Case HistoriesCase Histories

Case 1: Positive Newborn Metabolic

ScreenHistoryHistory: : NMS test result shows elevated Phenylalanine NMS test result shows elevated Phenylalanine

(0.75 umole/l; normal <0.125)(0.75 umole/l; normal <0.125) Term pregnancyTerm pregnancy

Normal P/L/DNormal P/L/D BWt 3.1 kg, BWt 3.1 kg, Normal neonatal courseNormal neonatal course

Questions:1. Describe briefly what your initial

counselling to parents would be.2. What investigations would you under take

to confirm diagnosis?

Results of investigations

Results Plasma

PHE=1.2umole/l; tyrosine = 0.05 umole/l

Urine organic acids increased PPA,PLA,PAA

Questions1. What other

tests need to be done to be sure this baby needs diet treatment?

2. What is the basis of the diet treatment?

Metabolism of Metabolism of PhenylalaninePhenylalanine

PHE

Dietary Protein Body

Protein

Tyrosine

PPA

PLA PAA

Phenylactetyl- glutamine

BH4

qBH2

(PAH)6-pyruvoyl-

BH4

GTP

PHE levels in the Newborn with PKU

PHE

Days of Age

1 2 3 4

NORMAL

RANGE

Screening Possible

Untreated Phenylketonuria

Signs / Symptoms Signs / Symptoms mental retardationmental retardation hypopigmentation eczema-like rasheczema-like rash autistic-like behaviorautistic-like behavior autosomal recessiveautosomal recessive high bloodhigh blood

phenylalanine levelsphenylalanine levels

PKU: Diagnostic work-up

Confirm that PHE level is elevated Rule out biopterin deficiency

disorders Urine pterin levels Dihydrobiopterin reductase activity Biopterin load test (optional) If present start DOPA/carbiDOPA/5HTP

If BH4 disorder not diagnsosed & PHE above 0.4 mM/l, start low PHE diet

Natural Foods

Contains some normal nutrients

and all those being restricted

Nutritional Treatment of PKU

Diet has two components:

Must meet all nutritional needs + limit intake of restricted nutrients to amts sufficient for growth

Medical Formula

Contains all nutrients

except those being restricted

+

Case 2: Hepatomegaly with Abnormal Liver Pathology

History: The pathologist in your hospital calls to discuss

an abnormal liver biopsy result 8 month old boy with (R) abdominal mass

extending down into the iliac fossa Seen by Oncology re: ? Tumor; taken to OR

for open biopsy

QuestionWhat do you see in the following biopsy?

Case 2: Case 2: patient & liver biopsy & liver biopsy

Questions

1. What types of disorders might cause this appearance?

2. What further historical information may be of help?

3. What further studies should you request from the pathologist?

Questions1.1. What types of disorders might cause this appearance?What types of disorders might cause this appearance?

• Glycogen storage disorders (types 1a & 1b, 3, 6 lysosmal storage disorders (Gaucher, Niemann-Pick, MPS, oligosaccharidoses, tyrosinemia, B-oxidation disorders (MCAD, LCHAD, VLCAD)

2.2. What further historical information may be of help?What further historical information may be of help?• Symptoms of hypoglycemia (relationship to fasting

including timing)• Mother indicates baby can only go about 2-4 hours

without a “bottle”

3.3. What further studies should you request from the What further studies should you request from the pathologist?pathologist?

• PAS staining +/- pretreatment with diastase• Electon microscopy

Diagnostic testingDiagnostic testing

Fasting challenge +/- feeding challenges

Enzyme assays Need fresh liver Need to choose specific enzymes to

target based on history Molecular testing

Now have bank of mutations but expensive

GSD-0

GSD-IV

GSD-1a&b

GSD-V, GSD-VI, GSD-IX

GSD-II ( lysosomal)

GSD-III

GSD-VII

GSD-X, GSD-XII, GSD-XIII

GSD-XI (LDH)

LIVER

MUSCLE

Glycogen Storage Disease IBControlled Fast

Time After Feed (min)Time After Feed (min)

00 3030 6060 9090 120120 150150 180180

GlucosGlucosee

(mM)(mM)

4.54.5 4.64.6 4.24.2 4.04.0 4.04.0 3.83.8 2.12.1

LactateLactate

(mM)(mM)1.21.2 1.61.6 1.81.8 1.71.7 1.81.8 2.12.1 5.65.6

GSD IA &IBClinical features

early onset hypoglycemia

lactic acidosis hepatomegaly Fanconi syndrome hyperuricemia hyperlipidemia

Diagnosis controlled fast (test

BS & LA) enzyme (liver biopsy) DNA testing

Therapy provide 5 - 10 mg

glucose/kg/min continuous .nocturnal

infusion of CHO as polycose or formula

frequent meals during days

corn starch days &/or nights

don’t over treat with CHO

Neutropenia in Type IB prophylactic antibiotics GCSFEmergency protocols for

illness, surgery etc.

