High-purity large-scale preparation of stannsoporfin

Reference to related requests

p00001The present application claims the priority benefit of the provisional United States patent application No. 60 / 849,641, filed on October 4, 2006, and of the provisional United States patent application No. 60 / 904,601, filed on February 28, 2007.

Field of the Invention

p00002The present invention relates to methods for synthesizing stansoporphine (tin mesoporphyrin IX dichloride (IV)) in large quantities and with high purity.

Background

p00003Stansoporphine, or tin (IV) mesoporphyrin IX dichloride, is an inhibitor of the heme-oxygenase enzyme. Stansoporphine has been proposed for therapeutic use in various diseases, such as childhood hyperbilirubinemia (U.S. Patent 4,657,902; U.S. Patent 4,668,670; WO 94/28906) and psoriasis (U.S. Patent 4,782,049). Due to its pharmaceutical utility, the procedures for preparing stansoporphine are of great interest.

p00004Childhood hyperbilirubinemia (also known as childhood jaundice or neonatal hyperbilirubinemia) occurs in a newborn when the liver is unable to conjugate bilirubin to excrete it at a rate proportional to the formation of bilirubin. Bilirubin comes from the release of hematine as part of the physiological conversion of fetal hemoglobin to adult at birth. The heme-oxygenase enzyme oxidizes hematine to biliverdin; The enzyme biliverdin reductase then reduces biliverdin to bilirubin. Bilirubin in elevated serum levels is a neurotoxic substance. In adult humans, the liver quickly converts bilirubin into a conjugated excretable form. In newborn humans, the liver is still developing and liver uptake and conjugation is not as efficient as in adults. In addition, hemolysis can be carried out at a relatively faster rate than in adults. All these factors can lead to excess bilirubin in the child. For some children, high serum bilirubin levels can have harmful physiological consequences. Bilirubin is yellow and children with excess bilirubin have a yellow pigmentation in their skin and in the whites of their eyes.

p00005Children who have very high levels of serum bilirubin have the risk of developing kernicterus, a rare but potentially devastating neurological disorder that can result in serious lifelong complications and complications such as athetosis, hearing loss, Vision problems and dental problems. (See the global disease control and prevention centers Web.cdc.gov/ncbddd/dd/kernicterus.htm.). Therefore, children should undergo careful controls after birth and therapeutic intervention should begin if a child's bilirubin level is excessive. The American Academy of Pediatrics published clinical practice guidelines to evaluate newborns to detect hyperbilirubinemia and treat newborns at risk; see Pediatrics 1 14: 297-316 (2004). As the costs of medical care have increased in the United States, seemingly healthy newborns and their mothers are discharged quickly, some only 24 to 48 hours after birth. However, it is believed that this practice may have contributed to an increase in cases of kernicterus that had been virtually eliminated in developed countries; see Hansen TWR, Acta Paediatr. 89: 1155-1157 (2000)). Because premature discharge can delay the detection of jaundice and hyperbilirubinemia in children, it is preferable to apply effective means to quickly treat hyperbilirubinemia. The unique medical condition of the newborn also requires that all means of treatment be as safe as possible since side effects that are tolerable in adults may be completely unacceptable in neonates.

p00006Currently approved and commonly used hyperbilirubinemia treatments include phototherapy and exchange transfusion. Phototherapy involves irradiating the newborn with light in the range of 430 to 490 nm (blue light). The light converts bilirubin into lumirubin and photobilirubin, which is excreted more rapidly by the child and thus can result in a reduction in bilirubin levels.

p00007Stansoporphine (tin mesoporphyrin IX dichloride (IV)) was shown to have therapeutic value for treating hyperbilirubinemia; see Valaes et al., Pediatrics 93: 1-11 (1994) and Kappas et al., Pediatrics, 95: 468-474 (1995). Other indications in which stansoporphine can be used is disclosed in U.S. Patent 4,692,440 (to increase the rate of hematine excretion), WO 89/02269 (to counteract the toxicity of cancer therapy), U.S. Patent 4,782,049 ( to treat psoriasis) and other publications.

p00008In U.S. Patent No. 6,818,763, the publication of U.S. Patent Application 2004/0210048 and U.S. Patent Application No. 11 / 096,359 disclose methods for synthesizing stansoporphine.

p00009WO 03/101999 A2 discloses a process for the preparation of metal halide compounds of mesoporphyrin. WO 2004/045546 A2 discloses water soluble mesoporphyrin compounds and the processes for their preparation.

p00010However, it is still desirable to develop procedures for producing high purity stansoporphine, due to the therapeutic advantages of using a substance as pure as possible and also due to the rigorous requirements of regulatory agencies.

p00011The present application discloses procedures for synthesizing stansoporphine at a level of purity not hitherto achieved, as well as large-scale pure stansoporphine preparations. In the present application a new method of inserting tin and other metals into the porphyrin rings is also disclosed. This new procedure can significantly reduce the time required for the synthesis of stansoporphine.

Description of the invention

p00012The present invention relates to a process for preparing stansoporphine according to claim 1.

p00013The process of the invention makes it possible to obtain a substance composition comprising high purity stansoporphine on a large scale (or in bulk). The process of the invention makes it possible to obtain a substance composition comprising high purity stansoporphine on a large scale (or in bulk) when produced in a single batch, that is, a high purity stansoporphine on a large scale (or in bulk) in a single lot. The high purity stansoporphine can be at least about 97% pure, at least about 98% pure, at least about 98.5% pure, at least about 99% pure, at least about 99.5% pure or at least about 99.8% pure. The amount of any single impurity in the high purity stansoporphine may be less than about 0.1%, less than about 0.09%, less than about 0.08% or about 0.07% or less; In another embodiment, any single impurity is a product related impurity. The amount of large-scale stansoporphine (in bulk) may be at least about 10 grams, at least about 25 grams, at least about 50 grams, at least about 100 grams, at least about 200 grams, at least about 500 grams, at least about 1.0 kg, at least about 2.0 kg or at least about 5.0 kg. In one embodiment, the high purity stansoporphine as described in various ways above, is produced in a single batch.

p00014The process of the invention allows obtaining a large-scale amount of stansoporphine that is at least about 97% pure, at least about 98% pure, at least about 98.5% pure, at least about 99% pure, at least about 99.5% pure or at least about 99.8% pure and has no impurities in an amount greater than about 0.2% and more preferably, it has no impurities in an amount greater than about 0.15% and even more preferably it has no impurities in an amount greater than about 0.12%. In one embodiment, high purity stansoporphine is produced in a single batch.

p00015In further embodiments, the amount of palladium impurities present in a large-scale amount of high purity stansoporphine is less than about 20 ppm, less than about 15 ppm, less than about 10 ppm or less than about 5 ppm. In one embodiment, high purity stansoporphine is produced in a single batch.

p00016The invention encompasses a process for preparing large-scale high purity stansoporphine according to claim 1, which comprises the following steps: a) exposing a metal hydrogenation catalyst to a hydrogen atmosphere to form a prehydrogenated catalyst; and b) contacting the hemin with the prehydrogenated catalyst and maintaining the hemin and the catalyst in one or more combinations of temperature, hydrogen pressure and a sufficient time to extract the iron from the hemin and reduce the vinyl groups of the hemin to ethyl groups, to thereby form mesoporphyrin IX. The invention encompasses a process for preparing large-scale high purity stansoporphine according to claim 1, which comprises the following steps: a) exposing a metal hydrogenation catalyst to a hydrogen atmosphere to form a prehydrogenated catalyst; b) contacting the hemin with the prehydrogenated catalyst and maintaining the hemin and the catalyst in one or more combinations of temperature, hydrogen pressure and sufficient time to extract the iron from the hemin and reduce the vinyl groups of the hemin to groups ethyl, to thereby form mesoporphyrin IX; and c) reacting mesoporphyrin IX with a tin (II) oxide to form stansoporphine by using a controlled rate of oxidation. In one embodiment, the metal hydrogenation catalyst comprises palladium, palladium on carbon, platinum on carbon, nickel or a nickel-aluminum catalyst. In another embodiment, the metal hydrogenation catalyst is palladium. In another embodiment, the metal hydrogenation catalyst is palladium on carbon. The process can produce a large-scale amount of high purity stansoporphine in a single batch.

p00017In another embodiment, the invention encompasses a process for preparing large-scale high purity stansoporphine comprising the following steps: a) exposing a metal hydrogenation catalyst to a hydrogen atmosphere to form a prehydrogenated catalyst; b) contacting the hemin with the prehydrogenated catalyst and maintaining the hemin and the catalyst in one or more combinations of temperature, hydrogen pressure and sufficient time to extract the iron from the hemin and reduce the vinyl groups of the hemin to groups ethyl, to thereby form mesoporphyrin IX; and c) reacting mesoporphyrin IX with a tin (II) oxide to form stansoporphine. In one embodiment, the metal hydrogenation catalyst comprises palladium, palladium on carbon, platinum on carbon, nickel or a nickel-aluminum catalyst. In another embodiment, the metal hydrogenation catalyst is palladium. In another embodiment, the metal hydrogenation catalyst is palladium on carbon. The process can produce a large-scale amount of high purity stansoporphine in a single batch.

p00018In another embodiment, the invention encompasses a process for preparing mesoporphyrin IX, which comprises the following steps: a) exposing a palladium on carbon catalyst to a hydrogen atmosphere to form a prehydrogenated palladium catalyst; and b) contacting the hemin with the prehydrogenated catalyst and maintaining the hemin and the catalyst at one or more combinations of temperature, hydrogen pressure and a sufficient time to extract the iron from the hemin and reduce the vinyl groups of the hemin to groups ethyl, to thereby form mesoporphyrin IX. In further embodiments, step b) is carried out at about 80 to 100 ° C, preferably at about 85 to 90 ° C, with hydrogen pressure at about 50 to 70 psi, preferably at about 55 to 60 psi, for about 1 to 3 hours , preferably about 1 to 1.5 hours; then at about 40 to 60 ° C, preferably at about 45 to 50 ° C, with hydrogen pressure at about 50 to 70 psi, preferably at about 55 to 60 psi, for about 18 to 48 hours, preferably about 24 hours.

