- Non-steroid Anti-inflammatory Drugs
- Gene Therapy
- Pseudomonas vaccines
- Dextrans and Other Mucolytics
- Synercid, compassionate use antibiotic
- Gentamicin and Stop Mutations
- CF Airway Epithelia Fail to Kill Bacteria
- Inhaling Buffered L-arginine
- Colchicine and Chemo Agents
- Small Molecular Weight Solvents
- Azithromycin and Pseudomonas aeruginosa
- Nebulized Heparin
- Cathelicidin Peptides
- A month before I started the l-arginine my PFTs were: FVC: 1.81 (35.7%); FEV1: 1.09 (25.4%)
- Five days after starting l-arginine: FVC: 2.41 (47.5%); FEV1: 1.27 (29.6%)
- One month after starting l-arginine: FVC: 3.22 (61.5%); FEV1: 1.79 (40.5%)
- On 12-11-96 (almost 8 mo. later): FVC: 3.84 (75.8%); FEV1: 2.20 (51.3%)
- On 05-29-97 (fourteen months later): FVC: 3.62 (71.4%); FEV1: 2.11 (49.2%) — this was a few days after stopping IV antibiotics..
Source: S.D. Freedman, M.H. Katz, E.M. Parker, M. Laposato, M.Y. Urman, and J.G. Alvarez, “A membrane lipid imbalance plays a role in the phenotypic expression of cystic fibrosis in cftr -/- mice,” PNAS, Vol. 96, Issue 24, November 23, 1999, 13995-14000
Docosahexaenoic acid, or DHA (not to be confused with DHEA!), is an essential fatty acid. The work of Steven D. Freedman, Juan G. Alvarez, and their colleagues at the Harvard Medical School, has recently shown that cftr-knockout mice have a 3-fold decrease in membrane-bound DHA and a 3-fold increase in membrane-bound arachidonic acid (AA), another essential fatty acid. Arachidonic acid and DHA compete with each other through esterification for placement as membrane phospholipids. AA tends to stimulate both inflammation and mucus secretion. The researchers are unclear how this lipid imbalance is linked to the genetic mutation causing CF. However, the lipid imbalance was most pronounced in the ileum, pancreas, and lungs of the cftr knockout mice, with the heart also being affected. The brain and kidneys of the mice seemed not to be severely affected by this lipid imbalance. This is preliminary evidence that CF pathology is related to the degree of imbalance between AA and DHA. The organs with the greatest imbalances are also the organs most affected by CF. The researchers are able to rule out intestinal malabsorption or hepatic dysfunction as the cause of the imbalance, and so somehow, then, the lipid imbalance must be related to faulty CFTR function.
What was most heartening was that this lipid imbalance could be reversed and normalized by oral administration of DHA. (Though fish oil is high in DHA, it also contains EPA which competes with DHA. The researchers thus used pure DHA.) Oral admnistration of DHA reversed pancreatic and ileal pathology in these mice. (Luminal diameter and villus height were normalized.) In addition, exaggerated neutrophil infiltration into te CF lung in response to Pseudomonas LPS was normalized. If these results could be duplicated in humans, this would be a tremendous therapeutic breakthrough, as DHA would represent one of the few therapies that would act to help prevent the deterioration of a CF person’s body.
Though other omega-3 fatty acids, such as alpha-linoleic acid or EPA, can lower membrane-bound AA (an omega-6), neither normalizes DHA levels and thus cannot reverse pathology. Only DHA both lowered AA and increased DHA levels, which resulted in the normalization noted above. The Harvard Medical School team intends to conduct clinical trials in Summer 2000, using a concentrated dose of DHA in CF patients. Genzyme Corporation is helping to develop the DHA product they will use. Previously, the team had used DHA produced by Martek, Inc., which sells DHA to health food stores under the trademark Neuromins. Dosages on the order of almost 1 gram DHA per day per 10 kilograms body weight are being contemplated. However, since it is possible to damage the liver if too much DHA is used, CF persons are strongly cautioned not to take such dosages until safety (and efficacy) can be demonstrated in these trials. We should know more in the Fall of 2000.
Non-steroid Anti-inflammatory Drugs
A study was completed at Rainbow Babies and Children’s Hospital (one of the university hospitals at Case Western Reserve University) on the use of high dose ibuprofen to counter inflammation in the CF lung. As covered in the January, 1996 issue of the IACFA Newsletter (issue 39) page 9: “The recently completed Ibuprofen study suggests that this non-steroidal anti-inflammatory can reduce the progression of lung disease in CF. Over the study’s four-year term, the rate of decline in FEV1 was nearly halved for persons receiving Ibuprofen, as compared with those taking a placebo.
However, the immunosuppressive effects of Ibuprofen are extremely dose-dependent, and must be carefully individualized for each patient. Further, ‘low’ doses of Ibuprofen – quantities typically used to alleviate mild aches and pains – actually stimulate neutrophil migration to some tissue types, and thus self-medication is absolutely not indicated, and is ‘possibly dangerous at a low dose’.”
The March 30, 1995 issue of the New England Journal of Medicine (Vol. 332, No. 13, pp. 848-54) reported on the same study. The positive effects are seen in the under 13 years old group, not in the over 13 age group (presumably due to more advanced disease in the older group). *Low doses* of ibuprofen are worse than none; *high doses* are associated with renal side effects. The point to be made is that ibuprofen, if proven beneficial, must be administered throughout life. This puts rather stringent requirements on toxicity or adverse reactions. The dose used is uncomfortably close (perhaps “dangerously close” would be more accurate) to (slightly below) that which produces renal side effects. In brief, DON’T SELF-MEDICATE; THIS STUFF MIGHT DO YOU REAL HARM!
With assistance from the Cystic Fibrosis Foundation, the authors of the study have formed the Cystic Fibrosis Ibuprofen Laboratory (at the Division of Clinical Pharmacology of the CWRU School of Medicine). The CF Ibuprofen Laboratory was formed to provide ibuprofen analyses and therapeutic recommendations for cystic fibrosis patients. It facilitates physicians in performing an Ibuprofen Test, which is used to determine the appropriate and accurate dose of ibuprofen for individual patient ibuprofen therapy, following the guidelines established during the course of this study. CF Ibuprofen Laboratory World Wide Web page:
The website provides instructions to physicians and patients about ibuprofen dosing, gives brief methodology on how plasma ibuprofen levels are determined, gives a sample test report, and also provides information about how to get in touch with them. There are links to other areas of the Internet relevant to Cystic Fibrosis.
email: [email protected]
Cystic Fibrosis Ibuprofen Laboratory Division of Clinical Pharmacology Dept. of Medicine, University Hospitals of Cleveland Case Western Reserve University School of Medicine Cleveland, Ohio, U.S.A.
The work of John Lloyd-Still and his colleagues in Chicago has shown that high dose ibuprofen causes intestinal ulcers in about 50% of subjects (he used a rat model ).  When rats not only received high dose ibuprofen, but also received pancreatic enzymes, the severity of those ulcers increased significantly. There have been reports on the cystic-l list of children developing blood in the stools after receiving high dose ibuprofen therapy. However, John Lloyd-Still discovered that the use of ursodeoxychoic acid (Urso) in rats who were also receiving high dose ibuprofen as well as pancreatic enzymes helped the situation tremendously. Only 14% of the rats developed ulcers of the intestine, and none of the ulcers were severe.  These are issues you should discuss with your doctor before going on high dose ibuprofen therapy.
 Kimura RE, Dy SA, Uhing MR, Beno DW, Jiyamapa VA, Lloyd-Still JD, “The effects of high dose ibuprofen and pancreatic enzymes on the intestine of the rat,” J Pediatr Gastroenterol Nutr 1999 Aug;29(2):178-83
 Lloyd-Still J, Kimura R, Beno D, Uhing M, Jiyanapa , “Ursodeoxycholic acid (Urso) prevents severe ibuprofen enteropathy,” Pediatr Pulm, Supplement 19, Sept 1999, abstract #487
(direct transfer of the normal CFTR gene with liposome or adenovirus vectors to the airway epithelial cells)
Gene therapy attempts to put a piece of functioning DNA into the nucleus of epithelial cells. Once in place, the DNA will make ‘wild type’ or normal ‘cftr’ or cystic fibrosis transmembrane conductance regulator proteins. The defective cftr continues to be made as well. Thus, gene therapy will work on all cf types equally well.
