NPC Diagnostic Tools and Managementgdrc-cong.tums.ac.ir/images/slide/slide2/2-ashrai.pdf ·...

NPC Diagnostic Tools and Management

Mahmoudreza AshrafiProfessor of Pediatric Neurology

Children’s Medical CenterPediatrics Center of Excellence

Tehran University of Medical SciencesTehran - Nov / 2019

Challenges in NP-C Detection and Diagnosis

Efforts to increase awareness of NP-C among clinicians are still needed .

Clinical history and a detailed examination are the initial steps undertaken before considering further

diagnostic procedures.

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The 7 key discriminatory signs and symptoms of NP-C

Useful resources:

For more information on symptoms of NP-C and to use the NEW NP-C SI, please go to: www.npc-si.com

NP-C

Prolonged, unexplained neonatal jaundice or cholestasis

Isolated unexplained splenomegaly

(historical and/or current) ± hepatomegaly

Vertical supranuclear gaze palsy (VSGP)

Gelastic cataplexyPre-senile cognitive decline and/or dementia

Psychotic symptoms (hallucinations, paranoid

delusions ± thought disorder)

Parent, sibling or cousin with NP-C

(two items combined)

Presenter

Presentation Notes

NP-C should be highly suspected when 2 or more of these 7 key discriminatory signs and symptoms, or VSGP alone, are present in your patient* If NP-C is suspected, the full completion of the NP-C SI >4y should be used to confirm suspicion and to obtain additional clinical information * Because only 7 signs and symptoms are assessed, this list does not reflect a comprehensive clinical picture of NP-C and should not be used as an isolated screening tool

http://www.npc-si.com/

Challenges in NP-C Detection and Diagnosis

Two steps to achieving early diagnosis of NP-C

Patients with suspected NP-C should undergo a comprehensivemedical history and general physical examination plus neurological assessment .

Recognitionof signs and symptoms by

paediatricians, neurologists, hepatologists, endocrinologist

psychiatrists and ophthalmologists and acting on

that suspicion

Confirmationof diagnosis using detailed physical and neurological assessment followed by

specialist tests

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Initial assessments of NP-C • Routine laboratory biochemistry profiles including standard

blood biochemistry, plasma lipids and unconjugated bilirubin are generally normal in NP-C patients, but can be altered in those with hypersplenism or cholestatic liver disease.

• Mild thrombocytopenia can sometimes be seen in patients with splenomegaly.

• Decreased plasma LDL / HDL-cholesterol and increased plasma triglycerides have been reported in NP-C patients .

• While plasma transaminases are generally normal, aspartate aminotransferase (ASAT) activity can be elevated, but generally returns to normal over time.

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Initial assessments of NP-C • Examination of tissue samples by light microscopy can identify

the presence of characteristic (but not specific) foam cells and/ or sea-blue histiocytes in the bone marrow, spleen, liver, lung and lymph nodes .

• ASM activities in NP-C have been reported as being normal or slightly elevated in leukocytes .

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Imaging in the Diagnosis of NP-C

• MRI or CT scans of the brain are usually normal until the late stages of NPC , and imaging is considered nonspecific .

• MRI in patients with slowly progressing disease may show cerebral atrophy and /or marked atrophy of the superior/anterior cerebellar vermis .

• Other visible abnormalities may include thinning of the corpus callosum and increased signal from the centrum semiovale, reflecting secondary demyelination .

• Patients with early-infantile neurological disease onset may show pronounced white matter signal hyperintensity in T2weighted MRI images.

Cerebellar atrophy

Slight white matter abnormalities

Tedeschi et al. J Neurol Neurosurg Psychiatry 1998; Galanaud et al. Mol Genet Metab 2008

Brain magnetic resonance imaging in NP-C

Current laboratory diagnostic techniques


Laboratory-based tests are the mainstay of diagnostic procedures in NP-C. 1- Cell biology-based tests

1-1.Fillipin Staining1-2. Cholesterol esterification test

2- Biochemical biomarkers2-1.Chitotriosidase activity2-2.Oxysterol analysis2-3.Lysosphingolipids2-4. Bile acid metabolites

3- Genetic testing

4- VOG (Video-Occulo-Graghy)

Filipin Staining Test

① The primary biomarker diagnostic test is Filipin test , that developed 30 years ago , but has been extensively used, and until recently was considered as the gold standard assay for NP-C diagnosis .

