Keywords

1 Introduction

The inhalation of medicinal compounds has been tested and documented in the peer-reviewed scientific literature as a delivery method to rid a patient of Mycobacterium tuberculosis (M. tb) infection for over a century [1,2,3,4,5,6,7,8,9,10,11,12,13,14,15,16,17,18,19,20,21,22,23,24,25,26,27,28,29,30,31,32,33,34,35,36,37,38,39,40,41,42,43,44,45,46,47,48,49,50,51,52,53,54,55,56,57,58,59,60,61,62,63,64,65,66,67,68,69,70,71,72,73,74,75,76,77]. M. tb infections have ravaged humanity for at least ten thousand years [76]. TB is spread through the air by someone expelling water droplets infected with M. tb through a cough or sneeze. Water droplets containing M. tb can float in the air for several hours, making it possible for someone else nearby to inhale the bacteria. The most common form of TB, pulmonary TB (PTB), occurs when the bacteria attack the lungs. Extrapulmonary TB (EPTB) occurs when the bacteria infect other parts of the body, including the brain [11]. Contradictory to PTB, EPTB is rarely transmitted to others. The combination of isoniazid (INH), rifampin (RIF), and other first- and second-line anti-TB medications, are widely used to treat M. tb infections and can take six to nine months to complete the regimen. Combination therapy reduces the risk of the bacteria becoming drug-resistant (DR) [4]. DR-TB is a type of infection in which the bacteria become resistant to the primary drugs used to treat TB. This condition can arise from improper use of medications and regimens and not completing the full course of treatment. Totally DR-TB and extensively drug-resistant (XDR) TB are the most severe forms of M. tb infections. Bacille Calmette-Guérin (BCG) is a vaccine used for TB but has limited efficacy. BCG is recommended for children and adults at an increased risk of infection or exposure to TB disease.

Infection by M. tb does not necessarily result in immediate disease expression. When a subject is infected, granulomas are formed around the bacterium [84] to isolate and prevent its growth. The human body is efficient in containing outbreaks of TB after infection by sealing the disease in lesions. Although the TB is contained, the body cannot eradicate it, so latent TB infection (LTBI), or inactive TB, is expressed once the immune system is compromised. This can be observed in the syndemic relationship between TB and HIV; HIV drastically weakens the immune system, causing a TB outbreak to proliferate. Tuberculin-based skin tests are used to test for TB in travelers. There are two types of TB tests that are economical and routinely utilized by medical professionals: the skin test and the blood test. A positive test indicates that the patient has been infected with the M. tb but does not determine whether or not it is latent or active TB. Further diagnostic tests are needed to determine whether TB is latent or active in the patient [10]. Lifestyle, as well as environmental factors, play a significant role in TB recovery. People with active TB self-quarantine to avoid infecting others. Patients may be recommended directly observed therapy (DOT), which requires you to take your medicine in front of your doctor several times a week. DOT helps prevent DR; however, it tends to add staggering costs to the patient.

2 History

Before the first mass production of penicillin in the 1940s and the introduction of antibiotics used to treat M. tb infections in the 1950s through the 1970s, there was no established treatment for the disease. The following three papers give an insight into some of the inhalation treatments tested. In a paper published in 1898, a medical professional had up to 2000 TB patients under his care. The infectious disease caused by M. tb was typically advanced in the patients [1]. The inhalation treatment was focused on the application of a vapor composed of:

  • eucalyptol (C10H18O), which is a cyclic ether and a monoterpenoid, also called 1,8-cineol;

  • oil of cloves (oil extracted from the clove plant, Syzygium aromaticum found in Southeast Asia, and is composed of β-caryophyllene, α-humulene, eugenyl acetate, and eugenol; and other ingredients

A quote from the author summarizes the results:

“symptoms have improved in all cases, though some have improved significantly more than others”.

These were a most unfavorable class of patients to treat, being in the advanced stage of the tubercular process with hemorrhages, emaciation, night sweats, anemia, and, in fact, scarcely able to breathe at all. In the absence of a specific symptom for TB, we believe that with proper apparatus and skillful and continued administration, much is to be hoped for in this class of patients by inhaling the antiseptic vapor. The historical literature does not explain why this promising approach was abandoned.

Benzol was the name for benzene and could be inhaled as a vapor. To put the study in perspective, penicillin was the first mass-produced antibiotic, which happened in the 1940s. The concept of an atom was not fully understood, emphasizing the Nobel prize in physics awarded to James Chadwick to discover the neutron in 1935. Benzene is now recognized by IARC as a human carcinogen that causes leukemia and non-Hodgkin lymphoma. The study did report changes in changes in the function of kidneys, liver, and hearts muscles in the mice, and the paper concluded [2]:

that with further study of this rather specific poison, we may get a little nearer the solution of some of the unsolved problems of infection and immunity.

