt.% Metacel in the following ratio of components in the drug mixture, mg:
Rifampipin - 140 - 160
Ethambutol - 270 - 280
Isoniazid - 70 - 80
Pyrazimide - 400
Pyridoxine - 10 - 15
The applicant selected a set of drugs (rifampicin, ethambutol, isoniazid, pyrazimide), which is used in the most preferred course of combination therapy. Pyridoxine, a precursor to the formation of pyridoxalphosphate, one of the derivatives of vitamin B _6, is also included in this kit.

Metacel is the brand name for cellulose methyl esters containing 14-30% methoxy groups. The substance is a fine crystalline white powder, easy to mold and tablet. Cellulose methyl esters are used as thickeners in a number of industries (in the textile industry, printing, cosmetology, and the food industry for thickening juices, pastes, etc.)
The applicant is not aware of the use of metacel as a carrier for combined anti-TB drugs. The proposed combination of active principles and the carrier, their ratios are new for combined anti-TB drugs, selected experimentally and affect the final identified properties of the drug.

 The applicant discovered unexpected (from the point of view of the prior art) the following properties of the proposed drug.

 1. Data on the pharmacokinetics of rifampicin in the proposed drug showed that the maximum concentration is achieved after 3-4 hours and is 7 μg / ml, which is sufficient for a high bacteriostatic effect. The pharmacokinetics curve is smoother, which is an advantage, since the high value of the bacteriostatic effect lasts longer. This proves that in this combination preparation the WHO requirements regarding the requirement for the bioavailability of rifampicin in combination anti-TB drugs are met.

 2. Chemotherapy of experimental tuberculosis of mice with rhizobutol in a dose (70 mg / kg of animal weight), 2 times less than the established therapeutic dose (used in the prototype - 140 mg / kg of animal weight), showed the treatment efficiency (80.1%), almost such the same as in the prototype (82.6%). This suggests the presence of a synergistic effect in the components of rhizobutol, possibly due to the reciprocal effect of the bacteriostatic effect and the general toxicity of the drug with prolonged administration, other possible interdependent processes in vivo (see, for example, Kiselev V.I. Kanamitsin sulfate - inducer of hesa Gene Escherichia coli. Molecular Genetics, Microbiology and Virology, 1989, 5, pp. 41-43). It is interesting that the bacteriostatic effect in vitro, when the general toxicity of the drug cannot be taken into account, is achieved only at the same concentration in the nutrient medium of both rhizobutol and the nutrient mixture of the prototype, as well as isoniazid, the most active and widely used in the clinic of tuberculosis the drug.

 3) The study of hematological and biochemical parameters as indicators of toxicity with long-term use of rhizobutol indicates that it does not have toxic properties.

 The active substances in a joint presence are stable during manufacturing and storage, the conditions for tableting and encapsulation are standard, the processes can be carried out on standard domestic equipment.

 The invention is illustrated by the following examples: example 1 describes the compositions used in examples 2-5, example 2 describes the pharmacokinetics of rifampicin in the composition of rhizobutol; Example 3 describes the chemotherapeutic efficacy of rhizobutol in experimental tuberculosis in mice; example 4 shows hematological and biochemical parameters as indicators of the toxicity of the drug; Example 5 is devoted to the clinical treatment of rhizobutol in a limited group of patients with a diagnosis of focal tuberculosis.

 Example 1

 Obtaining compositions.

 To 150 mg of rifampicin, 275 mg of ethambutol, 75 mg of isoniazid, 400 mg of pyrazimide, 10 mg of pyridoxine were added 100 mg of metacel (corresponding to 10% of the total weight of the composition). The mixture is thoroughly mixed on the mixer for 15 minutes, dried in a shelf type dryer (Münsterman), passed through a granulator RF-3056 and tabletted on a RTM-41 rotary tablet press.

The result is a composition of the following composition, which is used in examples 2-4, mg:
Isoniazid - 75
Rifampicin - 150
Ethambutol - 275
Pyrazinomide - 400
Pyridoxine - 15
Compositions with different concentrations of the components are also prepared.

 Example 2

 Pharmacokinetics of rifampicin in the composition of rhizobutol.

The determination of rifampicin in the blood plasma of mice was performed using reverse phase HPLC. For this, a Gilson M302 pump (USA), an Rheodyne 7125i injector (USA), and a Carlo Erba microUVIS20 photometric detector (Italy) were used. The analysis was carried out using a column packed with XTerra RP ₈ reversed-phase sorbent 150x4.6 mm (Waters, USA) using 40% acetonitrile in 0.1 M ammonium acetate buffer solution, pH 4.6, as a mobile phase at a flow rate 1.2 ml / min. Detection was carried out at 254 nm.

Samples for analysis were prepared according to the following method: animal blood plasma was obtained after centrifugation at 2500 rpm for 20 min, then it was extracted with 5 volumes of a mixture of CHCl ₃ : MeOH = 4: 1, the extractant was dried in a stream of argon, the dry residue was dissolved in 0 2 ml of the mobile phase.

 The amount of rifampicin in plasma samples was determined by the peak area corresponding to the antibiotic release time. For this, a calibration curve was constructed on the basis of the results of HPLC analysis of aliquots of a rifampicin solution prepared using an exact sample of the substance. The analysis data are presented in table 1.

 Example 3

 Chemotherapeutic efficacy of rhizobutol in experimental tuberculosis of mice.

