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Diss Factsheets

Administrative data

Description of key information

Two GLP in vivo studies were conducted in guinea pigs to test the antigenicity and delayed hypersensitivity of AF0150 when injected intradermally.

AF150 is Tetradecafluorohexane embeded in lipid microspheres to allow injection and increase half-life by avoiding rapid elimination by exhalation. In the two studies presented hereby, AF150 is injected intradermally, mimicing a virtual 100% dermal absorption and an increased contact time of tetradecafluorohexane with cells involved in sensitization.

Therefore, these two in vivo studies are considered relevant to assess sensitization potential to tetradecafluorohexane.

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Referenceopen allclose all

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
November 1996
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail
Qualifier:
no guideline available
Principles of method if other than guideline:
20 male guinea pigs were assigned into 4 groups (5 animais/group) and treated. Different dilutions of serum samples collected in ASA procedure were intradermally injected into each animal with 0.05 ml per injection site and at least 1 cm apart between sites. At approximately 4.5 hours following serum injection, the animais in Groups 6, 7, and 8 received 1 ml/kg of Evan's Blue/AF0150 mixture (0.1% Evan's Blue and 20 mg/ml AF0150) by IV injection, and the animais in Group 9 received 0.5 ml/animal of Evan's Blue/BSA mixture (0.1% Evan's Blue and 10 mg/mi BSA). Approximately 30 minutes after IV injection, the diameter of blue dye staining area at each intradermal injection site was measured. Clinical signs, body weight and pathology were not observed.
GLP compliance:
yes
Type of study:
intracutaneous test
Justification for non-LLNA method:
Exeisting in vivo data relevant to provide information on sensitization potential of tetradecafluorohexane.
Species:
guinea pig
Strain:
other: Crl:(HA)BR
Sex:
not specified
Details on test animals and environmental conditions:
albino guinea pigs, Crl:(HA)BR strain, were 5-10 weeks old at initiation of treatment with
body weight of 470-645 g (for PCA). Standard procedures were followed for housing, handling, feeding and care of the animais. The animais were acclimated for 5 days before initiation of treatment.
Route:
intradermal
Vehicle:
water
Concentration / amount:
20 mg/mL
Day(s)/duration:
33
Adequacy of induction:
not specified
No.:
#1
Route:
intradermal
Vehicle:
water
Concentration / amount:
20 mg/kg
Day(s)/duration:
5h
Adequacy of challenge:
not specified
No. of animals per dose:
5
Details on study design:
AF0150 and BSA Preparation: AF0150 (200 mg fill vial) was reconstituted with 10 ml of sterile water to a concentration of 20 mg/ml and used within 30 minutes of preparation. 10 mg/ml of BSA was prepared by adding saline to 0.1 g BSA to a total volume of 10 ml. Saline (0.9% NaC1) was used as negative control. For induction where applied, AF0150 was mixed with FCA (Freund's Complete Adjuvant) at the ratio of 1:1 (v/v).
Procedure for Passive Cutaneous Anaphylaxis (PCA) (Table 2): 20 male guinea pigs were assigned into 4 groups (5 animais/group) and treated as shown in Table 2. Different dilutions of serum samples collected in ASA procedure were intradermally injected into each animal with 0.05 ml per injection site and at least 1 cm apart between sites. At approximately 4.5 hours following serum injection, the animais in Groups 6, 7, and 8 received 1 ml/kg of Evan's Blue/AF0150 mixture (0.1% Evan's Blue and 20 mg/ml AF0150) by IV injection, and the animais in Group 9 received 0.5 ml/animal of Evan's Blue/BSA mixture (0.1% Evan's Blue and 10 mg/mi BSA). Approximately 30 minutes after IV injection, the diameter of blue dye staining area at each intradermal injection site was measured. Clinical signs, body weight and pathology were not observed.

The serums were collected from animais sensitized with AF0150 or BSA or saline in ASA study. PCA Reactions were measured based on the diameter of blue dye staining area at serum injection sites.
Challenge controls:
Group
,., Treatment Dilutions Administered
6 Group 1 serum 1:1, 1:10, 1:100, 1:1,000, and 1:10,000
7 Group 3 serum 1:1, 1:10, 1:100. 1:1,000, and 1:10,000
8 Group 4 serum 1:1, 1:10, 1:100, 1:1,000, and 1:10,000
9 Group 5 serum 1:100, 1:1,000, and 1:10,000

