1. Rosanna Coppo
Torino
Steroid resistant nephrotic syndrome:
still a therepeutic challenge

2. Idiopathic Nephrotic Syndrome
minimal change
GN
IgMGN
mesangial
proliferative GN
focal segmental glomerulosclerosis
In all these cases
steroid resistance is the most strong predictor
of progression

3. corticosteroids
Nakayama, 2002

4. Clinical Practice Guideline for
Glomerulonephritis
WWW.KDIGO.ORG

5. CHAPTER 4: STEROID-RESISTANT
NEPHROTIC SYNDROME IN
CHILDREN
4.1: EVALUATION OF CHILDREN WITH SRNS
4.1.1: We suggest a minimum of 8 weeks treatment with
corticosteroids to define steroid resistance. (2D)
4.1.2: The following are required to evaluate the child with SRNS
(Not Graded):
• a diagnostic kidney biopsy;
• evaluation of kidney function by GFR or eGFR;
• quantitation of urine protein excretion.
Kidney Disease: Improving Global Outcomes
WWW.KDIGO.ORG

6. Podocytopathies

8. 14 types
Genetically determined
steroid –resistant Idiopathic NS
2-5%
of all the cases

10. Treatment
of children and adults with
idiopathic steroid-resistant NS
ALKYLATING AGENTS

11. Cochrane Database Syst Rev 2010
Alkylating agents in idiopathic
steroid-resistant NS in children
9 RCTs involving 449 children: RR of persistent NS
• Oral Cyclophosphamide+P
vs Prednisone
RR 1.01 (0.74-1.36)
• IV CPA vs oral CPA
RR 0.09 (0.01-1.39)
• Azathioprine+P
vs Prednisone
RR 1.01 (0.77-1.32)
no significant effect of alkylating drugs
on RR of persistent NS

12. Treatment and outcome
of children and adults with
idiopathic steroid-resistant NS
ALKYLATING AGENTS
CYCLOSPORIN

13. CHAPTER 4: STEROID-RESISTANT
NEPHROTIC SYNDROME IN
CHILDREN
4.2: TREATMENT RECOMMENDATIONS FOR SRNS
4.2.1: We recommend using a calcineurin inhibitor (CNI) as initial therapy for
children with SRNS. (1B)
4.2.1.1: We suggest that CNI therapy be continued for a minimum of 6
months and then stopped if a partial or complete remission of proteinuria
is not achieved. (2C)
4.2.1.2: We suggest CNIs be continued for a minimum of 12 months when
at least a partial remission is achieved by 6 months. (2C)
4.2.1.3: We suggest that low-dose corticosteroid therapy be combined
with CNI therapy. (2D)
Kidney Disease: Improving Global Outcomes
WWW.KDIGO.ORG

14. Cyclosporin treatment decreases
glomerular utrafiltration coefficient
15
3,4
10
8,60
5
1,8
4,2
2,1
Saline
0
2,40
CyA
Glomerula volume nl
Kl (ul/min mmHg cmH20)
Kf (nl/min mmHg)

15. CyA
Meyrier’s hypothesis:
CyA is lipophilic and bind sto a lipidic complex associated to the
slit diaphragm, limiting the protein leak

17. Synaptopodin (protein which stabilyzes cytosckeletal actin and is
co-localyzes with calcineurin)

18. Synaptopodin is instable and undergoes lysis due to cathepsin
with disappearance of cytoskeleton actin stress fibers

19. The interaction between synaptopodin and serine/treonine
binding protein maintains F-actin stability, normal cytoskeleton
structure and absence of proteinuria
F
actin
ser / treo BP
Dephosphorylation
synactopodin
synactopodin
P
ser / treo binding protein
sin
Cyclosporin
sin
sin

20. Cochrane Database Syst Rev 2010
Cyclosporin in idiopathic
steroid-resistant NS in children
3 RCTs: 49 children:
RR of persistent NS
Cyclosporin (CyA) vs PL
RR 0.64 (0.47-0.88)

21. Treatment of steroid-resistant NS
Cyclosporin A
Evidence-based recommendations
Treatment
MCGL
Level of
evidence
Grade
Comments
4
D
Possible benefit from
pooled case series; no
significant benefit in RCT
– small numbers
1
A
Beneficial
Cyclosporin (at
least 6 months)
FSGS
Cyclosporin (at
least 6 months)