Case 3: 18 month boy with hepatomegaly and

obtundationHistory: stuporous on admission found pale & sweaty and unarousable by parents Had been ill for about 18 hours with no

significant intake Seizure in ambulance

Initial studies: Blood sugar = 0.2 mM/l, Na+=145, K+ =3.5, Cl- =104,

TCO2 = 10 Urinalysis = Normal All other testing including lactate, NH4 &

LFE’s normal

Key observationsKey observations

Severe hypoglycemia with hepatomegaly and no ketonuria on setting of history of prolonged fasting

Needs urgent treatment of hypoglycemia Route? How much glucose?

? Significance of no ketones in urine ?diagnostic testing

Trifunctional protein

VLCAD,MCAD, SCAD

Diagnostic InvestigationsDiagnostic Investigations

Plasma acylcarnitnes suggest Medium Plasma acylcarnitnes suggest Medium Chain Dehydrogenase deficiency (MCAD)Chain Dehydrogenase deficiency (MCAD)

Plasma free carnitine levels low while Plasma free carnitine levels low while acylcarnitines highacylcarnitines high

1414C- palmitic acid oxidation in leucocutes C- palmitic acid oxidation in leucocutes quite reducedquite reduced

Molecular diagnosis indicates Molecular diagnosis indicates homozygosity for the common caucasian homozygosity for the common caucasian mutation.mutation.

MS/MS Analysis of plasma MS/MS Analysis of plasma acylcarnitines (?= MCAD)acylcarnitines (?= MCAD)

270 300 330 360 450 480 510 540

50

100

% Intensity C2

INTERNAL STANDARDS

C3C4

C8

C16

C18:1C16

C3

C2

C14:1

390 420

C8

C6

C10

C10:1

MS/MS PS 85

Phases of Glucose HomeostasisPhases of Glucose Homeostasis

1.1.Glucose absorptive phaseGlucose absorptive phase: 3 - 4 hrs after : 3 - 4 hrs after glucose ingestion (high insulin) glucose ingestion (high insulin)

2.2.Post absorptive/early starvationPost absorptive/early starvation: 3-12 hrs: 3-12 hrs

glucose (from hepatic glycogen) to brain, glucose (from hepatic glycogen) to brain, RBC, renal medullaRBC, renal medulla

3.3.Early / Intermediate StarvationEarly / Intermediate Starvation: 14+ hrs: 14+ hrs

gluconeogenesis & (later) lipolysis gluconeogenesis & (later) lipolysis

Treatment

Avoid fasting L-carnitine if free carnitine low Emergency protocol & letter

Sick day management Admission to ER/hospital to maintain

blood glucose with IV infusion to prevent excessive lipolysis the would overload the B-oxidation pathway

Case 4: Neonatal PresentationCase 4: Neonatal Presentation

5 d.o. male5 d.o. male Well for 72 hrs then Well for 72 hrs then

became lethargic, fed became lethargic, fed poorly, began vomiting & poorly, began vomiting & developed alternating developed alternating flaccidity & opisthotonic flaccidity & opisthotonic posturing.posturing.

Became comatoseBecame comatose Developed hyperpnea and Developed hyperpnea and

respiratoy alkalosis respiratoy alkalosis progressing to respiratory progressing to respiratory failurefailure

O/E: hepatomegaly, O/E: hepatomegaly, hypothermiahypothermia

Test ResultsTest Resultso NormalNormal: CBC, ‘lytes’, : CBC, ‘lytes’,

bld glucose, lactic bld glucose, lactic acid, urinalysisacid, urinalysiso not acidoticnot acidotico not ketoticnot ketotico not hypoglycemicnot hypoglycemic

o NH3 (350 umole/l) NH3 (350 umole/l)

? Differential ? Differential DiagnosisDiagnosis

? Further testing? Further testing

Case 4: Investigation ResultsCase 4: Investigation Results

5 d.o. male5 d.o. male Well for 72 hrs then Well for 72 hrs then

became lethargic, fed became lethargic, fed poorly, began vomiting & poorly, began vomiting & developed alternating developed alternating flaccidity & opisthotonic flaccidity & opisthotonic posturing.posturing.

