SLE therapies Flashcards
1
Q
what do these small molecule drugs tend to target?
A
- unlike targeted-therapies, these drugs have broad mechanisms of action and target multiple cell types
- tend to have broad immunomodulatory effects, targeting inherent cellular processes rather than specific cytokines
- often have off-target effects
- good for diseases where we don’t fully understand the pathogenesis
2
Q
how are SLE patients broadly managed?
A
Determine prognosis – mild disease vs severe (Not specific)
- all patients should avoid sunlight, exercise, stop smoking
- Topical steroids good for rash
Mainstay of therapy depends on severity of disease
- Most patients have antimalarials and mild immunosuppressants e.g. for mild joint symptoms and rash, tend to use HCQ alone
- As severity increases with major organ involvement e.g. kidney disease, neurological disease, need potent immunosuppression cyclophosphamide, MMF, biologics
Pick drugs based on severity of disease
- Strong immunosuppression for severe, mild immunosuppression for less severe
3
Q
what is hydroxychloroquine?
A
Anti-malarial drug used for >50 years in SLE
Especially helpful for:
Skin disease, Joint disease, Thrombosis? (new evidence)
generally recommended for all SLE patients
4
Q
is HCQ useful in reducing organ damage?
A
Organ damage
- Irreversible change in organ e.g. liver fibrosis, end-stage kidney disease, lung fibrosis
- In a large multi-ethnic prospective cohort, hydroxychloroquine is associated with reduced rate of organ damage and death
HCQ reduces risk of organ damage and death
5
Q
what is HCQ mechanism of action?
A
2 parts:
1. prevents presentation of self-antigen on cellular surfaces
- Direct effect on TLRs
6
Q
how does HCQ reduce self-antigen presentation?
A
HCQ is accumulated into lysosomes and directly changes pH
- this pH change affects TLR signalling and antigen processing - less supported theory
HCQ is lysosomotropic, so accumulates in lysosomes and endosomes – interferes with processing of cellular material – reduces effects of autolysosome so affects autoantigen processing – reduces presentation of self/autoantigens on surface of cells via lysosomal pathway
- modifies effects of lysosomal enzymes
7
Q
how does HCQ directly interfere with TLR signalling?
A
OHCHQ can accumulate in endosomes which express TLR7,8,9
- these TLRs are important in SLE pathogenesis - genetic defects in TLR7 is risk allele for SLE development
- HCQ changes in endosomal pH and interferes with recognition of DNA/RNA immune complexes by TLRs
- therefore HCQ reduces TLR signalling and reduces production of cytokines, particularly IFN1
As well as change pH of endosome, HCQ binds ssDNA and directly interferes with binding of nucleic acids to TLR9 – prevents direct TLR activation
- Reduces IFN1 production
- Reduces self-antigen presentation
- Reduce autoantibody activity to RNA/DNA
8
Q
what other cell signalling pathway does HCQ interfere with?
A
There are nucleic acid sensors in the cytoplasm too, not just endosomes
- cGAS recognises cytoplasmic nucleic acids – HCQ also affects this
9
Q
how can HCQ have immunomodulatory function on a cellular level?
A
Different mechanisms of HCQ on broad immunomodulation
- Reduces antigen presenting cells and B cell antigen processing
10
Q
what is the main problem with HCQ?
A
Retinopathy – probably related to accumulation of drug in lysosomes of retinal cells
- Causes damage to macula (centre of retina) - most important for visual acuity
- Causes dark central area with light area around – bulls eye maculopathy
- Some patients have increased risk depending on dose and duration of drug
- Poor kidney function means HCQ can’t be cleared well - increased risk
- Tamoxifen also causes increased risk – used in breast cancer
11
Q
what can cause increased risk of HCQ toxicity?
A
Duration of treatment and cumulative dose increase toxicity
Duration:
- Retinal problems never occur before 5 years of treatment
- between 5-10 years = greatest retinal problems
- Longer duration = less problematic – likely because the people who were predisposed to getting renal toxicity may have already had it by now and so stopped drug
Cumulative dose: by the time of taking a kilo of HCQ over the course of life - retinal problems
12
Q
how is HCQ toxicity monitored?
A
Patients who take HCQ for 5 years or more, their retinas are monitored with CT scan on annual basis
- Any abnormality = stop drug before problems
- CT scan detects retinal issues before it affects vision – very sensitive
13
Q
what are the HCQ screening guidlines?
A
- dose based on weight
- baseline screening, and then annually after 5 years of treatment
- if they have risk factors they are screened from the beginning
- no treatment for retinopathy, just stop the drug
14
Q
is HCQ cardio-toxic?
A
- Reports of arrhythmia in patients receiving OHCHQ as part of COVID-19 studies - when combined with azithromycin, HCQ has increased risk of arrhythmia by prolonging QTc interval, but rare - but the doses in COVID studies were much higher than in RA or SLE, so not too worrying
- Direct cardiotoxicity (demonstrated on histology) is very rare - Can cause direct cardio myopathy, causing heart failure - Rare but specific appearance on histology
15
Q
summary of HCQ:
A
Hydroxychloroquine is an “anchor” drug in the
management of SLE
Modifies TLR signalling and reduces presentation of self-antigen
Cumulative risk of retinal toxicity
16
Q
what approach should be taken for treating severe SLE e.g. active renal lupus?
