1. Clinical failure and its management
David W. Denning
Director, National Aspergillosis Centre
University Hospital South Manchester
[Wythenshawe Hospital]
The University of Manchester

2. Problems with antifungal therapy
Drug toxicity
Drug interactions and low blood levels

3. Common reasons for stopping therapies
Drug toxicities
Common reasons for stopping therapies
Itraconazole Nausea Ankle swelling Peripheral neuropathy Fatigue Voriconazole Feeling ill Confusion/hallucinations/poor concentration Photosensitivity

4. Itraconazole concentrations in phase 2 studies
Denning et al, Am J Med 1994;97:135

5. Itraconazole concentrations in relation to timing of samples
Tucker et al, J Am Acad Dermatol 1990;23:

6. Optimising itraconazole levels – aim between 5 and 17 mg/L
Lestner et al, Clin Infect Dis 2009; 49:928

7. Itraconazole for ABPA in CF
Itraconazole often poorly absorbed and variable penetration into CF sputum
Sermet-Gaudelus, Antimicrob Ag Chemother 2001;45:1937.

8. Generic itraconazole (Sandoz)
Pasqualotto, Int J Antimicrob Ag 2007; 30:93

9. Approval of itraconazole by the FDA and Europe in 1991

10. Voriconazole - metabolism
98% metabolised by liver
Primarily metabolised by CYP2C19 and CYP3A4, less by CYP2C9.
Cirrhosis / prior alcohol abuse and elderly likely predictors of slow metabolisers. Also genetic polymorphism of CYP2C19.
Low levels likely in children, oral therapy and unpredictable.
Usual dosing 150 – 300mg twice daily
Voriconazole datasheet

11. Random voriconazole concentrations in adults receiving 3mg/Kg BID
100,000
Possible toxicity
10,000
1000
Log 10 [Concentration (µg/L)]
Very small children may metabolise voriconazole very fast and need dose escalation to ?7-10mg/Kg BID or 200mg BID
100 10 1 70
140
210
280
days after first dose
Data from Denning et al, Clin Infect Dis 2002;34:563

12. Voriconazole levels in children
Pasqualotto et al, Arch Dis Child 2008;93:578

13. Cytochrome P450 interactions
Fluc
Itra
Posa
Vori
Inhibitor
2C19 +
+++
2C9 ++
3A4
Substrate
Dodds Ashley & Alexander. Drugs Today 2006;41:393.

14. New section on drug interactions which you can search very quickly14

15. Problems with antifungal therapy
Drug toxicity
Drug interactions and low blood levels
Azole resistance, intrinsic and acquired

16. Chronic cavitary pulmonary aspergillosis (CCPA) in HIV February 2005
32 yr old from Malawi, on HAART Rx - haemoptysis - Aspergillus precipitin titre 1/16 CT scan shows 2 large cavities with aspergillomas, with additional lesions (October 2005)
Surgical removal would require a pneumonectomy So treated with itraconazole 16

17. CCPA in HIV February 2007
On HAART Rx, with low viral load, CD4 count >200 - New haemoptysis - Aspergillus precipitin titre 1/32 CXR & CT scan showed expansion of inferior cavity
MICs A. fumigatus Feb 2007 Itraconazole = >8.0mg/mL Voriconazole = 0.5 mg/mL Posaconazole = 1.0 mg/mL
February 2007
April 2007 17

18. CCPA in HIV - low itraconazole concentrations
Itraconazole concentrations Nov mg/L Dec mg/L March mg/L July mg/L Feb mg/L
Do low concentrations of antifungal predispose to the development of resistance? 18

19. microtitre, RPMI 2% glucose 35°C 48 hrs
Test inoculum
AF72
AF91
2x106/mL
microtitre, RPMI 2% glucose 35°C 48 hrs
Denning et al, JAC 1997;40:401

20. confirmation in vivo Strain 5 (AF 72) G54 CYP51A mutation
AmB 5mg/Kg
AmB 5mg/Kg
Itra 75mg/Kg
Itra 75mg/Kg
Itra 25mg/Kg
controls
Strain 5 (AF 72) G54 CYP51A mutation
Strain 6 (AF 91) M220 CYP51A mutation
Denning et al, JAC 1997;40:401