② It is the most sensitive and specific test and should be conducted in specialized expert centers.

③ Skin fibroblasts are cultured in lipoprotein-deficient serum and then exposed to a pulse of LDL-enriched medium , fixed and stained with Filipin.

④ Filipin is a fluorescent polyene antibiotic, which specifically binds to cholesterol but not to esterified sterols.

⑤ In NPC-positive cultures , these cells show a reduced ability to esterify cholesterol after loading with exogenously derived LDL-cholesterol .

⑥ Cholesterol-filled perinuclear vesicles of NPC cells are observed under fluorescence microscopy .


① Early studies showed that in fibroblast cultures from normal subjects, fewer than 10% of cells stain positive, while in those from typical NP-C patients, nearly all cells display multiple strongly fluorescent perinuclear vesicles .

② Studies in large series of patients indicate that this typical pattern is observed in 80–85% of cases with NP-C .

③ Subdivision of the typical staining pattern into ‘classical’ and ‘intermediate’ profiles has been proposed to reflect a variation in the percentage of positive cells (80 to 100% of cells) and their level of fluorescence .


Advantages

>80% of NP-C cases display a typical (‘classical’or ‘intermediate’) staining profile

Similar results in NPC1 and NPC2

Typical staining profile is unambiguous and easy to interpret—with clinical findings it supports confident diagnosis

Only one condition, mucolipidosis II/III (I-cell disease) is known to mimic a typical/classical NP-C filipin profile

Longest experience in a broad clinical setting

Limitations

Invasive skin biopsy and cell culture

Turnaround time : 5-7 weeks

Technical requisites / Expertise in interpretation

Variants makes problem- 15% of cases

False positive: 10% of NL subjects, NP-A/B, MEGDEL syndrome , Wolman Dis, SLOS, Tangier Dis, CDG

Final interpretation needs genetic studies





3- Genetic testing


Lysosphingolipids① It has recently become apparent that plasma levels of

Lysosphingolipids are increased in patients with LSDs such as Fabry , Gaucher , GM2 gangliosidoses , Krabbe and Niemann-Pick disease , making the lysosphingolipids potentially powerful biomarkers for both diagnosis and monitoring of treatment effects in their respective LSDs .

② The lysosphingolipids, lysosphingomyelin (SPC) and glucosylsphingosine (GlcSph), appeared to be elevated in the plasma of NP-C patients.

Richard W. D. Welford et al . PLOS ONE | December 5, 2014

Lysosphingolipids① The levels of lyso-SM-509 in blood reflect the burden of

the NP-C disease and it can be used for the easy diagnosis of NP-C patients and for the monitoring of the disease progression.

② Determination of lyso-SM-509 is performed by multiple reaction monitoring mass spectrometry (LC/MRM-MS) in plasma, serum, EDTA blood and dried blood spots (DBS).

③ By combining the results with the levels of lyso-SM-465 the type A/B and type C patients can clearly be distinguished.

④ Lyso-SM-509 has a sensitivity of 100 % and specificity of 98.15 % for the diagnosis of NPC.

Claudia Cozma, et al . Molecular Genetics and Metabolism 120 (2016) S17–S145

Lysosphingolipids

Advantages

A powerful tool for differentiating patients with NP-C and NP-A/-B from control subjects

Minimally-invasive small volume of blood

Multiplex assay can identify other LSDs , meet demands; faster turnaround time

Cost savings versus filipin testing

Limitations

Lyso-SM-509 does not clearly distinguish NP-C from NP-A/-B

By combining the results with the levels of lyso-SM-465 the type A/B and type C patients can clearly be distinguished

Does not have sufficient sensitivity as an NP-C biomarker

Experience in clinical laboratory setting is still limited





3- Genetic testing


Genetic testing

• Genetic analysis is necessary to confirm a diagnosis of NP-C following abnormal results with disease biomarkers .