A
Treating for active lupus
- Strong induction medication to induce period of remission
- Then maintained with weaker doses or lower immunosuppression
similar to cancer treatment
17
Q
what is cyclophosphamide?
A
Derivative of mustard gas – nerve toxin
- alkylating agent
- Forms inter- and intra-strand crosslinks within DNA at guanine N-7 position – crosslinking and cell death
- Toxic drug
- Metabolised into multiple compounds, some of which have biological activity
18
Q
what compound of cyclophosphamide is concerning?
A
Acrolein is a normal derivative of cyclo, excreted from kidneys and accumulates in bladder
- Can inflame bladder wall in dehydrated people, causing bleeding in bladder lining - haemorrhagic cystitis
- Drugs can bind acrolein in the bladder and help expel it
- This is usually self-limiting
- But, high dose of cyclo during life span can increase risk of bladder cancer – toxic drug, but sometimes is the only thing that works
19
Q
is cyclophosphamide effective in SLE? what are the limitations?
A
Compared to steroids alone, cyclophosphamide:
- Reduces development of renal scarring
- Reduces risk of relapse
- Reduces risk of permanent kidney damage and lupus relapse
Increased risk of infertility as it kills oocytes – related to total exposure and increasing age
- Low dose regime used now to reduce toxicity:
- Low-dose pulsed/intermittent regime preferred – as effective as high dose pulsed but
- Less infertility
- Fewer in infections
20
Q
how does cyclophosphamide work?
A
Inhibits cell replication
- Preferentially effects lymphocytes in autoimmune conditions
21
Q
what is mycophenolate mofetil (MMF)?
A
Prodrug of mycophenolic acid (MPA), MMF is the active agent
Inhibitor of inosine monophosphate dehydrogenase (IMPDH) - 2 isoforms:
- Type 1 isoform of IMPDH in most cell types
- Type 2 isoform is preferentially expressed in lymphocytes and proliferating cells
- MPA is 5x more potent in inhibiting type 2 isoform
22
Q
what are the effects of MMF on lymphocytes?
A
reduces proliferation
reduces Ig production by plasma cells
reduces lymphocyte/monocyte recruitment to sites of inflammation
23
Q
what do replicating cells require?
A
For replicating cells, purines are needed e.g. adenosine and guanosine
- these need to be phosphorylated to generate ADP, ATP for RNA, and deoxy versions for DNA
24
Q
what are the pathways of purine biosynthesis?
A
Guanosine nucleoside forms GTP in RNA, or dGTP in DNA - guanosine can be formed via 2 pathways:
- De novo pathway: synthesised directly from ribose-5-phosphate to inosine MP
- Salvage pathway from guanine using PRPP enzyme
25
how was MMF developed from purine biosynthesis?
In people with defect in PRPP, they have Lesch-Nyhan syndrome – associated with abnormal brain development but normal lymphocytes
- Shows that the salvage pathway isn’t essential for lymphocyte function or development
In people with adenosine deaminase (ADA) deficiency, there are reduced B and T cell numbers
- ADA moves adenosine reversibly to form Inosine MP - this is essential for normal lymphocyte development
- if there is accumulation of dADP, this is toxic to cells, so need to reduce its accumulate
MFA inhibits IMPDH (this enzyme moves inosine MP to guanosine)
- this means that the inosine MP pathway can only go towards adenosine production, leading to dADP accumulation, and impairs guanosine production via the de novo pathway, leading to impaired proliferation of lymphocytes
26
is MMF effective in SLE?
MMF studied mainly in kidney lupus, also in systemic
- MMF is as good as cyclophosphamide (53% response)
- 56% response rate for MMF and much less toxic than cyclophosphamide
- MMF can induce remission in lupus nephritis over 24 weeks
27
is MMF used in maintenance therapy?
MMF induces remission and maintains it
- Compared MMF to azathioprine after cyclophosphamide for induction therapy
- Rates of flare worse in AZA (25%) and more cytopenia compared to MMF (flares 19%)
- MMF is a key drug in maintaining remission
28
how may azathioprine and MMF have disease-specific effects?
In other kidney diseases e.g. vasculitis (inflammation of blood vessels, cause renal disease), AZA is better than MMF
- MMF better in kidney disease of SLE compared to vasculitis – different diseases
29
summary of cyclophosphamide and MMF
Cyclophosphamide an alkylating agent, used in the management of severe SLE, especially renal and neuropsychiatric disease
MMF inhibits IMPD, preferentially inhibiting nucleic acid synthesis in lymphocytes
MMF is as effective as cyclophosphamide in inducing renal remission, and is less toxic
both of these target DNA of replicating immune cells
30
what is azathioprine?
