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Wednesday, December 30, 2009

Adventures in Hematology

Many of you know that I threw two blood clots in 2009 (I have a hunch I threw one in 2000 as well, but that was never checked). They were both associated with PICC lines.

Being the spit-fire that I am, I began a journey to figure out if these clots were simply just the standard PICC line clot, or if there was something else going on.

My first stop was a no brainer: to my OBGYN to get a non-estrogen birth control Mirena IUD (medication with estrogen can make you more likely to throw clots). I have been on estrogen birth control since 1995 to control loooooooooooooong periods and to help with hemoptysis (although it never really seemed to work. i was probably going to try seasonale, as less frequent periods has helped a few CFer's at my clinic with their frequency of hemoptysis).

My next stop was two different hematologists. Many of you know that I'm involved in healthcare not just personally with CF but also professionally. I have seen that it's simply essential to see specialists for the issues that I have - there is no possible way that a CF pulmonologist can be up to date on all the latest and greatest info for every part of our body (CF pulmo's are even up to date on CF related stuff). I'm healthcare reform's worst nightmare - I see an allergist for my allergies, and ENT for my sinuses and ears, a gastro for my reflux, an OBGYN for my lady parts, a pulmo of course for my CF (and many, at that - Denver, Minnesota, Stanford, Cincinnati), an opthamologist for my eyes, a dermo for my skin, and a dentist for my teeth. Only the best for my health - why not see an expert to ensure you have the latest and greatest for the body?

So I went to see the first Hematologist at the beginning of December who said simply my CF doc had me on too much Lovenox for too long to treat my DVT and she thought going off the estrogen BC pills was a good idea (and commented that CFer women can't really get pregnant anyway.... to which I corrected her that women with CF CAN get pregnant and besides, that's not why I'm on BC in the first place). Other than that - she recommended prophilactic Lovenox (low dose) next time I get a PICC to prevent a clot. She didn't want to draw blood and do tests because she said we'd treat my clot throwing propensity the same way regardless of the results.

Fare enough.

On to Hematologist #2 about 3 weeks later (at a totally different medical center across town). This doc was pretty convinced that stopping my estrogen BC pills would have a HUGE impact on my clot-throwing propensity and asked me why my pills weren't stopped back in January when I threw my first clot (to which I answered, I have no clue, it didn't even occur to me and of course my CF care team didn't even mention it... grrrrrrr.... and I only see my OBGYN once a year and once I did see him in Nov this was brought to my attention).

Hem #2 also wanted to draw a bunch of blood (seriously, I thought I was going to pass out it was the most blood that's ever been taken from me) to run a boat-load of tests. He said I could have some genetic blood clotting issues, and although we most likely wouldn't treat me any differently with a genetic issue, it would be good to alert the rest of my fam of this gene. Well since my enormous family hasn't even been tested for the CF gene, I have a feeling this wouldn't phase them either but what the hey, let's test!

Fast forward two weeks to yesterday and I get a call from Hem #2 with some interesting test results:

  1. Everything came back normal including the test for the most common blood clotting genetic disorder Factor V Leiden (Up to 30% of patients who present with deep vein thrombosis (DVT) or pulmonary embolism have this condition). So good news
  2. I do, however, have a mutation (heterozygous for this mutation) on the gene right next to the Factor V Leiden gene which, according to Hem #2, has only been seen in the literature once. The clinical signficance of this mutation is unknown, but evidently the labs and geneticists at this medical center are going bananas at this mutation I have. So I was sent the lab to measure my Activated Protein C Resistance Ratio, which, from what I gather, measures if my blood likes to clot a lot.

Naturally I have a HUGE amount of questions regarding why this genetic mutation is so exciting to the geneticists (and as luck would have it, my Hem #2 is heading to Antarctica for 3 weeks today so I can't really discuss more with him for a bit), so I've found out the name of the geneticist and I have emailed him to set up an appt to discuss what's going on.

Bottomline, all of this testing could lead to absolutely no new info (it's possible that this gene mutation means absolutely nothing clinically)- but at least I know I investigated the issue thoroughly and I'm not missing an opportunity to treat this potentially deadly issue of DVT (my clinic has had 2 patients die from PE that originaled as a DVT from the arm).

But it's my health, and my life, and I'll be as proactive as I want to be. I deserve it.

Here's to a healthy and happy 2010!

