Cefic-lri Programme | European Chemical Industry Council

B14-RIFM: Skin Sensitisation – Chemical Applicability Domain of the Local Lymph Node Assay (LLNA)

Principal Investigator

Dr. Anne Marie Api

Research Institute for Fragrance Materials, Inc.

50 Tice Boulevard

Woodcliff Lake, NJ 07677

E-mail: AApi@rifm.org

Tel: +1 201 689 8089

Fax: +1 201 689 8090


David W Roberts PhD, C.Chem., Eur.Chem., FRSC, d.w.roberts@ljmu.ac.uk

Grace Patlewicz PhD, DuPont, Grace.y.tier@dupont.com

Jon F Lalko PhD, RIFM, Jlalko@rifm.org



Since the publication of the OECD AOP for skin sensitisation in 2012, there have been even greater efforts to anchor the development and evaluation of new in vitro and in chemico assays to key events within the AOP. Much of that type of evaluation is likely to use the Local Lymph Node Assay (LLNA) as its benchmark. Whilst the LLNA has been through an extensive validation relative to the incumbent assay of the time, the Guinea Pig Maximisation Test (GPMT), to date there has been no systematic evaluation of the chemical applicability domain of the LLNA, i.e. an exercise to define in chemical terms which classes of chemicals are well predicted, which are liable to be wrongly predicted and which are unpredictable by this in vivo method. “Wrongly predicted” in this context is being defined to encompass false negatives, false positives, as well as false relative potencies – i.e. if chemical X has a lower EC3 (concentration of chemical that when given as per the LLNA protocol stimulates a threefold increase in cell proliferation in the lymph node) than chemical Y, thus more potent when human and/or guinea pig evidence indicates the contrary, this would be construed as a deficiency in the LLNA.

The chemical domain of the LLNA is particularly important when considering outcomes of new in vitro test methods for sensitisation in order to be able to determine whether a given result is reasonable or whether the outcome is impacted by uncertainties in the LLNA which is typically used as a basis of comparison.

From a practical perspective, the applicability domain could be defined in terms of the chemicals that were used in the original validation exercises: compounds whose relevant chemical properties (such as reaction mechanism, hydrophobicity, reactivity) were not represented in the validation set are therefore deemed to be outside of the applicability domain.

The aim of this project therefore is to define the range of chemicals for which the LLNA result can be considered a reliable (quantitative) indicator of the chemical’s potential to sensitise, can therefore be carried forward for subsequent risk assessment purposes and/or be relied on as the benchmark when developing or evaluating new in vitro assays.

Related Publications


Skin Sensitisation – Chemical Applicability Domain of the Local Lymph Node Assay by Dr. Dave Roberts, Univ. of Liverpool, UK, Cefic-LRI Annual Workshop 2015

Correlation between experimental human and murine skin sensitization induction thresholds (2014) by Api A.M., Basketter D. and Lalko J.

Skin Sensitisation – Chemical Applicability Domain of the Local Lymph Node Assay (LLNA), presented at the 16th Cefic-LRI Annual Workshop 2014


Timeline: July 2014 > June 2015

LRI funding: € 60.000

Cefic-Lri Programme Responsible Care

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