p00019In another embodiment, the invention encompasses a rapidly filterable mesoporphyrin IX dihydrochloride preparation, where at least about 10 grams can be filtered in less than about 90 minutes, less than about 60 minutes, less than about 45 minutes, less than about 35 minutes, less than about 25 minutes or less than about 10 minutes, from a solution where the amount of solvent is present in at least about a 50-by weight ratio with respect to the amount of mesoporphyrin dihydrochloride IX. In another embodiment, the invention encompasses a rapidly filterable mesoporphyrin IX dihydrochloride preparation, where at least about 1,000 grams can be filtered in less than about 1 day, less than about 12 hours, less than about 6 hours, less than about 4 hours, less than about 3 hours or less than about 2 hours, from a solution where the amount of solvent is present in at least about a 50-by weight ratio with respect to the amount of mesoporphyrin dihydrochloride

p00020IX. In one embodiment, the solvent is a mixture of water, hydrochloric acid and formic acid; The hydrochloric acid can be hydrochloric acid at approximately 31% before mixing.

p00021In another embodiment, the invention encompasses a process for preparing a readily filterable mesoporphyrin IX dihydrochloride preparation, comprising the step of adding an aqueous solution of hydrochloric acid to a format solution of mesoporphyrin IX in formic acid. In one embodiment, the concentration of hydrochloric acid in the aqueous solution is about 0.5 to 2.0 N. In another embodiment, the concentration of hydrochloric acid in the aqueous solution is about 0.75 to 1.25 N. In another embodiment, the concentration of hydrochloric acid in the aqueous solution is about 1.0 N.

p00022In another embodiment, the invention encompasses a method for inserting tin into mesoporphyrin IX, which comprises reacting mesoporphyrin IX with a tin salt in the absence of a proton scrubber.

p00023In another embodiment, the invention encompasses a method for inserting tin into mesoporphyrin IX, which comprises reacting mesoporphyrin IX with a tin salt at a controlled rate of oxidation. In one embodiment, mesoporphyrin IX is reacted with a tin salt in a reaction vessel that has an air space and the oxidation rate is controlled by introducing an oxygen-containing gas into the air space of the container. reaction. In another embodiment, the oxygen-containing gas introduced into the air space of the reaction vessel has approximately 3 to 22% oxygen in an inert gas such as nitrogen. In another embodiment, the oxygen-containing gas introduced into the air space of the reaction vessel is air. In another embodiment, the oxygen-containing gas introduced into the air space of the reaction vessel has approximately 4 to 15% oxygen in an inert gas such as nitrogen. In another embodiment, the oxygen-containing gas introduced into the air space of the reaction vessel has about 5 to 10% oxygen in an inert gas such as nitrogen. In another embodiment, the oxygen-containing gas introduced into the air space of the reaction vessel has approximately 6% oxygen in an inert gas such as nitrogen. In another embodiment, the oxygen-containing gas introduced into the air space of the reaction vessel is approximately 6% oxygen in nitrogen.

p00024In any of the above-mentioned embodiments, the large-scale amount of high purity stansoporphine can be produced in a single batch.

p00025In any of the above-mentioned embodiments, the reagents, intermediates and / or products may be subjected to additional purification steps. In some embodiments, further purification comprises treating the reagent, intermediate or product with diatomaceous earth and / or activated carbon. In one embodiment, the treatment of the reagent, intermediate or product with diatomaceous earth and / or activated carbon comprises dissolving or suspending the reagent, intermediate and / or product in a solvent, adding diatomaceous earth and / or activated carbon, distilling off diatomaceous earth and / or activated carbon and recover the reagent, intermediate or filtrate product. In some embodiments, further purification comprises grinding the reagent, intermediate or product with hot acid, such as about 0.1 to 6N HCl in water, preferably about 3N HCl in water. In some embodiments, one, two or three stages for treating with diatomaceous earth, treating with activated carbon and grinding with hot acid are performed sequentially, in any order and can be repeated as desired.

p00026The invention encompasses a process for inserting tin into a porphyrin compound or its salt through the use of tin (II) oxide. In another embodiment, the porphyrin compound is a mesoporphyrin or its salt or a protoporphyrin.

<dl><dt /><dd>or its salt or a hematoporphyrin or its salt. In another embodiment, the porphyrin compound is mesoporphyrin IX or its salt. In another embodiment, the porphyrin compound is mesoporphyrin IX dihydrochloride. In another embodiment, the resulting product is tin porphyrin (IV) or its salt. In another embodiment, the resulting product is tin mesoporphyrin</dd></dl>

<dl><dt>(IV) </dt><dd>or its tin salt or protoporphyrin (IV) or its salt or tin hematoporphyrin (IV) or its salt. In another embodiment, the resulting product is tin mesoporphyrin IX (IV) or its salt.</dd></dl>

In another embodiment, the invention encompasses a process for inserting tin into a porphyrin compound or its salt by supplying a porphyrin compound or its salt, supplying tin oxide (II) and contacting tin oxide ( II) with the porphyrin compound or its salt under acidic conditions, whereby tin oxide

<dl><dt>(II) </dt><dd>it is inserted into the porphyrin ring to provide a tin porphyrin compound (IV). In another embodiment, the tin (II) oxide is dissolved or suspended in acetic acid or formic acid, preferably acetic acid. In another embodiment, the porphyrin compound or its salt is dissolved or suspended in formic acid or acetic acid, preferably formic acid. In another embodiment, the equivalent proportion of the total amount of tin oxide</dd></dl>

<dl><dt>(II) </dt><dd>used with respect to the total amount of porphyrin compound or its salt used is about two to six, preferably about four. In another embodiment, the solution or suspension of porphyrin compound or its salt is added dropwise to the tin (II) oxide solution. The drip addition can be carried out over a period of about three to nine hours, preferably more than about six hours. The tin (II) oxide solution or suspension is maintained at a temperature of about 25115 ° C, preferably about 50-75 ° C, more preferably about 60-65 ° C. </dd></dl>

In another embodiment, the invention encompasses a process for inserting tin into a porphyrin compound or its salt (such as mesoporphyrin IX or its salt, such as mesoporphyrin dihydrochloride IX) by supplying a mesoporphyrin compound or its salt (such as mesoporphyrin IX or its salt, such as mesoporphyrin dihydrochloride IX), the supply of tin (II) oxide and contacting tin (II) oxide with the mesoporphyrin compound or its salt (such as mesoporphyrin IX or its salt, as mesoporphyrin dihydrochloride IX) under acidic conditions, whereby the tin oxide (II) is inserted into the porphyrin ring (such as a mesoporphyrin ring IX) to produce a tin porphyrin compound (IV). In another embodiment, the tin (II) oxide is dissolved or suspended in acetic acid or formic acid, preferably acetic acid. In another embodiment, the mesoporphyrin compound or its salt (as mesoporphyrin IX or its salt, as mesoporphyrin IX dihydrochloride) is dissolved or suspended in formic acid or acetic acid, preferably formic acid. In another embodiment, the equivalent proportion of the total amount of tin (II) oxide used with respect to the total amount of mesoporphyrin compound or its salt (such as mesoporphyrin IX or its salt, as mesoporphyrin IX dihydrochloride) used is approximately two to six, preferably about four. In another embodiment, the solution or suspension of the porphyrin compound or its salt (such as mesoporphyrin IX or its salt, as mesoporphyrin dihydrochloride IX) is added dropwise to the tin oxide (II) solution. The drip addition can be carried out over a period of about three to nine hours, preferably more than about six hours. The tin oxide solution or suspension

<dl><dt>(II) </dt><dd>it is maintained at a temperature of about 25-115 ° C, preferably about 50-75 ° C, more preferably about 60-65 ° C. After completion of the drip addition, the reaction mixture can be maintained at a temperature of about 60 to 65 ° C for about 18 to 24 additional hours. The reaction mixture can be cooled and filtered after the additional reaction time.</dd></dl>

p00027In another embodiment, the invention encompasses a process for producing a tin porphyrin compound (IV) or its salt comprising a) preparing a solution or suspension of a non-metalated porphyrin compound or its salt; b) preparing a solution or suspension of tin (II) oxide, where steps (a) and (b) can occur in any order or simultaneously; and c) contacting the tin oxide (II) solution or suspension with the solution or suspension of the non-metalated porphyrin compound or its salt under suitable conditions to form the tin porphyrin compound (IV) or its salt. The solution or suspension of tin (II) oxide and the solution or suspension of the non-metalated porphyrin compound or its salt can be prepared independently with formic acid or acetic acid; for example, the solution or suspension of tin (II) oxide can be prepared with acetic acid and the solution or suspension of the non-metalated porphyrin compound or its salt can be prepared with formic acid. The unmetalated porphyrin compound is selected from mesoporphyrins, protoporphyrins, hematoporphyrins and their salts, such as mesoporphyrin IX or its salt, as mesoporphyrin dihydrochloride IX. The contact step c) may comprise adding the solution or suspension of the non-metalated porphyrin compound or its drip salt to the solution or suspension of tin (II) oxide under suitable conditions to form the porphyrin compound (IV) or its Salt. The drip addition can be completed between about 3 to 9 hours, like about 6 hours. The tin oxide (II) solution or suspension can be maintained at a temperature of about 60 to 65 ° C during drip addition. After the completion of the addition of the solution or suspension of the non-metalated porphyrin compound or its drip salt to the solution or suspension of tin (II) oxide, the reaction mixture can be maintained at a temperature of about 60 to 65 ° C for approximately 18 to 24 additional hours. The reaction mixture can be cooled and filtered after the additional reaction time. In another embodiment, the process for producing a tin porphyrin compound (IV) or its salt is performed in the absence of a proton scrubber or proton sponge.

