madman
Super Moderator
ABSTRACT
Sustained release of lipophilic compounds can be achieved with oil depots. These parenteral formulations are generally injected into the vastus lateralis and deltoid muscle. It is known that the absorption rate differs between these two muscles. The reason for this is not fully understood. The aim of the current study was to investigate the fate of an oil depot in different tissues to elucidate whether the disappearance rate of oil is the cause of observed differences in absorption rate. A study with healthy volunteers was conducted to determine 1.0 mL oil depots in the vastus lateralis and deltoid muscle for two weeks. The spatial distribution of the oil depots was determined using MRI. Additionally, a study in rats was conducted to microscopically examine the oil immediately and after 31 days of injection. All rats were injected with a 0.1 mL oil depot with and without benzyl alcohol (BOH), a commonly used excipient in oil depots.
In humans, it was shown that all oil depots were equal in volume and surface area directly after injection. Moreover, the disappearance rate for all oil depots was similar; within one week there was no depot visible anymore by MRI. This is in contrast to the depots in rats, which were still microscopically visible after 31 days. It is concluded from these observations that the oil is dispersed to small droplets in the course of time. The resulting increase in the surface area does not lead to an increase in absorption rate, however.
The results of this paper show that the variation in drug absorption as found for the two muscles is not caused by a distinction in surface areas or disappearance rates of the oil depots. Therefore, it is argued that the local tissue drainage (e.g. lymph flow) plays a considerable role in drug absorption from oil depots, whereby the lymph flow differs between the muscles.
1. Introduction
Long-term drug treatment is optimized using sustained delivery of drugs. This is applied for a number of diseases, such as hormone-related (Edelstein and Basaria, 2010; Morgentaler et al., 2008) and psychiatric disorders (Covell et al., 2012; Novakovic et al., 2013; Uchida et al., 2013; van Weringh et al., 1994). This method of drug delivery can be achieved with the use of parenteral oil formulations. Long-acting parenteral injections are administered intramuscularly (i.m.) or subcutaneously (s.c.) (Prettyman, 2005).
A considerable number of oil depots are registered for clinical use. These formulations are composed out of lipophilic compounds, dissolved in vegetable oils (arachis, sesame, or castor oil). A commonly used additive is benzyl alcohol (BOH), which enhances the solubility of the lipophilic compound in the oil decreases the oil viscosity to ease the administration, and provides some local anesthesia. Examples of oil depots registered on the marked contain nandrolone decanoate (ND) (Bagchus et al., 2005; Minto et al., 1997; Wijnand et al., 1985), testosterone undecanoate (Morgentaler et al., 2008), estradiol valerate (Düsterberg and Nishino,1982) or haloperidol decanoate (van Weringh et al.,1994). In all cases, the drug substance is compounded as a lipophilic prodrug. These oil depots are administered only once every 2– 3 weeks.
In recent years, new insights into the fundamental mechanisms of drug absorption from an oil depot have been obtained (Kalicharan et al., 2016c). Here, it is necessary to distinguish the drug release out of the oil depot and the absorption into the systemic circulation. After injection of the oil depot, the inactive prodrug (e.g. nandrolone decanoate) is released into the aqueous phase (interstitial fluid). This drug release is described by mass transport models, in which the release is determined by the drug partition coefficient (log P), the concentration gradient between the oil and aqueous phase, and the surface area of the oil depot. It has been assumed by Shaffer et al.that an injected oil depot forms a spherical shape in muscle tissue (Shaffer, 1929). In theory, a 0.5 mL injected oil depot would result in a spherical object with a surface area of 304.6 mm2. In our previous study, we showed with MRI that the in situ surface area of a 0.5 mL administered oil depot is approximately 750 mm2 (Kalicharan et al., 2016a). This measured surface area is much larger due to the spatial distribution of the oil liquid throughout the muscle fibers. Obviously, the shape was definitely not spherical but stretched.