Became comatoseBecame comatose Developed hyperpnea and Developed hyperpnea and

respiratoy alkalosis respiratoy alkalosis progressing to respiratory progressing to respiratory failurefailure

O/E: hepatomegaly, O/E: hepatomegaly, hypothermiahypothermia

Test ResultsTest Resultso NormalNormal: urine amino : urine amino

acids & organic acidsacids & organic acidso LowLow: urea, arginine, : urea, arginine,

ornithine, ornithine, o HighHigh: citrulline (1.21 : citrulline (1.21

mM) mM) glutamine (1.4 glutamine (1.4 mM) mM) asparagine asparagine

? Probable Diagnosis? Probable Diagnosis

Detoxification of NH3 by Urea Cycle Benzoate Dietary Protein Gut Bacteria Endogenous Protein Catab Buphenyl

NH4 PAA

NH4 + CO2 GLN

Carbamoyl Phosphate GluA PhAcGluNH2

Hippuric Acid

Ornithine Citrulline

MITOCHONDRION Aspartate

Urea

Arginine Argininosuccinic Acid

Fumarate

(CPS)

(OTC)

(ASAS)

(ASAL)

(Arginase)

Approach To Differentiation of Urea Approach To Differentiation of Urea Cycle Disorders Causing Neonatal Cycle Disorders Causing Neonatal

HyperammonemiaHyperammonemiaHyperammonemiaHyperammonemia

Before 24 hrsBefore 24 hrs After 24 hrsAfter 24 hrs

Metabolic acidosisMetabolic acidosis

PretermPreterm Fullterm Fullterm YesYes NoNo

plasma citrullineplasma citrulline

THANTHAN PDH PDH P PAA AA abs/trabs/tr >1000uM>1000uM GAIIGAII MMA MMA 100-300uM100-300uM

etc.etc. etc.etc. Ur. Orotate Ur. Orotate

uOAuOA low highlow high ASAASA

CPS OTCCPS OTC CitrullinemiaCitrullinemia

Approaches to Therapy of Urea Cycle Disorders

Acute Mgmt(based on NH3 level)

NPO Dialysis ( prefer.

Hemodialysis) IV: CHO (6–8 mg

Glc/kg/min) Lipid (3 gm / kg)

Alternate Pathway Therapy Oral (Phenylbutyrate +

L-Arg) IV (Phenylacetate +

benzoate + L-arginine

Chronic Mgmt Low protein diet

–1.0 to 1.5 gm/kg/d

-Cyclinex (ess. AA’s)

(up to 50 % of prot)

Phenylbutyrate (Buphenyl)

(450-650mg/kg/d) Arg / ornith /

citrulline Regular monitoring

Case 5: Case 5: 5 year old girl with hearing loss and 5 year old girl with hearing loss and

macrocephalymacrocephaly Relatively normal Relatively normal

global development global development otherwiseotherwise

Family history Family history negative & parents negative & parents unrelated unrelated

S/S hepatomegaly, S/S hepatomegaly, mild contractures-mild contractures-hands, knees & hands, knees & elbowselbows

HM: Radiologicaal studyHM: Radiologicaal study

Case History: HMCase History: HM

You were asked to see this girl for You were asked to see this girl for assessment regarding macrocephaly assessment regarding macrocephaly and hearing lossand hearing loss

Initial investigations showed an Initial investigations showed an abnormal urine metabolic screen that abnormal urine metabolic screen that positive for both CTAB & Toluidine bluepositive for both CTAB & Toluidine blue

All other initial metabolic studies All other initial metabolic studies normalnormal

What would you do next?What would you do next?

““Coursened Features”Coursened Features”Laboratory Screening Laboratory Screening

ParadigmParadigm

Course facies

MPS in urine

Yes

No

MPS Storage Disease

Uronic acid

MPS by TLC

Enzyme Assays

Total HexMLS II or III

Oligosaccharidoses

Enzyme Assays

Oligoscreen

How do you measure How do you measure mucopolysaccharide mucopolysaccharide

excretion?excretion?Uronic acidUronic acid GAG GAG

electrophoresielectrophoresiss

What are the useful GAGS in diagnosing a

particular MPS disorder?

How is this testing done?