 To determine the chemotherapeutic efficacy of rhizobutol, experimental reproduction of hematogenously disseminated tuberculosis was carried out on 85 C57BL / 6 mice; laboratory animals were obtained from the Central Animal Nursery of the Russian Academy of Medical Sciences. The experiment included quarantined mice, standardization by weight (20-25 g) and gender. Infection was carried out by a two-week virulent culture of M. tuberculosis H37Rv by intravenous (into the tail vein) administration of a suspension of mycobacteria at a dose of 2-10 CFU (colony forming units) in 0.5 ml of physiological saline. All experimental animals were divided into 5 groups, depending on the chemotherapy regimen used, 17 animals in each therapeutic group.

 Treatment of animals was started 14 days after infection, when multiple foci of tuberculosis infection were macroscopically determined in the lungs.

 Chemotherapy was carried out daily (except Sunday) for 2 months. The drugs were administered orally.

 A macroscopic evaluation of the effectiveness of each chemotherapy regimen was expressed by an efficiency index calculated using a special formula developed by A. N. Torgunova (see Methods of Experimental Chemotherapy (Practical Guide). Ed. 2, edited by G.N. Pershin. M: Medicine , 1971).

 Microbiological examination included smear-smear bacterioscopy and culture. The growth rate of crops was taken into account on a four-point scale proposed by G. N. Pershin. (see Order 558 of the Ministry of Health of the USSR of 06/08/1978 M., 1978, guidelines, Appendix 2).

 1 month of treatment.

 A macroscopic examination of the internal organs of mice that did not receive chemotherapy (group 2) showed pronounced signs of tuberculosis inflammation (large foci of tuberculosis infection in the lungs). The index of damage to the internal organs of mice in this group was 2.8 U (see table. 2).

 A macroscopic picture of the internal organs of animals treated with a mixture of the prototype preparations in the studied dose (group 3) corresponds to a lesion index of 1.8 units, i.e. in the lungs of experimental animals, single foci of tuberculosis infection were determined (see Table 2).

 Macroscopic examination of the internal organs of mice treated with rhizobutol for 1 month (groups 4 and 5) also showed the presence of tuberculosis infection. The lesion index was 1.5 and 1.5 units, respectively.

 The results of bacterioscopy and the study of inoculum obtained from mice that participated in the experiment for 1 month showed the presence of mycobacteria in all studied parenchymal organs (see table 3).

 2 months of treatment.

 A macroscopic examination of the internal organs of mice in the control group that did not receive chemotherapy showed marked signs of tuberculosis inflammation (large necrotic foci of tuberculosis infection in the lungs) by the end of the experiment. The index of damage to the internal organs of mice of this group was 3.5 U (see Table 3).

 The picture of damage to the parenchymal organs of mice of group 3 is close to that in groups 4 and 5, the index of damage in these cases is 0.6, 0.5, and 0.7 units, respectively, i.e. pronounced foci of infection were not detected. The effectiveness of chemotherapy in these groups is 82.6, 85.7 and 80.1%, respectively (see table 2).

 A decrease in the therapeutic dose of rhizobutol by 2 times from the maximum led to a decrease in efficiency by only 5.6%.

 A study of fingerprints and inoculum in the groups treated with the drug mixture and rhizobutol for 2 months revealed a significant positive effect of chemotherapy with a dose of 140.0 mg / kg. The results of microbiological studies of parenchymal organs showed sterilization of the test material from mycobacterium tuberculosis.

 At a dose of 70 mg / kg rhizobutol, bacterioscopy data did not reveal the presence of mycobacteria in smears from pathological material, and the growth of mycobacteria in crops was insignificant.

 Data on the inoculation of mycobacteria from the spleen of infected animals reflect the picture of the course of chemotherapy as a whole. The results are shown in table 4.

By the end of the experiment (2 months of treatment), no colony of mycobacteria was obtained from the spleens of mice of group 4. In groups 3 and 5 of mice treated with rhizobutol at a dose of 70.0 mg / kg, the mycobacterial inoculation rate amounted to several tens of colonies per organ of the infected mouse, which is an indicator of the effectiveness of therapy. In the control group of mice (group 2), the numerical indicator of seeding was 10.6 • 10 ⁷ colonies per infected organ of the mouse, which indicates an extensive process of infection of all organs of several survivors by the end of the experiment.

 Thus, as a result of an experimental study, high chemotherapeutic efficacy (85.7%) of the new anti-tuberculosis drug rhizobutol was established. The use of the test drug in half dose also gave a very high therapeutic efficacy result - 80.1%, which will allow the use of rhizobutol for half-dose therapy and thus further reduce the toxic effects of treatment.

 Example 4

 Toxic effects observed with prolonged use of anti-TB drugs.

 The experimental conditions are described in example 3.

 The data are given in tables 5 and 6.

 A comparative study of hematological and biochemical parameters as an indicator of toxicity with prolonged use of rhizobutol and a drug mixture according to the prototype indicates that rhizobutol, in contrast to the prototype, does not have toxic properties (see tables 5 and 6), even at a dose of 140 mg / kg.

 Example 5

 Clinical therapy with a limited number of patients with rhizobutol.

 The composition of Example 1 was used containing 20 mg of pyridoxine.

 The group consisted of 5 patients, treatment with rhizobutol was prescribed at a dose of 140 mg / kg per day, after 3 weeks X-ray, biochemical and hematological studies were carried out. In all cases, there was a positive trend, there were no complaints, including neurological symptoms.

Claims (1)

A combined anti-tuberculosis drug containing, as an active principle, a mixture of rifampicin, ethambutol and isoniazid and an inert filler, characterized in that it additionally contains pyrazinomide and pyridoxine, and as a filler 2-15 weight. % metacel in the following ratio, mg:
Rifampicin - 140-160
Ethambutol - 270-280
Isoniazid - 70-80
Pyrazinomide - 400
Pyridoxine - 10-15

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