22. An example of CyA nephrotoxicity.
Francois H, et al
Am J Kidney Dis. 2007;49:158-61.
Renal tolerability of CyA is reasonably good
when the dosage is low
Meyrier A, Expert Opin Pharm 2005

23. Long Term CyA treatment in SRNS
GE-ITALIAN STUDY Adult and Children
Ghiggeri 2004
• 55 steroid –resistant
NS treated with CyA
NS remission (partial or total): 20 patients
Mean follow-up: 81 months
Renal biopsy after 5 years of treatment:
no tubular or interstitial fibrosis

24. CHAPTER 4: STEROID-RESISTANT
NEPHROTIC SYNDROME IN
CHILDREN
4.2: TREATMENT RECOMMENDATIONS FOR SRNS
4.2.1: We recommend using a calcineurin inhibitor (CNI) as initial therapy for
children with SRNS. (1B)
4.2.1.1: We suggest that CNI therapy be continued for a minimum of 6
months and then stopped if a partial or complete remission of proteinuria
is not achieved. (2C)
4.2.1.2: We suggest CNIs be continued for a minimum of 12 months when
at least a partial remission is achieved by 6 months. (2C)
4.2.1.3: We suggest that low-dose corticosteroid therapy be combined
with CNI therapy. (2D)
Kidney Disease: Improving Global Outcomes
WWW.KDIGO.ORG

25. CHAPTER 4: STEROID-RESISTANT
NEPHROTIC SYNDROME IN
CHILDREN
4.2.2: We recommend treatment with ACE-I or ARBs for children with
SRNS. (1B)
4.2.3: In children who fail to achieve remission with CNI therapy:
4.2.3.1: We suggest that mycophenolate mofetil (2D), high-dose
corticosteroids (2D), or a combination of these agents (2D) be
considered in children who fail to achieve complete or partial
remission with CNIs and corticosteroids.
4.2.3.2: We suggest that cyclophosphamide not be given to
children with SRNS. (2B)
Kidney Disease: Improving Global Outcomes
WWW.KDIGO.ORG

26. Steroid-, cytotoxic- and cyclosporin-resistant
desperate NS:
rescue therapy
Steroid therapy : different doses,
different forms
 Cyclosporin late response late and
sustained effect,
cyclosporin dependancy, risk of toxicity
 Other calcineurin-inhibitors:
Tacrolimus

27. 22 children with steroid-resistant NS
(9 MCD, 11 FSGS, 2 MP)
TAC 0.1 mg/Kg/day (5-10 μg/ml TL)
Total remission10/13 resistant also to Cyclophosphamide(CP)
2/4 resistant also to Cyclosporine (CyA)
Side effects:diarrhea and hypertension (3 withdrawn)
TAC is an effective therapeutic modality for SRNS,
including children non responsive to CP and CyA

28. Mean time to remission : 60 days

29. Long-term outcome of children with steroid-resistant
nephrotic syndrome treated with tacrolimus
Roberti I, Vyas S. Pediatr Nephrol 2010, 25: 1117-24
19 children with steroid-resistant NS
(10 FSGS, 5 other forms)
Complete remission:11/19 (58%) , partial in 6
Mean time to remission : 8 weeks
Remission sustained during follow-up in 58%.
Among FSGF remission 50% ,
in 40% non responders, ESRF.

30. 41 children with steroid-resistant NS
TAC 0.1-0.2 mg/Kg or CsA 5-6 mg/Kg for 1 year
Alternative day steroids and ACE-i
(10 FSGS, 5 other forms)
At 6 mo Complete remission:TAC 85%, CsA 80%
RR for relapse after 1 year : OR 4.5 (TAC better)
Cosmetic negative effects in CsA only

31. Steroid-, cytotoxic- and cyclosporin-resistant
desperate NS:
rescue therapy
 Steroid therapy : different doses, different forms
 Cyclosporin late response late and sustained
effect,
cyclosporin dependancy, risk of toxicity

Other calcineurin-inhibitors:
Tacrolimus
 Purine synthesis inhibitors:
Mycophenolate

32. MMF and prednisone in steroid-dependent NS
Bagga A Am J Kidney Dis 2003
19 Children previously treated with P, oral CP,
and still cortico-dependent NS:
MMF 30 mg/Kg/day for 2 years associated with low tapering
doses of Prednisone. FU: 18 months
Frequency of relapses from 6.6 to 2 each year p<0.0001:
MMF was effective as steroid-sparing agent
75% reduction of relapses
After withdrawal relapse in 68% of the cases