Once released from the oil depot, the prodrug must be hydrolyzed into its active (parent) compound to become therapeutically active. It is generally known that hydrolysis of prodrugs with an ester bond can occur via chemical or enzymatic routes. Recently, it was concluded that carboxylesterases are responsible for the fast conversion of the prodrug (Kalicharan et al., 2016b). It was shown that prodrug hydrolysis occurs in human whole blood, but it is absent in human plasma and serum. Since the interstitial fluid has a similar composition as serum, hydrolysis at the injection site is not expected to occur either. Surrounding tissue cells contain the appropriate esterases, but because of the poor tissue permeation, the lipophilic compound cannot reach these local enzymes. Therefore, it was argued that the inactive prodrug is drained via the lymphatic system to the systemic circulation where it can be hydrolyzed into the parent compound (Kalicharan et al., 2016b, 2016c). The experiments indicated that the carboxylesterases are located in blood cells. It was shown that the prodrug hydrolysis did not start immediately, but after a delay of approximately half an hour, the time needed for permeation into the cells. The total time delay between the moment of injection and the appearance of the active substance in the systemic circulation (overall lag time) is a result of lymph transport and permeation into blood cells.
The rate at which the drug substance enters the bloodstream is determined by the factors that have been described; The so-called absorption rate constant (ka) was found to be mainly determined by the partition coefficient of the prodrug and the site of injection (Kalicharan et al., 2016c). The three mentioned absorption variables (lag time, ka, and site of injection) are summarized in Fig. 1 for ND. It has been shown that there is a correlation between the lag time and ka for nandrolone after injection in three different muscles (Kalicharan et al., 2016c). The explanation for the differences in kinetic parameters is still lacking. Fig. 1 also shows that an ND oil depot administered in the subcutaneous tissue results in a low nandrolone absorption rate constant and a relatively short lag time.
Although the mechanism of drug absorption from an oil depot has become somewhat more clear, there are still some phenomena that should be studied. First of all, the oil depot formulations and volumes in Fig. 1 were equal for the i.m. injection, and therefore, it is likely that the different ka’s are caused by factors within the body. For example, the shape and hence the surface area of the oil depot may differ. Also, although it is still unknown how rapidly an oil depot (with the prodrug) disappears from different muscles after injection, it is possible that the disappearance rate of the oil differs between these muscles. In this respect, it should be noted that there were some remarkable results in the previous study in which the spatial distribution of an administered oil depot was determined: a significant variation in oil depot disappearance rate was noticed (Kalicharan et al., 2016a). The period in which the oil depot seemed to disappear from the biceps branchii varied between 2 and 14 days. Until now, all pharmacokinetic studies on oil, depots show at least 2 weeks of sustained levels of the active substance (Bagchus et al., 2005; Jann et al.,1985; Minto et al.,1997; van Weringh et al., 1994; Wijnand et al., 1985), indicating that the functionality of sustained drug release is maintained during this period.
In this current study, the surface areas and disappearance rates of oil depots injected in the vastus lateralis and deltoid muscle were determined in situ using MRI. Although this technique was suitable to visualize oil in tissue, there is a possibility that it exhibits a too low sensitivity to detect oil that is spread out in the tissue (Kalicharan et al., 2016a). Therefore, this paper covers also a histology study in rats to determine whether the oil is still present in tissue after some prolonged time. Histological studies of human and rat muscles show a similar structure of tissue (Armstrong et al., 1983; Fridén et al., 1981) and it can therefore be assumed that the observations in rats have significance for human tissue.
The aim of the current study was to investigate the fate of an oil depot in different tissues in order to elucidate whether the disappearance rate of oil is the cause of observed differences in absorption rate. Both the MRI and histology study should provide some clarity in the rate of absorption of active substances from oil depots.
3.3. Mechanisms of drug absorption
The fact that the oil is obviously still present in the tissue explains the continuing exposure of the active substance. However, an interesting aspect of the formation of small droplets is an increase in surface area. Mass transfer models state that the surface area of the oil depot is relevant for the release rate of the compound. An increased surface area should then lead to a higher release. This implies that a larger injected volume of oil results in a higher release rate of dissolved compounds due to the larger surface area of the depot. The obvious explanation for this is that the surface area of the separate droplets is apparently of minor importance. In this respect, it can be argued that the drug release rate is determined by the outer layer of the assembly of droplets, encapsulated at the injection site (as seen in Fig. 3F); Consequently, the droplets in the center of the fragmented oil depot have a minor contribution to the drug release rate.