AZA used in transplant, IBD
- Metabolised to active compound 6-mercaptopurine
31
what affects AZA metabolism? how does this affect clinical use?
Genetic predispositions affect how AZA is metabolised - pharmacogenetics
- TPMT thiopurinemethyltransferase) metabolises AZA
- TPMT is under allelic control
- 10% of population have reduced TPMT activity due to genetic variation – less AZA metabolism – more toxicity
- Normal practice to measure TPMT activity assay before giving drug
- Genetic pharmacokinetics
1/300 have double null TPMT alleles – high risk of toxicity, so AZA is not given
32
does AZA have a wide therapeutic index?
Therapeutic index = distance between no effect and toxicity when in circulation
- Drug levels within this will likely be effective with no toxicity
- Narrow index = drug levels have a narrow window – easy to be toxic or sub-therapeutic, e.g. 6-merc - not ideal
- AZA has wider therapeutic index than 6-merc
33
how is AZA metabolised? (2 pathways)
AZA is cleaved to 6-merc (active form)
Then 6-merc goes down 2 pathways:
1. development of 6-MMP, driven by xanthene oxidase
2. development of 6-thioguanine nucleotides (6-TGNs), thio analogues of guanine, mono, di and tri-phosphates (TGMP, TGDP, TGTP), driven by HPRT
Can measure the metabolite levels in the clinic
34
what are the activities of the AZA metabolites? how does this affect clinical use?
Both compounds biologically active but have different effects:
- 6-TGNs are important in inhibiting DNA synthesis and cell proliferation - immunosuppression activity
- TGNs reduce T cell migration and survival, reduce macrophages iNOS production and reduce adhesion molecule expression on endothelial cells
- 6-MMP doesn’t cause immunosuppression but can cause liver inflammation
Need to balance these pathways, so that patients go down 6-TGN route, and less so to 6-MMP
- but not too much 6-TGN as it can be toxic
35
how are 6-TGNs immunosuppressive?
Mechanism 1: (broad)
- Incorporated into DNA of proliferating cells
- Stops IL-2 mediated T cell proliferation
- Induces T cell apoptosis
Mechanism 2: (complex, affects activation of Tmem cells) - interferes with CD28 signalling
- 6-TGTP competes with GTP binding Rac1 downstream of CD28
- 6-TGTP leads to ↓ of Bcl-XL (which is anti-apoptotic)
- Predominantly inhibits CD45RO memory T cells after CD28 activation
36
where has AZA been used clinically?
SLE (typically non-renal)
RA
Vasculitis
Inflammatory bowel disease
Transplant
Multiple sclerosis and myasthenia gravis
37
how can AZA be optimised for use in patients?
6-TGNs have clinical effect but can cause BM suppression
6-MMP causes liver inflammation and lacks clinical effect
20% patients (6-MMP shunters) favour 6-MMP pathway – drug won't work
- Xanthene oxidase can be inhibited by allopurinol and increases activity of HPRT to drive pathway to 6-TGN
- This can also reduce AZA dose to avoid liver toxicity
- combination of low dose AZA (25%) and allopurinol favours 6-TGNs over 6-MMP - not common
- Need to balance this to avoid BM suppression
38
has AZA with allopurinol been successful?
been mostly used in IBD
- Allo with AZA = 6-TGN increases – almost doubles the 6-TGN metabolites and reduces 6-MMP compared to AZA alone
39
what are calcineurin inhibitors?
Cyclosporin A and tacrolimus most commonly used - inhibit calcineurin
- Cyclosporin binds cyclophilin A
- Tacrolimus binds FKBP
40
what is the mechanism of action of calcineurin inhibtors?
When APCs signal to T cells via MHC-TCR, this increases calcium IC levels which activates calcineurin-calmodulin complex, which dephosphorylates and drives NFAT activation to transcribe IL-2
- Calcineurin also leads to degradation of IκB (which frees up NFκB → cytokine production
Inhibition of calcineurin means less dephosphorylation of NFAT and less IL-2 production
- T cells are particularly susceptible to calcineurin inhibition
41
what are the effects of calcineurin inhibitors?
↓ T cell activation (↓ TNFα, IL-1β, IFNγ, IL6, IL10)
↓ B cell activation, class-switching and IgG production
May also independently reduce protein loss by kidneys by stabilising podocytes – used in kidney disease independently of T cells) - favoured in treatment of kidney disease during inflammation of glomerulus
42
why is cyclosporin difficult to use/manage?
Narrow therapeutic index:
- Poorly absorbed – transport by P-glycoprotein (P-gp: transporter that sits on surface of enterocytes in bowel)
- As drugs are absorbed from lumen of bowel across enterocytes into blood, p-gp secretes drug from enterocytes back into bowel, so cyclosporin poor bioavailability.
- Drugs which inhibit p-gp can enhance cyclosporin absorption, but can result in toxicity
- Metabolised in liver by CYP3A into >30 metabolites which have biological effects – hard to manage
- also, it interacts with many other drugs
43
how can cyclosporin overlap with other drugs?