Amazingly enough, this new piece of DNA does not need to be incorporated into one of the 23 human chromosomes to function. It operates independently. Unfortunately, most cells do not duplicate themselves so the ability to make wild type cftr stops when the cell dies. It has been estimated that treatments would be required every three weeks or so.
The current thinking is that only a few percent of the epithelial cells lining the lung need be modified. Although this may be the case, the fact that cftr is expressed mainly in submucosal cells and only sparingly in epithelial cells suggests that treating epithelial cells may not be sufficient. Although much is known about cf, it is not understood how a defective chloride channel formed by cftr leads to chronic lung infection by those bacteria peculiar to cf, pseudomonas aeruginosa, staphylococcus aureous, and haemophilus influenza.
There are basically two methods being investigated for delivery of DNA to the nucleus. One method uses a ‘deactivated’ cold virus. Apparently a virus works by attaching to a cell wall and transferring genetic material into the nucleus of the host. The virus DNA uses the tools of the host to manufacture it’s own proteins required to replicate itself. When the host cell dies, copies of the virus are released. The viral method is very efficient, about half of those attached to the cell wall will successfully inject DNA into the nucleus.
The problem with a viral vector is that although the virus does not replicate itself to cause infection, some of the original viral protein is required to deliver the cftr DNA. Our immune system can have antibodies which respond to them. Each successive treatment may illicit an increasing immune response. This response may cause more damage than cf itself. This problem has been experienced in trials. In addition, there is a chance the virus may learn to replicate.
The second delivery method is to wrap the cftr DNA in a large liposome, or fat molecule. Liposomes enter the cell through the process of endocytosis, the cell wall absorbs them. The problem with this method is that only a small fraction of the DNA makes it’s way into the nucleus of the cell, about 1 in 1000 (but see the news report below).
Trials of both methods look encouraging. To date, they have demonstrated that a normal chloride channel can be produced in epithelial cells lining the nose. They measure this with the potential difference and ion current across the cell wall. To my knowledge, no one has demonstrated that gene therapy reduces the morbidity of cf, however, they are working very hard to do just that.
From: Gene Therapy Weekly via NewsPage :”Correction of Chloride Transport Defect in Cystic Fibrosis Airway Cells by Cationic Lipid-Mediated Gene Transfer.”
According to an abstract submitted by the authors to the 1995 International Conference of the American Thoracic Society, held May 20-24, 1995, in Seattle, Washington, “Cationic lipids offer several theoretical advantages over viral vectors for cystic fibrosis (CF) gene therapy.
However, cationic lipids described to date are relatively inefficient in mediating gene transfer to airway epithelial cells in vivo. To address this limitation, over 50 novel cationic lipids of diverse structures were synthesized and screened for their ability to mediate high efficiency gene delivery into these cells in vivo. Plasmid expression vectors containing appropriate regulatory elements for robust expression in human airway cells were also developed. Using an optimal formulation composed of the lipid #53 complexed with the vector pCMVHI-(beta) (harbors CMV promoter/enhancer, a hybrid intron and (beta)-galactosidase reporter gene) transfection of greater than 70% of an immortalized CF airway cell line (CFT-1) in vitro could be achieved. Transfection of confluent and polarized CFT-1 and Fischer rat thyroid (FRT, express low cAMP-stimulated current) cells with lipid #53 and a CFTR-encoding plasmid (pCF-1) restored cAMP-stimulated chloride current in these cells when assessed using the Ussing chamber assay. Short-circuit currents (Isc) of 2.7 +/- 0.4 and 10 +/- 1.8 FA/cm(2) for CFT-1 and FRT cells respectively, were obtained. Transfection of primary CF nasal epithelial cells corrected the cAMP-mediated Isc to 2.5 SLA/cm(2). To address the efficacy of these formulations in vivo, the lipid:DNA complexes (containing either pCMVHI-CAT or pCF-1) were also instilled intranasally into the lungs of BALB/c mice and Sprague-Dawley rats. Expression of up to 200 mg of CAT per mouse lung was achieved. Rat tracheal epithelial sheets that had been transfected with #53:pCF-1 exhibited a significant increase in cAMP-induced Isc over those from control animals. These results suggest that cationic lipid-mediated gene transfer may be clinically efficacious for the treatment of CF.”
AUTHORS: C. Jiang, J. Marshall, C. Siegel, J. Nietupski, R. Ziegler, M. Cherry, S. Fang, D. Harris, N.S. Yew, A.E. Smith and S.H. Cheng. Genzyme Corp. Framingham, Massachusetts.”
From: BIOWORLD 3/9/95: ORPHAN STATUS GRANTED TO TARGETED GENETICS’ AAV FOR CYSTIC FIBROSIS
Targeted Genetics Corp. was granted orphan drug status for any formulation of adeno-associated virus (AAV) vector for cystic fibrosis (CF).
The Seattle company received approval from the Recombinant DNA Advisory Committee of the National Institutes of Health for clinical testing of its gene therapy approach in September. The company filed an investigational new drug application in early January, and expects to start trials in May or June, said Alberta Garvin, the company’s manager, corporate communications.
Orphan drug status, given to diseases affecting fewer than 200,000 people in the U.S., allows seven-year exclusivity to the first sponsor of an orphan drug granted marketing approval.
Targeted Genetics’ trial will involve 16 patients with CF and mild lung disease at Johns Hopkins Children’s Center in Baltimore. They will be treated during an 18-day stay at the center, then followed one year.
Being granted orphan status “solidifies our position with regard to AAV as it relates to CF,” Garvin said. The company’s AAV vector is designed to deliver a normal copy of the CF transmembrane conductance regulator gene to lung cells. The Phase I trial is designed to assess the safety of the product and test its expression in the cells of the nose and lungs. – Jim Shrine
Description of a Gene Therapy Phase 1 trial
In the following account Massachusetts General Hospital in Boston, MA is abbreviated MGH. MGH/Genzyme Gene Therapy Phase 1 trial: proposal
History: 1981: first description of cellular defect (there is a problem with fluid flow across epithelial membranes, leaving the contents dehydrated) 1989: Gene discovered 1991: CFTR discovered 1991: Genzyme discovers that when CFTR gene is inserted in a cell in vitro (in a test tube in the lab), the defect was corrected, as shown by micro electrode work. recently: gene therapy work on in vitro nasal polyps and limited work on people.
Current ideas: 1) use gene therapy to add a CFTR producing gene 2) use pharmacologically active drugs to change the ion flux without altering the gene. (amiloride, UTP, ATP) 3) protein therapy; putting CFTR in? (long way off) CFTR is a protein. It regulates Na+ transport. It is messed up in CF and NA+ cannot leave the cell. There are *many* different CF mutations, and they muck things up at different levels. In the Delta F508 mutation, CFTR is made but does not reach the cell wall; in some other less severe forms some CFTR does get to the wall.
the Target: – The lung. (no other affected organs) – In vivo approach needed (can’t take the lungs out!) – there are 10ª10, 10ª11 (10 to 100 American billions) of epithelial cells – few cells replicate, which makes many DNA splicing techniques impossible. – However, it may be necessary to fix only 5 or 10% of the cells for effective Na+ channel regulation.
For gene therapy to work, we must insert a gene into the nucleus. The thing that does this is called a “vector.” MGH/Genzyme are looking at an adenovirus (common cold) vector.
adenovirus advantages: – cold virus been around for a long time. It’s really good at getting into cells, especially bronchial epithelial cells. – established medical history as a vaccines – efficiently infects non-reproducing cells – has demonstrably transferred genes – has a large insertion capability (or “cassette”) of 36,000 base pairs. This is necessary because the CF gene is big.
disadvantages: – possible inflammatory response from dose – denuded (no longer “cold inducing”) virus might still induce a response – patients may become immune to the treatment! – some low level of undenuded viruses could cause trouble difficulty getting to the important cells deeper in the lung and into the mucosal glands, which are below the surface. – virus may pose a threat to health care workers/public at large.
Can it work?
Past work: The literature provides a picture of a mouse lung in which a adenovirus vector had been used to insert a gene that dyed blue. This slide showed excellent dispersion of blue dye throughout the mouse’s tissue.
Second, the first Phase 1 nasal treatments (At Iowa) in 3 patients demonstrated that a CFTR gene was inserted into nasal cells and that it did correct the defect, as determined by micro electrode work. This was presented in 1993.