• Genetic analysis is necessary to confirm a diagnosis of NP-C following inconclusive filipin staining results .

• Genetic analysis is necessary to confirm a diagnosis of NP-C following even clear-cut results from filipin staining or Lyso 509 .

• The identification of two alleles with demonstrated disease-causing mutations in NPC1 or NPC2 confirms the diagnosis of NP-C, with supporting mutation analysis in at least one parent.

• About 380 distinct NPC1 and 22 NPC2 mutations are already known and are recorded in mutation databases, with only a few of them being common or recurrent.

• Missense and nonsense mutations constitute the vast majority of known NP-C mutations .





3- Genetic testing


EyeSeeCam Video-Oculography (VOG)

EyeSeeCam VOG provides a measurement for quantifying eye movements.

Detailed saccade analysis: eye position and velocitytraces over time for horizontal (right, left) and vertical (up, down) saccades

Detailed saccade findings for NPC

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NP-C / Management Once the diagnosis of NP-C is confirmed ,

the clinician is confronted with several challenges .

Educating the patient and family

Symptomatic treatments or Supportive Measures

Substrate reduction therapy or Targeted therapy

Treatments in development

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NP-C Management / Education Essentially all patients with NP-C die prematurely, although

disease progression rates and life expectancy vary greatly, primarily dependent on the age at onset of neurological symptoms.

The majority of patients die between 10 and 25 years of age .

While the clinical phenotypes of NP-C are highly heterogeneous , patients at the terminal stage of the disease are typically incapable of voluntary movement , tube-fed and intellectually impaired .

Historical data indicate that in the majority of patients death is often related to bronchopneumonia, most likely due to repeated aspiration following progressive dysphagia .

M.C. Patterson et al. / Molecular Genetics and Metabolism 106 (2012) 330–344

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NP-C Management / Education

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NP-C Management /Education

Educating the patient and family is a process that cannot be accomplished in a single meeting , since it is not possible to apprehend new information in the state of emotional turmoil that accompanies the delivery of such a diagnosis .

It is important that family and community resources be identified and linked to the family early in the course of the illness .

Pediatrician , family physician or general practitioner are central in promoting the best care of patients .

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NP-C Management / EducationOptimal support

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Substrate reduction therapy or Targeted therapy


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NP-C / Management Symptomatic Treatment

Symptomatic treatment consisted of non-pharmacological and pharmacological therapies .

Non-pharmacological therapies includingphysical , occupational and speech therapy , enteral feeding and hearing aids .

Pharmacological therapies such as AEDs , Trihexyphenidyl , Levodopa , Beta blockers , Clomipramine ,Antipsychotic drugs , Donepezil and Memantine can be helpful in managing the symptoms of the patients .

Symptom Treatment

Seizures Antiepileptic drugs

Cataplexy TCA and SSS stimulants

Dystonia and tremor Anticholinergic drugs

Movement restriction Physical therapy

Feeding problems Gastrostomy

Sleep disorders Melatonin and sedatives

Constipation Regular bowel programme

Respiratory involvement Physiotherapy, bronchodilators, antibiotics

Symptomatic Treatment

Mol Genet Metab 2009; 98: 152-165

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Specific therapy or Targeted therapy


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NP-C / Management

Patients with NPC have impaired intracellular lipid trafficking , which leads to the accumulation of lipids in the perinuclear lysosomes , that in turn , causes the build-up of excess lipids in various tissues .

Platt FM , et al J Cell Biol 2012 ;199(5)Madra M , et al Clin Lipidol . 2010 ;5(3)

Rosenbaum AI , et al J Neurochem . 2011 ;116(5)

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NP-C / ManagementSpecific Therapy

Treatment strategies based on the hypothesis that cholesterol is the offending metabolite were first proposed in the early 90's.

The combination of hypocholesterolemic drugsand a low-cholesterol diet seemed to partiallyreduce the cholesterol load in liver, but noamelioration of the neurological disease wasseen in patients after 2 years of treatment .

Schiffmann R: Niemann-Pick disease type C. From bench to bedside. JAMA 1996, 276:561-564.