Studies of this nature, before the modern era of antibiotics, emphasize the desperation in seeking treatments using toxic compounds that were very poorly understood.

In a 1935 paperback entitled “Inhalation Therapy Technique” by W. Collison [3], there is a chapter dedicated to inhalation therapy in PTB. First, the author outlined some oils and liquids cannot be used for inhalation therapy because the large particle sizes generated allow minimal penetration of the substance deep into the lungs. Likewise, there are medical sprayers used in that time period for treating the nasal cavity or throat maladies that do not provide the parameters needed for the mist to penetrate deep into the lungs. An Apneu Inhaling Apparatus was used to deliver the vapor to a mask that covers the patients’ faces and mouths. There is a discussion on the parameters that impact the size of the particles, as small as “1/5000th of an inch.” The apparatus is used to deliver compounds such as adrenalin, camphor, menthol, and creosote. The substances are delivered with a purpose; adrenalin is used to reduce congestion by dilating the bronchial tubes; camphor is stated to stimulate blood circulation and breathing; thymol and menthol are antiseptic; creosote is a disinfectant; pine and cypress soothed the inflamed mucous membrane, etc. A typical prescription is described as three minutes of adrenalin therapy, a three-minute rest, a three-minute camphor therapy, a three-minute rest, followed by a three-minute adrenalin dosage. While not described as a cure, it did help patients feel better or “obtain relief.”

3 Modern Work

Currently, there are three types of inhalers used for lung infection treatments:

  1. i.

    Small-volume nebulizer (SVN), it generates a liquid into an aerosol, and the droplets are propelled by a compressed gas, typically air or pure oxygen;

  2. ii.

    Dry powder nebulizer delivers the medication as a powder or small solid particles; and

  3. iii.

    The pressurized metered-dose inhaler (MDI), it is the most common type.

Although not placed in the same category of inhalers, anesthetics are gases that can be inhaled but are not used directly to treat a medical condition. There are some unsuccessful ventures mentioned in the literature in which gases such as carbon dioxide were used unsuccessfully.

For a medication to function properly, it should have the ability to reduce or eliminate the bacterial load contained within a macrophage without eliminating a large percentage of the macrophages residing within the lungs. Often heavy doses of antibiotics have to be given to patients to overwhelm physiological barriers such as macrophages and granuloma to eliminate the M. tb. This results in significant side effects for the patient.

We have identified the following pharmacokinetic and pharmacodynamics aspects related to the administration of inhaled agents with medicinal activity for a viral infection that need to be considered if a large fraction of the droplets will reach the desired region of the lungs and be effective in reducing the bacterial load in the lungs and entering the bloodstream to eliminate its presence throughout the body (Box 1).

Box 1

A sequential outline of the biological and chemical processes that could impact the drug delivery efficiency when inhalation is utilized

Some electronic vaporization processes can potentially cause a fraction of the medication or molecule to react or degrade;

The medication is absorbed in the mouth or throat;

The medication enters the digestive tract;

The medication is absorbed in the trachea;

The medication is absorbed in the upper lungs;

The medication is exhaled;

If a solid, the medication does not completely dissolve;

The medication penetrates the granuloma model (Sarcoidosis) intact;

If a prodrug, it has to encounter a reactive site (i.e., enzyme);

The medication enters the macrophage;

Depending on the MOA, the medication has to penetrate or disrupt the mycolic acid membrane;

The medication can undergo unwanted protein binding;

The medication enters the bloodstream and is distributed, metabolized, and excreted without interacting with any M. tb;

The medicine is evenly distributed through the alveolus;

The medication undergoes hydrogen bonding to an unwanted species as an H donor or acceptor;

The water solubility of the medication limits its mobility;

The size (high molar mass) of the medication limits its mobility; and

The composition of the inhaled medicinal particle is such that the dissolution and delivery process of the medication falls within acceptable parameters needed to effectively treat the patient.

For inhalation to meet each of these processes efficiently, arguably the most important parameter is the size of the particle. It should be small, with a diameter less than 200 nm, with values in the 50–70 nm range considered optimal to fully penetrate the alveoli in the lower lung. Capreomycin is a second-line antibiotic used to treat DR strains of M. tb. There are two drawbacks to its current administration, as a tablet or an injection:

  1. i.

    it involves an injection which can be problematic for several reasons, especially for children and emaciated adults with low muscle content; and

  2. ii.

    it necessitates regular visits to a health care facility.