p00028Tin (IV) mesoporphyrin produced by any of the procedures described above may undergo additional purification steps. In some embodiments, further purification comprises treating tin (IV) mesoporphyrin with diatomaceous earth and / or activated carbon. In one embodiment, the treatment of tin (IV) mesoporphyrin with diatomaceous earth and / or activated carbon comprises dissolving or suspending tin (IV) mesoporphyrin in a solvent, adding diatomaceous earth and / or activated carbon, remove by Distillation of diatomaceous earth and / or activated carbon and recover tin mesoporphyrin (IV) from the filtrate. In some embodiments, further purification comprises triturating tin (IV) mesoporphyrin with hot acid, such as about 0.1 to 6N HCl in water, preferably about 3N HCl in water, at a temperature of about 60 to 95 ° C, preferably about 80 at 95 ° C, more preferably about 85 to 90 ° C. In some embodiments, one, two or all three stages for treating with diatomaceous earth, treating with activated carbon and grinding with hot acid are performed sequentially, in any order and can be repeated as desired.

p00029The process of the invention allows to provide high purity stansoporphine in a large scale (or bulk) amount, where said high purity stansoporphine is stable for at least about three months or at least about six months in storage. In another embodiment, the high purity stansoporphine produced in large quantities is prepared as a single batch. In another embodiment, the process of the invention allows to provide high purity stansoporphine in a large scale (or bulk) amount, wherein said high purity stansoporphine retains its high purity for at least about three months or at least about six months in storage. . In another embodiment, the storage conditions are about 25 ° C and about 60% relative humidity. In another embodiment, the storage conditions are about 40 ° C and about 75% relative humidity. In another embodiment, the stansoporphine is stored in a polyethylene bag. In another embodiment, the stansoporphine is stored in a polyethylene bag inside another polyethylene bag. In another embodiment, the stansoporphine stored in two bags is stored in a high density polyethylene drum. In another embodiment, each poly bag has a thickness of about 4 mils (about 4/1000 of an inch or about 0.1 millimeters).

p00030The process of the invention allows stansoporphine to be provided for use in a method for treating childhood hyperbilirubinemia, which comprises administering stansoporphine to a patient in need of such treatment, where stansoporphine was produced with high purity and on a large scale. In another embodiment, stansoporphine is produced as a single batch.

p00031The procedure to prevent childhood hyperbilirubinemia comprises administering stansoporphine to a patient in need of such prevention, where stansoporphine was produced with high purity and on a large scale. In another embodiment, stansoporphine is produced as a single batch.

Detailed description of the invention

p00032In one embodiment of the present invention, stansoporphine is prepared in large quantities and with high purity. In another embodiment of the present invention, stansoporphine is prepared in large quantities with high purity, where the large quantity is prepared as a single batch. Stansoporphine (tin mesoporphyrin IX dichloride (IV)); Chemical Abstracts registration number 106344-20-1) is also known by the trade name Stanate®, which is a registered trademark of InfaCare Pharmaceutical Corp., Plymouth Meeting, Pennsylvania. Stansoporphine has the following structure:

p00033It has the molecular formula C34H36Cl2N4O4Sn and has a molecular weight of 754.29.

p00034The terms "large quantities", "large scale" or "bulk" refer to at least about 10 grams. Other quantities for large-scale stansoporphine production are at least about 25 grams, at least about 50 grams, at least about 100 grams, at least about 200 grams, at least about 500 grams, at least about 1.0 kg, at least about 2.0 kg or at least about 5.0 kg.

p00035The term "single lot" refers to the amount of the specified product being synthesized only once. The single batch generally occurs after a reaction (or series of reactions) is carried out once (note that a single preparation of the compound subjected as a whole to one or more reactions repeatedly, as repeated purifications, is considered a single lot). Therefore, the single batch excludes multiple preparations of a compound made on separate occasions or in divided quantities that are then combined.

p00036The term "high purity" refers to a preparation that meets the following two criteria: 1) the total level of purity is at least about 97%, that is, the desired product (stansoporphine) represents at least 97% of the preparation; and 2) any individual impurity present related to the product is present in an amount less than about 0.1% of the preparation. Purity is preferably measured by HPLC analysis. The "product-related" impurity is an impurity that requires characterization by the guidelines of the United States Food and Drug Administration; therefore, the components of the drug product such as water are not considered an impurity.

p00037The term "non-metalated porphyrin" refers to a porphyrin that lacks a metal ion coordinated by one or more pyrrole nitrogens. "Metalated porphyrin" is a porphyrin that has a metal ion coordinated by at least one pyrrole nitrogen.

p00038The term "intermediate oxidation state" refers to an element, such as a metal, that is present in an intermediate oxidation state between its neutral state (no charge or zero oxidation state) and its more highly oxidized state. By way of non-limiting example, iron generally forms oxidation states of (0), (II), and (III); the oxidation state (II) (ferrous state) is an intermediate oxidation state.

p00039The purity of the preparation is important for the use of the compound as a pharmaceutical product. The general level of purity may be at least about 97%, at least about 98%, at least about 98.5%, at least about 99% or at least about 99.5%. A high purity preparation is also defined, as set forth above, as a preparation with the additional condition that any individual impurity present is present in an amount less than about 0.1% of the preparation. (Note that the total amount of impurities can exceed 0.1% - for example, an impurity can be present in 0.08% and another in 0.07%, which amounts to a total of 0, 15% - but when measured individually, no impurity is present in amounts equal to or greater than about 0.1%). In another embodiment, any individual impurity present is present in an amount less than about 0.09%. In another embodiment, any individual impurity present is present in an amount less than about 0.08% or less. In another embodiment, any individual impurity present is present in an amount of about 0.07% or less. Water may be present in the preparation even in significant amounts (at least about 1% to 5%), but it is not considered an impurity. Other residual solvents, such as acetone, formic acid and acetic acid, are also not considered impurities, especially if they are within the permitted levels described in the guidelines of the international conference on the harmonization of technical requirements for the registration of pharmaceutical products for human use, harmonized tripartite guideline ICH - Impurities: Guideline for residual solvents, Q3C (R3), Stage 4 version, November 2005 (www.ich.org/LOB/media/MEDIA423.pdf) or below levels.

p00040In an alternative embodiment, stansoporphine has no impurities present in an amount greater than about 0.2% and more preferably has no impurities present in an amount greater than approximate

p00041mind 0.15% and even more preferably has no impurities present in an amount greater than about 0.12%.

p00042The current synthesis produces stansoporphine that meets the two criteria listed above regarding high purity (total purity of at least about 97%, without treating water or residual solvents as impurities and any individual impurity present is present in an amount about 0.1% or less). The second criterion in relation to the level of individual impurities is of interest due to regulatory requirements. The United States Food and Drug Administration generally requires a detailed characterization of impurities at a level equal to 0.1%, while impurities present at a level below 0.1% need not be characterized in detail unless that have unusually potent pharmacological or toxic effects at a level lower than 0.1% (see Guidance for Industry: AN-DAs: Impurities in Drug Substances, US publications Department of Health and Human Services, Food and Drug Administration, Center for Drug Evaluation and Research (CDER), November 1999; available at www.fda.gov/cder/guidance/2452fnl.htm; and Guidance for Industry, Q3A Impurities in New Drug Substances, United States Department of Health and Human Services, Food and Drug Administration, Center for Drug Evaluation and Research (CDER) and Center for Biologies Evaluation and Research (CBER), February 2003 ICH , Revision 1, available at www.fda.gov/cder/guidance/4164fnl.pdf). By complying with the threshold conditions established by the Food and Drug Administration, the high purity material has significant advantages over the material of lower purity from the regulatory point of view.

p00043Another advantage of the invention as described herein is the expected reproducibility of the synthesis, which provides the ability to generate repeated lots of high purity stansoporphine in large quantities. Another advantage of the invention as described herein is the ability of the process and the product to meet the requirements of good manufacturing practices (GMP), as defined by law, standards or in the requirements of regulatory agencies in several countries (for example, good manufacturing practices indicated in the code of federal provisions of the United States, title 21, sections 210 and 211).

p00044Another advantage of the invention as described herein is the production of high purity stansoporphine in bulk quantities in a single batch, with concomitant advantages of increased homogeneity, lower synthetic cost and relative ease of characterization.

p00045Synthesis of high purity stansoporphine

p00046Because porphyrins are light sensitive compounds, starting materials, intermediates, products and solutions or their suspensions should be protected from exposure to light and stored in a dark location in containers that protect them from light.

p00047The synthesis of stansoporphine proceeds with the hemin (protoporphyrin chloride IX of iron (III)) as a starting material. The quantities required for large-scale synthesis are obtained from porcine red blood cells. Hemin DMF is obtained in Harimex (Loenen, The Netherlands); The material is used without purification before use (the purity with which it is provided is greater than approximately 98% by HPLC). The hemin is heated in an organic solvent with a hydrogenation catalyst on carbon under a hydrogen atmosphere. This reductive stage serves both to extract the Fe ion from the porphyrin ring and to reduce the vinyl groups of protoporphyrin IX to ethyl groups (which thereby converts protoporphyrin IX into mesoporphyrin IX), as indicated in the following scheme.

p00048H2, Pd / C, HCOOH

p00049A preferred hydrogenation catalyst is palladium on carbon, used in an amount of about 0.0135 to 0.0165 equivalents, preferably about 0.015 equivalents. Other suitable catalysts may be used, including metallic particles of palladium, platinum on carbon, metallic particles of platinum, nickel or a nickel-aluminum catalyst, provided that the amounts of catalyst in the product meet the pharmaceutical specifications. The nickel-aluminum catalyst can be RANEY nickel (RANEY is a registered trademark of WR Grace &amp; Co., New York, New York). A preferred organic solvent is formic acid.

p00050It was found that pretreatment of the Pd / C catalyst with hydrogen gas prior to the addition of the hemin to the reaction reduces the impurities of the palladium and thereby contributes to the total purity of the final stansoporphine product. Without prior hydrogenation of the catalyst before the addition of hemin, palladium residue levels of 50 ppm were detected in the product, which is significantly above the specifications of the residual palladium product of less than about 20 ppm. With the previous hydrogenation stage, the residual palladium was reduced to undetectable levels (residual palladium of less than about 5 ppm). Therefore, the improved synthesis provides residual palladium levels in the tin mesoporphyrin IX dichloride product.

