Focal segmental glomerulosclerosis
Medical Grand Rounds
Harbor
Irvine
USC
Jeffrey Kopp, MD
Primer on the National Institutes of Health
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extramural
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Training.nih.gov
NIH Program Level
in Nominal Dollars and Constant Dollars
“Doubling”
FY17 initiatives
Cancer moonshot
Precision medicine: million person cohort
Brain research
Lecture objectives
• Review common causes of nephrotic syndrome
• Appreciate that focal segmental glomerulosclerosis
is set of syndromes, with distinct mechanisms and
therpaies
• Understand the role of APOL1 genetic variants in
causing FSGS and related diseases
Causes of end-stage kidney disease in adults
most are diseases of the glomerulus

Diabetes
40%

Hypertension
30%

Glomerulonephritis
20%

Cystic
10%
ESKD causes more premature deaths than
most cancers
Incident ESKD cases by cause
Kidney diseases
ESKD
cases
Diabetes
48,157
Hypertension
29,105
Glomerulonephritis
9,978
FSGS, including
AIDS nephropathy
3,449
Myeloma kidney
1,215
Lupus nephritis
1,104
Cystic kidney diseases
Total
2,613
~100,000
USRDS 2008
Glomerular filtration barrier = endothelial cell +
glomerular basement membrane + podocyte
Renal blood flow ~1000 ml/min
Renal plasma flow ~600 ml/min
Filtration fraction ~15-20%
Glomerular filtration ~100 ml/min =
150 L/d
Tubular reabsorption ~99%
Glomerular ultrastructure (electron microscopy)
Urinary space
3
Blood
2
1
Model of slit diaphragm molecular assembly
Tryggvasson, NEJM 2006
Nephrotic syndrome: diagnostic features
• Proteinuria >3.5 g/d or spot urine protein/creatinine ratio ≥2 g/g
• Reduced serum albumin
• Hyperlipidemia
Serum albumin
5
4
3
2
1
0
0
12
24
36
Months
• Edema
48
60
Nephrotic diseases are podocyte diseases
Fogo, JCI 2011
Podocyte depletion > segmental glomerular scar
> global gomerulusclerosis
 GBM contacts Bowman capsule
(synechia)
 Glomerular filtrate is misdirected into
paraglomerular space, which enlarges
and traps additional capillary loops,
stressing additional podocytes
 Accumulation of matrix proteins
(mesangial cell?)
 Remaining capillary loops hypertrophy,
further stressing podocytes
Kriz, Kidney Int 1998
Mechanisms of glomerular injury: podocyte injury in context
Category
Specific mechanism
Clinico-pathologic syndromes
Autoantibody against glomerular antigen
ANCA vasculitis
Ag/Ab complexes depositing in glomerulus
Lupus nephritis
Membranous nephropathy
IgA nephropathy
Complement deposition
Post-infectious glomerulonephritis
Lupus nephritis
Membrano-proliferative GN
Other deposition
Immunoglobulin light chain
Serum amyloid protein
Amyloid nephropathy
GBM
Genetic, ab
Alport, Goodpasture
Endothelial injury
Complement, toxin, other
Thrombotic microangiopathies
Podocyte dysfunction (e.g.nephrin)
Genetic focal segmental
glomerulosclerosis (FSGS)
To be discovered!
Minimal change disease
Primary FSGS
Excessive glomerular load (e.g. obesity) or
insufficient glomerular number (e.g. premature
birth) Podocyte dysfunction
Adaptive FSGS
Podocyte dysfunction
Viral FSGS (HIV, CMV…)
Glomerulonephriitis
Podocyte injury
Focal segmental glomerulosclerosis
Barisoni
Map of FSGS, 1957
Terra incognita
Rich, Bulletin Johns
Hopkins Hosp, 1957
FSGS syndrome: six mechanistic groups
Primary: cytokine, immune activation
Mendelian genes >30
Virus: HIV, CMV, B19?, EBV?
?