Wednesday, December 23, 2009

Lung transplant, H1N1 patients bypass ventilators with ECMO

Lung transplant, H1N1 patients bypass ventilators with ECMO

Liesbeth Stoeffler used to run marathons. By 2007, she could barely walk a block without taking a break. She began avoiding stairs and taking taxis instead of walking the eight blocks to work.
Before being diagnosed in 1995 with cystic fibrosis, an inherited disease in which the body produces unusually thick mucous that clogs the lungs and digestive tract, Stoeffler, now 50, was a vice president for AllianceBernstein, an investment management firm in Manhattan. She skied, did yoga and traveled.

Stoeffler eventually received a lung transplant and is now on the mend. But she might not have survived without a device that is proving to be a bridge to lung transplants and a possible alternative to ventilators.

ECMO (extracorporeal membrane oxygenation) devices have been around for about 20 years. Now simplified and portable, they are rising in popularity and even have a role in treating swine flu, or H1N1, patients.

Stoeffler, who had to resign from her firm and was forced to go on oxygen 24 hours a day, is one whose life was changed by the device.

Stoeffler found herself hospitalized on June 1 this year. That day, she went for routine blood work. Her oxygen levels were low, and she was very sick, says David Lederer, Stoeffler's pulmonologist. She was admitted to New York Presbyterian Hospital-Columbia's intensive care unit and placed on the ventilator.

Patients on ventilators are often partially sedated and unable to speak. There is also a high mortality rate for patients with cystic fibrosis on the ventilator, Lederer says.

Ordinarily, air is drawn into the lungs naturally when breathing. But a ventilator pushes air into the lungs instead, often causing lung damage from the pressure differences, says Matthew Bacchetta, Stoeffler's surgeon and director of the adult ECMO program at New York Presbyterian Hospital-Columbia.

"Patients usually have poor outcomes and die on the ventilator or never make it to transplant because they are so deconditioned," Bacchetta says.

He and Lederer decided to replace the ventilator with an ECMO device. Instead of pushing air into the lungs, ECMO machines bypass the lungs and directly oxygenate the blood.
With Stoeffler, the hospital used the device earlier than in most lung-failure cases and removed all "unnecessary" parts of the machine to make Stoeffler more comfortable and allow her to move, Lederer says. The device typically has two entrances into the body: one in the neck and one in the groin. But the team used only one entrance, the jugular vein in her neck, which allowed her to get up and walk.

"(Bacchetta) and I and the team decided to use ECMO in this novel fashion because we knew that once she was on that ventilator, her time was quite limited," Lederer says. But with ECMO, the team was able to improve her oxygen and carbon dioxide levels.
"You would never even know that this woman had these two tubes connected to the side of the neck," says Christina Restivo, Stoeffler's close friend.

On ECMO, Stoeffler could have most liquids and foods, which helped her gain weight and strength. She could eventually sit up, talk and even use her laptop and iPhone to document the events. "About five days into it, she told me it was the best she'd felt in years," Bacchetta says.
Bacchetta says ECMO can function as a bridge to transplant or a bridge to recovery; his patients have been on it for as long as three weeks. A young patient "literally dying" of swine flu in the spring was placed on ECMO to be transported to New York Presbyterian Hospital-Columbia and remained on the device until he recovered, he says.

"That's a patient who would've died otherwise," Bacchetta says. "We've had several other patients without this who have died."

ECMO could be used for pneumonia, pulmonary fibrosis and chronic obstructive pulmonary disease, Lederer says. It could be used during the swine flu outbreak, especially if hospitals are short of ventilators, he says.

For Stoeffler, ECMO allowed her to regain enough strength to remain eligible for a transplant.
She had double lung transplant surgery July 20. She returned home Sept. 4, without oxygen support.

"The ECMO was the bridge between my respiratory failure and the transplant," Stoeffler says. "If I had been intubated (a tube down the throat) like everybody else, I would've been sick. I don't think I would've survived that."

She recently started jogging. Though she has some muscle and weight to gain, Lederer says, there is a chance she could be running marathons again. But for now, she can walk around her apartment and talk without resting every few seconds. And she is free of the oxygen tank.

Melatonin improves sleep and reduces nitrite in the exhaled breath condensate in CF

J Pineal Res. 2010 Jan;48(1):65-71.

Melatonin improves sleep and reduces nitrite in the exhaled breath condensate in cystic fibrosis - a randomized, double-blind placebo-controlled study.

de Castro-Silva C, de Bruin VM, Cunha GM, Nunes DM, Medeiros CA, de Bruin PF.
Department of Medicine, Universidade Federal do Ceará. Rua Prof. Costa Mendes 1608 - 4 Andar, Fortaleza, Ceará, Brazil.

