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Toxicological information

Skin sensitisation

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Administrative data

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Remarks:
Human Repeat Insult Patch Test
Type of information:
experimental study
Adequacy of study:
key study
Study period:
4 January to 16 March 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail
Cross-reference
Reason / purpose:
reference to same study
Reference
Endpoint:
sensitisation data (humans)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
4 January to 16 March 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail
Type of sensitisation studied:
skin
Study type:
study with volunteers
Principles of method if other than guideline:
MARZULLI-MAIBACH METHOD: The Marzulli-Maibach test is a reference test to study the sensitizing potential of a product. The investigational product was applied under maximized conditions (occlusion) during 3 weeks (Induction phase). After a two-week resting period, the investigational product was re-applied on the same site (Challenge phase). If the investigational product was allergenic, it induced a cutaneous reaction during the challenge phase.
GLP compliance:
no
Remarks:
Good clinical practice
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Robertet / 1968538
- Appearance: Orange-Brown Solid

Type of population:
general
Ethical approval:
confirmed and informed consent free of coercion received
Subjects:
- Number of subjects recruited: 115
- Number of subjects exposed: 102
- Sex: 26 men and 76 women
Clinical history:
The subjects were included according to the following criteria:
Inclusion criteria:
The voluntary subjects satisfied the following criteria:
Female or male,
Aged from 18 to 70 years old,
Not showing tattoo, scars, pigmented areas, important pilosity and dermatological lesions on the back and not receiving any anti-inflammatory or anti-allergic medication,
Subjects with Social Security (health insurance) coverage (according to the requirements of the MACEDONIAN REPUBLIC),
Able and willing to give a written informed consent.
 
Non-inclusion criteria:
The voluntary subjects did not satisfy the following criteria:
Who had applied a care product on the sites of application during the 24 hours before the beginning of the study,
Who had not washed the sites of the application with water 2 hours before the beginning of the study,
Pregnant woman or in the process of breast feeding,
Who had taken part to a patch test in the 15 days before the beginning of the study,
Who had tattoos on the back,
Who had scars on the back,
Who showed pigmented areas on the back,
Who had a very important pilosity on the back,
Controls:
A semi occlusive patch receiving no product was applied in the same conditions and was considered as a non-treated control.
Route of administration:
dermal
Details on study design:
LOCATION: The study was carried out at the clinical unit of Skopje in MACEDONIA.

DESIGN: The study was blinded and randomized.

TYPE OF TEST(S) USED: Patch test (epicutaneous test)

ADMINISTRATION
- Type of application: Semiocclusive
- Description of patch: Trumed® Inc (size: 1.9 cm x 1.9 cm i.e. 3.61 cm2)
- Frequency: Induction phase: 3 times a week during 48 hours or 72 hours; Challenge phase: once during 48 hours
- Duration: Induction phase: 3 weeks; Duration Resting phase: 14 days; Challenge phase: 1 week
- Concentrations: Undiluted (8% (3646 µg/cm2) EtOH/DEP(25/75))
- Volume applied: 160 µL
- Removal of test substance: After removal of the patches, the sites were wiped with a cotton pad soaked with warm distilled water.

CONDITIONS OF USE:
INDUCTION PHASE:
- During the Induction Phase, the investigational product was applied on a predetermined site under semi occlusive patch (the semi occlusive patches were applied, on a right split back, in a zone limited by the spine of the scapula in top, the last coast in bottom, the dorsal rachis in inside and a vertical fIctitious line passing by the lower end of the scapula outwards). A semi-occlusive patch receiving no product was applied in the same conditions and was considered as a non-treated control. Each time, the patches (treated and non-treated) remained applied during 48 or 72 hours before removal and replaced on the same site.
- The investigator performed:
A clinical examination before each application of the investigational products; the application of the patches (treated and non-treated) on Days 1, 3, 5/6*, 8, 10, 12/13*, 15, 17, 19/20* and 22**.
At each visit, the patches were replaced by one with the same investigational product and applied on the same site (except in case of occurrence of a reaction making the application of new patches impossible).
* Subjects no. 104, 105, 106, 107 and 108 began the study 2 days later, so they did not have any visit at Day 5, Day 12 and Day 19, but at Day 6, Day 13 and Day 20.
** Additional application day for subjects no. 3, 13, 67, 69, 89, 95, 97 and 103, as they did not have any application nor reading performed one day during the induction phase.

RESTING PHASE:
- During the 14-day Resting Phase, no product was applied on the sites used during the Induction phase or on the contralateral split-back which was used during the Challenge phase.

