Laboratoire Philippe Auguste1 Robert Nataf M.D. Laboratoire Philippe Auguste 119Avenue Philippe Auguste 75011 Paris France Tel: (33)1.43.67.57.00

Laboratoire Philippe Auguste 1

Robert Nataf M.D.Laboratoire Philippe Auguste119 Avenue Philippe Auguste75011 Paris Francehttp://www.labbio.netTel: (33)1.43.67.57.00 Fax: (33)1.43.79.00.27Email : [email protected]

Evidence for Mercury Toxicity in Autism through Urinary Porphyrin

Profiling

ADHDAsperger’s syndrome

PDDAutism

http://www.labbio.net/







Urinary Porphyrin Levels: A Biomarker of Environmental Toxicity


What are the porphyrins?

Porphyrins are cruciform compounds synthesized by all the cells of the living world which constitute the active sites of the hemoproteins which transport oxygen Hb & Mb, ensure the energy production Cytochromes A3, B, C and the detoxication of xenobiotic, Cytochromes P450.


Polychorinated Biphenyl (PCB) UroPArsenic (As) 7cxPAluminium (Al)

Mercury (Hg) 5 cxP PcP

CoP

Lead (Pb) CoP

URO-D = Uroporphyrin Decarboxylase COP-O Coproporphyrin oxidase


Mercury (Hg), Lead (Pb), Arsenic (As), Polychlorinated Biphenyls and certain pesticides , are now known to cause many neurodevelopmental disabilities such as

- Learning or Intellectual Retardation,

- Dyslexia,

- Attention Deficit

- Hyperactivity disorders

- Autism.

Toxic chemicals in the environment


Hemogloblin & Myoglobin

Respiratory Chain Cytochromes :

CYT A3, CYT B, CYT C and CYT C1

Cytochrome P450 of Phase I detoxification


8 enzymes ensured in mitochondrion and the cytoplasm, the synthesis of porphyrins starts from the glycine and the succinyl-CoA. They have a different sensitivity to heavy metals and derivative organiccompounds which allows to find the intoxication type.

PORPHYRINS SYNTHESIS


Delta Aminolevulinoic acid (ALA)

Porphobilinogen

7-carboxyporphyrinogen III

6-carboxyporphyrinogen III 5-carboxyprophyrinogen III

Coproporphyrinogen III

Protoporphyrinogen IX

Protoporphyrinogen IX

Heme


Xenobiotic compounds


7-carboxy

Uro

Uro 7-carboxy Xenobiotic Impacts

04/12/2006

HPLC-UV+Fluorescence (nmol / urinary gCr )

patients reference

60 10-30

15,9 1,5-4,0

3,2 0,5-2,0

10,7 1,6-3,0

12,9 <7

257 40-80

Pentacarboxy porphyrin (5cxP)

Coproporphyrins I & III (cP)

P orphyrins

U r i n a r y p o r p h y r i ns

Uroporphyrins I &III (UP)Heptacarboxy porphyrin (7cxP)Hexacarboxy porphyrin (6cxP)

Precoproporphyrin (Preco)

LABORATOIRE PHILIPPE AUGUSTE119 Ave Philippe Auguste 75011 Paris France Tel (33) 1 43 67 57 00 fax (33) 1 43 79 00 27 Email : [email protected]

0

10

20

30

40

50

60

70

80

90

100

uP 7cxP 6cxP 5cxP prcP cP

nmol/g urinary Cr

norme max

average slightly increased rate increased rate very high rate


Uro 7-carboxy Xenobiotic Impacts

7-carboxy

Uro04/12/2006


patients reference

95 10-30

29,2 1,5-4,0

6,4 0,5-2,0

26,5 1,6-3,0

53,0 <7

699 40-80



Porphyrins





0

10

20

30

40

50

60

70

80

90

100


nmol/g urinary Cr

norme max



Aluminium (Al)


Arsenic (As)


Lead (Pb)


Mercury (Hg)


8-CP 7-CP 6-CP 5-CP 4-CP 2-CP Heme

KICP

HgHg

CPOXUROD

CPOX4

Role of Mercury , CPOX and UROD in Heme biosynthetic pathway

James S. Woods et al.. Texicology Letter 2005….