p00051(IV) less than about 20 ppm of palladium, preferably less than about 15 ppm of palladium, more preferably less than about 10 ppm of palladium, even more preferably less than about 5 ppm of palladium. The prior hydrogenation of the catalyst can be performed under a hydrogen atmosphere of about 15 to 75 psi (about 1 to 5 bar; about 100,000 to

p00052500,000 Pascals), preferably approximately 30 to 50 psi (approximately 2 to 3.5 bar; approximately

p00053200,000 to 350,000 Pascals), more preferably about 40 psi (about 2.75 bar or 275,000 Pascals). The temperature for the prior hydrogenation of the catalyst may vary between about 25 and 60 ° C, preferably about 35 to 50 ° C, more preferably about 40 and 45 ° C. The period of time for the prior hydrogenation of the catalyst may range from about 2 to 48 hours, preferably from about 6 to 24 hours, more preferably from about 8 to 16 hours, even more preferably about 12 hours.

p00054Therefore, generally the catalyst is first added to the chemical reactor, followed by formic acid solvent (for example, about 17.5 to 22.5 parts of solvent, preferably about 20 parts of solvent). Before the solvent is added, hydrogen can be evacuated and the reactor can be filled with a nitrogen atmosphere for safety reasons. After completing the addition of formic acid, the nitrogen atmosphere is replaced by a hydrogen atmosphere at, for example, approximately 40 pounds per square inch (approximately 2.75 bar or 275,000 Pascals). The temperature is then adjusted to about 35 to 50 ° C preferably about 40 to 45 ° C for about 8 to 24 hours, preferably about 12 hours, before the introduction of the hemin starting material into the reactor. The previously hydrogenated catalyst suspension is then cooled and then hemin (in solvent) is added to the reactor. The hydrogen atmosphere is evacuated during the introduction of hemin for safety reasons and only the associated hydrogen is left in the Pd / C catalyst. The reactor is pressurized again at about 30 to 35psi with hydrogen and the reaction is stirred at about 20 to 25 ° C for about 30 minutes. The reaction is then heated to approximately 80 to 100 ° C, preferably to approximately 85 to 90 ° C, with vigorous stirring and the hydrogen pressure is increased to approximately 50 to 70 psi (approximately 3.4 to 4.8 bar or approximately 340,000 to 480,000 Pascals ), preferably about 55 to 60 psi (about 3.8 to 4.2 bar or about 380,000 to 420,000 Pascals). The reaction temperature is maintained for about 1 to 3 hours, preferably about 1 to 1.5 hours. The reaction is then cooled to about 40 to 60 ° C, preferably to about 45 to 50 ° C and the hydrogen pressure is maintained and hydrogenation continues for about 18 to 48 hours, preferably 20 to 30 hours, more preferably about 24 hours.

p00055The reaction is then cooled and depressurized with hydrogen evacuation from the reactor. Diatomaceous earth (such as HYFLO SUPERCEL, registered trademark of Celite Corp., Santa Barbara, California), activated carbon (such as DARCO KB, registered trademark of NORIT Americas, Inc., Marshall, Texas) and solvent are added to the reactor . The suspension is filtered and the filter cake is washed with solvent. This treatment is used to extract residual iron and residual palladium from the material.

p00056The filtrate is concentrated by vacuum distillation (which can be carried out at room temperature or at lower temperatures, such as about 10 to 15 ° C) to extract excess solvent. A precipitant is then added, for example, an ether such as methyl t-butyl ether (MTBE), for a period of at least about 30 seconds to at least about 3 hours, preferably for a period of at least about 1 hour to the solution concentrated. When MTBE is added, it can be added in about 17.5 to 22.5 parts, preferably in about 20 parts.

p00057The suspension can be cooled to a temperature of about -15 to -30 ° C, preferably at about -20 to -25 ° C.

p00058The suspension is filtered and the filter cake is washed with an organic solvent, such as ethers, which include methyl t-butyl ether (MTBE), diethyl ether or diisopropyl ether. After the end of the filtration and the cake is rinsed, the material is then dried in a vacuum oven at a temperature not exceeding about 60 ° C, for example about 45 to 60 ° C.

p00059When prepared by using formic acid as a solvent, the resulting product, mesoporphyrin IX, precipitates as a format salt; This is the preferred form for the isolation of mesoporphyrin IX after the hydrogenation step. After the additional stages of purification, the mesoporphyrin IX format is converted into a hydrochloride salt. This step provides additional purification of the intermediary. Likewise, it has been shown that the presence of proton scrubbers such as format (or other organic anions, such as acetate) during the next stage of tin insertion results in higher levels of impurities than if such scrubbers had been excluded. Therefore, it is preferred to replace the format anion of mesoporphyrin IX format with an anion less capable of purifying protons or regulating the solution during the tin insertion stage; Such anions include chloride and other halide anions such as bromide or iodide.

p00060When the intermediate isolated from the hydrogenation stage is the mesoporphyrin IX format, it is placed in a reaction vessel with diatomaceous earth, activated carbon and formic acid (for example, with approximately 10% w / w diatomaceous earth, approximately 20% w / w activated carbon and approximately 10 parts formic acid) for further purification. The suspension is stirred at, for example, about 20 to 30 ° C, preferably at about 20 to 25 ° C, for about 1.5 to 2.5 hours. The suspension is then filtered and the filter cake is washed with formic acid, for example, about 5 parts of formic acid. Then the resulting filtrate solution is concentrated to approximately a volume of 5 to 6 parts. Another vessel is loaded with purified water and 31% hydrochloric acid to prepare approximately 15 parts of approximately 1N hydrochloric acid. Approximately 6 parts of this HCl solution are transferred to the container containing about 6 parts of the filtrate, preferably at a temperature of about 20 to 25 ° C for a period of at least about 60 minutes. The solution is then seeded with mesoporphyrin IX dihydrochloride (available from the above synthesis) and stirred, preferably for at least about 2 hours. The remaining 9 parts of 1N hydrochloric acid are transferred to the vessel with vigorous stirring, preferably for a period of at least 60 minutes. The suspension is further stirred at about 20 to 30 ° C, preferably at about 20 to 25 ° C, for about 2 to 3 hours. It is then filtered and washed with purified water. The product is dried on the filter under a stream of nitrogen.

p00061In the above processes, the previous step was carried out by redisolving the solid mesoporphyrin IX format into formic acid and then the formic acid solution was added to the hydrochloric acid to convert the mesoporphyrin IX format into mesoporphyrin IX dihydrochloride. However, it was found that filtration of the mesoporphyrin IX dihydrochloride produced was quite slow on a pilot plant scale and required up to five days to complete and subsequent drying on the filter took between about two to three weeks. An improvement in the process was developed; As described above, the 1N hydrochloric acid solution is added to the formic acid solution of mesoporphyrin IX format. This was found to result in mesoporphyrin IX dihydrochloride that can be filtered more quickly. The addition of the bead material can also be carried out during the process, for example, at the beginning of the addition of 1N HCl to the formic acid solution of mesoporphyrin IX format or during the addition of 1N HCl to the formic acid solution of format of mesoporphyrin IX, such as when approximately 10%, approximately 20%, approximately 30%, approximately 40%, approximately 50%, approximately 60%, approximately 70% were added, about 80% or about 90% of 1N HCl to the formic acid solution of mesoporphyrin IX format. Preferably, as in the process described above, the bead material is added after 40% of the 1N HCl was added to the formic acid solution of mesoporphyrin IX format. The addition of the account material can also help in the formation of a product that can be filtered more quickly. Since the mesoporphyrin IX dihydrochloride that results from these process improvements can seep much faster, in the order of hours or even minutes instead of days, significant savings are achieved in terms of time and costs. Therefore, in another embodiment, the time for filtration of at least about 10 grams of mesoporphyrin IX dihydrochloride is reduced to less than about 90 minutes, less than about 60 minutes, less than about 45 minutes, less than about 35 minutes, less than about 25 minutes or less than about 10 minutes. In a further embodiment, the time for filtration of at least about 1000 grams of mesoporphyrin IX dihydrochloride is reduced to less than about 1 day, less than about 12 hours, less than about 6 hours, less than about 4 hours, less than approximately 3 hours or less than approximately 2 hours.

p000621) HCOOH 2) H2O, HCl

p0006310 Conversion of mesoporphyrin IX hydrochloride to stansoporphine (tin mesoporphyrin IX (IV)) by treatment with tin (II) salt, which is not an embodiment of the process of the invention.

p00064Mesoporphyrin IX hydrochloride is then treated with a tin (II) salt, such as SnCl2 in an organic solvent, such as acetic acid, under oxidation conditions, which provides the desired product, mesoporphyri dichloride.

p00065fifteen na IX of tin (IV) (stansoporphine). For example, mesoporphyrin IX dihydrochloride and tin (II) chloride are placed in a container and acetic acid is added at about 20 to 30 ° C, preferably at about 20 to 25 ° C. Suspended reagents are stirred for at least about 30 minutes. The mixture is heated with vigorous stirring under an inert atmosphere (such as nitrogen or argon) at reflux.

p00066Once the reflux has started, an atmosphere of approximately 6% oxygen in nitrogen is introduced into the air space of the vessel. The gas can be from about 3% to about 22% oxygen; approximately 6% is preferred to minimize the risk of explosion. The mixture is refluxed du

p00067for approximately 100 to 130 hours. It was found that the use of 6% oxygen in an atmosphere of nitrogen in the air space instead of spraying or blowing the gas mixture through the liquid is advantageous for increasing the yield of tin mesoporphyrin IX dichloride (IV ). Tin (II) can enter the porphyrin ring to form complexes with the nitrogen and can also leave the porphyrin ring.