CHALLENGE PHASE:
- For the challenge phase, a symmetrical zone of controlateral split back (left) is used as the zone tested for the induction phase.
- During the challenge phase (Day 34/36*), the investigational product was applied under semi occlusive patch on the induction sites and on the contralateral sites. A semi occlusive patch receiving no product was applied in the same conditions and was considered as a non-treated control. The patches (treated and non-treated) remained applied during 48 hours and removed afterwards.
- A series of reading were practiced immediately (Day 36/38*), 24 hours (Day 37/39*), and 48 hours (Day 38/40*) after the patches removal.
* Subjects no. 104, 105, 106, 107 and 108 began the study 2 days later, therefore their challenge phase was performed on Day 34, Day 36, Day 37 and Day 38 instead of Day 36, Day 38, Day 39 and Day 40.

EXAMINATIONS
- Grading/Scoring system:
The clinical examination was carried out on the back on the treated zones according to the following scale:
Erythema (E)
0: no reaction
1: very slight reaction: pale pink erythema, homogeneous, not well delimited
2: slight reaction: clear pink erythema, homogeneous, well delimited, without oedema
3: moderate reaction: clear red erythema, homogeneous, well delimited, with or without slight oedema
4: severe reaction: dark red erythema, homogeneous, well delimited with or without an obvious oedema

Vesicles (V)
0: no reaction
1: obvious vesicles

Pruritus (P)
0: no reaction
1: obvious itching

Desquamation (visual and tactile evaluation) (D)
0: no reaction
1: fine desquamation
2: moderate desquamation
3: severe with large squamas

OTHERS:
Constraints of the study:
- During the Induction Phase, the patches application areas had not to be wet.
- During the repeated readings of the challenge phase, no wash, except for the daily personal hygiene habits, or any application of any kind of product was allowed.
- During the whole test, sun or UV voluntary exposure was not allowed.
Results of examinations:
SYMPTOMS
- During the induction phase, no sign was observed on the control site and the investigational product site.
- During the challenge phase, no sign was observed on the control site and on the investigational product site, neither on the homolateral side nor on the controlateral site.
- In this study, no adverse event or serious adverse event was observed by the investigating dermatologists.

NO. OF PERSONS WITH/OUT REACTIONS COMPARED TO STUDY POPULATION
- Number of subjects with positive reactions: 0
- Number of subjects with negative reactions: 102
- Number of subjects with irritating reactions: 0

DETERMINATION OF THE SENSITIZING POTENTIAL:
No significant clinical manifestation of intolerance or allergy was observed by the investigator.

DETERMINATION OF THE IRRITATION INDEX Z:
The investigational product presents an irritation index of: Z = 0.000; the investigational product is therefore considered as practically non-irritant.
Note: Z is calculated on the parameters erythema (including oedema) and desquamation; Z is calculated on the data of the induction phase.

Table 7.10.4/1: Induction phase – results

Days

Investigational product

% of subjects - reaction

Semi-Occlusive control

% of subjects - reaction

Day 1

Normal examination

0

Normal examination

0

Day 3

No reaction

0

No reaction

0

Day 5/6*

No reaction

0

No reaction

0

Day 8

No reaction

0

No reaction

0

Day 10

No reaction

0

No reaction

0

Day 12/13*

No reaction

0

No reaction

0

Day 15

No reaction

0

No reaction

0

Day 17

No reaction

0

No reaction

0

Day 19/20*

No reaction

0

No reaction

0

Day 22

No reaction

0

No reaction

0

Day 24**

No reaction

0

No reaction

0

* Subjects no. 104, 105, 106, 107 and 108 began the study 2 days later, so they did not have any visit at Day 5, Day 12 and Day 19, but at Day 6, Day 13 and Day 20.

** Additional application day for subjects no. 3, 13, 67, 69, 89, 95, 97 and 103 as they did not have any application nor reading performed one day during the induction phase.

Table 7.10.4/2: Challenge phase – results

Days

Sites

Investigational product

% of subjects - reaction

Semi-Occlusive control

% of subjects - reaction

Day 34/36*

H

Normal examination

0

Normal examination

0

C

Normal examination

0

Normal examination

0

Day 36/38*

H

No reaction

0

No reaction

0

C

No reaction

0

No reaction

0

Day 37/39*

H

No reaction

0

No reaction

0

C

No reaction

0

No reaction

0

Day 38/40*

H

No reaction

0

No reaction

0

C

No reaction

0

No reaction

0

H: homolateral site;C: contralateral site

* Subjects no.104, 105, 106, 107 and 108 began the study2days later, therefore their challenge phase was performed on Day34,Day 36,Day37and Day38instead of Day36,Day38,Day39and Day 40.