Positive control substance(s):
yes
Remarks:
BSA
Positive control results:
the serum prepared from BSA-induced animais resulted in a positive PCA reactions at 9 of the 10 injection sites with 1:100 serum dilutions and at one site with 1:1000 serum dilution.
Key result
Reading:
other: area of blue dye staining at the injection sites
Hours after challenge:
5
Group:
negative control
Dose level:
0 mg/kg
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
No measurable area of blue dye staining at the injection sites
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
other: area of blue dye staining at the injection sites
Hours after challenge:
5
Group:
test chemical
Dose level:
20mg/kg
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
No measurable area of blue dye staining at the injection sites
Remarks on result:
no indication of skin sensitisation
Reading:
1st reading
Hours after challenge:
5
Group:
positive control
Dose level:
1:100
No. with + reactions:
9
Total no. in group:
10
Clinical observations:
positive PCA reactions
Remarks on result:
positive indication of skin sensitisation
Reading:
1st reading
Hours after challenge:
5
Group:
positive control
Dose level:
1:1000
No. with + reactions:
1
Total no. in group:
10
Clinical observations:
positive PCA reactions
Remarks on result:
positive indication of skin sensitisation
Interpretation of results:
other: No antigenicity
Executive summary:

No systemic or interdermal antigenicity was observed after a 33d induction phase.

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Remarks:
Guinea-Pig Maximization test
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
January 22 - february 24, 1997
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
not specified
GLP compliance:
yes
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
Exeisting in vivo data relevant to provide information on sensitization potential of tetradecafluorohexane.
Species:
guinea pig
Strain:
other: Crl:(HA)BR
Sex:
not specified
Details on test animals and environmental conditions:
34 albino guinea pigs, Crl:(HA)BR strain, were 4-8 weeks old at initiation of treatment with body weight of 400-501g. Standard procedures were followed for housing, handling, feeding and care of the animals.
Route:
intradermal and epicutaneous
Vehicle:
water
Concentration / amount:
20mg/mL / intra-dermal 0.1mL ; patches 2x4 cm
Day(s)/duration:
9
Adequacy of induction:
highest technically applicable concentration used
No.:
#1
Route:
epicutaneous, semiocclusive
Vehicle:
water
Concentration / amount:
20 mg/mL
Day(s)/duration:
2
Adequacy of challenge:
not specified
No. of animals per dose:
The animais were divided into 3 groups:
- irritation screening group (4)
- AF0150 group (20)
- control group (10).
Details on study design:
Table I. Dermal Reaction Scales
= No reaction
1 = Scattered mild redness
2 — Moderate and diffuse redness
3 = Intense redness and swellinu.
Positive control substance(s):
no
Positive control results:
There were no concurrent positive control groups included in this study. It is difficulty to judge the true negative results. In the context of the US-FDA NDA 21-191, the sponsor provided a separate report of a positive control study (using 2,4-dinitrochlorobenzene) in an Appendix. The study was conducted within 6 month of AF0150 study and followed the same procedure. A strong positive dermal reaction was found in guinea pigs induced and then challenged with 2,4-dinitrochlorobenzene.
Reading:
1st reading
Hours after challenge:
48
Group:
test chemical
Dose level:
20 mg/mL
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
no dermal reactions
Remarks on result:
no indication of skin sensitisation
Interpretation of results:
GHS criteria not met
Executive summary:

AF0150 alone or co-administrated with FCA had no significant dermal reactions using the guinea pig maximization test (Magnusson and Kligman Assay). However, with lack of a concurrent positive control in the same study (although positive control study was conducted separately at different time), it is difficulty to conclude that AF0150 has no contact hypersensitivity in guinea pigs.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not sensitising)
Additional information:

Two GLP in vivo studies were conducted in guinea pigs to test the antigenicity and delayed hypersensitivity of AF0150.

The antigenicity (IMUS-021-TOX) was evaluated with active systemic anaphylaxis (ASA) and passive cutaneous anaphylaxis (PCA) methods. Animals were first induced by subcutaneous injection of 2 mg AF0150 with and without Freund's Complete Adjuvant (FCA). Two weeks following induction, blood samples were collected and meantime the animals received IV injection of 20 mg/kg AF0150 (35-fold PCD) for challenge (ASA test). Imunoactivity of serum from the blood samples were assayed with a skin test in a separate guinea pig groups (PCA test).

AF0150 did not induce significant active systemic anaphylactic or passive cutaneous anaphylactic reactions.

The delayed hypersensitivity (IMUS-029-TOX) of AFOI 50 was tested in guinea pigs using Magnusson and Kligman maximization assay (guinea pig maximization study). The animals were subjected to intradermal injection of AF0150 (20 mg/mL) with and without FCA for induction, followed by skin patching (with filter papers saturated with 20 mg/mL AF0150) for challenge. The dermal reactions were recorded following AF0150 skin patching, and no significant findings were observed. However, no concurrent but separate positive control group was included in the study to validate the assay system.

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not sensitising)

Justification for classification or non-classification

No indication of antigenicity nor delayed hypersensitivity was observed in two in vivo studies considered relevant for sensitization potential of tetradecafluorohexane.