Others have shown that a small amount of adenovirus containing fluid can be safely inserted in the lung with a bronchoscope. 18 people have received some form of gene therapy in the US; there has been only one bad reaction to any gene therapy efforts so far. This person was sick for several days, but recovered.
The MGH Study:
MGH and Genzyme are applying for permission to begin a phase 1 (safety) gene therapy trial. They have currently received approval from the Harvard Gene Therapy oversight committee, and the NIH (?) Gene Therapy review board in Washington. Still pending is F.D.A. approval. This appears reasonably likely.
The KEY OBJECTIVE OF THIS STUDY IS TO DETERMINE THE SAFETY OF ADENOVIRUS GENE THERAPY! The MGH study will use both a bronchoscope (as used by NIH’s Ron Crystal) and an aerosol. As aerosols have not yet been used in any gene therapy study, extensive precautions are being taken to make sure that health care workers and the public are not exposed to any of the adenovirus vectors.
There will be 8 groups of 5 patients each. 3 of each group will get treatment via bronchoscopy, and 2 will be treated using aerosols. Each group will receive a different dose. The dose will be increased by a factor of 3.33 each time, for a range of 8 x 10ª6 vectors at the low end and 2.5 x 10ª10 at the high end. The latter is a hefty dose: it will be concentrated in a small area in the bronchoscope and therefore has about 42 vectors/ cell. Using the aerosol and the same dose, the concentration will be lower, about 6 vectors per cell. Treatments will be staggered so that if any adverse effects are seen, the study will be ended.
They are using a new nebulizer called the Miniheart, which has several nice features. It makes small drops (2 micron) which penetrate the lung better; it automatically stops nebulizing when the patient stops inhaling. The nebulizer has been modified with HEPA filters to remove any viral particles from the exhaled air. The whole thing is done in an enclosed tent. The whole tent is in a negative pressure room. This is to prevent any infection of other people.
MGH will probably have enough people in its own pool of patients for the study. – you must be 18 years of age – no cepacia patients, or patients with other atypical mycobacteria, can enroll in the study – you must demonstrate prior exposure to the cold virus in real life.
Once safety of this approach has been demonstrated, there is a possibility that a second efficacy (how well it works) trial will be initiated. It was emphasized that gene therapy will not reverse any lung damage, nor will it aid any of the other affected organs. It is of utmost importance that you keep you lungs as free of scar tissue as is possible to get the maximum effect from any future gene therapy work! So stick with those treatments!
The death of Jesse Gelsinger in September of 1999 is a reminder of the downside of gene therapy. Jesse was in a trial to correct a genetic defect using an adeno-virus as the vector. Jesse died of multiple organ failure from the infusions he was receiving. Gene therapy has been hampered by the lack of a stable and harmless vector. However, on the plus side, the year 2000 has seen the first cure of a genetic disease with gene therapy. SCID-X1 is a disease which virtually knocks out the a person’s immune system. If you remember the “bubble boy” of many years ago, this was the disease that he had. A French team, using a defective retrovirus-derived vector, transfected patients with the disease. Ten months later, two patients were, in effect, cured, with normal levels of immune system activity.  Gene therapy would work very effectively in cystic fibrosis, as the most common mutation is a point mutation — that is, only one amino acid is missing in the entire gene. Estimates are that if between 10-50% of cells were corrected in a CF person, the person would revert to carrier status in terms of their health outlook. We eagerly await the development of techniques similar to that used by the French team that can cure cystic fibrosis.
 Cavazzana-Calvo M, et al., “Gene therapy of human severe combined immunodeficiency (SCID)-X1 disease,” Science 2000 Apr 28;288(5466):669-72
Chimeraplasty is a form of genetic therapy, but is different enough from the types of genetic therapies described in the previous section that it warrants its own. Kimeragen, Inc. is the driving entity behind this therapeutic approach. Their website is http://www.kimerageninc.com We will quote from the first page of their technology review link on that website:
“Gene “Repair” Therapy
“DNA is a linear molecule composed of four kinds of nucleotides, and the sequence of these nucleotides, organized into units called genes, encodes the genetic information of an individual. Most human disease is based, at lest in part, on this genetic code. The DNA sequence of a gene is arranged in a precise order and deviations from this order, called mutations, often cause disease. In some cases, inherited mutated genes are directly responsible for the dysfunction (inherited disorders) while, in other cases, spontaneous mutations predispose an individual to disease resulting from the genetic change (acquired diseases). Hemophilia, muscular dystrophy, cystic fibrosis, and sickle cell disease are examples of inherited disorders, while many forms of cancer and heart disease fall in the latter category.
“A number of gene therapy methods have been developed, most using some form of viral delivery system, to introduce functional new genes, or gene segments, into cells in an attempt to treat disease. This traditional gene therapy approach attempts to restore gene function by the addition of the correct gene or gene fragment, without the removal or correction of the resident mutant gene. Traditional gene therapy ha not shown clinical and commercial viability to date.
“Kimeragen’s technology is not gene therapy in the classic sense described above. In fact, the fundamental scientific approach is very different from all other gene therapy strategies. Kimeragen’s technology repairs the mutated DNA in its natural position in the gene, restoring normal gene function. This procedure effects a precise, site specific genetic correction while the rest of the genome is left unaltered. The repaired gene is then regulated and expressed by the normal genetic and cellular mechanisms.
“A novel, synthetic molecule, the chimeraplast, was developed by Kimeragen to harness the cell’s normal DNA repair system to change a targeted nucleotide or nucleotides within a gene. The chimeraplast, an oligonucleotide composed of both DNA and RNA, is designed to specifically bind to the target DNA sequence and create a mismatched base-pair. This mismatched base-pair acts as a signal to attract the cell’s repair enzymes to the site and replace, insert, or delete the designated nucleotide(s) within the gene. Once the process is completed, the new genetic information is expressed by the normal cellular mechanism. This technique, chimeraplasty, can be used to repair mutant genes, alter genes, or interrupt normal gene function. The activity of chimeraplasts is not restricted by cell cycle, expression, transcription or other known cellular or genetic activities. The data demonstrate that chimeraplasty is technically feasible, efficient, and commerically applicable within plant, animal, human and bacterial cells.”
The chief scientific officer of Kimeragen is Dr. R. Michael Blaese, formerly chief of clinical gene therapy at the National Human Genome Research Institute of the National Institutes of Health. We were told that Dr. Eric Alton of the United Kingdom has been approached by Kimeragen to develop chimeraplasty techniques for use with cystic fibrosis patients.
Kimeragen has seen some success of its therapies. For example, they were able to induce correction of hemophilia B in dogs. The most striking success has been to cause the skin cells of albino mice to begin to secrete melanin, and thus color the skin of those mice. ( http://www.jeffersonhealth.org/news/1998/112998.html ) Kimeragen is also doing trials on a genetic liver disease which strikes Amish children.
The exciting thing about chimeraplasty is that it offers a vision of a one-time cure. If close to 50% of the body’s cells could express functional cftr, cystic fibrosis persons would attain the health status of cystic fibrosis carriers. The chimeraplasts are delivered via IV over a several day period. But after that period, some proportion of your genes are actually repaired and will function normally. And when those cells reproduce, they will reproduce with the repaired gene. Just imagine!
In cystic fibrosis, the ion transport system is abnormal. Since chloride (Cl-) cannot be effluxed easily, it attracts sodium (Na+). Defective CFTR leads to defective ENaC function; translated, this means that since the chloride channel (CFTR) is virtually absent, the sodium channel (ENaC) becomes uninhibited and allows for increased Na+ absorption by the cell. Water follows sodium and chloride, so fluid absorption in CF cells is higher, leading to a dehydration of extracellular mucus in CF as well as copious and salty sweat.
But what if you could block the sodium channel, and try to stimulate Cl- efflux from other Cl- channels besides the CFTR? This is the approach taken by those who advocate the use of amiloride and UTP (uridine triphosphate). Amiloride blocks the sodium channel, and UTP stimulates Cl- secretion from a Ca2+ chloride channel. In vitro, this has looked like a very promising approach.
However, in vivo, the situation appears more complicated. A French team recently completed a multicenter randomized double-blind placebo-controlled trial of nebulized amiloride in cystic fibrosis patients.  Their trial included 137 patients, and these folks nebulized three times a day for six months. At the end of the trial, the French team assessed whether there was any significant change in clinical parameters such as PFT scores, bacterial counts, cough, nutrition, need for antibiotics, etc. They found no benefit to amiloride whatsoever. However, it does not appear that the study included the UTP element. This may be because UTP’s effects are very transient, and they might not have been able to find a form of UTP that would last long enough so you wouldn’t have to be continuously taking it.