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NP-C / ManagementTargeted therapy or SRT

Since glycolipid storage appears to contribute to at least some of the neuropathologic features, an iminosugar inhibitor of glucosylceramide synthase (miglustat, also known as N-butyl-deoxynojirimycin), later approved for substrate reduction therapy ofmild to moderate type 1 Gaucher disease), wasadministered to npc1 mutant mice and cats.

It resulted in delayed onset of the neurological symptoms in both species, and a 20% longer survival of the mice .

Zervas M, Somers KL, Thrall MA, Walkley SU: Critical role forglycosphingolipids in Niemann-Pick disease type C. Curr Biol 2001,

11:1283-1287.

ZAV153

Miglustat (Zavesca): Substrate Reduction Therapy• Miglustat, a small iminosugar molecule, inhibits the

action of glucosylceramide synthase involved in the production of the glycosphingolipid (GSL) substrate that accumulates in NP-C

Zavesca SmPC, http://emc.medicines.org.uk/; accessed Feb 2009

glucosylceramide synthase

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Controlled clinical trial was thus initiated inneurologically symptomatic patients, first in adolescents and adults(12 years and above) then inchildren (4-12 years).

Miglustat was approved for the treatment of progressive neurological manifestations in pediatric and adult patients with NP-C in the EU in 2009 , and has since been approved for this indication in a number of further countries.

Actelion, Miglustat (Zavesca) Summary of Product Characteristics, EMA (EudraPharm) (2010)

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Miglustat is able to cross the blood–brain barrier [1], and was shown to reduce GSL accumulation and cellular pathology in the brain, delay onset of neurological symptoms, and prolong survival during pre-clinical studies [2].

[1] A. Treiber, O. Morand, M. Clozel, The pharmacokinetics and tissue distribution of the glucosylceramide synthase inhibitor miglustat in the rat, Xenobiotica 37 (2007) 298–314.[2] M. Zervas, K.L. Somers, M.A. Thrall, S.U. Walkley, Critical role for glycosphingolipids in Niemann–Pick disease type C, Curr. Biol. 11 (2001) 1283–1287.

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It may also indirectly modulate intracellular calcium homeostasis related to sphingosine storage a suspected initiating factor in the pathogenesis of NPC1 – through its effects on glucosylceramide levels [1-2-3].

[1].L. Liscum, S.L. Sturley, Intracellular trafficking of Niemann–Pick C proteins 1 2: obligate components of subcellular lipid transport, Biochim. Biophys. 1685 (2004) 22–27.

[2].E. Lloyd-Evans, F.M. Platt, Lipids on trial: the search for the offending metabolitein Niemann–Pick type C disease, Traffic 11 (2010) 419–428.

[3].E. Lloyd- et al , Niemann–Pick disease type is a sphingosine storage disease that causes deregulation of lysosomal calcium, Nat Med. 14 (2008) 1247–1255..

The recommended dose of miglustat for the treatment of adult and adolescent patients with NP-C is 200 mg three times a day .

There is limited experience with the use of miglustat in patients with NP-C under 4 years of age , and there is no experience with the use of Zavesca in patients over the age of 70.

Dosing in patients between 4 and 12 years should be adjusted according to body surface area .

Miglustat, Zavesca®

Gastrointestinal disturbances :

Gastrointestinal effects are the chief tolerability issue with miglustat, and can usually be managed with symptomatic therapy (e.g., loperamide) or dietary modification (initiation of a diet low in lactose and other carbohydrates) ; to taking Zavesca between meals, temporary dose reduction may be of help in some patients .Loperamide :

2-5 Y 1 mg Tid6-8 Y 2 mg Bid 8-12 Y 2 mg Tid

Miglustat, Zavesca®

• All patients with neurological, psychiatric or cognitive manifestations at diagnosis should be offered miglustat therapy based on potential improvements in, or maintenance of, quality of life.

• Patients without neurological manifestations should not receive miglustat therapy, as some can remain asymptomatic for considerable periods of time [1,2,3].

Miglustat : Whom and when to treat

[1] M. Sevin, et al , The adult form of Niemann–Pick disease type C, Brain 130 (2007) 120–133L

[2] Dvorakova, J. et al , Subclinical course of adult visceral Niemann–Pick type C1 disease. A rareor underdiagnosed disorder? J. Inherit. Metab. Dis. 29 (2006) 591.