Increased cyclosporin: high dose steroids, warfarin, macrolide antibiotics
Decreased cyclosporin: anticonvulsants, anti-TB treatment
20% of SLE patients have anti-phospholipid sYndrome with stickly blood and increased risk of blood clots, so given warfarin to thin blood
Warfarin, antibiotics and steroids can increase cyclosporin levels in blood
44
is tacrolimus better than cyclosporin?
tacrolimus is better
- isn't affected by p-gp
- wider therapeutic window
- fewer active metabolites
45
what are the common adverse effects of calcineurin inhibitors?
GI pain, diarrhoea
Heart burn
Increase diabetes risk
Increase blood pressure
Excessive hair growth
Swollen gums – gingival hypertrophy
Toxic to kidneys:
- Can reduce protein leakage but reduces kidney function
- Complex effects
- Reduces nitric oxide and activates renin-angiotensin system – constricts blood vessels inside kidneys, which can reduce blood flow to kidneys and induce ischaemia and inflammation of kidneys
not used in patients with kidney impairment
46
summary of AZA and calcineurin inhibitors
Azathioprine modifies T cell adhesion and migration
Efficacy and toxicity balance of azathioprine is affected
by TPMT activity and “shunting”
Allopurinol can favour 6-GTNs but need to watch for bone marrow suppression
Calcineurin inhibitors inhibit transcription downstream of TCR activation
Modulates B and T cells and possible podocytes
Ciclosporin especially has narrow therapeutic index
47
how are Tregs affected in SLE?
Patients with SLE have reduced Treg numbers
- Variations in Treg populations and how they are defined by surface markers
- Reduced % of CD25highFoxP3+ Treg in SLE but functionally normal
- Increase in other Treg populations: CD25lowFoxP3+ and CD25-FoxP3+
- % CD4+CD25high inversely related to disease activity: with increasing disease activity, the fewer Tregs there are
- Maybe they are more sensitive to apoptosis
CD4+CD25+ Tregs lower in active lupus compared to inactive SLE
- In inactive SLE, Treg levels are similar to healthy controls
No differences in Tregs in sjorgens – this is an SLE-specific change
48
what is IL-2?
IL-2 is important for the development of CD25+CD4+ Treg cells - they depend on IL-2
- Up-regulates CTLA-4 (negative regulator of T cell function)
- Required for differentiation of T cells into TH2
(may also have some immunostimulatory properties)
- Low dose of IL-2 may have an immunomodulatory role: Use low dose IL-2 to induce Treg formation
- At high doses, can be immunostimulatory
49
how has recombinant IL-2 (rIL-2) been studied in mice?
low dose rIL-2 reduces disease activity in lupus-prone mice
- Both anti-CD25 and anti-IL-2 antibodies exacerbate disease in lupus mouse models
- (3 injections of low dose rIL-2 to (NZB x NZW) F1 mice):
- Increases number of CD25highFoxP3+ Tregs
- Increases Treg proliferation (BrdU marker) in the spleen
- Reduces proteinuria – improves renal disease in mice with reduced protein in urine
- Improves survival
50
has IL-2 been successful in humans?
RCT of low dose IL-2 in 60 SLE patients - early study
- 60 patients randomised to IL-2 or placebo
- IL-2 (1x106 IU) alternate days for 2 weeks then 2 week break (x3)
- Primary endpoint of SRI-4 at 12 weeks
- Gave IL-2 in 2 week blocks given every other day – weird dose regime
- Then gave 2 week break
- Grey bar = 2 weeks treatment, white bar = 2 weeks rest - Off-on treatment regime
- Improved SRI-4 in 12 weeks, but not statistically significant, but still big difference in response rate 55% vs 30%
51
what happens biologically with the off-on treatment regime of IL-2 in humans?
- Tregs go up in treatment, and down in break
- To maintiain high levels of Tregs, persistent IL-2 dose is needed
- Similar effects on NK cells
- No. CD4 and CD8 cells didn’t change
Specific way of increasing Treg numbers but needs to be maintained
52
what are the effects of IL-2 on CD8 cells?
Required for
- CD8+ T cell function
- Development of memory CD8+ T cells
- Activation-induced cell death (AICD) → down-regulation of expanded T cell clones: Via up-regulation of Fas, FasL and Fas-associated death domain (FADD) which leads to activation of caspase-8 – reduces T cell expansion with low-dose IL-2, independently of its effect on Tregs
53
summary of IL-2 in SLE:
IL-2 is essential for the development of regulatory T cells (but may have other immuno-regulatory functions)
SLE patients have lower % of Tregs
Treg can be induced by IL-2 in vivo and ↓ disease activity in mouse models of SLE
2 small RCTs did not meet primary endpoints but may be a problem with dosing and/or trial design
54
are are Ikaros and Aoilos?
Zinc-finger protein transcription factors
- expressed in common lymphoid progenitors (CLP)
Ikaros: IKZF1 regulator of CLPs, acts as a checkpoint for pro B cell → pre B cell (early B cell dev)
Aoilos: IKZF3 needed for memory B cell and plasma cell formation (late B cell dev)
55
what are CELMoDs?