p00068However, tin (IV) that is not yet bound to the nitrogens in the porphyrin ring cannot enter the ring to bind to the nitrogens. To generate tin mesoporphyrin IX (IV), the tin ion (II) must enter the porphyrin ring and then undergo oxidation to tin (IV) in situ. Excessively rapid oxidation of the tin (II) ion will cause the insertion action to stop, which can significantly decrease the yield. Therefore, adequate control of the oxidation rate is necessary. The introduction of oxygen into the mixture through the interface between the solvent and the oxygen / nitrogen atmosphere of the air space provides this control and leads to a reasonable reaction rate with a good production of the final product.

p00069Optionally, samples of the reaction mixture can be taken during the tin insertion stage by lowering the temperature to about 50 to 70 ° C, preferably about 55 to 60 ° C by extracting a sample and returning the reaction to reflux.

p00070After the tin insertion stage, the reaction mixture is cooled and WFI (water for injection) is added. The suspension is then filtered and the filter cake is washed with water for injection. The filter cake is then placed under vacuum for a minimum of 4 hours to extract residual water.

p00071Conversion of mesoporphyrin IX dihydrochloride into stansoporphine (tin mesoporphyrin IX (IV)) by treatment with tin (II) oxide according to the invention

p00072SnO, AcOH

p00073Tin can also be inserted into the mesoporphyrin IX ring by treating mesoporphyrin IX dihydrochloride with tin (II) oxide. This reaction can proceed until its completion in a period of time as short as 2 hours, compared to the four days to three weeks required for the insertion of tin by using the procedure of the tin (II) salt described above. A solution / suspension of mesoporphine dihydrochloride IX in a suitable solvent, for example, formic or acetic acid, is added to a solution / suspension of tin (II) oxide in a suitable solvent, for example, acetic acid or formic acid. An exemplary procedure is described below and in the Examples.

p00074Mesoporphyrin IX dihydrochloride is dissolved / suspended in formic acid at room temperature. Since the solution or suspension will have an intense purple color, it is advantageous to spray mesoporphyrin IX dihydrochloride in a powder as fine as possible to aid dissolution.

p00075Tin (II) oxide is suspended in acetic acid at room temperature and stirred. After prolonged agitation, the tin oxide suspension can be transformed into a gel, which does not appear to adversely affect the reaction. The gel separates once the addition of mesoporphyrin IX dihydrochloride begins. The amount of tin (II) oxide is approximately two equivalents to approximately six equivalents per equivalent of mesoporphyrin IX dihydrochloride; preferably, approximately four equivalents of tin (II) oxide are used per equivalent of mesoporphyrin IX dihydrochloride. (In this reaction, the equivalent ratio is the same as the molar ratio).

p00076The tin (II) oxide solution is maintained at a temperature of about 25-115 ° C, preferably about 50-75 ° C, more preferably about 60-65 ° C. The mesoporphyrin IX dihydrochloride solution is then added over a period of approximately three to nine hours, preferably over a period of approximately six hours. The solution of mesoporphyrin IX dihydrochloride can be found at room temperature during the addition or can be maintained at a temperature of about 5075 ° C, such as about 60-65 ° C during the addition. The reaction mixture is maintained at about 25-115 ° C, preferably about 50-75 ° C, more preferably about 60-65 ° C, for about an additional 2 to 48 hours, preferably about an additional 16 to 30 hours, more preferably about 18 to 24 additional hours, such as approximately 18 additional hours or approximately 24 additional hours. After the additional reaction time, the suspension is cooled to room temperature (about 20-25 ° C), stirred for at least about 5 minutes, preferably at least about one hour and filtered.

p00077General insertion of metals into porphyrins through the use of metal oxides

p00078The procedure used for the insertion of tin into the porphyrin rings through the use of metal oxides can also be applied to the insertion of other metals through the use of metal oxides. Particularly useful metal oxides are metal oxides where the metal cation of the metal oxide is in an intermediate oxidation state. The process can be used for porphyrin compounds or their salts, which include, by way of non-taxation, mesoporphyrin or its salt, mesoporphyrin IX or its salt, mesoporphyrin dihydrochloride IX, protoporphyrin or its salt, hematoporphyrin or its salt or deuteroporphyrin or its salt , to provide the metalated porphyrin compound (or its salt).

p00079Metal oxides that can be used include, but are not limited to, tin oxide, zinc oxide, copper oxide, cadmium oxide, cobalt oxide, chromium oxide, iron oxide, aluminum oxide, titanium oxide, oxide nickel, manganese oxide, silver oxide, gold oxide, vanadium oxide, platinum oxide, antimony oxide, arsenic oxide, tin oxide (II), zinc oxide (II), copper oxide (I ), copper (II) oxide, cadmium (II) oxide, cobalt (II) oxide, cobalt (III) oxide, cobalt (IV) oxide, Co3O4, chromium (II) oxide, chromium (III) oxide, chromium (IV) oxide, chromium (V) oxide, chromium (VI) oxide , iron (II) oxide, iron (III) oxide, Fe3O4, aluminum (III) oxide, titanium (II) oxide, titanium (III) oxide, titanium (IV) oxide, nickel (II) oxide ), manganese oxide (II), manganese oxide (III), manganese oxide (IV), manganese oxide (VII), silver oxide (I), silver oxide (II), gold oxide (I) , gold (III) oxide, vanadium (II) oxide, vanadium (III) oxide, vanadium (IV) oxide, vanadium (V) oxide, platinum (II) oxide, platinum (IV) oxide, oxide of antimony (III), antimony oxide (IV), antimony oxide (V), arsenic oxide (III) or arsenic oxide (V).

p00080Other porphyrin and tetrapyrrole compounds may also be metallized by the use of methods described herein, which include, by way of non-taxation, porphyrins such as deuteroporphyrins and deuteroporphyrin IX 2,4bis (ethylene glycol) (8,13-bis (l, 2 -dihydroxyethyl) -3,7,12,17-tetramethyl-21H, 23H-porphine-2,18-dipropionic). Additional porphyrin compounds that can be metalated by the use of the methods described herein include, but are not limited to, coproporphyrin, cytoporphyrins, ethioporphyrins, hematoporphyrins, mesoporphyrins, phyllophyrins, protoporphyrins, pyrroporphyrins, rhodoporphyrins, uroporphyrins and phytoporphyrins. An exhaustive list of porphyrin compounds is provided at www.chem.qmul.ac.uk/iupac/tetrapyrrole/; The porphyrins described therein are incorporated herein by reference as porphyrins that can be metalated by the use of the methods described herein.

p00081Purification of tin mesoporphyrin IX dihydrochloride (IV): hot acid crushing

p00082At this point, the crude tin (IV) mesoporphyrin IX dihydrochloride is then crushed with hot acid to extract the impurities. The material is resuspended in hydrochloric acid (approximately 0.5 N to 2.0 N, preferably 1 N) and the temperature rises to approximately 75 to 100 ° C or approximately 80 to 100 ° C, preferably approximately 85 to 950 ° C, more preferably at approximately 85 to 90 ° C, for approximately one to two hours with moderate agitation. The suspension is then cooled to about 20 to 30 ° C, preferably to about 20 to 25 ° C and filtered; The filter cake is washed with purified water and dried on the filter under a stream of nitrogen.

p00083Purification of tin mesoporphyrin IX dihydrochloride (IV): high pH treatment

p00084The material of the hot acid crushing stage is combined with diatomaceous earth, activated carbon, water and ammonium hydroxide. The temperature is adjusted to about 20 to 30 ° C, preferably to about 20 to 25 ° C and then preferably stirred for about 1 to 2 hours. A sample is taken to ensure that the pH is approximately 9 or higher. The mixture is then stirred, preferably for an additional 1 to 2 hours. The mixture is then filtered. Any material that remains in the filter is washed with water; The filter cake is discarded.

p00085Reacidification of tin mesoporphyrin IX dihydrochloride (IV):

p00086The filtrate is then transferred to a mixture of 31% acetic acid and hydrochloric acid and the mixture is set at about 20 to 30 ° C, preferably at about 20 to 25 ° C. The resulting suspension is stirred, preferably for about 15 minutes, samples are taken to ensure that the pH is less than or equal to about 1 and then stirred again, preferably for an additional 1 to 2 hours. The suspension is filtered and the filter cake is washed with water followed by extracting the residual water under vacuum.

p00087At this stage, a sample of the filter cake is taken to detect the residual starting material, mesoporphyrin dihydrochloride IX. If the level is greater than about 0.1%, the high pH treatment followed by the reacidification is repeated as necessary (for example, an additional 1, 2 or 3 times).

p00088Additional crushing with hot tin mesoporphyrin IX dichloride acid (IV)

p00089The filter cake of the previous step is resuspended in a mixture of about two parts by weight of water for injection (WFI) and about one part by weight of 31% HCl, at about 20 to 30 ° C, preferably at about 20 at 25 ° C. Under moderate stirring, the mixture is adjusted to approximately 80 to 100 ° C, preferably to approximately 85 to 90 ° C, for approximately 6 to 48 hours, preferably approximately 12 to 24 hours, more preferably approximately 16 to 18 hours, followed by cooling to approximately 20 to 30 ° C, preferably at about 20 to 25 ° C, for at least about 1 hour. The suspension is filtered, the filter cake is washed with an aqueous solution of hydrochloric acid (for example, approximately 1 part of 31% HCl to 25 parts of water for injection (WFI) w / w) and dried under a stream of nitrogen (at 50 ° C or less, approximately).

p00090The final hot acid treatment serves to readjust the form of stansoporphine to monomer. In the neutral solution, stansoporphine is in a monomer-dimer equilibrium; The strong acid treatment causes the balance to shift sharply to the monomer form.

p00091Development work on the synthesis of stansoporphine indicates that for optimal results, the hydrogenation catalyst should be pre-hydrogenated before the introduction of the hemin starting material; Isolation of mesoporphyrin IX dihydrochloride from the form of mesoporphiarin IX in formic acid should proceed through the addition of the HCl solution to the formic acid solution; the presence of proton scrubbers should be avoided during the tin insertion stage; and the introduction of oxygen during the tin insertion stage should proceed by introducing the oxygen / nitrogen mixture into the air space of the reaction, instead of spraying or blowing the gas through the solution. With these optimal parameters in mind, other variables such as temperature, reaction time, reagent concentration and order of reagent addition can be handled in a certain way, for example, the concentration and reaction time can vary between approximately one 50 and 200% of the indicated values ​​or within about 75 and 150% of the indicated values ​​and the temperature may vary from about 5 to 10 ° C of the indicated values, such that the variation does not result in a large-scale synthesis of stansoporphine with a purity less than the high purity defined herein. The purification and precipitation steps can be repeated as necessary to maintain the high purity of the large-scale stansoporphine preparation.