So, at the moment, even though the approach seemed very promising, it does not yet seem to be either feasible or effective. Perhaps if a long-lasting form of UTP were found, the two substances together might make a difference.
 Pons G et al., “French multicenter randomized double-blind placebo-controlled trial on nebulized amiloride in cystic fibrosis patients,” Pediatr Pulmonol 2000 Jul; 30(1):25-31
Vaccination Against Pseudomonas aeruginosa
There are at least three research teams investigating the possibility of vaccinating CF persons against Pseudomonas aeruginosa (PA). This would be an important step forward, as 68% of CF persons are colonized with PA by age 17, and when PA develops a biofilm around it (“mucoid” PA), it is very difficult to eradicate and may serioulsy compromise one’s health.
The first team of researchers is located in Uppsala, Sweden.  This team uses eggs that contain antibodies to PA. This is done by injecting hens with a killed PA solution. After first receiving intensive antibiotic therapy, patients are asked to gargle with the yolks of these special eggs every night after brushing their teeth. The results are very impressive. Fourteen patients have been studies over a period of five years, and not one has become chronically colonized with PA. When on rare occasion a positive culture is noted, aggressive antibiotic therapy is used, with the result that persistent colonization does not occur. Fourteen patients not using this therapy were also tracked over 27months as a control. During that time period, 23% of the sputum cultures for the group not gargling with the eggs was reported. The percentage for those who were gargling with the eggs was 2%. One benefit of this approach is that these antibody-rich eggs do not seem to stimulate the human immune system, which might result in harmful inflammation.
The second group is working in Bern, Switzerland.  They use a “polyvalent polysaccharides-toxin A conjugate vaccine” against PA. Patients are immunized yearly in this fashion. After 9 years of observation, only 28% of these vaccinated patients became colonized with PA, compared with 72% of patients in an unimmunized control group. Only local and mild adverse reactions were noted.
The third group is working at Cornell University.  This team incubates bone marrow-derived dendritic cells with PA, which led to uptake of the PA by these cells. Mice were then immunized with these cells by injection. When the lungs of these mice were challenged with PA infection, they survived longer than unimmunized mice. The unimmunized mice died within 72 hours, but almost 50% of the immunized mice survived for at least 2 weeks.
It looks fairly promising, then, that a relatively effective vaccine for PA might be on the horizon within the next several years. Work in Europe appears to be far advanced over work in the USA, however.
 HO Kollberg et al., CF Centre, Uppsala, Sweden, “More than Five Years Without Any New Chronic P. Aeruginosa Infections at Uppsala CF Centre,” poster presented at the 13th North American Cystic Fibrosis Conference, Seattle, Washington, October 1999
 AB Lang et al., University of Bern, Bern, Switzerland, “Longterm protection against Pseudomonas aeruginosa (PA) infections in patients with cystic fibrosis (CF): Results of a 9-year vaccine study,” poster presented at the 23rd European Cystic Fibrosis Conference, The Hague, Netherlands, June 1999
 S Worgall et al., Weill Medical College of Cornell University, New York, New York, “Dendritic cells pulsed with Pseudomonas protect against lethal pulmonary Pseudomonas infection in mice,” poster presented at the 13th North American Cystic Fibrosis Conference, Seattle, Washington, October 1999
Dextrans and Other Mucolytics
New Mucolytics Under Investigation
CF persons have more viscous mucus than normal persons. Though the precise cause of this is still under dispute, all agree that the higher viscosity is harmful. The thicker mucus prevents the cilia of the lung from beating normally, and results in reduced clearance of mucus. Stagnant mucus provides fertile breeding grounds for bacteria and fungi. Plugs of mucus may block off alveoli of the lung, resulting in decreased oxygenation. Therefore, it is seen as desirable to thin the mucus of CF persons.
Standard mucolytic agents used with CF persons include Mucomyst and Pulmozyme or Dnase. The former works by breaking apart disulfide bonds of the solids in the mucus; the latter works by enzyme action to break apart the DNA of the dead cells within the mucus. Some research reports an increase in elastase from the use of Pulmozyme/DNase, which is not good – but these reports are under dispute.
However, new mucolytic agents are under investigation. Dextran or dextran sulfate are being researched for use in cystic fibrosis. Significant decreases in mucus viscoelasticity have been reported from its use in experiments with dogs.  A variant of n-acetylcysteine (NAC), which is the primary component of Mucomyst is also under investigation. It has been named Nacystelyn (or NAL). Researchers view it as a drug that would be used in concert with Pulmozyme/DNase, as not only is it a mucolytic, but also acts as an anti-protease, which would offset the possible elastase-releasing effect of Pulmozyme/DNase.  Another therapy with mucolytic potential is that of inhaled or ingested glutathione (GSH). Indeed, NAC is a precursor of GSH, so it is possible that Mucomyst and NAL work by increasing GSH levels in the lung. GSH is a potent mucolytic, as it cleaves disulfide bonds readily. 
In sum, in addition to current mucolytic agents, there are several other promising candidates on the horizon.
 Sudo E., et al., “Effect of Dextran Sulfate on Tracheal Mucociliary Velocity in Dogs,” Pediatric Pulmonology, Supplement 19, September 1999, abstract #262
 Malfroot, A. et al., “Randomized Multicentric Double Blind Study of Tolerability and Efficacy of a DPI Nacystelyn Versus Placebo in Cystic Fibrosis Patients Treated by RhDNase for at Least Three Months,” Pediatric Pulmonolgy, Supplement 19, September 1999, abstract #281
 See Glutathione section under this heading.
Synercid, compassionate use antibiotic
For an article about this, see Appendix L, Longer Articles.
Gentamicin and Stop Mutations
Gentamicin is an aminoglycoside antibiotic. One of the properties of this type of antibiotic (if and only if it has a 2′ amino group) is that it increases the frequency of erroneous insertion of nonsense sequences as DNA is being “read” during protein production. Now this might seem like a bad thing. However, for CF patients who have “stop mutations,” this is actually a very good thing. By inserting this nonsense, gentamicin actually suppresses the stop mutation! The nonsense allows the cell to finish reading the DNA instructions for making the CFTR protein.
Testing of this therapeutic approach is in its infancy. However, a research study undertaken by the CF centers of Israel (where stop mutations are abundant) showed that application of gentamicin drops three times a day to the nasal epithelium of CF patients greatly increased chloride transport. Indeed, in 7 of the 9 patients studied, normal chloride transport was achieved. In 2 patients, normal NPD (nasal potential difference) scores were obtained. This demonstrates that stop mutations would – if the “stop” could be suppressed – produce normal, functional CFTR. 
Of course, one would hope this approach would work not only for the nasal epithelium, but for the entire body of a CF person with stop mutations. If this were the case, these individuals would, in effect, be “cured” of cystic fibrosis. Even though their genetic code would still be mutated, the mutation would be overcome by the use of gentamicin.  We have heard of a few doctors, primarily in Europe, that are already prescribing daily doses of gentamicin for their CF patients with stop mutations. However, no formal study of the effects of such a maintenance therapy have yet been published, so the jury is still out. Nevertheless, this looks like a very promising approach!
 E. Kerem et al., “Topical gentamicin rectifies abnormal nasal PD measurements in CF patients carrying stop mutations,” The Netherlands Journal of Medicine, June 1999, Vol. 54, Supplement, p. S84
 K. Keeling et al., “Structural Features of Aminoglycosides that Mediate the Suppression of Premature Stop Mutations Within Disease Models,” Pediatric Pulmonolgy, Supplement 19, September 1999, p, 239
CPX is an alkylxanthine currently in Phase II trials. In vitro experiments show that “CPX causes increases in expressed delF508-CFTR protein in the cells, in its level of glycosolation, in its cellular distribution, and in the chloride channel function of the properly trafficked delF508-CFTR.”  It appears that CPX binds to the mutated first nucelotide binding fold of the delF508 CFTR protein. In some fashion, this binding allows more delF508-CFTR to make it to the cell membrane. At the membrane, CPX also acts to increase the open time of the channel created by the CFTR protein.