Polymorphism Influence of Cop-Ox on Mercury toxicity reflected by Porphyrin Profile

CPOX

Wild Cop-Ox 85% Cop-Ox

mutation N272H exon 4

15%


CoproPrecopro

5-carboxy

5-carboxy Precopro Copro Mercury Impacts

04/12/2006


patients reference

24 10-30

8,6 1,5-4,0

1,8 0,5-2,0

13,7 1,6-3,0

41,6 <7

405 40-80



Porphyrins





0

10

20

30

40

50

60

70

80

90

100


nmol/g urinary Cr

norme max



CoproPrecopro

5-carboxy

5-carboxy Precopro Copro Mercury Impact

04/12/2006


patients reference

20 10-30

6,0 1,5-4,0

1,1 0,5-2,0

12,0 1,6-3,0

33,5 <7

394 40-80



Porphyrins





0

10

20

30

40

50

60

70

80

90

100


nmol/g urinary Cr

norme max




Saline injection

1st DMPS injection

2nd DMPS injection

3rd DMPS injection

Effects of consecutive DMPS injections on urinary porphyrin concentration in MMH-exposed rats


Porphyrin

Total Hg Hg 2+ CH3Hg+

Penta 0.927 0.984 0.876

Precopro 0.961 0.884 0.987

Copro 0.998 0.989 0.984

Correlation coefficient (r)

Linear Relationship of Prechelation Urinary Porphyrin Concentrations with Urinary Mercury Concentrations

Porphyrin

Total Hg Hg 2+ CH3Hg+

Penta 0.967 0.996 0.879

Precopro 0.895 0.987 0.762

Copro 0.989 0.908 0.984

Correlation coefficient (r)


S.D. Pingree, P.L.Simmonds, K.T. Rummel, J.S. Woods,Toxicological Sciences 61, 234-240 (2001)

• Mercury exposure elicits characteristics changes in the urinary porphyrin excretion Pattern.

• Urinary porphyrin levels in MMH-exposed rats were highly correlated with kidney mercury levels and declined with consecutive DMPS treatments.

• Prechelation urinary porphyrin levels are highly correlated with Postchelation urinary.

• Prechelation urinary porphyrin levels are highly correlated with Renal mercury Contents.

Porphyrins Measures the Kidney Mercury content

and Mercury Body Burden

during Prolonged Methylmercury Exposure in Rat


Heavy metal toxicity, particularly mercury, has been implicated in the etiology of autism and related spectrum disorders.

Despite several studies addressing heavy metal exposure through elemental analysis and epidemiology, direct evidence for mercury toxicity in autism is lacking. We report a study of urinary porphyrins, markers of heavy metal exposure, in 269 children with neurodevelopmental disorders. The specific porphyrin profile in a subset of children with autistic syndrome points to mercury toxicity in this disorder.

Mercury & Autism


COPRO (autist)

COPRO (control)

PRECOPRO (control)

PRECOPRO (autist)

5-carboxy (autist)

5-carboxy(control)

Control

Autist

5cxP PrP Copro


CoproPrecopro

5-carboxy

04/12/2006


patients reference

24 10-30

8,8 1,5-4,0

1,8 0,5-2,0

13,7 1,6-3,0

41,6 <7

405 40-80



Porphyrins





0

10

20

30

40

50

60

70

80

90

100


nmol/g urinary Cr

norme max



CoproPrecopro

5-carboxy

04/12/2006


patients reference

20 10-30

6,0 1,5-4,0

1,1 0,5-2,0

12,0 1,6-3,0

33,5 <7

394 40-80



P orphyrins





0

10

20

30

40

50

60

70

80

90

100


nmol/g urinary Cr

norme max



A study on urinary porphyrin levels in 256 children with neurodevelopmental and

related disorders including 106 with autistic disorder.