33. MMF in steroid-resistant NS
Author
N
cases
Regimen
Efficacy
Day CJ
(Wolverhampt,
UK, NDT 2002)
7
adults
MMF (1 gx2)
Complete
remission 6/7
1/7 partial r.
MP pulses (15 mg/kg/week x 4-8)
ACE-i/ARB
MMF (250-500 mg/m2)
Proteinuria
(6-24 months)
72% below
baseline
p<0.01
Montané (Miami,
9
US, Ped Nephrol childr.
2003)
Mendizabal S
(Spain, Ped
Nephrol 2005)
Ulinski (Lyon,
Ped Nephrol
2005)
no response to CP and CyA
27
5 SRNS MMF (1200 mg/m2)
1/5 remission
Relapse after
withdrawal.
CyA with GFR impairment:
2g/1.73 m2
0/4 remissions
9
4SRNS

34. In FSGS
MMF should be used for > 6 months,
No RCT is available.
The rate of relapse is high.

35. CHAPTER 4: STEROID-RESISTANT
NEPHROTIC SYNDROME IN
CHILDREN
4.2.2: We recommend treatment with ACE-I or ARBs for children with
SRNS. (1B)
4.2.3: In children who fail to achieve remission with CNI therapy:
4.2.3.1: We suggest that mycophenolate mofetil (2D), high-dose
corticosteroids (2D), or a combination of these agents (2D) be
considered in children who fail to achieve complete or partial
remission with CNIs and corticosteroids.
4.2.3.2: We suggest that cyclophosphamide not be given to
children with SRNS. (2B)
Kidney Disease: Improving Global Outcomes
WWW.KDIGO.ORG

36. Steroid-, cytotoxic- and cyclosporin-resistant
desperate NS:
rescue therapy
sporadic case reports
Permeability factor (PF)
V.Savin 1993
PF is a small anionic protein
that binds to Prot A and
has analogies with
Immunoglobulins
Plasmapheresis

37. Plasmapheresis and protein A
immunoadsorption
Dantal et al (N Engl, 1994)
In native and in recurrent FSGS in grafted kidneys:
• Effect often limited in time, with relapse at withdrawal
• High cost / often limited benefits
• In cases with antiproteinuric response the
progression to ESRF is only partially limited

38. Recurrence of FSGS
on transplanted kidney
Plasmapheresis or
Immunoadsorbance
on A Protein
+ cyclophosfamide:
70% reduction in proteinuria
Lyon and Miami Protocol

39. Steroid-, cytotoxic- and cyclosporin- resistant NS:
sporadic case reports of
rescue therapy
B cells
as new target
for NS treatment?
Anti-CD 20
chimeric MoAb
Rituximab

40. Rituximab is a
Chimeric Monoclonal Ab:
Mouse IgG variable region
directed against CD20 Ag
+
Human IgG1 constant
region

41. A review of the current use of rituximab in autoimmune diseases
Gurvan HM Int Immunopharmacol 2008 Nov
Pharmacological effects:
B lymphocytes depletion
Reduced expression of
activated T cell markers

42. B and T
collaboration
T lymphocytes and NS
Hodgkin’s disease
Allergy
Viral infections
In vitro evidences
Shaloub’s hypothesis:
permeabilizing
T lymphokine
Permeability Factor (Ig part?)
B cells activated in relapse

43. Francois H, Daugas E, Bensman A, Ronco P.
Unexpected efficacy of rituximab in multirelapsing minimal
change nephrotic syndrome in the adult
Am J Kidney Dis. 2007;49:158-61.
Rituximab
375 mg/m2 x4
persistent
remission

45. Results
• Always effective in 15/15 proteinuria-free
patients
• Remission was induced in 3/7 NS
• In 19/22( 85%) one or more concomitant
immunosuppressive treatment was stopped

46. RTX was repeated in 12 patients who responded,
when CD19 count was >1% of total lymphocytes

48. Collaborative study (London, Tokyo, Toronto, Turin)
G1) steroid-dependent NS : (28 cases)
Complete remission: 61%
G 2) steroid-resistant NS (27 cases)
Complete remission: 22%
G3 post transplant recurrrence (15 cases)
Complete remission: 40%
Side effects 26%
Skin rash,bronchospasm, hypotension

50. Rituximab used in > 500.000 subjects
Adverse events:
Allergic reactions, anti chimeric Antibodies.
35
30
% HACA positive
-
25
20
15
10
5
0
0
3
6
9
12