Cereblon 3 Ligase Modulatory Drugs - induce targeted protein degradation
- Rely on E3 ubiquitin ligase complex which induce degradation of other proteins via ubiquitin ligation
- Small molecules CELMoDs bind E3 complex to other proteins to ubiquitinate the protein and induce degradation
- Preferentially targets 2 transcription factors: IKZF1 (Ikaros)
IKZF3 (Aiolos) (neosubstrates), where CELMoD binds the TFs and marks for degradation via:
→ ubiquitination → degradation of the TFs
56
how are Ikaros and Aoilos implicated in SLE?
SNPs in IKZF1 associated with SLE susceptibility, malar rash, and cytopenia
A SNPs in IKZF3 associated with SLE susceptibility in Chinese patients
SNPs in both associated with SLE susceptibility
57
what is Iberdomine in SLE?
Iberdomide (CELMoD) reduces development of B cells:
- Inhibits TLR7- and IFN-mediated B cell activation
- Inhibits TLR7-mediated differentiation of plasmablasts
- Reduced CD19 and CD20 B cells, no effect on NK cells or CD4 cells, small increase in CD8 cells – predominantly effects B cells
- Stops differentiation of plasmablasts
- Tablet because it’s a small molecule
58
what is the pDC-IFN axis in SLE? how can it be targeted?
pDC produces IFN1, which affects B cells, plasmablasts
- Stimuli for IFN1 production is NETs
- Anti-netosis drugs e.g. metformin – changes activity of neutrophil enzymes to reduce NET formation
- Other drugs to target pDCs
- Or target how IFN signals in target cell – anifrolimab blocks IFN signalling
- IFNa has positive feedback loop back onto pDCs for more production – anifrolimab inhibits this too
59
how can Bcl-2 inhibitors be used in SLE?
Non-selective pDC depletion
- Bcl-2 is anti-apoptotic protein important for lymphocyte survival
- Inhibiting anti-apoptotic protein depletes B cells, T cells, pDCs and plasmablasts
- BCL-2 is increased in SLE, correlating with disease activity
- Venetoclax is beneficial in patients with plasmacytoid dendritic cell malignancies e.g. pDC lymphoma
- venetoclax may be good for SLE treatment
60
has Bcl-2 inhibition been sucessful in preclinical studies??
- Lupus mice have increased BCL-2 expressed in pDCs
- applied Bcl-2 inhibitor to pDCs ex vivo
- ex vivo from mice: the pDCs, when stimulated by TLR9 agonist CpG, don’t produce IFNa post-BCL-2 inhibition, indicating that they have been depleted by the drug
Treat human pDCs with Bcl-2 inhibitor, no. pDCs fall in a dose-dependent manner, while no. myeloid DCs doesn’t change – specific target of Bcl-2 in pDCs
- pDCs express CD123: This falls after the drug
Still being developed
61
what is BDCA2? how can it be targeted?
- BDCA2 is a type-C lectin specific to pDCs
- Antibodies bound to BDCA2 are rapidly internalised and presented - Causes BDCA2 to be internalised and reduce IFN1 production by pDC in response to stimuli e.g. viruses, SLE serum
Anti-BDCA2 antibodies result in BDCA internalisation and significant dose-dependent ↓ in type 1 IFN production, regardless of stimuli:
Viruses
CpG-DNA
Anti-dsDNA/plasmid DNA immune complexes
SLE serum
62
how does anti-BDCA2 work?
Binds BDCA2 on pDC surface
- Causes internalisation into endolysosome
- Inhibits TLR7 and 9 signalling in endosome – reduces IFN1 production
- Doesn’t kill pDC, just prevents IFN1 production
63
should anti-BDCA2 be combined with HCQ?
HCQ already inhibits TLR signal and IFN1 production, so is anti-BDCA2 even needed?
Stimulated PBMCs with TLR9 agonist (CpG) or TLR7 agonist (R848)
- HCQ alone significantly impairs TLR9, but not TLR7 – HCQ binds DNA and impairs TLR9, but can't bind RNA so may not inhibit TLR7
– logic for anti-BDCA2 – synergistic effect to block TLR9, but separately blocks TLR7
Looked at blood levels of HCQ in patients:
- Blood HCQ negatively correlates with ability to produce IFNa in response to TLR9 stimulation, but not in response to TLR7 stimulation - this happens in patients, not just in vitro
- Logical to give anti-BDCA2 also to inhibit TLR7
64
has anti-BDCA2 been successful in trials?
(Litifilimab) – 2 phase 2 RCT
- Humanised IgG1 anti-BDCA2 monoclonal antibody causes internalisation
- Primary outcome – swollen and tender joints
- Significantly reduces swollen joints compared to placebo
- significantly reduces skin disease as early as week 8 compared to placebo
65
what is a kinoid?