Studies by Garcia-Contreras et al. [21, 24] developed a low-density particle that was produced by a sprayed dry technique that contained capreomycin. The study produced pharmacokinetics data when the particles were inhaled by guinea pigs with an optimum dose of 14.5 mg/kg, which for a 70 kg adult would translate to a dose of 1.015 g. It was argued that if the approach was applied to humans, it would eliminate injections and lower side effects. A scanning electron micrograph supplied an image that indicated the particles are in the range of approximately two to four micrometers in diameter. Alveolar and interstitial macrophages, which can be M. tb reservoirs in the lung, have 17.1 and 13.2 µm diameters for nonsmokers, 23.7 and 11.3 µm for smokers, and 23.7 and 11.8 µm for chronic obstructive pulmonary disease patients. The medicine has to penetrate the macrophage before acting on the bacterium. Significantly reducing the size of the capreomycin particles, coupled with a coating that might make the particle appear as cellular debris and/or a nutrient to the macrophage, might increase the uptake rate.

Antibiotics used to treat DR strains of M. tb can result in significant side effects, including ototoxicity and nephrotoxicity. Barberis et al. [76] compared capreomycin to amikacin for up to 192 days for patients with MDR-TB and up to 735 days with XDR-TB. Their study revealed that amikacin was up to five times more likely than capreomycin to result in severe ototoxicity. Amikacin had less hypokalemia (low potassium levels in serum) than capreomycin. Both sets of patients, those given amikacin or capreomycin, experienced a similar increase in the creatinine levels.

In a follow-up publication [24], the same group measured pharmacokinetic (PK) parameters related to the same capreomycin particles being inhaled by guinea pigs. This study focused on doses of 20 mg/kg. The capreomycin concentration in bronchoalveolar fluid and lung tissue of the animal was up to one-hundred times greater than in the plasma when compared to guinea pigs that received the medication via injection.

DR strains of M. tb have become more problematic worldwide. The treatments for MDR, XDR, and TDR have more severe side effects when compared to latent or active TB. Attempts to develop a method that could deliver the antibiotics more efficiently seem to be a natural progression in order to penetrate the lungs. Capreomycin is a second-line TB drug that belongs to a group of medications called glycosides and has been on the market since 1979. It is administered by injection daily for two to four months and then reduced to two or three times per week but varies with the patient’s condition. Patients who suffer from trypanophobia or fear of injections and those with very low muscle mass, such as a child or an adult with a chronic condition, often will quickly stop taking the medication. Administering capreomycin via an inhalation route not only removes the use of injections but should also lower the dose and, subsequently, the side effects.

In a rare study involving human patients [44], capreomycin was formulated as micrometer-sized particles, which was produced by a dry spray technique. An aerosol approach was used to deliver the micron-sized particles to the lungs. This was a phase 1 trial using 20 healthy adults with the goal of measuring several pharmacokinetic parameters. This study incorporated a relatively simple but efficient method for the patients to self-medicate, a tremendous improvement over visiting a medical office on a daily basis for injection or an IV. The single daily doses administrated were 25, 75, 150, or 300 mg doses of capreomycin, with five patients in each dosage group. The doses were 25 mg of capreomycin with 5 mg lysine serving as an excipient. The 300 mg administration required the patient to sequentially self-administer twelve doses of the medication.

The patients had their blood sampled for capreomycin four times before the antibiotic was delivered and at eight points (1, 2, 4, 6, 8, 12, and 24 h) after delivery. A sample of the mean area under the curve (AUC) values over a finite time interval for each of five patients was measured; 969 (h ng/ml) for the 25 mg dose group; 3555 (h ng/ml) for the 75 mg group; 7019 (h·ng/ml) for the 150 mg dose group; and 19,959 (h ng/ml) for the 300 mg dose group. The Cmax values measured were 169, 569, 972, and 2315 ng/mL, correspondingly. The published in vitro MIC value for capreomycin treating M. tb was 2 μg/ml (or 2000 ng/ml). The Cmax for the 300 mg dose group was the only value that was above the MIC value.