The current study shows that there is no reason to assume that differences in the spatial distribution of the oil (Table 3) are responsible for the rate of absorption or lag time that has been found for the tissues (Fig. 1). Therefore, it must be concluded that the cause must be found elsewhere in the body. Several body parameters have been suggested to be relevant during drug absorption (Prettyman, 2005; Zuidema et al., 1994, 1988): type of tissue, blood and lymph flow in tissues, physical activity of the tissue and the local activation of the immune system.
Recently, Darville et al. reported that the immune system is activated after parenteral administration (Darville et al., 2016). It is generally known that the immune system is triggered by foreign material. This might be the mechanism of oil disappearance after injection. And although not studied by us, it is very likely that this activation is related to the amount of foreign material. This could be the reason why the larger amount (1.0 mL) of administered oil in this study disappears earlier than the lower amount (0.5 mL) of oil in the previous study (see Fig. 2).
Fig.1 shows that not only does the drug absorption rate constant and lag time between i.m. injected oil depots differ, but also the subcutaneous from the intramuscular injection. It has already been argued that drug absorption of lipophilic prodrugs starts with uptake via the lymphatic system. Subsequently, once transported through the lymphatic system, the prodrug enters the vena cava superior and the systemic circulation. Here, it is hydrolyzed. With the findings of the current study, we conclude that the cause of absorption variables cannot be found in the fate of the oil after injection. At the same time, once in the systemic circulation, there is no difference either. This means that the cause must be found in between. Therefore, the difference in drug absorption rate constant and lag time from s.c. or i.m. injected oil depots can be explained by different absorption path lengths via lymph flows and tissue-specific lymphatic vessel lengths.
4. Conclusion
In this paper, the rate of disappearance of oil depots from the vastus lateralis and deltoid muscle is similar. The scanning method with MRI is only sufficient to determine the parameters of interest on the same injection day. After one day of injection, histological studies in rats showed that the oil is dispersed and that a part of the oil remains at the site of injection for 31 days. This was not seen with MRI analysis. It is concluded that differences in drug absorption cannot be explained by drug product properties such as the surface areas or mentioned disappearance rates of oil depots. Therefore, body factors such as the activated immune system can influence this with macrophages. Speculatively, the lymph flow and path length at the injection site is argued to be the dominating factor in drug absorption of the released compounds from oil depots.
Sustained release of lipophilic compounds can be achieved with oil depots. These parenteral formulations are generally injected into the vastus lateralis and deltoid muscle. It is known that the absorption rate differs between these two muscles. The reason for this is not fully understood. The aim of the current study was to investigate the fate of an oil depot in different tissues to elucidate whether the disappearance rate of oil is the cause of observed differences in absorption rate. A study with healthy volunteers was conducted to determine 1.0 mL oil depots in the vastus lateralis and deltoid muscle for two weeks. The spatial distribution of the oil depots was determined using MRI. Additionally, a study in rats was conducted to microscopically examine the oil immediately and after 31 days of injection. All rats were injected with a 0.1 mL oil depot with and without benzyl alcohol (BOH), a commonly used excipient in oil depots.
In humans, it was shown that all oil depots were equal in volume and surface area directly after injection. Moreover, the disappearance rate for all oil depots was similar; within one week there was no depot visible anymore by MRI. This is in contrast to the depots in rats, which were still microscopically visible after 31 days. It is concluded from these observations that the oil is dispersed to small droplets in the course of time. The resulting increase in the surface area does not lead to an increase in absorption rate, however.
The results of this paper show that the variation in drug absorption as found for the two muscles is not caused by a distinction in surface areas or disappearance rates of the oil depots. Therefore, it is argued that the local tissue drainage (e.g. lymph flow) plays a considerable role in drug absorption from oil depots, whereby the lymph flow differs between the muscles.
1. Introduction
Long-term drug treatment is optimized using sustained delivery of drugs. This is applied for a number of diseases, such as hormone-related (Edelstein and Basaria, 2010; Morgentaler et al., 2008) and psychiatric disorders (Covell et al., 2012; Novakovic et al., 2013; Uchida et al., 2013; van Weringh et al., 1994). This method of drug delivery can be achieved with the use of parenteral oil formulations. Long-acting parenteral injections are administered intramuscularly (i.m.) or subcutaneously (s.c.) (Prettyman, 2005).