p00092Therapeutic use of stansoporphine for the treatment or prevention of childhood hyperbilirubinemia and other diseases

p00093Stansoporphine as produced by the process of the invention can be used for the treatment or prevention of childhood hyperbilirubinemia (childhood jaundice) (see US Patents 4,657,902 and 4,668,670; and WO 94/28906). Other methods for using stansoporphine are disclosed in US Patent 4,692,440 (to increase the rate of hematine excretion), WO 89/02269 (to counteract the toxicity of cancer therapy), Patent 4,782,049 (to treat psoriasis) and other publications. The treatment or prevention of childhood hyperbilirubinemia is achieved by dissolving stansoporphine in a pharmaceutically acceptable carrier. Preferably, stansoporphine is provided in a solution that can be regulated to maintain an adequate pH. The pH regulators that can be used include phosphate, citrate, gluconate, lactate, tartrate, glycinate, glycylglycinate, bicarbonate, carbonate, maleate or acetate, with sodium, potassium, magnesium, calcium or aluminum present as cation. Histidine and imidazole can also be used as pH regulators. Phosphate pH regulators, particularly the sodium phosphate pH regulator, are preferred. PH regulators must be pharmaceutically acceptable for use as injectable agents in neonates. The pH of the solution for administration is preferably between about 7.0 to 8.0, more preferably about 7.2 to 7.9, even more preferably about 7.4. The osmolarity of the solution is preferably physiological or near osmolarity; a preferred range is between about 280 mOsm / L and 310 mOsm / L. Stansoporphine is preferably administered by injection, more preferably by intramuscular injection. Stansoporphine is administered in an amount sufficient to treat or prevent childhood hyperbilirubinemia, generally approximately 4.5 mg / kg birth weight; by using lower doses of stansoporphine, such as 1.5 mg / kg birth weight or 3.0 mg / kg birth weight.

p00094The following examples are intended to illustrate the invention and are not intended to limit the invention in any way.

Examples

p00095Reference Example 1

p00096Exemplary synthesis of high purity stansoporphine

p00097Initial conversion of hemin to mesoporphyrin IX

p00098A 200L glass coated vessel, pressure calculated at 150 psi, is loaded with 0.6 kg of 5% palladium on carbon and 73 kg of formic acid. The reactor is pressurized with hydrogen at 60-65 psi with vigorous stirring and heated at 40-45 ° C for a minimum of 12 hours. The reaction is cooled to 20-25 ° C with moderate stirring, the hydrogen atmosphere is evacuated and the reactor is charged with 6.0 kg of hemin (DMF grade) and 73 kg of formic acid. The reactor is pressurized at 30-35 psi with hydrogen and stirred at 20-25 ° C for 30 minutes.

p00099The reaction is heated to 85-90 ° C with vigorous stirring. The hydrogen pressure then rises to 55-60 psi. The pressure and temperature are maintained for a period of 1-1.5 hours.

p00100The reaction is cooled to 45-50 ° C and hydrogenation is continued at 55-60 psi for 24 hours. The reaction is then cooled to 20-25 ° C, depressurized and samples are taken.

p00101The reaction is heated to 45-50 ° C, pressurized at 55-60 psi with hydrogen and stirred for an additional 6 hours. The reaction is then cooled to 20-25 ° C, depressurized and samples are taken again.

p00102Hydrogen is evacuated from the vessel that is then loaded with 3.0 kg of HYFLO SUPERCEL, 2.3 kg of DARCO KB and 42 kg of formic acid. The suspension is filtered and the filter cake is washed with 122 kg of formic acid.

p00103A portion of the filtrate is transferred to a 200L glass coated vessel, cooled to 10-15 ° C and distilled under vacuum to extract formic acid. Once the residual volume dropped to 25-35L, the rest of the filtrate is transferred and distillation continues to a residual volume of 25-30L.

p00104The reaction temperature is adjusted to 20-25 ° C and 89 kg of methyl tert-butyl ether are added for a minimum of 1 hour. The resulting suspension is stirred at 20-25 ° C for 2 hours before cooling to -25 to -20 ° C for a period of 4 hours.

p00105The suspension is filtered and washed with 12 kg of methyl tert-butyl ether. The intermediate product is dried in a vacuum oven at 60 ° C or less.

p00106Purification of the mesoporphyrin IX format with diatomaceous earth and activated carbon; conversion of mesoporphyrin IX format to mesoporphyrin IX dihydrochloride

p00107The intermediate is transferred to a 50L glass coated container with 10% w / w DARCO KB, 20% w / w HYFLO SUPERCEL and 10 parts formic acid. The suspension is stirred at 20-25 ° C for a period of 1.5-2.5 hours.

p00108The suspension is filtered in a second 50L glass coated vessel. The filter cake is washed with 5 parts of formic acid and discarded. The filtrate solution is distilled under vacuum at a residual volume of 5-6 parts.

p00109A third container is loaded with purified water and 31% hydrochloric acid to prepare 15 parts of 1N hydrochloric acid. Six parts of the filtrate solution are transferred to the reactor at 20-25 ° C for a minimum of 60 minutes. The solution is seeded with mesoporphyrin IX dihydrochloride and stirred for a minimum of 2 hours. The remaining 9 parts of 1N hydrochloric acid are transferred with vigorous stirring to a vessel for a minimum of 1 hour. The resulting suspension is stirred at 20-25 ° C for a period of 2-3 hours before isolation by filtration. The filter cake is washed with 4 parts of purified water. The intermediate product of mesoporphyrin IX dihydrochloride is dried on the filter under a stream of nitrogen.

p00110Conversion of mesoporphyrin IX dihydrochloride IX to tin (IV) mesoporphyrin dihydrochloride (stansoporphine)

p00111A 50L glass coated vessel with 1.57 kg of mesoporphyrin IX dihydrochloride, 1,862 kg of tin (II) chloride and 40.9 kg of acetic acid is charged at 20-25 ° C. The suspension is maintained at 20-25 ° C with moderate agitation for a minimum of 30 minutes.

p00112The mixture is heated under nitrogen with vigorous stirring to reflux (approximately 115 ° C). Once the reflux has been reached, an atmosphere of oxygen in nitrogen is introduced into the air space of the vessel. The reaction mixture is refluxed for a period of 100-130 hours.

p00113The reaction mixture is cooled to 55-60 ° C and samples are taken to detect residual mesoporphyrin; While waiting for the results, the reaction mixture is reheated to reflux. Once complete, the reaction is cooled to 60-70 ° C and charged with 15.7 kg of water for injection. The temperature of the suspension is adjusted at 20-25 ° C for 30 minutes and stirred for one hour.

p00114The suspension is filtered and the container and cake are washed with 6.3 kg of water for injection. Once the washing is finished, the cake is placed under vacuum for a minimum of 4 hours to extract the residual water.

p00115A 50L glass coated container is loaded with the wet filter cake, 22.4 kg of purified water and 3.7 kg of 31% hydrochloric acid at 20-25 ° C. With moderate stirring, the temperature of the mixture is adjusted to 85-90 ° C for a period of 1-2 hours and then cooled to 20-25 ° C. The suspension is filtered and the filter cake is washed with 6.3 kg of purified water. The product is dried on the filter under a stream of nitrogen and packaged.

p00116Purification of tin (IV) mesoporphyrin IX dichloride (stansoporphine) at high pH with diatomaceous earth and activated carbon

p00117A 50L glass coated vessel with 1,448 kg of tin (IV) mesoporphyrin IX dichloride, 0.194 kg of HYFLO SUPERCEL, 0.066 kg of DARCO KB, 14.5 kg of water for injection and 1.0 kg of hydroxide is charged of ammonium 26 Be. The temperature of the reaction mixture is adjusted to 20-25 ° C and stirred for a period of 1-2 hours. A sample is taken to verify that the pH is> 9. The mixture is then stirred for an additional 1-2 hours. The mixture is filtered through a glass receiver. The cake is washed with 2.9 kg of water and discarded.

p00118A second 50L glass-coated container is loaded with 38.2 kg of acetic acid and 2.6 kg of 31% HCl. The temperature is adjusted to 20-25 ° C. The filtrate of the glass receiver is transferred to the second 50L container for a minimum of 45 minutes at 20-25 ° C. The glass receiver and the transfer apparatus are washed with 2.1 kg of water for injection into the container. The resulting suspension is stirred at 20-25 ° C for 15 minutes before taking the sample to verify that the pH is ≤ 1. The suspension is then stirred for an additional 1-2 hours.

p00119The suspension is filtered and the container and cake are washed with 1.3 kg of water for injection. Once the washing is finished, the cake is placed under vacuum for a minimum of 4 hours to extract the residual water.

p00120A sample of the filter cake is taken for testing. If the residual starting material (mesoporphyrin dihydrochloride IX) is at an acceptable level, the reaction proceeds to the next stage, otherwise the treatment is repeated (i.e., the filter cake is dissolved again by the use of ammonium hydroxide as set forth above).

p00121Treatment of tin (IV) mesoporphyrin IX (stansoporphine) dichloride at a low pH to configure it in the monomeric form

p00122The wet filter cake is placed back in the 50L glass coated container which is then loaded with 20.4 kg of water for injection and 10.2 kg of 31% hydrochloric acid at 20-25 ° C. With moderate agitation, the temperature of the mixture is adjusted to 85-90 ° C for a period of 16-18 hours, followed by cooling to 20-25 ° C for a minimum of 1 hour. The suspension is filtered and the filter cake is washed with a premixed solution of 0.5 kg of 31% hydrochloric acid in 12.8 kg of water for injection. The product is dried on the filter at &lt;50 ° C under a stream of nitrogen and packaged.