51. Rituximab toxicity and adverse events:
progressive multifocal leukoencephalopathy (PML)
in 2 cases of SLE and one of RA
treated with multiple drugs , including Rituximab

52. 2 SLE, 1 RA, 2 autoimmune pancytopenia/thrombocytopenia
7 stem cells Tx, 26 purine analogues, 39 alkylating agents
52 lymphoproliferative disorders

53. The emerging role of podocytes

54. The interaction between synaptopodin and serine/treonine
binding protein maintains F-actin stability, normal cytoskeleton
structure and absence of proteinuria
F
actin
ser / treo BP
Dephosphorylation
synactopodin
synactopodin
P
ser / treo binding protein
sin
sin
sin

55. New treatments for
Nephrotic syndrome
protease inhibitors?

57. PROTEASE INHIBITORS
HIV
ANTI-RETROVIRAL DRUGS
transcription
viral proteins
viral capsids
PROTEASE
INHIBITORS

59. 8 patients in treatment with saquinavir 30 mg/Kg/day
• 6 males, 2 females
• Previous history of NS: 7.2 4.2 years
3 primary SRNS, 3 secondary SRNS
2 SDNS
• Previous treatments:
– Steroids, ACTH (8/8)
– Cyclophosphamide (4/8)
– Cyclosporine A (7/8), Tacrolimus (5/8), MMF (3/8)
– Plasma exchange (3/8)
– Rituximab (4/8)
• Median age at SAQ start-up : 13.5 (7-38) years
• Median duration of treatment: 14.7 (6-68) months

60. drug interaction
between saquinavir and calcineurin inhibitors
Pharmacokinetic
interactions
pGp
MDR2/3
CyP450
SAQ is substrate and inhibitor
Increase in
Cyclosporine A
and
tacrolimus
blood levels
SAQ is metabolyzed by
isoenzyme CYP3A4: modifies
CyA and TAC metabolism

61. Medium dosage of calcineurin inhibitors
administered together with saquinavir
• Cyclosporine A:
2 mg/kg/day
(vs 5 mg/kg/day before SAQ)
to maintain a blood levels of 100 ng/ml
• Tacrolimus:
0.01-0.06 (median 0.018) mg/kg/day
(vs 0.1 mg/kg/day before SAQ)
to maintain a blood levels of 3-5

63. Significant decrease in
cumulative steroid dosage
Medium dosage
of prednisone:
25.2 mg/kg/month
Prednisone mg/kg/month
35
Pre-saquinavir
Mean
mean
reduction
reduction
of 63%
of 56%
Post-saquinavir
100
30
75
25
p=0.03
20
50 %
15
10
25
5
0
0
12 months before SAQ
Last 12 months with SAQ
Medium dosage
of prednisone:
8.4mg/kg/month
Pre-SAQ
Post-SAQ

64. Nuclear binding of NF-kB p50- p65
in immortalized human podocytes
activated by LPS
without or with addition
of SAQ 10 and 20 µM.
Nuclear binding of NF-kB p50, p65
in immortalized human podocytes
activated by TNFα
with or without addition
of SAQ10 and 20 µM.

65. Conclusions
• The protease inhibitor SAQUINAVIR
provided with proteasome inhibitor activity:
1/2 primary SRNS
5/5 SDNS or secondary SRNS
Infrequent
relapsers
• This drug was active when associated with
calcineurin inhibitors, which had to be
reduced to less than one third of the original
dose to obtain safe blood levels.

66. saquinavir benefits:
hypotheses
Proteasome-inhibitor activity:
NF-kB hyperactivity inhibition of in circulating mononuclear cells
with decrease synthesis of a permeability factor (IPS downregulation)
Proteasome-inhibitor activity:
NF-kB hyperactivity inhibition in podocytes with
foot process rearrangement.
Direct impact on podocyte protein synthesis

67. saquinavir benefits:
hypotheses
Combined effect with low doses
of CNI and prednisone
NF-kB target for
Saquinavir (protease inhibitor)
Glucocorticoids (GCR binding to p65)
CNI (non competitive inhibitors of proteasome
& ubiquitinylation)

68. saquinavir benefits:
hypotheses
anti retroviral drug:
antiviral beneficial effect in NS?

70. A rescue therapy
may be tried
even in steroid resistant NS
CsA, TAC (MMF)
PE
Rituximab
new approaches :
Protease inhibitors
Antifibrotic drugs?

71. Thank you
Grazie
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