This week, I wanted to highlight a new case report published last week in Cell. It documents the successful phage therapy of a Mycobacterium abscessus infection, which led a 26-year old in Colorado with cystic fibrosis to receive a lung transplant one year after phage therapy. This is the first documented successful treatment of pulmonary M. abscessus infection with phage.
The study was published May 13, 2022 in Cell by Jerry Nick, Rebekah Dedrick and colleagues.
Quick snapshot
What was the patient dealing with?
A 26-year-old man with cystic fibrosis with advanced bronchiectasis and M. abscessus lung disease. For almost 5 years, the patient dealt with this infection (he also had chronic infection with P. aeruginosa and MRSA). In the year before phage, he was hospitalized 11 times. Because of this infection, he was denied lung transplant by three independent programs.
What was done?
A personalized phage therapy approach was used to treat the patient. Two phages, both lytic on the patient’s M. abscessus strain, were administered intravenously over 500 days.
They extensively monitored the patient’s response to phage therapy, and showed that the M. abscessus strain was reduced by the treatment and lysed by the phage. They even followed the genetic changes of the isolate over time.
They also looked at antibodies to phages and saw that even though some neutralization occurred, it didn’t block the phages from doing their job (even after more than a year of giving the patient the same phages, every day!). They also looked for phage resistance, but didn’t find any.
What was the clinical outcome?
The patient, who was not allowed to get a lung transplant before phage because his M. abscessus infection wasn’t under control (even after years of antibiotics), finally got his lung transplant! And no adverse effects of the phages were observed.
The nitty gritty: how was this done?
Identifying the bacteria
The patient’s strain was isolated over four years before phages were given. It was sequenced, which showed plenty of antibiotic resistance genes, but no prophages or plasmids.
What is M. abscessus?
Mycobacterium abscessus is a species of non-tuberculous Mycobacteria (NTM). NTM frequently colonize CF patients, and is commonly associated with poor prognosis due to antibiotic resistance.
Choosing and preparing the phages
The strain was sent to Graham Hatfull’s Mycobacterium phage genetics lab, which has a collection of thousands of phages, most of which were isolated by undergraduate students in the SEA-PHAGES program.
From there, they found two with lytic activity and chose those for therapy. The two winners were ‘host range mutants’ of phage BPs and D19. BPs was engineered to be lytic, and D29 was a naturally occurring lytic phage (though it seems to have derived naturally from a parent that was temperate). They each lysed the strain alone, and also in combination. One phage was actually the same one used in two previous phage therapy cases — clearly a high value phage!
Phages were actually identified 1500 days (~4 years!) before phage therapy started. Fortunately, M. abscessus is generally considered clonal, so it wasn’t surprising that the bug didn’t change too much in the patient over this time. (I am personally still surprised — it seems wild that this can be the case! Definitely bodes well for phage therapy, knowing that these kinds of phage-sourcing head-starts can be possible for some pathogens/some situations.)
Phages were produced in the Hatfull lab. According to their previously described method (here), they do this by growing phages on plates, CsCl-purifying the lysate, dialyzing into PBS, and checking for endotoxin levels with a kit (not an issue since Mycobacterium naturally doesn’t produce it, but still needs to be checked for). Lastly, they check sterility. For this case, they stored their phages separately at 10^11 and 10^10 PFU/mL each, and combined them into a fresh cocktail monthly.
How were the phages administered?
10^8 to 10^9 pfu/ml in PBS, by IV, twice daily. This continues today.
Were antibiotics administered concurrently?
Yes. Many!
Major outcomes
- Before phage therapy: 78% cultures were positive for Mycobacterium
- The year after phage: 41% were positive
- The most recent 280 days: 10% were positive
- Hospital free time increased
- Lung function didn’t change
- He was listed for a transplant a year after since his Mycobacterium infection was controlled
- Successfully transplanted on day 379 following phage therapy
- They looked at his removed lungs and couldn’t culture the Mycobacterium!
- Uncomplicated transplant recovery
- Normal lung function now
- Still on phage and antibiotics
In-depth tracking of M. abscessus over time
The authors had to get more creative than standard culture-based methods to show M. abscessus was actually decreasing over time, since culturing from airway samples is unreliable for this pathogen, especially when other pathogens are also present.
So they did lots of extra things, like monitoring urine for sugars present in Mycobacterium cell wall, and showing they decreased in response to treatment. They even looked at the patient’s lungs after they were removed during his transplant: they showed Mycobacterium couldn’t be cultured from any part of the lungs (though they did culture other bacteria like Pseudomonas and Mycobacterium avium).
How did the pathogen evolve over time?
They sequenced the M. abscessus 40 different times over the course of pre- and post treatment with first antibiotics alone and then with phage added.
Mainly they saw a dominant clone which is seen worldwide, and multiple sublineages from this same ancestor. They actually think it colonized the patient almost 3 years before it was ever cultured.
They mapped which mutations arose over time, so you can see how the phage and antibiotics seemed to select for mutations in certain genes, and how the M. abscessus population became more homogeneous the longer the patient got antibiotics and phage. However, it wasn’t a huge difference; the greatest distance between any 2 isolates was 11 SNPs.
Did antibiotic resistance increase?
No, antibiotic resistance (measured as number of classes of antibiotics the strains were resistant to) increased gradually over the years of antibiotic use, but did not increase as a result of phage.
Was phage resistance an issue?
Surprisingly no. Phage resistance was not detected for either phage over the course of treatment, even 8 months after treating with the phages.
Were there any adverse or immune reactions?
The patient’s serum before phage therapy actually generated antibodies against the phages, which increased gradually the course of treatment for one of the phages (though did not substantially inactivate it until several months after therapy, when the individual’s cultures were mostly negative anyway). However, the other phage was not neutralized by serum antibodies even after 500+ days of phage therapy.
Did the phages replicate in the body?
They used conventional PCR to look for phages in serum, sputum and explanted lung samples. Strangely, all samples were negative. Instead, the authors seemed to focus on tracking the Mycobacterium, and considered a spike in Mycobacterium DNA soon after phage treatment as a proxy for phage-induced lysis.
Where did the phages end up?
We don’t know where the phages ended up in this case. We know they weren’t detected anywhere they looked; in serum, sputum or in the explanted lung.
What was the process of getting approval to try phage therapy?
The investigators got permission from FDA to use these phages through the expanded access investigational new drug (IND) pathway for compassionate use. They also had to of course get informed consent from the patient, and get their treatment protocol approved by an Institutional Review Board.
Normally data collection is frowned upon by FDA in a compassionate use setting. But this patient was enrolled in two CF Foundation trials: PREDICT (NCT02073409)(HS-2798) and PATIENCE (NCT02419989)(HS-2833). This allowed for data and specimen collection during the case.
Conclusions
This is an exciting case report that shows successful use of phages to help someone get a transplant they were otherwise unable to get. It shows how phage therapy can be given over a LONG time and still not be thwarted by neutralizing antibodies or phage resistance. It’s also a great example of how new therapeutic monitoring approaches during therapy can be enlightening (for example we see here how the pathogen evolved in response to phage). This paper also showed creative ways of demonstrating the effects of phage, even when a pathogen is tough to culture.
Kudos to the groups involved, this was a massive feat, and the phage world will undoubtedly be influenced big time by the findings of this paper.
Further reading
Main source for this issue:
Nick, J. A., Dedrick, R. M., Gray, A. L., Vladar, E. K., Smith, B. E., Freeman, K. G., … & Davidson, R. M. (2022). Host and pathogen response to bacteriophage engineered against Mycobacterium abscessus lung infection. Cell.