In Phase I trials, delF508 CF patients were given up to 1000 mg of CPX to test for the safety of the substance. No adverse effects were noted. Furthermore, it was found that a dose of about 300 mg of CPX raised plasma levels of CPX to the point where the in vitro studies had found the desired changes to be occurring. However, only after the Phase II trials have been completed will we know if CPX actually improves the health of CF patients. There was no indication of health improvements having occurred during the Phase I trials, but remember that the purpose of Phase I trials is merely to test for the safety of the substance being used. The principal investigator on the CPX project is Dr. H.B. Pollard of the USU School of Medicine in Bethesda, Maryland.
 Zhang J et al., “CPX Affects Expression and Trafficking of DelF508-CFTR,” Pediatric Pulmonology, Supplement 19, September 1999, abstract #65.
CF Airway Epithelia Fail to Kill Bacteria
Because of Abnormal Airway Surface Fluid
Jeffrey J. Smith,* Sue M. Travis,+ E. Peter Greenberg,$ and Michael J. Welsh # *Department of Pediatrics +Department of Internal Medicine $Department of Microbiology Department of Physiology and Biophysics #Howard Hughes Medical Institute University of Iowa College of Medicine Iowa City, Iowa 52242
Despite an increased understanding of the cellular and molecular biology of the CFTR Cl~ channel, it is not known how defective Cl~ transport across airway epithelia causes chronic bacterial infections in cystic fibrosis (CF) airways. Here, we show that common CF pathogens were killed when added to the apical surface of normal airway epithelia. In contrast, these bacteria multiplied on CF epithelia. We found that bactericidal activity was present in airway surface fluid of both normal and CF epithelia. However, because bacterial killing required a low NaCl concentration and because CF surface fluid has a high NaCl concentration, CF epithelia failed to kill bacteria. This defect was corrected by reducing the NaCl concentration on CF epithelia. These data explain how the loss of CFTR Cl~ channels may lead to lung disease and suggest new approaches to therapy.
Cell, Vol. 85, 229-236, April 19, 1996, Copyright 1996 by Cell Press.
Inhaling Buffered L-arginine
L-arginine is one of the precursors of nitric oxide in the body. Nitric oxide, or NO, plays several important functions. Among other things, it is a bactericidal agent, and it is also a smooth muscle relaxant. Levels of exhaled NO have been found to be normal in CF patients. This is actually worrisome, because in many pulmonary diseases, exhaled NO is elevated – a sign that the body is defending itself properly.
One member of cystic-l, Richard Andrew Young, used inhaled L-arginine for a while. He had found an article by Dr. Clive Solomons on the potential of that therapy for cystic fibrosis patients. The first several times he used it, he got quite remarkable results. Over time, the results were not quite as striking, and eventually he ceased that therapy. Richard died in May of 1998. Nevertheless, there is still some interest in using inhaled L-arginine for cystic fibrosis. In addition to a group of doctors in Italy, Dr. Dawn Ericson at Harvard was interested in doing a study. Dr. Clive Solomons himself is still very interested in the therapy, and he can be contacted at [email protected]
What follows is Richard Young’s account of his use of inhaled L-arginine. Please note that Richard had to use a special form of the substance, available only from a chemical company. It is not possible to simply inhale the type of L-arginine available at a health food store!
While attending Brigham Young University in October, 1995 I found quite by accident an article in the February, 1971 issue of Pediatrics by Dr. Clive Solomons detailing a small scale study he had done using inhaled l-arginine in patients with cystic fibrosis. The article is: Solomons, Clive C., PhD, et al. “The Use of Buffered L-Arginine in the Treatment of Cystic Fibrosis.” _Pediatrics_ Vol. 47, no. 2. (Feb., 1971). Pages 384-391.
I began inhaling l-arginine at the end of March, 1996. I had an appointment at the CF clinic five days later, and my FVC was up about 10% and my FEV1 up slightly less. The most pronounced effect was much better oxygen saturation: 98% at room air, despite FVC of 49% and FEV1 of about 32%. After five weeks of inhaling l-arginine, my FVC had increased 28% to 63% and my FEV1 had increased 16% to 41%. For the previous 18 months, my FVC had been under 50% and my FEV1 under 30%. For the previous six months my FVC had been under 35%, my FEV1 under 25%. My small airway capacity had increased from 5% to 14%.
At the time I began inhaling l-arginine, I was on oxygen, was short of breath from walking at a normal pace any distance, had dropped out of school, was being evaluated for a lung transplant, and was unable to work. Within a couple of weeks, I was reenrolled in school, working eight hours a day on my feet and lifting, and no longer eligible for a lung transplant. I had much better exercise tolerance and increased lung functions gave me a better appetite, and made it much easier for me to do my secretion clearance treatments.
To make the solution, I dissolved 0.9g of l-arginine free-base and 15g of l-arginine hydrochloride in 300ml of purified water. (Solomons used tap water.) This mixture should have a pH of 7.4-7.6 and should be refrigerated. In Solomon’s study, it was inhaled four times a day for thirty minutes (I don’t do it quite as much or in such even doses).
100g of l-arginine hydrochloride lasts me eight to twelve weeks, and 100g of l-arginine free base should last over two to three years. The chemicals can be purchased from Advanced Scientific, in Florida, at (800) 524-2436. It costs $95 for 100g bottles of both, plus shipping. 1000g of l-arginine hydrochloride (enough for 2 years) costs only two or three times more than 100g. The chemicals also be obtained from Sigma Chemical. They supplied the chemicals for Dr. Solomon’s study. Their phone number (314) 771-5765 or (713) 256-1616. However, they will not sell to individuals, but will sell to institutions, medical researchers and laboratory supply companies. They also charged much less (as I remember). You may be able to order it from Sigma by asking a chemical supply shop to special-order it from them for you.
Sigma Chemical, which is still in business carried L-arginine hydrochloride (at least, that’s what they said on the phone) while I could only get arginine hydrochloride (no “L”) from Advance Scientific. I don’t know how important this difference is, though I do know that left-polarized amino-acids are more biologically efficient, though I don’t know if this applies to it’s mucolytic action (and, even when undifferentiated, I would imagine much of what I use is left-polarized, too).
It may not be wise to use the l-arginine found health food stores since it is almost always l-arginine free base (which is very caustic) derived from l-arginine hydrochloride.
16-24 hours after I began inhaling it I became extremely congested, more congested than when I’m sick, and was coughing up the most vile looking, foulest smelling mucus. Even when I’ve been sick, it doesn’t look that gross! And “coughing up” is an overstatement: mostly, it just slid out of it’s own accord. This continued for a couple of days. My breath smelled REALLY bad, too, but this also went away.
After inhaling it for a long time, I am sometimes short of breath for 15-30 minutes afterward, but have found that adding 0.5cc of Ventolin alleviates this(though I rarely do it).
The article says that L-arginine has a detergent effect and loosens mucus because of it’s calcium and metal-ion binding effects and by reducing fibril aggregation. The article also reported that staph. aureus and hemophilus disappeared from the cultures of those who inhaled l-arginine, although it did not affect the presence of pseudomonas. The authors expected that this treatment would work best as a preventative measure in patients who had not suffered much lung damage. Despite this, the patients who used it averaged a 10% increase in lung functions, as the l-arginine liquefied mucus plugs, although their lung functions stopped increasing after about three weeks. The most pronounced effect was increased arterial Po2.
The article also suggests oral use of L-arginine, but their suggested dose was 1g/kg/day, up to 25g, which is a lot of L-arginine. Oral use resulted in weight gain, better fat absorption, and 100% success in relieving ALL intestinal cramping. Since this study was done, drugs treating absorption problems in CF have become more effective and prevalent. The study also noted that, to achieve it’s preventative effects, it might be sufficient to take l-arginine orally, as a small amount ends up in the bronchial fluids.
Most CF doctor’s haven’t had a problem with using l-arginine (although most didn’t think it would help much, either). L-arginine has no side-effects, as was confirmed in a letter to Pediatrics (I can’t remember the date) by Dr. Solomons two or three years after he published his study.
What happened to L-arginine? Apparently this treatment was studied at the time but “lost” in the excitement over a related drug with similar effects, N-acetylcysteine (Mucomyst). (Which, incidently, has serious side-effects and is now rarely used in CF.)
A Dr. reading the Cystic-L mailing list found an article in Pediatrics 1975, vol. 55, p. 96 by an East German group which compared l-arginine with N-acetyl cysteine and found it inferior. They did not recommend it’s use. Solomons wrote a rebuttal in Pediatrics, 1976, vol. 76, p. 166 where he criticized the methodology of the East German study, arguing that the researchers should have dissolved the l-arginine in water, instead of sodium hydroxide, and that this could actually cause lung inflammation and that arginine needed to buffered by its own salt. The doctor who dug up this information concludes, “Since then I can find no reports either in the mainstream literature or main CF conference meetings.”