p00123Inventive Example 2

p00124Alternative stage of tin insertion through the use of tin (II) oxide as a source of tin

p00125The insertion of tin into mesoporphyrin IX to produce stansoporphine can be carried out through an alternative synthetic route by using tin (II) oxide as a reagent for the introduction of tin.

p00126A 1,000 ml round neck, three-necked round bottom dark flask, equipped with a magnetic stirrer, a Claisen head, an addition funnel, a thermometer, a condenser and a nitrogen insufflator with 8.4 g of oxide was charged tin (II) and 200 ml of acetic acid, at 20-25 ° C to form a gray suspension. The suspension was heated at 60-65 ° C under nitrogen.

p00127Another round bottom flask with a neck and 250 ml equipped with a stirrer was charged with 10 g of mesoporpphyrin IX dihydrochloride and 50 ml of formic acid. The mixture was stirred at 20-25 ° C for 30 minutes to effect dissolution, resulting in approximately 60 ml of a deep purple suspension / solution at 20-25 ° C. (Due to the color of the solution, it is difficult to observe the complete solution visually; mesoporphyrin IX dihydrochloride must be thoroughly crushed before the addition of formic acid).

p00128The mesoporphyrin IX dihydrochloride solution was charged to the addition funnel and added dropwise to the tin (II) oxide / acetic acid solution / suspension for a period of 6 hours, while maintaining the suspension / solution temperature of tin oxide / acetic acid (II) at 60-65 ° C. The volume in the flask is increased from 200 ml to 250 ml; The appearance of the reaction changes from a gray suspension (or white gel) to a purple suspension, to a red suspension.

p00129Once the addition is complete, the reaction is stirred under a nitrogen atmosphere at 60-65 ° C for an additional 18-24 hours. Then 100 ml of water is added by dripping for about 20-40 minutes, while maintaining the temperature at 60-65 ° C. The resulting red suspension (approximately 360 ml) was cooled to 20-25 ° C for about 30 minutes and stirred for a minimum of 1 hour and then filtered under reduced pressure (the total filtration time was approximately 10-20 minutes). The filter cake was washed with two 20 ml portions of water. The filtrate volume of approximately 400 ml was a dark red solution; The filter cake dough of approximately 50 g also had a dark red color.

p00130The wet filter cake was carefully split into pieces and reloaded into the reaction flask with 100ml of 1N HCl. The resulting dark red suspension was heated at 85-95 ° C for 1 hour. The suspension was then cooled to 20-25 ° C and filtered under reduced pressure (the total filtration time was approximately 20-30 minutes); The filtrate showed a dark red to brown color. The dark red filter cake was rinsed with two 20 ml portions of water, dried under a stream of nitrogen and further dried under high vacuum at 80-90 ° C for 24 hours. In several repetitions of the synthesis, the yield of the product varied from 16.5-21.2 g (70-90%).

p00131Example 3

p00132Analysis of high purity stansoporphine performed by exemplary synthesis by using tin (II) chloride as a source of tin

p00133Stansoporphine batches were prepared by using exemplary synthesis basically as described above in Example 1, as well as in the above procedures (see US Patents 6,818,763 and 2004/0210048).

p00134The basic HPLC analysis is performed by using a C-18 column (Zorbax Extend C-18, 4.6 x 150 mm, particle size of 3.5 um or the equivalent). The detector is configured at 400 nm. The solvents (acetonitrile, methanol and water) are of quality for HPLC. The mobile phase is 16% acetronitrile: 40% methanol: 44% 0.5M ammonium acetate, pH 5.15. (The ammonium acetate solution is prepared by dissolving 38.5 g of ammonium acetate in 440 mL of H2O and adjusting the pH to 5.15 with acetic acid. Both ammonium acetate and acetic acid are reactive Then 160 mL of acetonitrile and 400 mL of methanol are added; the mobile phase solution is mixed, filtered and degassed before use). The flow rate is 1.0 ml / minute. Stansoporphine samples and standards are prepared for injection at a concentration of 0.04 mg / mL in 1N NaOH. Since stansoporphine and related compounds are sensitive to light, stansoporphine-containing solutions, starting materials or impurity standards must be stored in opaque containers and their handling and analysis should be carried out under reduced light conditions. Standard samples and solutions must be used within 12 hours of preparation. 5 uL of analyte solution is injected and a runtime of 10 minutes is used. The retention time of stansoporphine is generally about 4.8 minutes. The column temperature is maintained at 60 ° C. After analysis, the column is washed with 80% methanol and 20% water for at least 1 hour at 1.0 mL / min.

p00135HPLC analysis for the quantification of impurities is performed by using an ACE 5 C-18 column, 4.6 x 250 mm, particle size of 5 um, with detection at 400 nm. The protection of light sensitive samples and standards is carried out as described above. The mobile phases used are A: 30% methanol, 70% water with 0.02M ammonium acetate, pH 9.1 and B: 80% methanol, 20% water with 0.02M ammonium acetate, pH 9 , 1 (mobile phase A is prepared by dissolving 3.0 g of ammonium acetate in 1,400 mL of water, adjusting the pH to 9.1 with NH4OH and adding 600 mL of methanol; mobile phase B is prepared by dissolving 3.0 g of ammonium acetate in 400 mL of water, adjusting the pH to 9.1 with NH4OH and adding 1,600 mL of methanol; the mobile phases are mixed, filtered and degassed before use). The samples are dissolved in 0.5% w / w TEA in water at a concentration of approximately 0.2 mg / mL. Standard samples and solutions must be used within 12 hours of preparation.

p00136The analysis is performed by using the following gradient conditions:

<dl><dt>Weather </dt><dd>%TO % B </dd></dl>

<dl><dt>0 </dt><dd> 100 </dd><dt>0 </dt><dd /></dl>

<dl><dt>50 </dt><dd> 70 30 </dd></dl>

<dl><dt>65 </dt><dd> 70 30 </dd></dl>

<dl><dt>90 </dt><dd> 0 100 </dd></dl>

<dl><dt>110 </dt><dd> 0 100 </dd></dl>

<dl><dt>111 </dt><dd> 100 0 </dd></dl>

<dl><dt>120 </dt><dd> 100 0 </dd></dl>

p001375 where concentrations change linearly between the points shown.

p00138Table 1 contains a comparison of the HPLC analysis of the product of the synthesis in progress, column C, compared to the analyzes of the products of the previous synthesis of column A and column B. The detected peaks are listed in order of retention time relative to stansoporphine, with the retention time of the

p0013910 Stansoporphine set to 1. The batch analyzed in column A was produced in an amount of 1.1 kg; The batch analyzed in column C was also produced in an amount of 1.1 kg.

p00140Table 1. Analysis of various stansoporphine preparations As seen in Table 1, the ongoing synthesis of stansoporphine in column C resulted in the material that resulted in a high purity product, of total purity> 99% and not containing no impurity in 0.1% or greater.

<dl><dt>Relative retention time </dt><dd>TO B C </dd></dl>

<dl><dt>0,33 </dt><dd> 0,06% </dd></dl>

<dl><dt>0,51 </dt><dd> 0,05% 0,05% 0,07% </dd></dl>

<dl><dt>0,55 </dt><dd> 0,06% </dd></dl>

<dl><dt>0,73 </dt><dd> 0,14% 0,05% 0,06% </dd></dl>

<dl><dt>0,76 </dt><dd> 0,07% 0,05% </dd></dl>

<dl><dt>0,83 </dt><dd> 0,05% </dd></dl>

<dl><dt>0,84 </dt><dd> 0,05% </dd></dl>

<dl><dt>0,92 </dt><dd> 0,26% 0,06% </dd></dl>

<dl><dt>0,95 </dt><dd> 0,30% 0,05% </dd></dl>

<dl><dt>0,96 </dt><dd> 0,22% </dd></dl>

<dl><dt>1 </dt><dd> 98% 99% 100% </dd></dl>

<dl><dt>1,05 </dt><dd> 0,09% </dd></dl>

<dl><dt>1,26 </dt><dd> 0,06% </dd></dl>

p00141Example 4

p00142Analysis of high purity stansoporphine prepared by exemplary synthesis through the use of tin (II) oxide as a source of tin

p0014310 Three batches of stansoporphine were produced by using the tin insertion stage as described in example 2. Analysis of the three batches indicated that the purity of the stansoporphine produced was 99.7%, and 99.6 % (the total stansoporphine content was 96.4%, 99.1% and 97.2%, respectively).

p00144fifteen The HPLC analysis was performed on a Zorbax Extend C-18 column, 4.6x150 mm and 5 μm thick. The eluents used were: A: 80% 0.05 M ammonium acetate, pH 5.15 with acetic acid: 20% acetonitrile; B: 90% methanol: 10% acetonitrile. The temperature used was 40 ° C. A flow rate of 1.2 ml / min was used. with a detection of 400 nm. The retention time of stansoporphine was 8.8 min., While that of mesoporphyrin IX was 23.1 min., With the use of the following gradients listed in Table 2.

p00145Table 2

<dl><dt>Weather </dt><dd>TO B </dd></dl>

<dl><dt>0,0 </dt><dd> 60 40 </dd></dl>

<dl><dt>10,0 </dt><dd> 25 75 </dd></dl>

<dl><dt>30,0 </dt><dd> 25 75 </dd></dl>

<dl><dt>31,0 </dt><dd> 60 40 </dd></dl>

<dl><dt>40,0 </dt><dd> 60 40 </dd></dl>

p00146More extensive analyzes of two stansoporphine batches produced by using the tin oxide insertion procedure were performed. These analyzes are detailed in table 3 (lot weight 0.840 kg) and table 4 (weight of lot 1.344 kg) below (where a / a indicates the proportion of HPLC peak area).

p00147Table 3 Table 4

<dl><dt>Proof </dt><dd>Method Results </dd></dl>

<dl><dt>Total purity </dt><dd>HPLC Total impurities <1% a / a; impurity at RRt 0.72 = 0.06% y / y; no other impurity> 0.05% a / a</dd></dl>