IFN vaccine/kinoid
- rIFNα2b coupled to a Th carrier protein (keyhole limpet haemocyanin)
- Give low dose of IFNa conjugated to carrier protein – drives anti-IFNa response
- Polyclonal response to other IFNa subtypes, where anti-IFN antibodies are produced – crossreactivity
- Polyclonal response against IFNα2b (and other IFNα subtypes)
66
have kinoids been successful in mice?
Lupus flare model:
- IFNα adenovirus induces renal disease in (NZB/NZW)F1 mice rapidly - these mice usually develop lupus too slowly (around 25 weeks), so IFNa shot increases
- Immunisation with IFN-K prior to adenovirus challenge reduced renal disease (histology and proteinuria) and death
67
has IFN-a kinoid (IFN-K) been successful in trials?
Trial in humans 184 patients
- Significant reduction in IFN gene signature in those vaccinated, but no difference in primary outcome at week 36
- Then did group analysis looking at if they were on steroids and if they had produced neutralising antibody
- Those with low-dose steroids and neutralising Ab had significant overall response
Generates anti-IFN antibodies, but some are neutralising and some are not
- May need to modify vaccine to drive neutralising Ab only
68
what are JAKs?
JAKs: 4 IC proteins
- JAK1-3 and TYK2
- Different cytokines that signal via receptors containing JAK have different combos
- IFNyR has JAK1/2
- IFNAR has JAK1/TYK2
69
why should JAK inhibitors be used in SLE? which should be inhibited?
Prevent IFN signalling
- Logic to have drug preferentially targeting JAK1 will work well and target all common cytokines
- JAK2 blockade won't impair IFN signal
Most drugs are specific for JAK1
- tofacitinib
70
have JAK inhibitors been successful in lupus mice?
Both tofacitinib (JAK1/3) and baracitib (JAK1/2) show benefit in mouse models of SLE
- Control: neutrophils stimulated to form NETs
- In lupus mouse with vehicle – large NETs formed
- When treated with tofa – partial reduction in NETs
- Tofa also reduces ANAs like anti-dsDNA, proteinuria and glomeruluonephritis and skin lesions
But:
- In mice, aortic ring contraction is improved - Small improvement in vascular function with tofa
- This could target SLE and CVD that drives death – but in humans, kinase inhibitors increase risk of CVD mortality and thrombosis – in mice, tofa works differently than in humans
71
has baricitinib been successful in SLE trials?
phase 2 study: 314 patients
- Endpoint: resolution of arthritis or rash at week 24
- Didn’t cause a big change in phase 2 – barely met endpoint
- Failed phase 3 trial
72
what are the adverse effects of JAK inhibitors?
Initial reports suggested increased thrombosis risk with JAK inhibitors
- 20% SLE patients have anti-phospholipid syndrome which increases risk of clotting, so JAK inhibitor shouldn’t be used as it would further increase risk
Increased risk of HZV in some studies in RA - Increases risk of shingles
Risk of serious infection increased, but not significantly
73
how can JAK inhibitors be improved?
broad JAK inhibition with Bari and Tofa isn't working
Use more selective target – TYK2
- use selective TYK2 inhibitor: Deucravacitinib
- Type 1 IFN blockade only, shouldn’t increase risk of thrombosis
- Also blocks IL-23, IL-10 and IL-12 – for psoriasis and IBD
74
has deucravacitinib been successful?
binds TYK2 and blocks signalling
- Oral allosteric inhibitor of Tyk2 (binds regulatory domain irreversibly to block signalling)
- Increased response rate in high dose of Deu (58%) compared to placebo (34%)
- Serological response: anti-DNA decrease, complement increase
viable therapeutic target as it reduces IFN1 signalling
75
how else can TLR signalling be disrupted?
TLR7 small molecule antagonists being developed – early phase
Anti-TLR7 antibodies
- Blocks TLR7 signalling
When TLR7 binds ssRNA and drives signalling, it activates myd88-IRAK complex:
- Drugs are structurally similar to MYD88 and act as decoy to prevent complex formation - being developed
- Direct inhibitor of IRAK4 being developed - blocks cascade
- Similar to CELMOD – other drugs developed to flag this complex for degradation
76
summar of IFN1 in SLE
Inhibition of type 1 IFNs is promising in lupus
- pDC depletion
- Induction of anti-IFN antibodies
- JAK inhibitors (non-selective for IFN)
77
what is the role of B cells in SLE? rationale for targeting B cells in SLE:
Failure of tolerance → autoreactive B cells
B cell hyperactivity: Increased levels of BAFF, IFN increases B cell responsiveness to TLR activation
Pathogenic effect of autoantibodies:
- Stimulation of IFN responses
- Immune complex deposition and inflammation
- anti-dsDNA directly drive inflammation
78
what are the biologic drug targets in SLE?
Antigen-presentation to autoreactive T cells which then activate autoreactive B cells
Targets:
- Anti-cytokines e.g. anti-IFNa
- Belimumab (anti-BAFF)
- Rituximab (anti-CD20) – depletes B cells
79
what is rituximab?