A considerable number of oil depots are registered for clinical use. These formulations are composed out of lipophilic compounds, dissolved in vegetable oils (arachis, sesame, or castor oil). A commonly used additive is benzyl alcohol (BOH), which enhances the solubility of the lipophilic compound in the oil decreases the oil viscosity to ease the administration, and provides some local anesthesia. Examples of oil depots registered on the marked contain nandrolone decanoate (ND) (Bagchus et al., 2005; Minto et al., 1997; Wijnand et al., 1985), testosterone undecanoate (Morgentaler et al., 2008), estradiol valerate (Düsterberg and Nishino,1982) or haloperidol decanoate (van Weringh et al.,1994). In all cases, the drug substance is compounded as a lipophilic prodrug. These oil depots are administered only once every 2– 3 weeks.
In recent years, new insights into the fundamental mechanisms of drug absorption from an oil depot have been obtained (Kalicharan et al., 2016c). Here, it is necessary to distinguish the drug release out of the oil depot and the absorption into the systemic circulation. After injection of the oil depot, the inactive prodrug (e.g. nandrolone decanoate) is released into the aqueous phase (interstitial fluid). This drug release is described by mass transport models, in which the release is determined by the drug partition coefficient (log P), the concentration gradient between the oil and aqueous phase, and the surface area of the oil depot. It has been assumed by Shaffer et al.that an injected oil depot forms a spherical shape in muscle tissue (Shaffer, 1929). In theory, a 0.5 mL injected oil depot would result in a spherical object with a surface area of 304.6 mm2. In our previous study, we showed with MRI that the in situ surface area of a 0.5 mL administered oil depot is approximately 750 mm2 (Kalicharan et al., 2016a). This measured surface area is much larger due to the spatial distribution of the oil liquid throughout the muscle fibers. Obviously, the shape was definitely not spherical but stretched.
Once released from the oil depot, the prodrug must be hydrolyzed into its active (parent) compound to become therapeutically active. It is generally known that hydrolysis of prodrugs with an ester bond can occur via chemical or enzymatic routes. Recently, it was concluded that carboxylesterases are responsible for the fast conversion of the prodrug (Kalicharan et al., 2016b). It was shown that prodrug hydrolysis occurs in human whole blood, but it is absent in human plasma and serum. Since the interstitial fluid has a similar composition as serum, hydrolysis at the injection site is not expected to occur either. Surrounding tissue cells contain the appropriate esterases, but because of the poor tissue permeation, the lipophilic compound cannot reach these local enzymes. Therefore, it was argued that the inactive prodrug is drained via the lymphatic system to the systemic circulation where it can be hydrolyzed into the parent compound (Kalicharan et al., 2016b, 2016c). The experiments indicated that the carboxylesterases are located in blood cells. It was shown that the prodrug hydrolysis did not start immediately, but after a delay of approximately half an hour, the time needed for permeation into the cells. The total time delay between the moment of injection and the appearance of the active substance in the systemic circulation (overall lag time) is a result of lymph transport and permeation into blood cells.
The rate at which the drug substance enters the bloodstream is determined by the factors that have been described; The so-called absorption rate constant (ka) was found to be mainly determined by the partition coefficient of the prodrug and the site of injection (Kalicharan et al., 2016c). The three mentioned absorption variables (lag time, ka, and site of injection) are summarized in Fig. 1 for ND. It has been shown that there is a correlation between the lag time and ka for nandrolone after injection in three different muscles (Kalicharan et al., 2016c). The explanation for the differences in kinetic parameters is still lacking. Fig. 1 also shows that an ND oil depot administered in the subcutaneous tissue results in a low nandrolone absorption rate constant and a relatively short lag time.
Although the mechanism of drug absorption from an oil depot has become somewhat more clear, there are still some phenomena that should be studied. First of all, the oil depot formulations and volumes in Fig. 1 were equal for the i.m. injection, and therefore, it is likely that the different ka’s are caused by factors within the body. For example, the shape and hence the surface area of the oil depot may differ. Also, although it is still unknown how rapidly an oil depot (with the prodrug) disappears from different muscles after injection, it is possible that the disappearance rate of the oil differs between these muscles. In this respect, it should be noted that there were some remarkable results in the previous study in which the spatial distribution of an administered oil depot was determined: a significant variation in oil depot disappearance rate was noticed (Kalicharan et al., 2016a). The period in which the oil depot seemed to disappear from the biceps branchii varied between 2 and 14 days. Until now, all pharmacokinetic studies on oil, depots show at least 2 weeks of sustained levels of the active substance (Bagchus et al., 2005; Jann et al.,1985; Minto et al.,1997; van Weringh et al., 1994; Wijnand et al., 1985), indicating that the functionality of sustained drug release is maintained during this period.