21. Development of international standards for susceptibility testing and breakpoints

22. Manchester azole MIC distributions
Itraconazole MIC (mg/L)
Voriconazole MIC (mg/L)
Posaconazole MIC (mg/L)
modified EUCAST method x 105 not x 105 cfu/mL 22

23. Azole resistance in A. fumigatus in Manchester 1997-20090% 7% 3% 5%
17%
14%
20%
Bueid, J Antimicrob Chemother 2010;65:2116. Howard et al, EID 2009; 15:1068 23

24. Clinical features of patients with azole resistant A. fumigatus
17 patients, 15 from UK, different cities
9 had CCPA, all with aspergilloma
3 had sputum isolate, with no treatment data
2 had ABPA
2 had IA
1 had Aspergillus bronchitis
13 of 14 patients had prior azole exposure
8 failed therapy and 5 failed to improve
(12 itraconazole, 1 voriconazole)
Howard et al, EID 2009; 15:1068 24

25. 25

26. Molecular detection of Aspergillus spp. in sputum
Laboratory result
ABPA
CPA
Normals
Culture positive for A. fumigatus
0/19
7/42 (16.7%)
0/11
qPCR positive for Aspergillus spp
15/19 (78.9%)
30/42 (71.4%)
4/11 (36.4%)
Denning et al. Clin Infect Dis 2011; 26

27. CF and Aspergillus cultures
Pre-sonication
Post-sonication
Baxter, unpublished

28. Routine culture cfu versus qPCR for Aspergillus Sputum and BAL
Sample BL AC PC VC JO
culture
qPCR
Sputum before 8
32.8 1 2
33.6
34.8
Ist trap 33
28.9
37.8
36.9
33.4
Ist wash (5-20mL) 38
30.3
33.5
BAL ( mL)
37.2
33.1
LLL BAL 12 34
LLL trap
32.7
RML BAL
neg
RUL BAL 10mL
RLL (120mL)
31.2
Sputum after 3
32.2
29.6
34.9
34.6
31.4
E. dermatiditis
Kirwan, AAA 2012 Abstract 28

29. Direct detection of resistance mutations in clinical specimens, without positive cultures
Laboratory result
ABPA
CPA
Normals
Culture positive for A. fumigatus
0/19
7/42 (16.7%)
0/11
qPCR positive for Aspergillus spp
15/19 (78.9%)
30/42 (71.4%)
4/11 (36.4%)
A. fumigatus CYP51A mutation detected directly from qPCR positive sample
6/8 (75%)
12/24 (50%) NT
Denning, Clin Infect Dis 2011;52:1123 29 29

30. Problems with antifungal therapy
Drug toxicity
Drug interactions and low blood levels
Azole resistance, intrinsic and acquired
Antifungal failure (without resistance/low azole blood levels etc)
Immune reconstitution or other ‘switching’ of immune response

31. Aspergillomas in CF
Turcios –

32. Felton, Clin Infect Dis 2010; 51:1383.

33. Second and third line antifungal therapy for ABPA and/or asthma
26 patients, ABPA (n = 21) or SAFS (n = 5).
All patients had failed itraconazole (n=14) or developed adverse events (n=12)
Chishimba et al, J Asthma . In press

34. Second and third line antifungal therapy for ABPA and/or asthma
26 patients, ABPA (n = 21) or SAFS (n = 5).
All patients had failed itraconazole (n=14) or developed adverse events (AEs) (n=12)
34 courses of therapy, 25 with voriconazole and 9 with posaconazole.
Voriconazole responses: 17/25 (68%) at 3 months, 15/20 (75%) at 6 months and 12/16 (75%) at 12 months,
Posaconazole responses: 7/9 (78%) at 3, 6 and 12 months for posaconazole.
18/24 (75%) discontinued oral corticosteroids, 12 of them within 3 months of starting antifungal therapy.
6/23 (26%) patients on voriconazole had AEs requiring discontinuation before 6 months compared to none on posaconazole (p=0.15).
4 relapsed (57%), 1 at 3 months and 3 at 12 months after discontinuation.
Chishimba et al, J Asthma . In press