Cystic fibrosis (CF) is a chronic progressive disorder characterized by repeated episodes of respiratory infection. Impaired sleep is common in CF leading to reduced quality of life. Melatonin, a secretory product of the pineal gland, has an important function in the synchronization of circadian rhythms, including the sleep-wake cycle, and has been shown to possess significant anti-oxidant properties.

To evaluate the effects of exogenous melatonin on sleep and inflammation and oxidative stress markers in CF, a randomized double-blind, placebo-controlled study initially involving 20 patients with CF was conducted. One individual failed to conclude the study. All subjects were clinically stable when studied and without recent infectious exacerbation or hospitalization in the last 30 days.

Groups were randomized for placebo (n = 10; mean age 12.1 +/- 6.0) or 3 mg melatonin (n = 9; mean age 16.6 +/- 8.26) for 21 days. Actigraphy was performed for 6 days before the start of medication and in the third week (days 14-20) of treatment. Isoprostane and nitrite levels were determined in exhaled breath condensate (EBC) at baseline (day 0) and after treatment (day 21). Melatonin improved sleep efficiency (P = 0.01) and tended to improve sleep latency (P = 0.08). Melatonin reduced EBC nitrite (P = 0.01) but not isoprostane.

In summary, melatonin administration reduces nitrite levels in EBC and improves sleep measures in clinically stable CF patients. The failure of melatonin to reduce isoprostane levels may have been a result of the low dose of melatonin used as a treatment.

PMID: 20025642 [PubMed - in process]

Tuesday, December 22, 2009

Comparison of inhaled mannitol, daily rhDNase, and a combination of both

Comparison of inhaled mannitol, daily rhDNase, and a combination of both in children with cystic fibrosis: a randomised trial.

Minasian CC, Wallis C, Metcalfe C, Bush A.Royal Brompton Hospital & Imperial College, London, United Kingdom;

BACKGROUND: Osmotic agents, such as inhaled dry powder mannitol may increase mucociliary clearance (MCC) by rehydrating the airway-surface liquid (ASL) and thus act as disease-modifying treatments in cystic fibrosis (CF). This is the first therapeutic trial of inhaled mannitol in children with CF. We have compared it to human recombinant deoxyribonuclease (rhDNase), the current best established mucolytic therapy.

METHODS: 38 children were recruited to an open crossover study. Subjects underwent an initial bronchial provocation challenge with dry powder mannitol. Those children with a negative challenge were randomly allocated to one of three consecutive twelve-week treatment blocks (inhaled mannitol alone, nebulised rhDNase alone and mannitol + rhDNase). The primary outcome was forced expiratory volume in 1 second (FEV(1)). A number of secondary outcome measures were also studied.

RESULTS: Twenty children completed the study. Bronchoconstriction and cough associated with mannitol administration contributed to the high attrition rate. The mean increase in FEV(1) following 12 weeks of treatment was 0.11 litres (6.7%) (p=0.055) for mannitol alone, 0.12 litres (7.2%) (p=0.03) for rhDNase alone & 0.03 litres (1.88%) (p=0.67) for rhDNase and mannitol. None of the secondary clinical outcomes were statistically significantly different between treatments.

CONCLUSIONS: Inhaled mannitol was at least as effective as rhDNase after 3 months treatment. There was a marked individual variation in tolerance to mannitol and in response to therapy however. Children who do not respond to rhDNase many benefit from a trial of inhaled mannitol. The combination of mannitol and rhDNase was not useful.

The trial was registered with ClinicalTrials.gov (Identifier NCT00117208).

CFTR Mutations in azoospermic and oligospermic men

Reprod Biomed Online. 2009;19(5):685-694.

Cystic fibrosis transmembrane conductance regulator mutations in azoospermic and oligospermic men and their partners.

Gallati S, Hess S, Galié-Wunder D, Berger-Menz E, Böhlen D.

Division of Human Genetics, Departments of Paediatrics and Clinical Research, Inselspital, University of Berne, CH-3010 Berne, Switzerland.

The objective of this study was to investigate the contribution of cystic fibrosis transmembrane conductance regulator (CFTR) to human infertility and to define screening and counselling procedures for couples asking for assisted reproduction treatment.

Extended CFTR mutation screening was performed in 310 infertile men (25 with congenital absence of the vas deferens (CAVD), 116 with non-CAVD azoospermia, 169 with severe oligospermia), 70 female partners and 96 healthy controls. CFTR mutations were detected in the majority (68%) of CAVD patients and in significant proportions in azoospermic (31%) and oligospermic (22%) men.