Before trying this treatment, I would urge you to get a copy of the original article. Any library should have it, or your doctor can get it for you. Also, in light of the above information, l-arginine should probably not be mixed with saline (which is what my own CF doctor did when he researched it himself — which produced marginal results). Be careful to be sure to mix it in the proper proportions, as inhaling it at the wrong pH could pose a problem. If you don’t have access to a scale, pocket-sized scales that can measure to 0.1g can be purchased for around $100 from scale or laboratory supply shops.
Update, May 1997
I still continue to use it. I took a month off in March, because I had some inflammation problems after using high-dose tobra, and was advised to stop inhaling anything but Ventolin for a month or so After being in the hospital at least once a year, I have not been back since I started l-arginine in 03-96. This is the first winter I have gone through in Utah without ending up in the hospital, though I did have to do home antibiotics.
Before starting l-arginine, my FEV1 had been below 40% for fourteen months, and around 20% for several months before. I was told this was irreversible, permanent damage, which was also the conclusion arrived when I was given a CAT scan for a tx evaluation. When I started the l-arginine, I had dropped out of school and was not working because I was too sick. Within two days I could walk long distances without being short of breath; within a week I was working; I switched to a job that involved standing all day and lifting; I then resumed school, able to climb the necessary hills and do well in the 6000ft altitudes the next fall.
Recent studies indicate that l-arginine may be useful in combatting inflammation and secondary pulmonary hypertension, which results from chronic lung disease, further reducing lung damage. Currently, a group in Italy is researching the subject on CF patients.
Richard Young, email: [email protected]
Colchicine and Chemo Agents
There was an interesting CF case in France, where a CF patient undergoing chemotherapy for cancer experienced a “remission” in his CF. The patient died several years later from the cancer, but his CF remained in abeyance. The French team monitoring his case wondered if the chemotherapy drugs had induced his body to express MRP, a channel that is significantly redundant with the CFTR channel. The MRP channel might be taking the place of his CFTR channel, providing for this remarkable remission. Toxins induce the production of MRP as a kind of “code blue” response to try and flush the toxin from cells.
Because chemotherapy drugs are so toxic, the French team decided to try colchicine, a mild toxin used as an anti-inflammatory agent to treat gout. They discovered that colchicine could induce some MRP. So they did a 6 month trial of colchicine therapy on CF patients. They received 1 mg of colchicine a day. FEV1 scores improved in 6 of 7 patients, and one patient who was on the lung transplant list, was taken off that list. Weight gain and reduced need for antibiotics was also seen. The drug was well tolerated, with mild diarrhea in 2 patients. We hope that further trials will be done to see if altering the dose makes the therapy even more effective.
Sermet-Gaudelas I, et al., “Interest of colchicine for the treatment of cystic fibrosis patients: Preliminary report,” Meds Inflamm 1999;8:13-15
Sodium 4-phenylbutyrate, also known as 4BPA, is used as a drug to treat conditions such as urea cycle disorder. Its trade name is Buphenyl. Recently, researchers have discovered that 4BPA might be useful in treating cystic fibrosis, as well. However, it is currently under investigation as a treatment for those who have the delF508 mutation only.
The delF508 mutation causes the CFTR protein to be misfolded. The cell somehow recognizes that the protein is misfolded, and sends a chemical “chaperone” to prevent the mutated protein from being trafficked to the cell membrane. Research has shown that Hsc70, a member of the heat shock protein family, is the (or at least one of the) suspected chaperones.
4BPA decreases the production of Hsc70. Researchers have shown that Hsc70 decreases in a dose dependent manner with the administration of 4BPA. Administration of 4BPA has also been shown to be associated with restored CFTR function in certain CF cell lines. It is hypothesized that the 4BPA is decreasing the Hsc70, which decrease then allows for more of the delF508 mutated protein to reach the cell membrane. Although the delF508 protein creates a channel that is not fully functional (its open time, stability, and survival time are shorter than normal), it is at least partially functional, which presumably accounts for the restored CFTR channel function noted in 4BPA experiments with CF cell lines. [1, 2]
However, there are two caveats. One is that research has shown that too high doses of 4BPA arrest cell growth and lead to accelerated cell death.  The other is that after a period of several days, cells appear to respond to the depression of Hsc70 production by overproducing Hsc70, leading to a type of boomerang effect. It may be necessary to take the drug for several days, and then cease taking the drug for several days, and so on.
Researchers at Johns Hopkins University in Maryland are currently spearheading the efforts to test 4BPA in cystic fibrosis. Having passed Phase I trials (demonstrating safety of use), the research team is now in Phase II trials (to determine the most effective dosage.)
 Rubinstein, R.C. “Regulation of HSC70 Expression by Sodium 4-Phenylbutyrate,” Pediatric Pulmonology, Supplement 19, September 1999, abstract 104.
 Choo-Kang, L.R., “Butyrate-Mediated Upregulation of DelF508 Expression in CF Airway Epithelial Cells,” Pediatric Pulmonology, Supplement 19, September 1999, abstract 271.
 McGrath, S.A., “The Effect of Sodium Phenylbutyrate on Cell Growth and Cell Cycle Progression on IB3-1 Cells,” Pediatric Pulmonology, Supplement 19, September 1999, abstract 272.
A Layman’s Explanation of GSH Augmentation Therapy for Cystic Fibrosis Patients
Valerie M. Hudson, Ph.D., Brigham Young University; revised version, 8 April 1999
GSH is the abbreviation for reduced glutathione. GSH is a very important substance in your body’s defenses. It is produced inside of cells, where it plays a role in neutralizing harmful substances. GSH is also exported from cells, and once outside of the cells in the extracellular environment, it plays several important roles as well. We will talk more about exactly what GSH does in a moment.
Normally, the cells make enough GSH for themselves, and also export GSH to outside of the cells. Some areas of the body need more extracellular GSH than others. One such place is the lungs. Your lungs import GSH from the rest of the body in order to have enough GSH to meet its needs.
However, a strange situation exists in a CF person’s body. To understand this situation, you must first understand that GSH is an anion, or a negatively charged substance. Cells have special anion transporters or channels that allow anions like GSH to be exported from the cells. The usual transporter in most cells is the channel created by the CFTR protein. This channel is missing or defective in CF persons. There is also a channel created by the MRP protein. It does everything the CFTR channel does, but unfortunately for CF persons, most cells do not normally express this channel. Only a few types of cells normally express the MRP channel, and these include liver cells, red blood cells, and also immune system cells.
So here is the strange situation: because CF persons have a missing or defective CFTR channel, most of their cells – the cells that do not express MRP – will not be able to export GSH very efficiently if at all. This leads, over time, to a severe deficit of GSH in the extracellular environment of a CF person. However, in the cells which do express MRP, GSH levels will be depleted, because those cells will be trying very hard to make up that big deficit outside of the cells. Got that? Regular cells will have high levels of GSH inside of them. MRP cells will have depressed levels of GSH inside of them. And the levels of GSH outside of the cells will be very low. This is the strange situation of a person with CF.
Let’s look at the effects of this strange situation on a CF person’s body:
1) Regular cells will have high levels of GSH inside of them.
No one yet knows what the consequences of this are. Stay tuned.
2) MRP-expressing cells will have depressed levels of GSH inside of them.
The MRP-expressing cells that are important to look at here are the immune system cells. The immune system consists of many different types of cells. These cells control the body’s defenses. The two main thrusts of body defenses (outside of the antibody system) are inflammation and neutralizing/killing harmful things, such as toxins or germs. Those two thrusts go hand in hand. When your body inflames, it is marshalling the killing forces and sending them to where the harmful things are. In the case of, say, a germ, the killing forces release oxidants and elastase, which pummel the germ in question. After pummeling it, the killing forces attempt to actually kill and clear away the germ from the body.
GSH is a very important part of all of this activity. The level of GSH inside of the immune system cells determines whether the cells go into inflammatory action or not. Low levels of GSH lead to inflammation. Normal levels of GSH switch off inflammation.
Furthermore, the level of GSH in the killer cells themselves determines whether the cells get the signal to “kill!” or not. Low levels of GSH in the killer cells lead to lack of killing action. Normal levels of GSH in the killing cells allow them to kill.