35. Inhaled amphotericin B35

36. Dose and reconstitution
Dose can be increased in 5mg/1ml stages up to 20mg/4mls twice a day or a maximum daily treatment dosage of 1mg/kg
Reconstitution:
10ml water for injection added to 50mg yellow powder (5mg per ml)
(2ml therefore yields 10mg dose)
Consider residual volume of nebuliser!
Residual volume of pari lc plus nebuliser is 1ml
At uhsm we use fill volume of 4ml to increase drug delivery to patient, so to the 2ml solution we add a further 2ml water for injection 36

37. Compressors Need servicing regularly!
To drive most nebulisers an output of at least 8 L/m is required
UHSM nebuliser service – provide a year’s worth of kit and do annual service and replacement of consumables
Annual service – air outlet filter changed, output measured directly by engineers (Econoneb compressors even when brand new vary between 8.5 and 9.5 lpm)
Patients have to change the white air inlet filter every 3 months 37

38. The Pari LC plus with exhaust filter
Features:
Fill volume 2ml-8ml
Delivers approx 65% respirable dose
Can go through the dishwasher
Can survive boiling in water
Once a week patient should boil for 15 mins with a few drops of detergent. Can put through dishwasher
Filter pad needs changing after every use
Wash in warm soapy water and allow to air dry after every use
Typically 4ml takes under 10 min to nebulise
Pretty difficult to break and lightweight
When nebulising patient should be in room alone with door shut and window open – the filter system is not perfect!! !(encouragement may be required to do thus if sub zero temperatures outside!)
Nebuliser chamber 38

39. We may change to LC sprints (cost is deciding factor ultimately)39

40. Comparison of Pari LC versus other nebulisers
Many studies using Fungizone/ non-liposomal Amphotericin B/ Amphotericin B deoxycholate report using doses that exceed our usual 10mg. However, these studies use inferior nebulising equipment which delivers a lower respirable dose fraction (e.g. Dubois et al 1995 used a 30mg dose of  which 5% or 1.5mg was deposited in the lungs. For a 30mg dose delivered using a Pari LC plus nebuliser (the one I use!), this would deliver 65% or 19.5mg. a 10mg dose delivered by a Pari LC plus would deliver 6.5mg.) Much less will be delivered to the lungs if a facemask rather than mouthpiece is used. 40

41. Another challenge – immune reconstitution41

42. Day 0
Day 7
Miceli, Cancer 2007;110:112; Caillot Eur J Radiol 2010;74:e172

43. Immune reconstitution in invasive pulmonary aspergillosis, in AIDS
Patient HB
Day +14, CD4 cells 84/uL
Patient HB
Day +42, after AmB and ITZ
Sambatakou, Eur J Clin Microbiol Infect Dis 2005;24:628

44. Immune reconstitution in invasive pulmonary aspergillosis, in AIDS
Patient HB
Day +64, CD4 cells 340/uL, on VRC
Patient HB
Day +87, day of death
Sambatakou, Eur J Clin Microbiol Infect Dis 2005;24:628

45. Several patients have increasing breathlessness with antifungal therapy Documented fall in DLCO in one patient Deaths in others. Mechanism unclear. Likely benefit from steroids, needs good antifungal cover. 45

46. Interferon gamma replacement
Both patients improved with γIFN
Kelleher, Eur Resp J 2006;27:1307

47. CPA treatment – IFN gamma?
Denning DW et al, Clin Infect Dis 2003; 37(Suppl 3):S

48. Management approach
Exclude low blood levels – be careful of large dose increases with voriconazole
Fungal cultures – test for resistance
Exclude or treat bacterial co-infection
Use IV therapy if patient very ill
Consider surgical resection, gamma IFN, inhaled AmB (if ABPA/SAFS),
Long term IV therapy for CPA feasible and partially effective.