Nataf R, Skorupka C, Amet L, Lam A, Springbett A, Lathe R, Porphyrinuria in childhood autistic disorder


Condition/diagnosis M F TotalMean age (yr)

M/F % total% ASD group

Allergy 5 3 8 7,3 1,67 3Asperger 10 1 11 10 10 4,1 5,8

Attention deficit 2 7 9 9,4 0,29 3,3Autism (autistic disorder) 79 27 106 6,4 2,9 39 55,5

Autism+epilepsy 7 2 9 9,3 3,5 3,3Cerebral palsy 6 6 12 8,3 1 4,4

Epilepsy 2 0 2 10 na 0,7Hyperactivity 27 2 29 9,1 13,5 10,7MR+ epilepsy 1 1 2 6 1 0,7

PDD-NOS 51 12 63 6,6 4,3 23,4 33Psychomotor retardation 1 3 4 7,3 0,33 1,5

Rett 0 2 2 2,5 0 0,7Control group 7 5 12 10,3 1,4 4,4 1

TOTAL 198 71 269 7,4 2,8

ASD= 71% of

total sample (M/F=3,34)

269 Study Subjects (2002-2004)


0

10

20

30

40

50

60

70

80

ALLERGYASPERGER

ATTENTION DEFICIT

AUTISM

AUTISM + EPILEPSYCEREBRAL PALSY

EPILEPSY

HYPERACTIVITYMR + EPILEPSY

PDD-NOS

PSYCHOMOTOR RET

RETT

CONTROL GouRP

Coproporphyrin (micromol/mol CRT)

n=2

n=2

n=2

n=4

CTRL MEAN

CTRL 2 x SD

Coproporphyrin levels in urines of children with neurodevelopmental and related disorders


0

5

10

15

20

25

ASP AUT

AUT+EPICTRL

COPRO/URO ratio

0

10

20

30

40

ASP AUT

AUT+EPICTRL

Coproporphyrin

)

Elevated urinary Coproporphyrin (COPRO) levels in ASD expressed as absolute values normalised to creatinine (left) or

as an internal ratio with uroporphyrin (URO) (right)


Precoproporphyrin and pentacarboxy porphyrin: Markers of heavy metal toxicity

0

2

4

6

8

0 2

0

2

4

6

8

0 2

Control ASP

(0.62)

(0.56)

Precoproporphyrin plotted against baseline uroporphyrin : the ratio is independent of age-related creatinine variation

0

1

2

3

4

5

6

7

8

0 2 4

AUT +EPI

0

1

2

3

4

5

6

7

8

0 2 4

AUT

(1.09)

(1.28)


0

1

2

3

4

5

ASP PDD-NOS AUT AUT+EPI CTL

Porphyrin ratios to control group values

UROP

7CXP

6CXP

5CXP

COP

PRECOP

Spectrum of mean porphyrin excess, expressed as a ratio of control group (CTL) value for the different porphyrin subtypes


Outcome of chelation

DMSA (Dimercaptosuccinic

Acid )

0

20

40

60

1 2 3

Coproporphyrin (millimol/mol creatinine)

1st 2nd 1st 2nd

Chelation(DMSA)

NoChelation

Non-ASDControl Group

Reduction in urinary coproporphyrin on chelation with DMSA


Conclusion

• Porphyrinuria is a reliable marker of environmental toxicity

• The increasing population exposure to heavy metals including mercury evidence of increasing prevalence of autistic disorder.


Urinary Porphyrins

To affirm the intoxication ex. toxic impact on the physiology of the organism. When a numerical biological value is not enough

To Orient the diagnosis etiologic by certain profiles to porphyrins (mercury).

To measure the overload through the intensity of the toxic impact exerted by the biologically active compartment thus pathogenic.

To Monitor the detoxication (chelation.) through the reduction of the toxic effect.


Urinary Porphyrin Levels: A Biomarker of Environmental Toxicity

Nataf R, Skorupka C, Amet L, Lam A, Springbett, Lathe R, Porphyrinuria in childhood autistic disorder, The Lancet Neurology (in press)


Thank you

Documents

Laboratoire Philippe Auguste1 Robert Nataf M.D. Laboratoire Philippe Auguste 119Avenue Philippe Auguste 75011 Paris France Tel: (33)1.43.67.57.00