Conclusions:
Under the conditions of the study, the investigational product FENUGREEK ABS 220275 1% (456 µg/cm2) EtOH/DEP(25/75) tested undiluted, under semi occlusive patch, can be considered as non-sensitizing.
Executive summary:

A cutaneous tolerance and sensitizing potential study of the investigational product, FENUGREEK ABS 220275 1% (456 µg/cm2) EtOH/DEP(25/75) tested undiluted was conducted by patches application, according to the Marzulli-Maibach sensitization method. The data of 102 female and male subjects aged from 19 to 63 years old were analyzed during the study. The subjects were willing and able to give a written informed consent. The study was carried out at the clinical unit of Skopje in MACEDONIA. The study was blinded and randomized.

Duration:Induction phase: 3 weeks (3 times a week during 48 hours or 72 hours); Duration Resting phase: 14 days; Challenge phase: 1 week (once during 48 hours)

 

INDUCTION PHASE: During the Induction Phase, the investigational product was applied (160 µL) on a predetermined site under semi occlusive patch (back). A semi-occlusive patch receiving no product was applied in the same conditions and was considered as a non-treated control. Each time, the patches (treated and non-treated) remained applied during 48 or 72 hours before removal and replaced on the same site.

RESTING PHASE: During the 14-day Resting Phase, no product was applied on the sites used during the Induction phase or on the contralateral split-back which was used during the Challenge phase.

CHALLENGE PHASE: During the challenge phase (Day 34/36*), the investigational product was applied under semi occlusive patch on the induction sites and on the contralateral sites. A semi occlusive patch receiving no product was applied in the same conditions and was considered as a non-treated control. The patches (treated and non-treated) remained applied during 48 hours and removed afterwards.

 

A clinical examination was performed by the investigating dermatologist during the Induction Phase on Days 1, 3,5/6,8, 10,12/13,15, 17,19/20and22/24,and during the Challenge Phase on Days34/36, 36/38, 37/39and38/40.

 

During the induction phase, no sign was observed on the control site and the investigational product site. During the challenge phase, no sign was observed on the control site and on the investigational product site, neither on the homolateral side nor on the controlateral site. In this study, no adverse event or serious adverse event was observed by the investigating dermatologists. No significant clinical manifestation of intolerance or allergy was observed by the investigator. The investigational product presents an irritation index of: Z = 0.000; the investigational product is therefore considered as practically non-irritant.

 

Under the conditions of the study, the investigational product FENUGREEK ABS 220275 1% (456 µg/cm2) EtOH/DEP(25/75) tested undiluted, under semi occlusive patch, can be considered as non-sensitizing.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2012
Report Date:
2012

Materials and methods

Test guideline
Qualifier:
no guideline followed
Principles of method if other than guideline:
MARZULLI-MAIBACH METHOD: The Marzulli-Maibach test is a reference test to study the sensitizing potential of a product. The investigational product was applied under maximized conditions (occlusion) during 3 weeks (Induction phase). After a two-week resting period, the investigational product was re-applied on the same site (Challenge phase). If the investigational product was allergenic, it induced a cutaneous reaction during the challenge phase.
GLP compliance:
no
Remarks:
Good clinical practice
Type of study:
other: Study with volunteers
Justification for non-LLNA method:
Available study on volunteers

Test material

Reference
Name:
Unnamed
Type:
Constituent
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Robertet / 1968538
- Appearance: Orange-Brown Solid

In vivo test system

Test animals

Species:
other: Human
Strain:
other: Not relevant
Sex:
male/female
Details on test animals and environmental conditions:
- Number of subjects recruited: 115
- Number of subjects exposed: 102
- Sex: 26 men and 76 women

Results and discussion

Positive control results:
No positive control

In vivo (non-LLNA)

Resultsopen allclose all
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
FENUGREEK ABS 220275 1% (456 µg/cm2) EtOH/DEP(25/75) lot 1968538
No. with + reactions:
0
Total no. in group:
102
Clinical observations:
No significant clinical manifestation of intolerance or allergy was observed by the investigator.
Remarks on result:
no indication of skin sensitisation
Reading:
other: Non performed
Group:
negative control
Remarks on result:
not measured/tested
Reading:
other: Non performed
Group:
positive control
Remarks on result:
not measured/tested