APOL1
72% AA FSGS
Features of/mimics all other groups
Medication: IFN, Lithium,
Bisphosphonate
Adaptive FSGS
Mismatch: glomerular load >
glomerular capacity
FSGS course: Proteinuria response to therapy
predicts long term outcome
Complete remission
(20% of cases)
Partial remission
(40% of cases)
No response
(40% of cases)
Troyanov..Cattran, JASN 2005
Pathologic approach to FSGS:
Columbia classification decision tree



≥1 glomerulus with hyalinosis or sclerosis
IF limited to IgM and C3
EM immune deposits limited to mesangium
Collapsing variant?
Tip lesion?
Cellular lesion?
Perihilar variant?
Not Otherwise Specified
(NOS) = classic FSGS
D’Agati, Fogo, Bruijn Jennette,
AJKD 2004
Clinical relevance of FSGS classification:
frequency and course
Variant
Frequency
(total N=192)
Collapsing
variant
11%
Tip lesion
17%
Cellular
lesion
3%
Perihilar
variant
26%
NOS
42%
Patients at risk …
Tip
n = 34
Perihilar
50
NOS
78
Collapsing 20
Thomas, KI, 2006
Tip
Perihiilar
NOS
Collapsing
26
35
47
12
Renal Survival (years)
22
21
33
5
19
12
17
3
11
4
6
3
Mechanistic approach to FSGS
Mechanism
Primary
Adaptive
APOL1
Mendelian
Genetic
Medication
Virus
Subtypes
Most cases: idiopathic
suPAR?
Cardiotrophin-like cytokine?
Mismatch hypothesis:
Decreased glomerular number/hypertrophic
capacity and increased metabolic load
(FSGS, HIVAN, arterionephrosclerosis)
Monogenic: >30 genes (inheritance AR, AD,
mitochondrial)
Pamidronate (collapsing)
Interferon a b or g, lithium
CNI, amphotericin (adaptive)
HIV, CMV, B19? EEBV?
Biopsy report returns FSGS
 Historical data: family history, birth history
(gestational age, weight)
 Renal biopsy findings
 Laboratory data: proteinuria, serum
albuminuria
 Response to therapy
Primary FSGS: characteristics
 May have a characteristic presentation
Distinct or even explosive onset
Heavy proteinuria (>4 g/d)
Low serum albumin (<3 or lower)
Marked hyperlipidemia
 Light microscopy: tip lesion or collapsing
glomerulopathy suggestive
 Electron microscopy: widespread foot process
effacement
 Role of circulating plasma factors
Recurrent FSGS after kidney transplant
 ~25%. Risk factors: FSGS onset <6 yr, rapid
progression to ESKD (<3 yrs), and to a lesser degree
white race, LD transplant
 Bioassay for the elusive factor (rat glomerular albumin
permeability in vivo) is not useful for patient care: not
sufficiently predictive
Candidate molecules: soluble urokinase-type
plasminogen activator receptor (SUPAR), cardiotrophin-
like cytokine factor-1 (CLCF1)
 Prevention: plasma exchange?
 Treatment: plasma exchange? Cyclophosphamide?
Rituximab?