Carrier frequency among partners of infertile men was 16/70, exceeding that of controls (6/96) significantly (P = 0.0005). Thus, in 23% of infertile couples both partners were carriers, increasing the risk for their offspring to inherit two mutations to 25% or 50%. This study emphasizes the necessity to offer extended CFTR mutation screening and counselling not only to patients with CAVD but also to azoospermic and oligozoospermic men and their partners before undergoing assisted reproduction techniques. T

he identification of rare and/or mild mutations will not be a reason to abstain from parenthood, but will allow adequate treatment in children at risk for atypical or mild cystic fibrosis as soon as they develop any symptoms.

Conference Findings for Lung Transplantation in CF

International leaders in transplantation and cystic fibrosis (CF) met in June 2008 in Lansdowne, Virginia in a roundtable conference to consider the state of the art in lung transplantation for CF. The meeting brought together different members of the pediatric and adult lung transplant community, experts in biostatistics, clinical research techniques, quality of life, and advocates of the CF community.

Lung transplantation for CF was first performed using a combined heart–lung transplant in 1983, with the first bilateral lung transplants for CF performed a few years later. Short-term success with lung transplantation prompted growth in the use of the procedure for CF and fostered hope in children and adults whose CF lung disease had progressed to end stage. According to the Organ Procurement and Transplantation Network, approximately 200 patients with CF, including approximately 25 children, undergo lung transplantation in the United States each year.

As with all transplantation, there are more candidates than available donors. Another issue with CF patients is the fact that lung transplant is not a cure for CF lung disease. Adult CF patients have a the median survival after lung transplantation of only 6.4 years. This makes choosing the time to list a patient with CF for a lung transplant crucial.

Reports intended to provide guidance to physicians and surgeons regarding the timing of listing began to appear in the early 1980s. One of the first and most widely referenced reports is from the Toronto Lung Transplant Program. Subsequent studies have attempted to refine those criteria.

The authors of the Toronto paper found that in their experience, patients with a forced expiratory volume in one second (FEV1) of 20% of predicted had a 70% chance of death within 2 years, but the chance of death halved for every 10 percentage-point increase in FEV1. Using that information, the authors concluded that CF patients should be considered candidates for lung transplantation when the FEV1 falls below 30% of the predicted value. The conference agreed that better clinical predictors of short-term mortality among patients with CF are needed as “prediction of the appropriate time to offer transplant for CF remains largely subjective.”
Lung transplantation is an option for selected pediatric and adult patients with advanced or severe lung disease that has failed to respond to standard therapy. Despite the steady increase in the number of candidates listed for lung transplantation, there have been few population-based studies on the benefits (including survival) of lung transplantation for adults and children with end-stage lung disease. The June 2008 Lansdowne Conference participants agreed that:
Conference participants agreed that:

Optimal care for CF children and adults, whether before or after lung transplantation, should remain a primary goal.

Lung transplantation should remain an option for selected patients with CF and advanced lung disease.

Multiple factors influence the outcome of lung transplant recipients including:

  • physiological variables
  • patient-specific variables
  • lung transplant and CF Center-specific practices
  • infectious agents

There is a need for evidence-based practice guidelines for lung transplant centers
A prospective, multicenter study on patients with CF and lung transplant is necessary to answer important questions on criteria for candidate selection and transplant practice guidelines

Friday, December 11, 2009

FDA panel: Gilead Sciences drug to treat deadly lung infection appears safe and effective

FDA panel: Gilead Sciences drug to treat deadly lung infection appears safe and effective

On 3:50 pm EST, Thursday December 10, 2009

WASHINGTON (AP) -- Federal health advisers are recommending the use of an experimental Gilead Sciences drug to treat a form of lung infection.
A Food and Drug Administration panel voted 15-2 in favor of the safety and effectiveness of the inhalable drug Cayston.
Cayston is intended to treat chronic lung infections in patients with cystic fibrosis -- an often fatal disease that affects the lungs and digestive system. Lung infections are the biggest killer of people with the disease.
The drug is designed to be given three times per day for 28 days.
The same panel of experts voted unanimously that Gilead has identified the appropriate dose for Cayston.
The FDA is not required to follow the group's advice, though it often does.

CFvoice.com video

Many of you know me as a pretty private person, but CFvoice.com has given me a great outlet to raise awareness of this disease.

This newest segment was a long time in the works.

Make sure to watch all the videos on the right hand side.


Tuesday, December 8, 2009

The effect of N-acetylcysteine on chloride efflux from airway epithelial cells

Cell Biol Int. 2009 Oct 20

The effect of N-acetylcysteine on chloride efflux from airway epithelial cells.

Varelogianni G, Oliynyk I, Roomans GM, Johannesson M.