4 The Future

The following is a proposed structure and administration route that could be applied for latent, active, and resistant M. tb strains. It utilizes electronic vaporization as a method to form an aerosol and deliver the medication. Our group has incorporated copper in selected cancer drugs and antibiotics for several reasons [78,79,80,81,82,83,84]. Metal–ligand complexes composed of Cu(II)-sucrose and Cu(II)-DALB (denatured albumin) were built to minimize unwanted interactions such as protein binding or the Cu(II) cation generating reactive oxidation species prematurely. Mostly, M. tb is impacted by the toxicity of copper metal or the copper ion (Cu(II), Cu(I), Cu(0)). We demonstrated that a copper-capreomycin complex has a higher efficacy against active and resistant strains of M. tb [84]. The MIC values were up to 200 times lower for the copper-capreomycin complex than for pure capreomycin. The complex was synthesized and characterized in our lab and tested at the National Institutes of Health (Bethesda, MD, USA) against active INH-R (isoniazid-resistant), RMP-R (rifampin-resistant), and OFX-R (Ofloxacin resistant) strains of M. tb. While adding the Cu(II) cation to the capreomycin molecule improves parameters related to Lipinski’s Rules, such as water solubility and the number of hydrogen bonds possible, it also lowered the MIC value significantly [83]. The presence of copper might raise toxicity concerns when administrated orally due to higher doses. Applying the complex directly to the lungs decreases the dose, lowers side effects associated with higher doses given by tablet, and can increase the effectiveness of the treatment.

The results of a time-of-flight mass spectrometry analysis (Fig. 1) illustrate that denatured human serum albumin (DALB), used as a drug delivery platform, can be electronically vaporized using a Propylene Glycol-Glycerol-Ethanol solvent and transported as a vapor through two feet of 0.3 cm inside diameter tubing [75]. Figure 2 provides a novel complex to deliver the copper-capreomycin molecule. The DALB structure is denatured using ethanol, allowing the glucose molecules and the copper (II)-capreomycin complex to attach (bond) to it. Glucose is added via a glycation reaction and included so that macrophages and M. tb recognize it as an energy source and increase the medication uptake rate. DALB may also be consumed by the macrophage because it is recognized as cellular debris, providing an easy entry to the location of the M. tb reservoir. Also, both the macrophage and M. tb, sensing amino acids, would consume the complex to fulfill its nutrient and energy needs. Copper strongly binds the amines on capreomycin (CAP) molecules and to the amines on the protein structure, serving as an atomic level connection. The copper (II) cation is highly toxic to M. tb and, because of its high metal-ligand stability constant, is somewhat protected by the protein from dissociating from the complex. The protein-glucose-copper-capreomycin complex (PGCC) is small enough to be engulfed/phagocytosed by local cells or to leave the lungs and be transported through the circulatory system. In pulmonary tuberculosis (PTB), a common form of the disease, M. tb enters the lungs and is consumed by a macrophage as a single complex would be the desired route.

Fig. 1
figure 1

A MALDI-TOF-mass spectra of human serum albumin. It was electronically vaporized and transported

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Fig. 2
figure 2

A proposed delivery method for the copper-capreomycin complex, a novel complex was developed specifically for patients infected by M. tb that uses a protein-glucose-Cu complex to deliver capreomycin via inhalation to the patients’ lungs

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There are review papers that take the reader back hundreds and even thousands of years and outline the impact that M. tb infections had on various cultures and time periods [76]. With modern chemical separation, synthesis, and analysis techniques providing a more detailed look at the compositions of medical treatments, this review focused on modern developments. There are two key messages taken away from this review:

  1. i.

    inhalation therapy should be examined in closer detail as a method of treatment for the different forms of TB; and

  2. ii.

    because of some unique features of M. tb, such as its waxy (mycolic acid) outer membrane and its ability to reside within a macrophage for long periods, treatments should be devised at a molecular level with these considerations in mind, rather than borrowing techniques and technologies from disorders that have a different set of biological and chemical conditions.

There is a significant crossover and adaption of inhalation technologies from other conditions such as cystic fibrosis and viral infections.

5 Conclusion

With a rise in antibiotic resistance, new techniques are needed to treat patients. This paper reviewed inhalation therapy first from a historical basis. Despite the fact that PTB is very common, there has been very little work using inhalation therapy that has been brought to clinical trials. For the future, with the right delivery mechanism, capable of making the small droplets needed to penetrate into the lower lungs, coupled with using a solvent mixture and a formulation that can attack the M. tb with several mechanisms of action, inhalation therapy may prove to be part of the solution.

Core Messages

  • Inhalation therapy offers the potential to transform the treatment of all levels of TB therapies.

  • Existing vaporization and inhalation technology can be adapted to drug delivery.

  • Millions of users have tested the units, and delivery parameters are well understood.

  • Ethanol and glycerol can be used for inhalation and delivery, having minimal impact on the patient.

  • We welcome collaboration with any group interested in delivering CuINH or CuCAP for pulmonary TB.