You can see the problem already, I’m sure. In CF, a person’s immune cells will be in a constant state of depleted GSH. The consequences? First, chronic inflammation. Those immune cells will never switch off the inflammation mode, because (at least in CF adults) they never have normal levels of GSH inside of them. Chronic inflammation is a hallmark of CF.
Second, immunodeficiency. The killer cells will be very ineffective killers because they never get a clear signal to kill. Because they aren’t effective killers, germs will remain and the body will recruit more and more killers because the germs aren’t being killed. (A CF person has about 400 times the number of neutrophils (one type of killer cell) than normal persons.) But the greater number of killers doesn’t do much good, because they can’t kill very well. Furthermore, chronic stimulation of these killer cells because of chronic inflammation leads the killer cells to “poop out.” This is another reason why the needed killing doesn’t take place. The germs get a big break. Colonization of the lung with germs and other nasties is another hallmark of CF.
The end result? A CF person is in a chronic state of inflammation — even over-inflammation — but cannot get rid of germs and other nasty organisms because of immunodeficiency. Bad news all around.
3) Over time, a severe extracellular deficit of GSH develops.
A mentioned above, GSH not only plays an important part inside of cells, it plays an important part outside of cells as well, in the extracellular environment. However, CF adults have only about 10% of normal GSH levels in the extracellular environment of their lungs. So what happens if, like in CF, a person over time develops a severe shortage of GSH in the extracellular environment?
One important consequence is that your mucus becomes thicker. GSH in the extracellular environment is used by the body to break down mucus and make it thinner. A severe shortage of GSH outside the cells will prevent that desired thinning action from taking place. Second, your lung surface will begin to lose its protective coating, called surfactant. Normal levels of GSH are needed for adequate surfactant in the lungs.
The third cluster of effects relate to the antioxidant capabilities of GSH. There are numerous antioxidants used by the body, but the main small molecule antioxidant is, in fact, GSH. In the case of CF, a deficit of GSH means a breakdown of antioxidant function. What does this mean?
Well, first, any oxidants you breathe in (like pollution) will be less likely to be neutralized.
But perhaps even more important than the oxidants you breathe in are the oxidants you produce yourself. Remember those immune system cells? One of the primary means of defense is polymorphonuclear neutrophils (PMN, or neutrophils for short). These neutrophils are killers, sent out to destroy bacteria and other nasties that get into your lungs. Now, the weapons the neutrophils use, and the debris left over from killing the nasties, are themselves harmful things. To simplify, the neutrophils cause the production of oxidants and the neutrophils also produce elastase. Oxidants can directly damage cells. Elastase eats elastin — the connective tissue of the body. The task for the body is to keep these killers around, while simultaneously preventing the oxidants and the elastase produced by their activities from damaging the body itself.
So, the body has a couple of systems to prevent that from happening. One is called the antioxidant system. This system finds and neutralizes oxidants so that they cannot attack your own cells. The second system is called the antiprotease system. Antiproteases tackle elastase and prevent it from eating your own connective tissue.
So in the normal body, everything is fine. The neutrophils kill nasties, and the antioxidant system cleans up the oxidants produced, and the antiproteases tackle the elastase produced and so the body is protected from the killing activity of the neutrophils.
So what happens when one of the body’s main antioxidants is in seriously short supply extracellularly? One effect of this is easy to see — oxidants produced by neutrophils will not be neutralized and these oxidants can then damage your cells directly. Ouch.
But there’s more. Oxidants inactivate antiproteases. The increased amount of un-neutralized oxidants attack the antiproteases, causing them to be less effective. Less elastase is neutralized by the body. So this un-neutralized elastase attacks the connective tissues of the body. Double ouch.
But there’s even more. Oxidants also damage the neutrophils themselves. For this reason, as well as the reasons outlined in the previous section, the neutrophils lose their killing ability to a significant degree. This results in two things: more nasties can stay in the lungs and not get killed, but ALSO the body makes a lot more neutrophils to overcome their lack of killing power. In CF, your body will make almost 400 times the normal amount of neutrophils to compensate for this. Triple and quadruple ouch.
Now, these almost 400 times more neutrophils are going to produce much more elastase and oxidants. Much more. And because of the messed up antioxidant system, this means that there will be even more killing of your body’s cells and connective tissue. More and more and more. It is a cascade — a vicious negative cycle that gets worse and worse. Ouch to the tenth power!
How do your lungs die in CF? Yes, in part from the harm done by the bacteria and other nasties which can now take up residence in your lungs. But ALSO because your body’s attempt to kill these nasties is in fact killing your own body. We call that “auto-immune destruction.” Indeed, scientists are beginning to think of CF as a disease with a strong autoimmune destruction component. So even if you are not colonized by any bacteria, your lung function will degenerate every day that you live. Every single day.
What to Do?
The therapeutic approach described is actually pretty simple. If you can’t get GSH out of the cells because of the genetic defect, why not just take more into your body, whether by inhalation, oral supplementation, or even IV? In other diseases, this has worked effectively to restore normal levels of GSH. Restoring that GSH will prevent the oxidant damage, prevent the elastase damage (by rescuing the antiproteases), and restore the killing capacity of the neutrophils — which will in turn cause the body to produce less (that is, more normal levels of) neutrophils. The inflammation switch is turned to “off.” Killer cells get the signal to “kill.” Your mucus is thinner, as it should be. Lung protective coating is restored. Your body stops the bacteria and your body stops killing itself and starts effectively protecting itself.
(It may also be desirable at the beginning of the treatment period to add something called a cysteine donor, such as Mucomyst (N-acetyl cysteine), as well, in order to help the MRP-expressing cells to up their level of GSH inside of themselves quicker. But the backbone of the approach remains the addition of GSH to the body directly.)
The only study we could find of CF and GSH was an in vitro study (a study done in cell plates). When they added GSH to CF sputum, an indicator of oxidant presence dropped by over 90%. An indicator of amount of neutrophils dropped by almost 50%. More recently, in July 1999, the report of an in vivo trial using a small number of CF patients was made by the same team. They found similar decreases in oxidant stress. A summary of their findings can be found at http://members.tripod.com/uvicf/gsh/gsh_abstract.htm
An old, unglamorous, unpatentable therapy — the addition of GSH — may in fact be extremely effective. A longer scientific essay on this therapeutic approach can be found on the UVICF website at http://members.tripod.com/uvicf/gsh/gshprospects.htm Sometimes old, unglamorous unpatentable therapies are overlooked by the scientific and pharmaceutical worlds. But they should not be overlooked. That was the purpose in writing the essay on the scientific rationale of this approach. Give it to your doctor and have a serious discussion about it.
For more information click here http://members.tripod.com/uvicf/research/glutathione.htm Also please note that as of July, 2000 three trials (one of oral GSH and two of inhaled GSH), are scheduled to take place soon in Europe and in the US.
Small Molecular Weight Solvents, and also ThapsigarginResearchers have long noted that it is possible to get delF508 CFTR protein to the cell membrane, where it is somewhat functional. Incubating cells at about 72 degrees Fahrenheit will do it, and so will incubating the cells in a variety of small molecular weight solvents, such as glycerol, DMSO (dimethyl sulfoxide), TMAO (trimethylamine oxide), D2O, myo-inositol, betaine, taurine, and others. Since many of these solvents are relatively harmless, researchers are investigating whether they could play a role in clinical care.
A second interesting approach is to prevent the cell from holding back delF508 from the cell membrane. Indeed, this is the same approach as 4BPA and CPX, both of which are described in other sections. Thapsigargin depletes intracellular levels of Ca2+. By doing so, it makes it harder for the cell to “tag” the delF508 protein as defective. In one in vitro trial using THAPS, 34% of the defective protein made it to the membrane, and there was a 15% increase in chloride secretion.  It remains to be seen whether THAPS could become a useful therapy in CF, but keep your eye on it.
 Egan ME, et al., A Small Molecule Approach to Increasing delF508-CFTR Surface Expression in CF Epithelial Cells,” Pediatr Pulmonol, Supplement 19, Sept 1999, abstract #274
Azithromycin and Pseudomonas aeruginosa
By age 17, almost 70% of cystic fibrosis persons culture Pseudomonas aeruginosa (PA). Some individuals are not terribly affected by the acquisition of PA, but many others are. In particular, when PA develops a biofilm, or becomes “mucoid”, it is very difficult to treat. Even synergistic combinations of antibiotics may not do much in terms of stemming or eradicating this pathogen. One reason for this is that cystic fibrosis cells, for some reason, allow for very tight adhesion of PA. This tight adhesion allows for infiltration of the epithelial barrier.