How do you measure How do you measure mucopolysaccharide mucopolysaccharide

excretion?excretion?Uronic acidUronic acid GAG electrophoresisGAG electrophoresis

Acid Hydrolysis to free

monosaccharide subunits

+

Colorimetric detection

of the uronic acid (A or B)

A B

Additional testing

Urine MPS excretion

Uronic acid by acid hydrolysis /carbazole

Uronic acid is 52 ug/ml

(Normal < 16.7)

Other methods

GAG electrophoresis ( HM

spec)

Diagnosis & Management

Mucopolysaccharidosis Type III San Filippo syndrome Four different enzyme deficiencies all leading to

inadequate breakdown of heparin sulfate She had deficiency of heparin-N-sulfatase (MPS-

IIIA)

No definitive treatment at present extensive social/psychological/educational

support appropriate for child with neurodegenerative

disorder

Case 6:Recurrent Abdominal Pain in 6

yo Male

History The mother of a six year old boy tells you that

her son has had three episodes of abdominal pain without any flu-like symptoms or other systemic problems.

During the last episode, blood but no bacteria or “pus” cells were seen in his urine. The urine did not smell, look cloudy or have an unusual color.

Her husbands had had similar episodes as a child but these had stopped once he followed the advice of a pediatrician to ”drink lots of water” at nights..

Your initial investigations showed that this boy had a normal physical exam, normal CBC.

‘lytes, urea & creatinine and aside from microscopic hematuria, a normal microscopic

& chemical urinalysis

What do you think is causing his abdominal pain & microscopic hematuria?

What genetic / metabolic disorders should you consider in your differential diagnosis list?

What specific testing would you suggest to investigate these possibilities?

Differential Diagnostic List

Cystinuria (basic aminoaciduria) Partial HGPRTase deficiency (uricosuria) GSD I (uricosuria) Primary hyperoxaluria (oxalic aciduria +/-

glycolic or glyceric acid) Idiop. Hypercalciuria (calcium oxalate or

urate) Hyperparathyroidism (calcium oxalate / urate) Adenosine phosporibosyl transferase

deficiency (dihydroxyadenosinuria)

Results of Special Results of Special TestingTesting

Unable to isolate characteristic stones in random urine specimen

Urine oxalic acid, calcium, uric acid / creatinine ratio all normal

urine oxypurine profile normal cystine, lysine, ornithine & arginine but

no other amino acids elevated in urine Plasma amino acid profile normal

What is your Diagnosis?

Cystinuria disorder of basic amino

acid transport involving renal tubule & GI mucosa

urolithiasis:poor

solubility of cystine when: urine concentrated

(cystine> 1200 uM/l) urine acidic

autosomal recessive 1: 7000

Types I, II & III (newer classification refers to type I as classical & others as non-classical)

Heterozygotes (type II) may excrete Cyst. Lys & Orn +/- arg but in reduced amts

Renal immaturity (< 1 yr) may cause “apparent” cystinuria in Type III carrier

Dibasic Amino Aciduria Normal: Cystine filtered at glomerulus but over 98

% reabsorbed in proximal renal tubule

Common renal tubular reuptake mechanism for dibasic amino acids (cystine,lysine, ornithine, arginine). All increased in urine but not blood if transporter is deficient

Nephrolithiasis: onset by 10 yrs (25-30 %) to 20 yrs (50-60 %) Cystine precipitates out in urine / renal filtrate at

concentrations above 1200 uM/l stones: hexagonal, golden-brown, grain size to staghorn size account for 1-3 % of all stones in adults (kids ?)

Subtypes based on amt excretion of cys/lys/orn but not arg in heterozygotes. Type III has similar defect in mucosal uptake

Molecular Genetics

Locus 1: 2p16.3-p21

Type I cystinuria rBAT protein SLC3A1 gene 40+ mutations Transmembrane

protein

Locus 2: 19q13.1

Non-Type 1 cystinuria Bo, +AT protein SLC7A9 gene 30 + mutations Complexes with rBAT

protein to form dibasic amino acid transporter

Phenotype/Genotype CorrelationCorrelation

Cystinuria I/I: two fully recessive SLC3A1 mutations no gut absorption kidney - high risk for nephrolithiasis

Cystinuria III/III two incompletely recessive SLC7A9 mutations stones in adults some gut absorption

Mixed types: I/III(A), I/III(B), II/II,

Treatment of Cystinuria dilution of urine:

drink up to 4.0 liters fluid / day (1.75 - 2.0 l/m2/24 hr) important to drink during the night time (water at

bedside) monitor urine cystine concentrations morning &

evening

“alkalinization” of urine: NaHCO 3 ( 1.5 - 2.0 mEq/kg/24 hr)

Medication: D-penicillamine, Thiola R (tiopronin) Surgical: lithotripsy etc.