<dl><dt>Water content </dt><dd>Karl Fischer coulometer Trace &lt;1% p / p </dd></dl>

<dl><dt>Residual Solvents -acetone </dt><dd>Chromatography (GC - air space) No detection &lt;0.1% w / w </dd></dl>

<dl><dt>Organic content - formic acid + acetic acid </dt><dd>HPLC 0.1% w / w </dd></dl>

<dl><dt>Inorganic content - palladium and iron </dt><dd>Inductively coupled plasma - optical emission spectroscopy Pd = 5 ppm Fe = 5 ppm </dd></dl>

<dl><dt>Inorganic content - free tin </dt><dd>Differential Pulse Polarography &lt;0.1 tin free </dd></dl>

<dl><dt>Inorganic content - tin </dt><dd>Inductively coupled plasma - optical emission spectroscopy 144500 ppm </dd></dl>

<dl><dt>Proof </dt><dd>Method Results </dd></dl>

<dl><dt>Total purity </dt><dd>HPLC Total impurities <1% a / a; impurity at RRt 0.72 = 0.06% y / y; no other impurity> 0.05% a / a</dd></dl>

<dl><dt>Water content </dt><dd>Karl Fischer coulometer Trace &lt;1% p / p </dd></dl>

<dl><dt>Residual Solvents -acetone </dt><dd>Chromatography (GC - air space) No detection &lt;0.1% w / w </dd></dl>

<dl><dt>Organic content - formic acid + acetic acid </dt><dd>HPLC No detection &lt;0.1% w / w </dd></dl>

<dl><dt>Inorganic content - palladium and iron </dt><dd>Inductively coupled plasma - optical emission spectroscopy Pd = 5ppm Fe = 67 ppm </dd></dl>

<dl><dt>Inorganic content - free tin </dt><dd>Differential Pulse Polarography &lt;0.1% tin free </dd></dl>

<dl><dt>Inorganic content - tin </dt><dd>Inductively coupled plasma - optical emission spectroscopy 165000 ppm </dd></dl>

<dl><dt>Inorganic content - chloride </dt><dd>Elemental analysis 103300 ppm </dd></dl>

p00148Example 5

p00149Stability of high purity stansoporphine preparations

p00150The long-term stability of the compound was studied under two different storage conditions: 25 ° C (+/- 2 ° C) and 60% relative humidity (+/- 5%); and 40 ° C (+/- 2 ° C) and 75% relative humidity (+/- 5%). Primary packing

p0015110 for the compound it was a 4 mils polyethylene bag and the secondary packaging for the compound was a 4 thousand polyethylene bag. The stansoporphine stored in two bags was placed in a high density polyethylene drum (CurTec®).

p00152Table 5 and Table 6 contain the stability data for the lot described in Table 3, under conditions of

p00153fifteen 25 ° C / 60% RH and 40 ° C / 75% RH, respectively. Table 7 and Table 8 show the stability data for the lot described in Table 4, under conditions of 25 ° C / 60% RH and 40 ° C / 75% RH, respectively. The data for the zero month time point was taken from the batch release analysis (the zero month time point represents the actual date the samples were placed in the stability test chambers). The samples were analyzed at approximately 3 months and approximately 6 months after the samples were placed in conditions

p00154twenty of storage.

p00155Table 5

<dl><dt>Proof </dt><dd>0 months 3 months 6 months </dd></dl>

<dl><dt>Appearance </dt><dd>Red powder free of visual signs of contamination Red powder free of visual signs of contamination Red powder free of visual signs of contamination </dd></dl>

<dl><dt>HPLC purity (total impurities) </dt><dd> 0,3 0,22 0,24 </dd></dl>

<dl><dt>HPLC purity (impurity peak at retention time RRt 0.72-0.73) </dt><dd> 0,06% &lt;0,05% 0,07% </dd></dl>

<dl><dt>HPLC test (w / w, solvent-free anhydrous base) </dt><dd> 100,7% 99,8% 98,4% </dd></dl>

<dl><dt>HPLC test (w / w without changing) </dt><dd> 100,4% 99,6% 98,2% </dd></dl>

<dl><dt>Water content (Karl Fischer, coulometer) </dt><dd>Trace &lt;1% &lt;1% (0.1%) &lt;1% (0.1%) </dd></dl>

p00156Table 6

<dl><dt>Proof </dt><dd>0 months 3 months 6 months </dd></dl>

<dl><dt>Appearance </dt><dd>Red powder free of visual signs of contamination Red powder free of visual signs of contamination Red powder free of visual signs of contamination </dd></dl>

<dl><dt>HPLC purity (total impurities) </dt><dd> 0,3 0,28 0,26 </dd></dl>

<dl><dt>HPLC purity (impurity peak at retention time RRt 0.72-0.73) </dt><dd> 0,06% &lt;0,05% 0,06% </dd></dl>

<dl><dt>HPLC test (w / w, solvent-free anhydrous base) </dt><dd> 100,7% 101,2% 99,8% </dd></dl>

<dl><dt>HPLC test (w / w without changing) </dt><dd> 100,4% 100,9% 99,6% </dd></dl>

<dl><dt>Water content (Karl Fischer, coulometer) </dt><dd>Trace &lt;1% &lt;1% (0.2%) &lt;1% (0.1%) </dd></dl>

p00157Table 7

<dl><dt>Proof </dt><dd>0 months 3 months 6 months </dd></dl>

<dl><dt>Appearance </dt><dd>Red powder free of visual signs of contamination Red powder free of visual signs of contamination Red powder free of visual signs of contamination </dd></dl>

<dl><dt>HPLC purity (total impurities) </dt><dd> 0,22 0,29 0,19 </dd></dl>

<dl><dt>HPLC purity (impurity peak at retention time RRt 0.72-0.73) </dt><dd> 0,06% 0,05% 0,05% </dd></dl>

<dl><dt>HPLC test (w / w, solvent-free anhydrous base) </dt><dd> 102,3% 102,1% 98,5% </dd></dl>

<dl><dt>HPLC test (w / w without changing) </dt><dd> 102,3% 102,0% 98,4% </dd></dl>

<dl><dt>Water content (Karl Fischer, coulometer) </dt><dd>Trace &lt;1% &lt;1% (0.1%) &lt;1% (0.1%) </dd></dl>

p00158Table 8

<dl><dt>Proof </dt><dd>0 months 3 months 6 months </dd></dl>

<dl><dt>Appearance </dt><dd>Red powder free of visual signs of contamination Red powder free of visual signs of contamination Red powder free of visual signs of contamination </dd></dl>

<dl><dt>HPLC purity (total impurities) </dt><dd> 0,22 0,29 0,24 </dd></dl>

<dl><dt>HPLC purity (impurity peak at retention time RRt </dt><dd>time 0.72-0.73) 0.06% &lt;0.05% 0.06% </dd></dl>

<dl><dt>HPLC test (w / w, solvent-free anhydrous base) </dt><dd> 102,3% 101,1% 97,8% </dd></dl>

<dl><dt>HPLC test (w / w without changing) </dt><dd> 102,3% 101,0% 97,7% </dd></dl>

<dl><dt>Water content (Karl Fischer, coulometer) </dt><dd>Trace &lt;1% &lt;1% (0.1%) &lt;1% (0.1%) </dd></dl>

p00159The disclosure of all publications, patents, patent applications and published patent applications referred to herein by means of an identifying citation are incorporated herein by reference in their entirety.

p00160According to a preferred embodiment of the composition of claim 1, the composition comprises at least about 100 grams of stansoporphine.

p00161According to another preferred embodiment of the composition of claim 1, said stansoporphine is at

p0016210 minus 99% pure; where preferably any individual impurity present is present in an amount less than about 0.09%; wherein said composition preferably further comprises at least about 100 grams of stansoporphine.

p00163According to a preferred embodiment of the composition of claim 1, said stansoporphine is a stansoporphine preparation in a single batch.

p00164In accordance with a preferred embodiment of the process of claim 4, in step a) palladium on carbon is exposed to a hydrogen atmosphere of about 30 to 50 psi at a temperature of about 45-50 ° C for about 8 to 16 hours .

p00165In accordance with a preferred embodiment of the method of claim 3, the reaction of mesoporphyrin IX with a tin (II) salt is carried out in the absence of proton scrubbers.

p00166According to a preferred embodiment of the process of claim 5, the oxygen-containing gas is approximately 6% oxygen in nitrogen.

p00167In accordance with a preferred embodiment of the method of claim 9, mesopor30 firmine IX dihydrochloride is readily filterable.

p00168According to a preferred embodiment of the process of claim 12, the solution or suspension of tin (II) oxide is prepared with acetic acid.

p0016935 In accordance with a preferred embodiment of the method of claim 12, the solution or suspension of the non-metalated porphyrin compound or its salt is prepared with formic acid.

p00170In accordance with another preferred embodiment of the method of claim 12, the nonmetalated porphyrin compound is mesoporphyrin IX or its salt.

p00171In accordance with another preferred embodiment of the process of claim 12, the unmetalated porphyrin compound is mesoporphyrin IX dihydrochloride.

p00172In accordance with a preferred embodiment of the method of claim 15, the drip addition is com

p00173Four. Five full between approximately 3 and 9 hours; where preferably the solution or suspension of tin (II) oxide is maintained at a temperature of about 60 to 65 ° C during the drip addition, where also preferably after the completion of the addition of the solution or suspension of the non-metalated porphyrin compound or its salt by dripping to the solution or suspension of tin oxide (II), the reaction mixture

p00174fifty it can be maintained at a temperature of about 60 to 65 ° C for an additional 18 to 24 hours.

p00175In accordance with another preferred embodiment of the process of claim 12, the step of contacting the solution or suspension of tin (II) oxide with the solution or suspension of the non-metaphorsed porphyrin compound or its salt under conditions suitable to form the tin porphyrin compound (IV) or its salt is carried out

p00176in the absence of a proton scrubber or proton sponge. 22