Adaptive FSGS: biopsy findings
 Large glomeruli
 Perhilar sclerosis: ≥50% of
sclerotic glomeruli have
sclerosis confined to hilar
quarter of glomeruli (sensitivity
and specificity unknown)
Obesity-related FSGS
 Only partial foot process
effacement, mean 40% vs 75%
but much overlap
 Mismatch: increased load
versus decrease increased
filtration capacity > increase
single nephron glomerular
filtration
Controls
Khambam..D’Agati, KI, 2001
Helal ..Schrier, NRN 2012
Adaptive FSGS: clinical features
 Premature or small for gestational age
 Nephrotic or sub-nephrotic proteinuria
 “Bigness” - obesity or tall/muscular
 May have preserved serum albumin
 RAAS antagonism: proteinuria may fall >50%
CNF, FSGS, DMS: 25 genes are implicated, functioning in 7
cellular compartments/functions
Genes
Cell Matrix
Slit
diaphragm
Cytoskeleton
Mitochondria
DNA repair transcription
Cell
signaling
Lysosome
2
5
5
7
6
2
1
NPHS1
ACTN4
INF2 *
WT1 *
PLCE1
NPHS2
INF2 *
NEIL1
TRPC6
CD2AP
AHRGP24
PTPRO
AHRGDIA
Nonsyndromic
* = appear > once
MYO1E
Syndromic
ITGB4
MYH9 Epstein,
Fechtner
LAMB2
Pierson
INF2 CharcotMarie-Tooth *
WT1 DenysDrash, Frasier *
tRNA –leucine
MELAS
LMX1B
Nail-patella
COQ2
SMARCAL1
Schimke
COQ6
NXF5
PDSS2 Leigh
Most recessive, 8 autosomal dominant,
1 mitochondrial DNA
ADCK
SCARB2
Action
mycoclonus
Cellular pathways and podocyte genetics: 7 known
and 5 pathways could be speculated
Cell signaling
Slit diaphragm
Cell/matrix interaction
Nucleus
Lysosome
Mitochondrion
ER stress –
unfolded protein response
Secretory pathway
Cytoskeleton
Endocytosis
Cell polarity
Autophagy
Genetic testing depends on clinical context
Setting (onset age)
Non-syndromic
Syndromic
Infant (<1 y) with nephrotic NPHS1, NPHS2, LAMB2, As suggested by
syndrome
WT1, PLCE1
clinical findings
Child 1-10 with SRNS/FSGS
NPHS2, LAMB2, WT1,
PLCE1
“
Child with SSNS/FSGS, with PLCE1, PTPRO
+ FH
- Child/adolescent 10-18
with SRNS
- Adult with FSGS and
family history
INF2, ACTN4
TRPC6, WT1
Adult lacking family history Generally not tested
“
“
Hinkes, Pediatrics 2007; Ismaiil Ped Nephrol 2009; Santin, CJASN 2011; McCarthy, CJASN 2013
Why test for FSGS genetic mutations?
• Extra-renal manifestations
• Prognosis
• Therapy. Generally steroids don’t work. Calcineurin
inhibitors may work. RAAS antagonist.
• Kidney transplant: FSGS recurrence is unlikely
• Family screening
• Every patient/family deserves a molecular diagnosis,
when feasible (Hildebrandt)
FSGS treatment algorithm
 Primary FSGS >2-3 g/d
despite RAAS antagonist +
 Primary FSGS, <2 g/d or
diuretic and preserved GFR
risk for treatment toxicity
(>40-50 ml/min/1.73m2 ?)
 Genetic FSGS
 Adaptive FSGS
 Daily prednisone
 CSA or tacrolimus
ACEI or ARB ± Aldo blocker  MMF
Blocking RAAS: Renin-angiotensin-aldosterone
and more
PTXF
mAb
PAI-1
Leptin
CTGF
ARB
Ang
Ang I
Angiotensin
Receptor I
Ang II
ACEi
Aliskeren
Renin
Angiotensin
converting
enzyme
PTXF
TSP-1
NEP
Aldo
Spironolactone
Eplerenone
TGFb1
ET-1
Bosentan
Pirfenidone
Tranilast
fresolimumab
FSGS: Time to remission (complete +
partial) with steroid Rx
 Dose 1 mg/kg up to 80 mg
 Continue 8 – 16 wk
 Proteinuria reduced by 50%:
slow taper
 Proteinuria not reduced: rapid
taper
 Alternatives