Type 1 anti-CD20: mAb targets CD20 on surface of B cells
chimeric – human and murine components – can have immune reactions
relocalises CD20 on surface of lipid rafts – can cause resistance
80
how does rituximab induce depletion?
complement-dependent cytotoxicity (CDC)
- activates complement cascade by Fc binding C1q = MAC formation and B cell depletion
ADCC
- Fc binds FcRIII on NK cells and other innate cells = B cell killing
Direct cell death (DCD) (minimal) - relocalises CD20 to lipid rafts
81
where is CD20 expressed?
Only on subpopulation of B cells
- CD20 normal role: unknown, doesn’t have natural ligand, binding to CD20 can affect calcium influx
- Use of rituximab to deplete CD20+ B cells leaves pro-B cells and stem cells in tact – can drive B cell repopulation
- Also doesn’t affect plasma cells, so antibodies against other antigens are preserved
- CD20 is on plasmablasts (after mature B cell, before plasma cell) which are the main autoantibody producers
82
is rituximab successful? why/why not?
response rate in patients is only 50%
- C1q deficiency or polymorphisms, or other complement issues e.g. C3, C4 can impair CDC – inherited complement disorders
- Polymorphisms in FcyRIII may modify ADCC effects
- CD20 is localised to lipid rafts – use of other medications like statins may change lipid raft structure
- CD20 is expressed in health, and we don’t know how its expression changes in disease – maybe in some diseases it is downregulated
- Soluble CD20 could mop up rituximab – some patients secrete CD20 in the blood
- Lupus is heterogeneous, so molecular and cellular differences may limit rituximab e.g. B vs T cell driven or innate driven
- Not all B cells expressed CD20 – long lived plasma cells may be producing autoantibodies which aren’t being affected
- Rituximab is chimeric, so some patients may develop neutralising antibodies against rituximab:HACA – human anti-chimeric antibdoies
- Reconstitution of plasmablasts can drive relapse following rituximab – if plasmablasts aren’t fully depleted they come back
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what was the EXPLORER trials and outcomes?
randomised trial of rituximab in lupus, using BILAG - failed:
- stringent outcomes which aimed to switch off disease - bar set too high
- small study - not good for heterogenous disease like SLE
- major placebo response and strong steroid use
- combined flare definitions (severe and moderate) even though these severities are different
- patients were immunosuppressed (standard of care) already, so hard to distinguish the benefit of rituximab
biological effects: increased C3/4, depleted B cells, reduced anti-dsDNA
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how can relapse occur with rituximab?
Relapse following RTX is associated with B cell repopulation
Partial reduction in plasmablasts associated with early relapse
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what is obinutuzumab? is it better than rituximab?
type 2 anti-CD20 mAb
- Afucosylated Fc portion – affects effector function of mAb
Type 2 antibodies
- Don’t localise CD20 onto lipid rafts
- Minimal CDC and more DCD and ADCC
- More ADP (phagocytosis)
- Low C1q can affect rituximab, so move to type 2 may be more beneficial
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what is an alternative approach to deplete B cells?
target CD19 present on majority of B cell subtypes
- E.g. autologous CAR-T cells target this in malignancies, and this could be approached in autoimmunity
- Targeting CD19 works in mouse models of lupus
- CD19 targeting would give broader coverage
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has CAR-T been successful in SLE?
Lymphodeplete and reinfuse CAR-T – proliferate and deplete B cells
- B cells repopulate but they are naïve – lose autoreactive B cells
- Lymph node biopsies showed also T cell depletion – unclear why
Works well for severe refractory SLE:
- Complete clinical remission in 4/5 patients in 3 months – prolonged drug free remission
- Reduces kidney disease – reduced protein in urine
- Complement increased, Anti-dsDNA reduces, Fatigue improved
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how could B cell activity be targeted?
BAFF (B cell activating factor)/Blys (B lymphocyte stimulating factor)
- Blockade of BAFF
- Mice lacking BAFF have reduced B cell development – BAFF is crucial for immune function
- Overexpression of BAFF in mice drives autoimmunity
- Spontaneous mouse models of lupus have high levels of circulating BAFF
-Role of BAFF not fully understood, but can control autoreactivity
- High levels can prevent B cell apoptosis - autoreactive B cell escape
- Concentrations of BAFF are higher in SLE patients compared to healthy volunteers and RA patients
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what is the signal pathway of BAFF?
BAFF exists as a soluble trimer or soluble 60-mer (60-mer has more biological action)
- Signals via BAFF-receptor, TACI and BCMA
- BAFF doesn’t exist in isolation: APRIL has some functions that overlap with BAFF
-APRIL signals via similar receptors
- Don’t understand how BAFF and APRIL contribute to autoimmunity development, and roles in SLE are unclear
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what drugs can target the BAFF/APRIL pathway?
Belimumab – targets soluble BAFF
Other agents target soluble and membrane form of BAFF
Atacicept targets both BAFF and APRIL - currently trialed
Targeting BAFF receptor
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what is belimumab?