Defective chloride transport in epithelial cells increases mucus viscosity and leads to recurrent infections with high oxidative stress in patients with cystic fibrosis (CF). N-acetylcysteine (NAC) is a well known mucolytic and antioxidant drug, and an indirect precursor of glutathione.

Since S-nitrosoglutathione (GSNO) previously has been shown to be able to promote Cl- efflux from CF airway epithelial cells, it was investigated whether NAC also could stimulate Cl- efflux from CF and non-CF epithelial cells and through which mechanisms.

CF bronchial epithelial cells (CFBE) and normal bronchial epithelial cells (16HBE) were treated with 1mM, 5mM, 10mM or 15mM NAC for 4h at 37 degrees C. The effect of NAC on Cl- transport was measured by Chloride efflux measurements and by X-ray microanalysis. Cl- efflux from CFBE cells was stimulated by NAC in a dose dependent manner, with 10mM NAC causing a significant increase in Cl- efflux with nearly 80% in CFBE cells.

The intracellular Cl- concentration in CFBE cells was significantly decreased up to 60 % after 4h treatment with 10mM NAC. Moreover immunocytochemistry and Western Blot experiments revealed expression of CFTR channel on CFBE cells after treatment with 10mM NAC.

The stimulation of Cl- efflux by NAC in CF airway epithelial cells may improve hydration of the mucus and thereby be beneficial for CF patients.

High rates of fracture among CFer's

Calcif Tissue Int. 2009 Dec 1.

The Prevalence of Osteoporosis, Osteopenia, and Fractures Among Adults with Cystic Fibrosis: A Systematic Literature Review with Meta-Analysis.

Paccou J, Zeboulon N, Combescure C, Gossec L, Cortet B.

Département Universitaire de Rhumatologie, Centre Hospitalier et Universitaire, Hôpital Roger Salengro, Lille, France, julienpaccou@yahoo.fr.

Observational studies have indicated a high but heterogeneous prevalence of low bone mineral density for adult patients with cystic fibrosis. Fracture complications were also described. The objective of this study was to determine the prevalence of osteoporosis, osteopenia, and fractures among adult patients with cystic fibrosis.

A systematic literature review was conducted using electronic databases. The keywords used were "cystic fibrosis [MeSH] AND bone density." Original studies were eligible if they reported the prevalence of osteoporosis and/or osteopenia and/or fractures in adult patients with cystic fibrosis. A meta-analysis of pooled proportions was performed. Heterogeneity was tested with the Cochran Q statistic, and in the case of heterogeneity a random effect model was used. Of 117 studies, 12 were selected, i.e., that represented a total of 1055 patients. Mean age ranged from 18.5 to 32 years (median: 28.2 years). Mean body mass index ranged from 19.9 to 22.4 (median: 20.7); 53.8% were men.

The pooled prevalence of osteoporosis in adults with cystic fibrosis was 23.5% (95% CI, 16.6-31.0). The pooled prevalence of osteopenia was 38% (95% CI, 28.2-48.3). The pooled prevalences of radiological vertebral fractures and nonvertebral fractures were 14% (95% CI, 7.8-21.7) and 19.7% (95% CI, 6.0-38.8), respectively.

In conclusion, this systematic literature review with meta-analysis emphasized the high prevalence of osteopenia and osteoporosis in young adults with cystic fibrosis. The prevalence of fracture was also high.

Scientists Restore Some Function to Cells from Cystic Fibrosis Patients

Scientists Restore Some Function to Cells from Cystic Fibrosis Patients

ScienceDaily (Dec. 7, 2009) — In an encouraging new development, a team led by Scripps Research Institute scientists has restored partial function to lung cells collected from patients with cystic fibrosis. While there is still much work to be done before the therapy can be tested in humans, the discovery opens the door to a new class of therapies for this and a host of other chronic diseases.

The results were published on December 6, 2009 in an advance, online edition of the high-impact journal Nature Chemical Biology.

"We are very excited by these results," said team leader Professor William Balch, a professor in the Departments of Cell Biology and Chemical Physiology and member of the Institute for Childhood and Neglected Diseases, who also receives support from the Skaggs Institute for Chemical Biology, all at Scripps Research. "Because we came at the problem of restoring cell function from a new perspective -- using biology to correct biology -- these findings have the potential to be game-changing."