[3] A.H. Fensom et al , An adult with a non-neuronopathic form of Niemann –Pick C disease,J. Inherit. Metab. Dis. 22 (1999) 84–86

Miglustat treatment can be initiated in patients aged < 4 years who have a confirmed diagnosis of NP-C and who have neurological symptoms.

In cases where severe neurological impairment is already present at diagnosis, particularly in very young patients, miglustat is less likely to provide substantial therapeutic benefits .

Miglustat : Whom and when to treat

M. Pineda, et al , Clinical experience with miglustat therapy in pediatric patients with Niemann–Pick disease type C: a case series

Mol. Genet. Metab. 99 (2010) 358–36.

Initiation of miglustat treatment in NP-CRecommendations on the diagnosis and management of Niemann-Pick disease type C. NP-C Guidelines Working Group, Wraith JE, Baumgartner MR, Bembi B, Covanis A, Levade T, Mengel E, Pineda M, Sedel F, Topçu M, Vanier MT, Widner H, Wijburg FA, Patterson MC. Mol Genet Metab. 2009

NP-C diagnosis with any neurological manifestation

Treat

Y

Familial history of NP-C

N

Estimated time of neurological symptom

onset known

Y

Monitor

N

NTreat at or

before estimated time

of onsetY

Due to the neurodegenerative nature of NP-C, disease stabilization or a reduced rate of disease progression are likely the best attainable goals for long-term disease-specific therapy [1].

Before disease-specific therapy with miglustat is commenced, patients and family members should be informed of treatment expectations based on findings from previous clinical studies.

Treatment goals with miglustat in NP-C

[1] J.E. Wraith, M.R. Baumgartner, B. Bembi, A. Covanis, T. Levade, E. Mengel, M.Pineda, F. Sedel, M. Topcu, M.T. Vanier, H. Widner, F.A. Wijburg, M.C. Patterson,

Recommendations on the diagnosis and management of Niemann–Pick disease type CMol. Genet. Metab. 98 (2009) 152–165.

In patients with early-infantile onset disease, it can take 6 months to 1 year to see discernable clinical benefits.

In patients with later-onset disease, it can take 2–3 years to see clinical benefits.

Patients with early-infantile onset NP-C, and those with severe dementia in the terminal stage of the disease are less likely to benefit from treatment with miglustat .

These clinical informative points increase compliance of patients and their parents .

Treatment goals with miglustat in NP-C

In general, miglustat therapy should be continued as long as patients continue to derive discernable therapeutic benefits with an acceptable profile of tolerability/safety.

Treatment should be stopped wherever severe gastrointestinal disturbances occur despite dose reduction, dietary modification and symptomatic treatment.

Discontinuation should be considered in patients who show progressive neurological deterioration leading to an unacceptable quality of life.

Stopping Miglustat Therapy

A clinical trial indicated that miglustat can stabilize neurological disease in children , juveniles and adults with NP-C over 12 months , and therapeutic benefits were manipulated up to 48 months during subsequent extension treatment .

Miglustat was well tolerated up to 66 months .

Decisions to alter or stop miglustat therapy should be based on individual patient characteristics and in consultation with patients and family members.

Stopping Miglustat Therapy

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Substrate reduction therapyor Targeted therapy


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NP-C / Treatments in Development Histone deacetylase inhibitor valproic acid Curcumin Imatinib

Cyclodextrin Vorinostat Arimoclomol

Additional data from clinical studies will be required to confirm these promising outcomes and assess the long-term effects these drugs have in humans.

8th Scientific Symposium on NP-C March 2016 Budapest

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Conclusion In the absence of any curative treatment, quality of

life represents a legitimate treatment goal in themanagement of NP-C and can be addressed by :1) Rigorous symptomatic treatment2) Miglustat therapy for existing neurological

manifestations.

Miglustat treatment effects on neurological manifestations should be evaluated on a regular basis (e.g. every 6 months); and continuation of therapy should be re-appraised after ≥1 year of treatment .

Thank youfor your Attention

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