80 or 120 mg QOD
Pulse oral dexamethasone:
25 mg/m2 d1-2-3-4, repeat
q28d X 8 cycles (Cho, submitted)
Rydel
AJKD 1995
ESKD incidence ~100,000 cases/yr:
Major African American health disparity
ESKD Cause
All causes
3.4
Diabetes
3.7
Hypertension-attributed
5.3
Glomerular disease
2.7
 Lifetime risk for ESKD:
WF 1.8%, WM 2.5%, BM 7.3%, BF 7.8%
Kiberd, JASN 2002
AA:EA
incidence
ratio
FSGS
4.5
AIDS nephropathy
20
USRDS, 2007 report
2004-2006 incidence rates,
adjusted for gender
Hypothesis
 The increased risk for idiopathic FSGS and HIV-associated collapsing
glomerulopathy (HIVAN) among African Americans is due to an African
origin genetic variant
Assumptions
 Ancestral populations from Africa, Europe, Asia differ in prevalence of FSGS
risk alleles
 Admixture mapping can find the gene by using SNPs that tag African vs
European origin genetic segments
Analysis: among cases, the causative gene should be in a genomic region that
preferentially came from Africa
C22
C22
Cases:
Preferentially
African segments
Controls:
African and
European
segments
Admixture mapping in African American FSGS identified a
peak of African ancestry: MYH9 ->APOL1
Kopp..Winkler, Nat Genet 2008
Kao…Parekh, Nat Genet 2008
FSGS association
S342G, I384M (G1)
180 African American FSGS cases
and 205 African American controls.
Genovese…Pollak, Science 2010
Tzur…Skorecki, Human Genet 2010
Controlling for rs73885319 (G1)
NYK388K (G2)
Controlling for G1 and G2
Apolipoprotein L1
Pays..Perez-Morga, Nat Rev Microbiol, 2014
Pore forming
Membrane
addressing
SRA
interacting
 7 exons, 3 isoforms, 398 – 414 amino acids
 Component of innate immune system, driven by IFN-a
Friedman..Pollak, Kidney Int, 2014
 Minor component of HDL subclass 3
 Binds phospholipids, induces autophagy
Wan..Hu, JBC 2008
The Apolipoprotein L gene family evolved
Figure 2
rapidly in primates
APOL1
APOL2.1
(Macaque)
(Orangutan)
Gorilla
Human
Island Siamang Gibbon
Macaque
(Baboon #2)
(Baboon)
APOL4
Baboon
Macaque
Patas Monkey
African Green Monkey
Pseudogenes in
parentheses
Human
(Gorilla)
(Orangutan)
Gorilla
Human
Chimpanzee
Orangutan
(Island Siamang Gibbon)
APOL7
(Baboon)
(Rhesus)
APOL3
0.96
1.00
1.00
1.00
1.00
Human
Chimpanzee
APOL2
Gorilla
Kikuyu Colobus
Purple-Faced Langur
Patas Monkey
Baboon
Douc Langur
African Green Monkey
Macaque
Hominoids
Old World Monkeys
New World Monkeys
Prosimians
0.1
1.00
APOL6
APOL5
(Lemur)
(Macaque)
(Baboon)
(African Green Monkey)
(Douc Langur)
(Kikuyu Colobus)
Smith and Malik, Genome Research, 2009
(Orangutan)
Gorilla
Chimpanzee
Human
Lemur
Gorilla
Human
Chimpanzee
Orangutan
Island Siamang Gibbon
White-Cheeked Gibbon
Pygmy Marmoset
White-Tufted-Ear Marmmoest
Tamarin
Squirrel Monkey
Owl Monkey
Titi Monkey
White-Faced Saki
Spider Monkey
Kikuyu Colobus
Douc Langur
African Green Monkey
Macaque
Baboon
APOL gene sets vary among primates:
A flexible tool kit for innate immuty
✚
Human
−
Chimpanzee
Ψ
Gorilla
Orangutan
Ψ
Macaque
Ψ
Baboon
Mouse
Serum lytic to
T.b.brucei
Ψ
✚
Ψ
−
Ψ
−
Ψ
✚
−
Adapted from Smith and Malik, Genome Res. 2009
APOL1 alleles: G1, red and G2, orange
Kopp..Winkler, JASN 2011
Trypanosoma brucei and African Sleeping Sickness
T.b.gambiense
T.b.brucei
T.b.