In this current study, the surface areas and disappearance rates of oil depots injected in the vastus lateralis and deltoid muscle were determined in situ using MRI. Although this technique was suitable to visualize oil in tissue, there is a possibility that it exhibits a too low sensitivity to detect oil that is spread out in the tissue (Kalicharan et al., 2016a). Therefore, this paper covers also a histology study in rats to determine whether the oil is still present in tissue after some prolonged time. Histological studies of human and rat muscles show a similar structure of tissue (Armstrong et al., 1983; Fridén et al., 1981) and it can therefore be assumed that the observations in rats have significance for human tissue.
The aim of the current study was to investigate the fate of an oil depot in different tissues in order to elucidate whether the disappearance rate of oil is the cause of observed differences in absorption rate. Both the MRI and histology study should provide some clarity in the rate of absorption of active substances from oil depots.
3.3. Mechanisms of drug absorption
The fact that the oil is obviously still present in the tissue explains the continuing exposure of the active substance. However, an interesting aspect of the formation of small droplets is an increase in surface area. Mass transfer models state that the surface area of the oil depot is relevant for the release rate of the compound. An increased surface area should then lead to a higher release. This implies that a larger injected volume of oil results in a higher release rate of dissolved compounds due to the larger surface area of the depot. The obvious explanation for this is that the surface area of the separate droplets is apparently of minor importance. In this respect, it can be argued that the drug release rate is determined by the outer layer of the assembly of droplets, encapsulated at the injection site (as seen in Fig. 3F); Consequently, the droplets in the center of the fragmented oil depot have a minor contribution to the drug release rate.
The current study shows that there is no reason to assume that differences in the spatial distribution of the oil (Table 3) are responsible for the rate of absorption or lag time that has been found for the tissues (Fig. 1). Therefore, it must be concluded that the cause must be found elsewhere in the body. Several body parameters have been suggested to be relevant during drug absorption (Prettyman, 2005; Zuidema et al., 1994, 1988): type of tissue, blood and lymph flow in tissues, physical activity of the tissue and the local activation of the immune system.
Recently, Darville et al. reported that the immune system is activated after parenteral administration (Darville et al., 2016). It is generally known that the immune system is triggered by foreign material. This might be the mechanism of oil disappearance after injection. And although not studied by us, it is very likely that this activation is related to the amount of foreign material. This could be the reason why the larger amount (1.0 mL) of administered oil in this study disappears earlier than the lower amount (0.5 mL) of oil in the previous study (see Fig. 2).
Fig.1 shows that not only does the drug absorption rate constant and lag time between i.m. injected oil depots differ, but also the subcutaneous from the intramuscular injection. It has already been argued that drug absorption of lipophilic prodrugs starts with uptake via the lymphatic system. Subsequently, once transported through the lymphatic system, the prodrug enters the vena cava superior and the systemic circulation. Here, it is hydrolyzed. With the findings of the current study, we conclude that the cause of absorption variables cannot be found in the fate of the oil after injection. At the same time, once in the systemic circulation, there is no difference either. This means that the cause must be found in between. Therefore, the difference in drug absorption rate constant and lag time from s.c. or i.m. injected oil depots can be explained by different absorption path lengths via lymph flows and tissue-specific lymphatic vessel lengths.
4. Conclusion
In this paper, the rate of disappearance of oil depots from the vastus lateralis and deltoid muscle is similar. The scanning method with MRI is only sufficient to determine the parameters of interest on the same injection day. After one day of injection, histological studies in rats showed that the oil is dispersed and that a part of the oil remains at the site of injection for 31 days. This was not seen with MRI analysis. It is concluded that differences in drug absorption cannot be explained by drug product properties such as the surface areas or mentioned disappearance rates of oil depots. Therefore, body factors such as the activated immune system can influence this with macrophages. Speculatively, the lymph flow and path length at the injection site is argued to be the dominating factor in drug absorption of the released compounds from oil depots.