The new study, performed in collaboration with a large number of cystic fibrosis investigators across the United States and Canada, showed that a compound called suberoylanilide hydroxamic acid (SAHA), which is already approved by the U.S. Food and Drug Administration as a treatment for lymphoma, can restore about 28 percent of normal function to lung surface cells with the most common, yet severe, cystic fibrosis mutation that results in complete loss-of-function in homozygous patients (those receiving a copy of the mutated gene from both parents).
"The results are very promising," said Balch. "We know that cystic fibrosis individuals with 15 to 30 percent of normal cellular function, as can occur with certain mutations, have milder cases of the disease and a more normal lifestyle than patients carrying a severe mutation. The added degree of function conveyed by SAHA or a compound like SAHA could make a tremendous difference to patients with acute disease."

A Life-Shortening Condition

Cystic fibrosis is an inherited disease that affects about 30,000 children and adults in the United States, and 70,000 worldwide, according to the Cystic Fibrosis Foundation.
In cystic fibrosis, patients produce thick, sticky mucus, which can clog the lungs and result in damaging inflammation and life-threatening infections. This thick mucus can also obstruct the pancreas and interfere with the proper digestion and absorption of food. Other symptoms include diabetes and infertility.

People with cystic fibrosis have mutations in the cystic fibrosis gene, which leads the body to produce a defective protein -- called the cystic fibrosis transmembrane conductance regulator (CFTR) protein -- which is normally found at the cell surface and is necessary for the proper movement of sodium and chloride (salt) in and out of cells. This process is necessary for the proper hydration of the lung, intestine, and pancreas.

Although more than 1,400 different mutations can lead to defects in CFTR, the most common mutation is a deletion of a phenylalanine residue at position 508 of the protein (DF508 CFTR). This mutation, which causes a severe form of the disease, is responsible for more than 90 percent of cystic fibrosis cases worldwide.

While 50 years ago few children with cystic fibrosis lived to attend elementary school, today it is not unusual for people with cystic fibrosis to live into their 30s and 40s. Most treatments available today focus on symptom management -- specifically clearing the airways through lung compression vests, antibiotics, inhaled medications, and anti-inflammatory drugs, as well as promoting proper nutrition through a healthy diet, substantive digestive enzyme supplements, and other dietary aids.

Unfortunately, still lacking are approved medications that address problems with the DF508 cystic fibrosis protein. (Balch notes that a potentiator drug (Vertex 770) affecting a very rare mutant population of cystic fibrosis patients (G551D CFTR) which is at the cell surface, but unable to mobilize chloride, did achieve significant clinical benefit in a recent clinical trial). A broadly effective therapeutic correcting DF508 CFTR protein delivery to the cell surface and restoring function would be a great boon for most cystic fibrosis patients and their families.
Such a development could alleviate the chronic need for the many drugs and therapies that attempt to mitigate the onslaught of symptoms and to enable patients to sustain a normal lifestyle.

Turning a Classical Approach on Its Head

In the new study, Balch took an original approach to correcting cystic fibrosis defects. This new approach grew out of a unique understanding of protein folding and misfolding that Balch had been working out for some time with Scripps Research colleague Jeffery Kelly, chair of the Department of Molecular and Experimental Medicine, Lita Annenberg Hazen Professor of Chemistry, and member of the Skaggs Institute for Chemical Biology. This perspective may also have implications for conditions as diverse as type II diabetes, arthritis, osteoporosis, and amyloid disease (including Alzheimer's).

While in many genetic diseases specific mutations within a particular gene cause the protein product of the gene to misfold, Balch and colleagues note that is not the end of the story. Critically, this defective protein must interact with the general biological machinery of the cell, which controls the protein folding and stability environment. This can contribute significantly to the protein's loss of function and a breakdown in tissue/organism function. This biological machinery controlling the folding and function environment of the cell is referred to as the proteostasis network and is central for life.

thinking and practices of the pharmaceutical industry that focuses on drugging single targets," Balch said. "This traditional view limits our ability to tackle pharmacologically many complex loss-of-function sporadic and inherited diseases which are really systems disorders. These diseases have multiple steps in the biological network that must be adjusted to regain a more normal function of the compromised protein and tissue."

In the case of cystic fibrosis, Balch suspected that the endoplasmic reticulum -- a compartment in the cell responsible for the synthesis of CFTR which normally works to protect the body by degrading potentially dangerous abnormal proteins -- could be viewed as doing its job too efficiently, eliminating mutant CFTR proteins that could still provide some function to the cell and tissue if given the opportunity.

In the new Nature Chemical Biology study, Balch and colleagues drew on their previous theoretical and experimental work to turn the classical model of drug development on its head. Rather than attempting to directly target or replace the mutant CFTR proteins present in cystic fibrosis patients -- an approach that had so far failed to yield dramatic new treatments for the DF508 disease -- Balch and colleagues sought instead to adjust the cell folding or maintenance proteostasis machinery of the cell to make a new cellular environment that would "work" with the mutant CFTR proteins.