Any other information on results incl. tables

Table 7.10.4/1: Induction phase – results

Days

Investigational product

% of subjects - reaction

Semi-Occlusive control

% of subjects - reaction

Day 1

Normal examination

0

Normal examination

0

Day 3

No reaction

0

No reaction

0

Day 5/6*

No reaction

0

No reaction

0

Day 8

No reaction

0

No reaction

0

Day 10

No reaction

0

No reaction

0

Day 12/13*

No reaction

0

No reaction

0

Day 15

No reaction

0

No reaction

0

Day 17

No reaction

0

No reaction

0

Day 19/20*

No reaction

0

No reaction

0

Day 22

No reaction

0

No reaction

0

Day 24**

No reaction

0

No reaction

0

* Subjects no. 104, 105, 106, 107 and 108 began the study 2 days later, so they did not have any visit at Day 5, Day 12 and Day 19, but at Day 6, Day 13 and Day 20.

** Additional application day for subjects no. 3, 13, 67, 69, 89, 95, 97 and 103 as they did not have any application nor reading performed one day during the induction phase.

Table 7.10.4/2: Challenge phase – results

Days

Sites

Investigational product

% of subjects - reaction

Semi-Occlusive control

% of subjects - reaction

Day 34/36*

H

Normal examination

0

Normal examination

0

C

Normal examination

0

Normal examination

0

Day 36/38*

H

No reaction

0

No reaction

0

C

No reaction

0

No reaction

0

Day 37/39*

H

No reaction

0

No reaction

0

C

No reaction

0

No reaction

0

Day 38/40*

H

No reaction

0

No reaction

0

C

No reaction

0

No reaction

0

H: homolateral site;C: contralateral site

* Subjects no.104, 105, 106, 107 and 108 began the study2days later, therefore their challenge phase was performed on Day34,Day 36,Day37and Day38instead of Day36,Day38,Day39and Day 40.

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
Under the conditions of the study, the investigational product FENUGREEK ABS 220275 1% (456 µg/cm2) EtOH/DEP(25/75) tested undiluted, under semi occlusive patch, can be considered as non-sensitizing.
Executive summary:

A cutaneous tolerance and sensitizing potential study of the investigational product, FENUGREEK ABS 220275 1% (456 µg/cm2) EtOH/DEP(25/75) tested undiluted was conducted by patches application, according to the Marzulli-Maibach sensitization method. The data of 102 female and male subjects aged from 19 to 63 years old were analyzed during the study. The subjects were willing and able to give a written informed consent. The study was carried out at the clinical unit of Skopje in MACEDONIA. The study was blinded and randomized.

Duration:Induction phase: 3 weeks (3 times a week during 48 hours or 72 hours); Duration Resting phase: 14 days; Challenge phase: 1 week (once during 48 hours)

 

INDUCTION PHASE: During the Induction Phase, the investigational product was applied (160 µL) on a predetermined site under semi occlusive patch (back). A semi-occlusive patch receiving no product was applied in the same conditions and was considered as a non-treated control. Each time, the patches (treated and non-treated) remained applied during 48 or 72 hours before removal and replaced on the same site.

RESTING PHASE: During the 14-day Resting Phase, no product was applied on the sites used during the Induction phase or on the contralateral split-back which was used during the Challenge phase.

CHALLENGE PHASE: During the challenge phase (Day 34/36*), the investigational product was applied under semi occlusive patch on the induction sites and on the contralateral sites. A semi occlusive patch receiving no product was applied in the same conditions and was considered as a non-treated control. The patches (treated and non-treated) remained applied during 48 hours and removed afterwards.

 

A clinical examination was performed by the investigating dermatologist during the Induction Phase on Days 1, 3, 5/6, 8, 10,12/13,15, 17,19/20 and 22/24,and during the Challenge Phase on Days 34/36, 36/38, 37/39 and 38/40.

 

During the induction phase, no sign was observed on the control site and the investigational product site. During the challenge phase, no sign was observed on the control site and on the investigational product site, neither on the homolateral side nor on the controlateral site. In this study, no adverse event or serious adverse event was observed by the investigating dermatologists. No significant clinical manifestation of intolerance or allergy was observed by the investigator. The investigational product presents an irritation index of: Z = 0.000; the investigational product is therefore considered as practically non-irritant.

 

Under the conditions of the study, the investigational product FENUGREEK ABS 220275 1% (456 µg/cm2) EtOH/DEP(25/75) tested undiluted, under semi occlusive patch, can be considered as non-sensitizing.