One very interesting discovery has been that azithromycin, sometimes called Zithromax, appears to be unusually effective in boosting the lung function of those colonized with PA. For example, in a study by Adam Jaffe et al at the Royal Brompton Hospital in London, lung function scores increased by about 11% in young adult patients colonized with PA who were put on long term AZM therapy (daily AZM for 3-12 months).  However, it seems that this effectiveness is not due to outright killing of the bacteria by this macrolide antibiotic. There appears to be another mechanism at work. Various hypotheses have been put forward as to how AZM works in cystic fibrosis patients colonized with PA: the AZM might be reducing inflammation, it might be reducing the propensity of PA to become mucoid, it might decrease mucus hypersecretion in airways, and it might reduce the adherence of PA to epithelial tissue, it might stimulate the expression of an anion channel with functions redundant to the cftr.
Other studies are beginning to be performed that examine the use of long term AZM therapy for CF persons culturing PA. Most of these use a protocol of 250 mg of AZM every other day for months, even years. Some studies find improvement of PFT scores of up to 21%.  In one study performed in Germany, rates of adhesion of PA to cystic fibrosis cells was reduced to near normal range when long term AZM therapy was used.  Another study finds that CF sputum becomes less viscous when long term AZM is used.  This may lead to improvement in mucociliary clearance, which would be useful in the fight against PA.
 Jaffe, Adam, et. al., “Long-term azithromycin may improve lung function in children with cystic fibrosis,” Lancet 1998 Feb 7; 351(9100): 420
 Anstead, MI et al., “Effect of Chronic Azithromycin on Lung Function in Cystic Fibrosis,” abstract presented at the 13th annual North American Cystic Fibrosis conference, printed in Pediatric Pulmonology, Supplement 19, September 1999
 Fischer JJ, et al., “Azithromycin reduced epithelial adherence of P. aeruginosa in patients with cystic fibrosis,” abstract presented at the 13th annual North American Cystic Fibrosis conference, printed in Pediatric Pulmonology, Supplement 19, September 1999
 Tai, S et al., “Effect of azithromycin (AZM) treatment on sputum rheology in cystic fibrosis patients,” abstract presented at the 13th annual North American Cystic Fibrosis conference, printed in Pediatric Pulmonology, Supplement 19, September 1999
Most persons are familiar with heparin as a blood-thinning agent used to flush IV tubing. However, heparin also has other properties. It is an anti-inflammatory, inhibits histamine release, and thins sputum by interfering with its ionic charge. A group of researchers in Liverpool, England, decided to test whether this combination of properties might mean that heparin could be useful in the treatment of CF.  Their sample was 6 B. cepacia colonized stable adult CF patients (mean age 27.7 years, 4 male). To these 6 patients they gave a nebulized solution of 25,000 IU’s heparin sulphate diluted with 5 mls normal saline once a day for 7 days. There were no adverse effects, and sputum IL6 and IL8 (markers of inflammation) decreased somewhat. The patients reported that it was easier to expectorate their sputum, and visual inspection of the sputum found it to be somewhat thinner. However, spirometry of the sputum did not change, and neither did sputum volume. There was no change in B. cepacia colony counts. Along this same line, other researchers are looking into nebulized lidocaine.
 M.J. Ledson et al., “Nebulised Heparin Therapy in B. Cepacia Colonised Adult Cystic Fibrosis Patients,” Pediatric Pulmonology, Supplement 19, September 1999, abstract 522.
One school of thought in cystic fibrosis research believes that the salt concentration of the airway secretions of CF patients is higher than that of normal persons. It has been found that certain pathogen-killing substances of the body, called beta defensins, are unable to do their job at higher levels of salt concentration. Thus, it is theorized that CF persons would be less effective at killing bacteria in the lung, and this might help explain the typical colonization of the CF lung that begins a few years after birth.
If you accept this premise (and it is still hotly debated), then one would wish to find a replacement for these defensins – a replacement that would still function in a high salt environment. That’s where cathelicidin peptides come in. Cathelicidin peptides are created by the bodies of humans and animals. These cationic peptides seem especially helpful in attacking gram negative bacteria, which are the most troublesome to CF patients. Researchers have examined cathelicidin peptides from a variety of mammals, including sheep, mice, rats, rabbits, and humans. (From humans, they examined peptides found in saliva, which need to be able to work in that sometimes high salt environment.) Typically, the experiments are in vitro, where a wide variety of bacteria are placed in dishes with a higher-than-normal salt environment, and then incubated with each of the peptides to see to what degree the bacteria are destroyed. There have been a few in vivo animal experiments where the lungs of the animals are purposefully infected and then the aerosolized peptide is administered. Once again, the degree to which the bacteria have been destroyed is the benchmark. Some of the animals have been non-CF; in one experiment, the animals were CF-knockout mice.
In the preliminary work done so far, two cathelicidin peptides stands out as being very effective against a range of bacteria even in a high salt environment. One is the sheep peptide SMAP29. This is one of the very few peptides that are even somewhat effective against cepacia. SMAP29 is very effective against Pseudomonas a., even mucoid type. The other is IB-367, which has passed Phase I trials and is now in Phase II trials. The promise of this research agenda is that one day these peptides would be used as a prophylactic antibiotic aerosol. That is, even before a CF person’s lungs were colonized by bacteria, inhalation of this peptide on a regular basis would prevent colonization in the first place. Since the average life expectancy of a CF person that never colonizes Pseudomonas or cepacia is 51 (as versus 29 or 30 for those who do), this would represent a significant step forward in CF care.
 Travis, SM et al., “Cathelicidin-Derived Bacterial Peptides: Airway Expression and Antipseudomonal Activity,” Pediatric Pulmonology, Supplement 19, September 199, abstract 337
 Tran, LT et al., “P-113D, A Protease-Resistant Peptide Derivative of Histatins, Has Potent Activity Against CF Pathogens,” Pediatric Pulmonology, Supplement 19, September 199, abstract 349
 Saiman, L et al., “Mutliply Drug Resistant Organisms from CF Patients are Inhibitied by Cathelicidin Peptides,” Pediatric Pulmonology, Supplement 19, September 199, abstract 540
 Brogden, KA et al., “Efficacy of SMAP29 in an Ovine Model of Pulmonary Infection and Its Potential for Treating P. Aeruginosa Infection in Patients with Cystic Fibrosis,” Pediatric Pulmonology, Supplement 19, September 199, abstract 542
 Brogden, KA, “Antimicrobial Activities of the Cathelicidins SMAP29 and SMAP34 Against Ovine Respiratory Pathogens and Pseudomonas Aeruginosa,” Pediatric Pulmonology, Supplement 19, September 199, abstract 543
 Fujii, CA et al., “In Vitro Antimicrobial Activity of the Protegrin Analog IB-367 Against Cystic Fibrosis Relevant Bacterial Pathogens,” Pediatric Pulmonology, Supplement 19, September 199, abstract 539
 Davidson, DJ et al, “Contrasting Antibacterial Profiles of Mouse and Human Beta Defensins Peptides May Contribute to Species Specific Lung Phenotypes in Cystic Fibrosis,” Pediatric Pulmonology, Supplement 19, September 199, abstract 552
Malnutrition is a perpetual problem in cystic fibrosis, and is often complicated by lack of appetite in the patient. Such loss of appetite only hastens deterioration of health. In other diseases where loss of appetite is a problem, such as AIDS and cancer, Megace (megestrol acetate) has been used with considerable success in improving appetite. The question was raised as to whether Megace would improve the appetites of CF patients.
In one study in Ireland, 17 CF patients were prescribed 320 mg a day of Megace for a 3 month period. Of the patients who completed the study, all had a weight gain, ranging from 2.2 kg to 15.2 kg, with a mean gain of 6.3 kg. 
However, Megace is a synthetic progestin. As such, it does produce undesirable side effects in men, such as the growth of breasts. In the Irish study, 3 patients (presumably men) dropped out because they disliked the side effects they were experiencing.
In conclusion, Megace does appear to be effective in stimulating the appetites of CF persons. However, the side effects may be troubling for some, especially for men.
 J. Dowsett, “The use of megestrol acetate (Megace) in the treatment of cachexia associated with cystic fibrosis,” The Netherlands Journal of Medicine, June 1999, Vol. 54, Supplement, p. S67.