In so doing, the scientists hoped that the mutated CFTR proteins, while not perfect, could now function more effectively in the cell, reducing the more severe effects of cystic fibrosis symptoms in the common DF508 variant.

Positive Results on Multiple Levels

To tweak the cellular machinery in this fashion, Balch and the team enlisted compounds that were known to inhibit a family of enzymes known as histone deacetylases (HDACs). Studies have shown that HDACs affect the packaging of the DNA in chromosomes and regulate gene expression. Based on earlier studies of the folding environment required for CFTR function published in the journal Cell, the scientists reasoned that altering HDAC function might also rebalance proteostasis networks in the cell to favor functional restoration.
For expertise in these compounds, Balch teamed up with Scripps Research colleague Professor Joel Gottesfeld and his group.

"Joel's lab and my lab worked very closely together on this," said Balch. "That's what's great about Scripps Research -- its collaborative nature. People just walk in next door and two hours later you are doing an experiment together!"

Working with Ray Frizzell and Joe Pilewski, cystic fibrosis investigators at the University of Pittsburg School of Medicine, the Balch laboratory treated human lung epithelial cells isolated from patients with the devastating ΔF508 mutation with known HDAC inhibitors. Intriguingly, the FDA-approved HDAC inhibitor SAHA was shown to be most effective in restoring surface channel activity -- one of the main markers of cystic fibrosis that is responsible for rehydration of the cells' surface. Control cells demonstrated negligible levels of channel activity, while SAHA-treated cells were restored to 28 percent of the normal level found in healthy individuals.
"It's a pretty solid rescue with some intriguing properties," commented Balch.

Using a bioinformatics approach led by the Gerard Manning laboratory at the Salk Institute of Biological Sciences, the team showed that the compound increased the functioning of mutated CFTR proteins at multiple levels in the proteostasis network. Not only were mutant CFTR proteins more protected from destruction in the endoplasmic reticulum, they were also more efficiently transported to the lung cell surface where they were found at comparable levels to that of wild-type (normal) CFTR. In addition, once at the surface of HDAC inhibitor-treated cells, mutant CFTR proteins were better able to resist destruction by additional degradation pathways than DF508 CFTR proteins in untreated cells.

"By rebalancing the proteostasis program to provide a more supportive cellular environment," said Balch, "the cells appear to treat the mutation more like a polymorphism [genetic differences that are responsible for individual diversity] rather than something dangerous needing to be completely eliminated."

Balch also likened this process to evolutionary adaptations to changes in protein structure, which support mutations providing a selective functional advantage.

The study showed that one specific HDAC -- HDAC7, one of 18 known human HDACs -- appeared to be largely responsible for the effects on the treated cells. Little is known about the function of HDAC7 in human physiology and efforts are currently under way by the laboratories of Balch, Gottesfeld, Manning, and Scripps Research Professor John Yates to understand its mechanism of action.

The Way Forward

Mindful that dosing would be a key issue in any attempts at drug development, the researchers also began to address dosing issues in the current study.
"We know that a compound won't make it into the clinic if patients have to take the equivalent of a cereal bowl of it several times a day," said Balch. "That's especially true because, given the nature of cystic fibrosis, this would need to be a sustained, life-long treatment to protect the patient from disease."

Remarkably, the team found that low doses of SAHA not only worked in cultures of cystic fibrosis lung cells, but also offered some significant advantages over acute doses. While acute doses of SAHA produced an increase in the surface channel activity the next day, the effects stopped soon after the drug was withdrawn. In contrast, much smaller doses began working efficiently after six to eight days and strong channel activity was observed after the drug was withdrawn, gradually declining over the following week. This feature is reminiscent of its potential mechanism of action, Balch said, perhaps involving chromatin remodeling leading to an altered, protective proteostasis environment in the lung cell that could be sustainable.
While thrilled with the results, Balch cautions that "there is much work to do" -- including further drug development, preclinical work, and clinical trials -- before any new therapy for cystic fibrosis becomes a reality using this approach. The FDA-approved drug SAHA, while initially approved as an acute dose regimen for cancer therapeutics, remains to be carefully examined for use in a low-dose, chronic treatment regimen that would be required for protecting cystic fibrosis patients from disease over a lifetime.