Fully human IgG1 mAb binding soluble BAFF
- Neutralise circulating BAFF
- Block BAFF interacting with BAFFR (Less clear how it interacts with the other receptors)
- Disrupts 60-mer formation to drive 3-mer formation (less active than 60-mer)
- first new licensed treatment for SLE in over 50 years
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what was the BLISS-52 trial?
Belimumab, SRI-4 at 52 weeks
- Responders from 32% on placebo to 39% with belimumab – tiny improvement
- No change in SLEDAI
Can we predict who will do better with belimumab?
- If more disease activity >10 SLEDAI, then patients did better with belimumab
- High anti-DNA and/or low C3/C4 did better
- Approved for these patients
- Less people eligible for drug
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how do BAFF levels in the blood change following B cell depletion?
BAFF levels in blood increase following B cell depletion with rituximab
- This is a homeostatic response – once B cell count is reduced, immune system increases BAFF to stimulate B cell repopulation
- Protein and RNA levels of BAFF are increased following rituximab
- This may drive relapse: Repopulated B cells following rituximab may be more likely to be autoreactive
- All flares that occur in patients that receive rituximab are not the same, and BAFF may have role in flares associated with rise in anti-dsDNA
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why should anti-CD20 anti-BAFF be used?
Combination therapy could be good: rituximab and belimumab together to prevent surge in BAFF levels following B cell depletion
- Give rituximab first to deplete B cells
- Then give belimumab to mop up BAFF
-This is being trialed in lupus: Combination reduced anti-dsDNA and no. flares
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what is the BEAT-lupus trial?
52 patients (26 rituximab + placebo, 26 rituximab + belimumab)
- Post hoc analysis suggests better response in patients with IgA2 anti-dsDNA antibodies
- reduced time to flare
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how is IFN1 implicated in SLE?
- Plasmacytoid DCs are the major produces of IFN1, B cells also produce, and keratinocytes (IFN-k)
- In SLE, no. pDCs low in blood and not loads in tissues, so other cells produce it too
- pDCs produce IFN1 in response to immune complexes binding FcRs and activating TLRs
- IFN1 induces netosis, activates APCs to interact with T cells, activates B cells
- IFN1 is potent anti-viral cytokine
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what stimuli can cause IFN1 production?
NETs
immune complexes
ROS
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how are IFN1 levels affected in SLE?
Type 1 IFN in SLE is increased
- IFN-stimulated gene score (ISG) – surrogate measure of IFN1
- ISG is increased in SLE patients and in other autoimmune diseases
- IFN1 levels in lupus associate with: Disease activity, Arthritis, nephritis, lymphadenopathy, Presence of autoantibodies (anti-dsDNA)
- Not sure if its IFN1 driving autoantibody production, or autoantibodies driving IFN1 production
- Haematological parameters – presence of lymphopenia
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what are the 3 main IFNs?
Type 1 and 3 important in lupus
Role of type 2 in lupus is less clear
- IFNy (type 2) functions differently to type 1 and 3 and signals differently
Type 1 IFN receptor has 2 subunits: 1 and 2
- IFNAR1/2
- IFNAR is receptor for multiple IFN1s e.g. IFNa, IFNb, IFNk
- Ligation of IFNAR results in activation of JAK1 and TYK2
- JAK leads to downstream phosphorylation of STAT which activates transcription of ISGs
- IFN-lambda is in kidney disease in lupus
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what is anifrolumab?
Human IgG1 anti-IFNAR mAb
- Induces internalisation of IFNAR and inhibits STAT1 phosphorylation
- Blocks signalling of all IFN1s, not just IFNa
- Broad effects because IFN1 affects many cells
- Inhibits pDC-mediated plasma cell differentiation
- Tested in 2 large phase 3 randomised control trials
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what were the trials for anifrolumab?
2 phase 3 trials – TULIP 1 and 2:
- Low dose wasn’t effective – found out in TULIP1
- Many non-responders in TULIP1 – found that people were on ibuprofen which can be taken for anything, but these people had to drop out – dumb
- Study became unblinding and changed rules for TULIP 2 where the ibuprofen rule was removed
- SRI-4 in TULIP 1 wasn’t being met, but were responding to secondary outcome BICLA
- During TULIP2, changed primary endpoint from SRI-4 to BICLA before unblinding – still unbiased
- Approved it in US and europe as it improved primary outcome (BICLA response) in TULIP2
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what are the challenges in lupus trials?
- Uncommon disease with marked clinical and cellular/molecular heterogeneity
- No good biomarkers for diagnosis or active disease
- “Standard of care” arm includes steroids and other immunosuppressants; testing “add-on” therapies
- Variable other immunosuppressant use, so hard to compare
- Imperfect response definition; e.g. is improvement in skin rash and renal disease the same?
- often have non-responders or high placebo response
- many components to measure which is hard to rationalise individually e.g. anti-dsDNA in 70% patients
- scoring system isn't consistent, and use of composite end points
- standard of care is often needed instead of placebo e.g. for kidney disease, so can't distinguish role of new therapy