rhodesiense
APOL1 is the Trypanolytic Factor of Human Serum
Alternate: Plasma membrane cation
channel. Thomson, PNAS 2015
Pays..Perez-Morga, Nat Rev Microbiol, 2006
APOL1 is under balancing selection
0 risk allele
G0/G0
(WT/WT)
1 risk allele
G0/G1
G0/G2
Homozygous
disadvantage
Heterozygous
advantage
Trypanosomiasis
(African sleeping sickness)
Resistance
2 risk alleles
G1/G1
G1/G2
G2/G2
Homozygous
disadvantage
Resistance
Chronic kidney disease
APOL1 nephropathies
Disease/setting
Phenotype/ effect size
Reference
HIVAN
OR 29 (US), 89 (SA)
Kopp..Winkler, JASN 2011
Kasembeli..Naicker, JASN 2015
FSGS
OR 17
Kopp..Winkler, JASN 2011
Arterionephrosclerosis > ESKD
OR 7
Lipkowitz..Kopp, KI 2012
Deceased donor genotype -> kidney
allograft survival
Allograft loss, HR 3.8
Reeves-Daniel..Freedman, Am J
Transplant 2011
Recipient genotype -> kidney
allograft survival
No effect
Lee..Chandrakar, Am J Transplant,
2012
Sickle cell
Proteinuria, RR 3.4
Ashley-Koch..Telen, Br J Heme 2011
Lupus
Collapsing glomerulopathy, OR 5.4
Larsen..Walker, JASN 2013
APOL1 FSGS can manifest clinically as
four different mechanistic forms
Primary FSGS
Adaptive glomerular
processes
Viral FSGS
Drug
Associated
FSGS
Can be steroid
responsive
Sickle cell disease
>CKD
(no biopsies)
HIVAN
OR: US 29
OR: South Africa 89
Interferon therapy
Kopp..Winkler, JASN
2011
Ashley-Koch..Telen, Br J
Hem 2011
Kopp. .Winkler JASN
2011
Kasembeil..Naicker, JASN
2015
Nichols..Friedman, KI
2014
Can recur after
kidney transplant
CKD associated with
prematurity
Unpublished
observation
Unpublished (CKID,
NEPTUNE)
APOL1-associated FSGS:
sharper incidence peak - spanning ~15-45 yr
APOL1 2-risk allele individuals with CKD tend to
progress quickly despite conventional therapy:
therapeutic need
Percent survival
100
80
60
0/1 RA
40
p = 0.001
2 RA
20
0
0
5
10
15
Years since FSGS onset
FSGS, NIH
Deceased donor
transplant, Wake Forest
FSGS, Controlled Trial
AASK: hypertension
Approximate lifetime risks for APOL1 nephropathy
~43 million African Americans
~5 million (~13%) have 2 APOL1 risk alleles
Among those, perhaps 20-25% (over 1 million) will develop a renal disease
or renal phenotype associated with APOL1 variants
FSGS
4%
5 million at-risk individuals:
estimated life-time risks
HIVAN
HA-ESKD
11%
OR 17
Lupus CG,
ESKD
HA-CKD
(not ESKD) ?
OR 7
Kopp..Winkler, JASN 2011
Lipkowitz..AASK Investigators, Kidney Int, 2012
Dummer..Kopp, Sem Nephrol, 2015
Nephropathy in type 2
diabetics: rapid progression
Proposed pathways of APOL1 variant toxicity:
mechanisms may lead to targeted therapy
NLRP12 suppression
NLRP3 activation
Cation channels
Amplification
of TLR4 and
IL1R signaling
Inflammasome
activation
G1 and G2 RNA
stem loops
Protein kinase R
activation
JNK/MAPK
IL-1b
Altered
endolysomal
trafficking and
lysosomal function
Reduced protein
synthesis
Podocyte injury & loss