First authors of the paper are Darren Hutt and David Herman of Scripps Research. In addition to Balch, Hutt, and Herman, other authors include: Jeanne Matteson, Ben Hoch, Wendy Kellner, Jeffery Kelly, J. R. Yates IIIrd and Joel M. Gottesfeld of Scripps Research; Ana Rodrigues and Gerard Manning of the Salk Institute for Biological Studies; Sabrina Noel, Joe Pilewski and Ray Frizzell of University of Pittsburgh School of Medicine; Andre Schmidt and Philip Thomas of the University of Texas Southwestern Medical Center; Yoshihiro Matsumura and William Skach of Oregon Health and Sciences University; Martina Gentzsch and John R. Riordan of University of North Carolina, Chapel Hill; Eric J. Sorscher of University of Alabama at Birmingham; and Tsukasa Okiyonad and Gergely L. Lukacs of McGill University. Frizzel, Skach, Thomas, and Lukacs are members of the Cystic Fibrosis Consortium, a team of investigators supported by the Cystic Fibrosis Foundation that are encouraged to collaborate and share results to accelerate the pace for a cure for cystic fibrosis. Finanical disclosure: Balch is a Consultant and equity holder in Proteostasis Therapeutics Incorporated (PTI), 200 Technology Square, Boston MA, whose goal is to develop drugs that correct human misfolding disease. Kelly is a founder and equity holder in PTI.

This research was supported by National Institutes of Health (NIH) grants HL79442 (WEB/JRY), GM42336 (WEB), and the Cystic Fibrosis Consortium (CFC)(WEB/JRY); NS055781 to JMG; AG03197 to GM; AG84567 to JWK; DK68196, DK72506, and the CFC to RAF and JP; UR98647 and the CFC to ES; DK075302, the Canadian Institutes of Health Research and the CFC to GLL; DK51818 and the CFC to WRS; DK23567 to JRR. D. Hutt was supported by fellowships from the Canadian Cystic Fibrosis Foundation and the Canadian Institutes of Health Research; D. Herman was supported by a fellowship from the Friedreich's Ataxia Research Alliance.

Genta To Support Initiation Of New Clinical Trial Using Ganite(R) As Treatment For Life-Threatening Infections In Patients With Cystic Fibrosis

Genta Incorporated (OTCBB: GETA) announced that the Company will supply Ganite® (gallium nitrate injection) for a new clinical trial that will be initiated in patients with cystic fibrosis (CF) who may develop serious infections.

Infection is the most common cause of death in CF patients. Frequently, these infections are due to bacteria known as Pseudomonas aeruginosa, and patients are commonly treated with years of antibiotic therapy to control such infections. However, prolonged antibiotic use has greatly increased resistance due to genetic mutations. In addition, Pseudomonas has evolved protective mechanisms known as "biofilms" that encase colonies of bacteria and prevent antibiotics from reaching infection sites.

Bacteria in biofilms are far more resistant to being killed. Recent information has suggested that gallium may be used as an anti-bacterial agent against Pseudomonas, in part due to its ability to disrupt biofilms. The initial clinical trial using a gallium compound for this purpose will involve patients with cystic fibrosis who will be treated at the University of Washington in Seattle, WA and the University of Iowa, Iowa City, IA. If initial results are promising, additional trials will be conducted with additional medical centers and a larger number of patients.

Genta has provided cross-reference to its Investigational New Drug (IND) exemption for Ganite®, and the Company will supply the drug at no cost to patients in the trial. The study is also supported by grants awarded by the Orphan Products Division of the Food and Drug Administration (FDA) and by the Cystic Fibrosis Foundation.

"Work from our center has established that gallium can be used as a 'Trojan horse' that interferes with how iron is used by Pseudomonas," said Dr. Christopher H. Goss, Associate Professor of Medicine at the University of Washington, who is the Principal Investigator of the trial. "Our preliminary data suggest that gallium exploits potentially vulnerable mechanisms in Pseudomonas by disrupting biofilms and killing antibiotic-resistant strains of bacteria. This study is the first formal pharmacokinetic and safety evaluation of gallium in patients with cystic fibrosis." "

This trial represents the rapid clinical translation of enormously promising observations," said Dr. Raymond P. Warrell, Jr., Genta's Chief Executive Officer. "Initial results from our first CF patient showed that systemic treatment achieved target gallium levels in sputum, and that these levels could be sustained for a prolonged duration. Pseudomonas infections in CF are exceptionally difficult to eradicate. If initial results of the IV drug in this trial are promising, future patients might also benefit from extended therapy that could be afforded by one of our oral gallium compounds."

Further background on this work from the Universities of Washington and Iowa can be accessed here. About Ganite® Ganite® is exclusively marketed by Genta and is approved in the U.S. for treatment of cancer-related hypercalcemia that is resistant to hydration. The potential use(s) described in the release are investigational and have not been approved. SourceGenta Incorporated