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2019 Society of Toxicology Annual Meeting

MB Research Labs - Protecting You Like One of Our Own.

Join Us at the 58th Annual Meeting of the Society of Toxicology and ToxExpo, by the waterfront in Baltimore, MD, March 10- 14, 2019

  • MB Research ToxExpo Booth # 4157
  • MB In Vitro & Alternative Toxicology Posters
  • About MB Research

    MB Research Labs, a Contract Toxicology Testing organization, serves the in vivo and in vitro toxicology testing needs of both government and industry.

    Complete support services including consultation, protocol development, quality assurance, analytical chemistry and archive facilities are integral components of studies performed at MB.

    Our technical and support procedures are in full compliance with OECD, FDA, and EPA-OCSPP. For more information about our capabilities, please click here.

    Industries Served:

    OECD, FDA, EPA, OCSPP. Regulatory Toxicity Testing.Regulatory Toxicology
    Chemical Toxicity TestingChemical Toxicity
    Cosmetics, Cosmetic Ingredients, Consumer Products Safety AssessmentsCosmetics & Personal Care

    MB conducts Good Laboratory Practice (GLP) compliant toxicology assays as well as low-cost screening studies for clients in the cosmetic, consumer product, chemical, biotech and pharmaceutical industries.

    Scientific Poster Presentations

    Wed 9:15 AM - 4:30 PM. Session: Ocular Toxicity, Abstract 2959, Poster Board P401
    E. Ditzel1, P. Vij2, D. Sergeyev2, B. Varsho2, G. DeGeorge2
    1. Ecolab, Eagan, MN
    2. MB Research Laboratories, Spinnerstown, PA

    The PorCORA is a 21-day ex vivo ocular assay, which can distinguish between a substance’s potential to cause severe (reversible) versus corrosive (irreversible) damage. We used the PorCORA’s ability to determine GHS category 1 versus category 2 (2A/2B) of a Fryer Cleaner and Degreaser (FC-D) mixture formula. The Bovine Corneal Opacity and Permeability (BCOP) In Vitro Irritancy Score (IVIS) for a prototypical FC-D was 24.3 and its amine-based cogeners ranged between 35.5 and 50.8. However, the BCOP assay does not directly address reversibility of ocular damage, which is a key component of the GHS classification scheme for ocular irritants. The PorCORA revealed complete healing of the cornea by study day 7, indicating reversible eye irritation and non-corrosivity. The reversibility of damage for this material suggests that including the PorCORA can more accurately distinguish between GHS Category 1 and GHS Category 2 ocular irritation classifications than the BCOP alone. Thus, the PorCORA, along with other alternative test methods for ocular irritation such as the BCOP, can be used to assess Cleaner and Degreaser class mixture formula induced eye damage as well as the ability of this damage to heal, without the use of live animal testing.

    Wed 9:15 AM - 4:30 PM. Session: Ocular Toxicity, Abstract 2961, Poster Board P403
    A. Brinkman1, P. Vij2, D. Sergeyev2, B. Varsho2, G. DeGeorge2
    1. SC Johnson, Racine, WI
    2. MB Research Laboratories, Spinnerstown, PA

    Through collaboration between SC Johnson and MB Research Labs (both of whom have a long history of using alternative methods to reduce dependency on animals in toxicity testing) a concentrated laundry detergent product mixture was evaluated in the porcine corneal opacity reversibility assay. The PorCORA is an ex vivo assay developed to predict the reversibility (healing potential) of ocular irritants. Historical bovine corneal opacity and permeability (BCOP) data classified the formula as a GHS Category 1 eye irritant. However, the BCOP assay does not address reversibility of ocular damage, which is a key component of the GHS classification scheme for ocular irritants. PorCORA results indicated that the ocular damage induced by exposure to the concentrated laundry detergent mixture was fully reversible. The inclusion of reversibility of damage as an endpoint suggests that the PorCORA can supplement BCOP assay results to distinguish between GHS Category 1 and GHS Category 2 ocular irritation classifications. Thus, the PorCORA, along with other alternative test methods for ocular irritation, can be used to assess consumer detergent product-induced eye damage as well as the ability of this damage to reverse (heal) following detergent mixture exposure to the eye without the use of animal testing.

    Tues 9:15 AM - 4:30 PM. Session: Ocular Toxicity, Abstract 2452, Poster Board P867
    B. Varsho, B. Yasso, M. Troese, E. Delacruz, G. DeGeorge
    1. MB Research Laboratories, Spinnerstown, PA, United States

    OECD Test Guideline 405 prescribes a weight-of-evidence (WoE) analysis and sequential testing strategy for the classification of acute eye hazards, including, “Perform validated and accepted in vitro or ex vivo ocular test(s).” Four tests qualify: three designed to identify severe eye irritation/corrosion (GHS Category 1) and one to identify non-irritants (GHS No Category). GHS Category 2 (eye irritant) classification is impossible using any single test. The EpiOcular™ Eye Irritation Test; EIT (OECD 492) classifies a substance to be No Category, or contrarily causes (uncategorizable) eye effects. Conversely, the Bovine Corneal Opacity and Permeability (BCOP) Test (OECD 437) is used for ruling in or ruling out Category 1 effects. By using a dual-assay/approach system – the combination of the EIT and BCOP test – we have determined, with a high degree of accuracy, GHS Acute Eye Hazard Category 2 chemicals that cause reversible eye irritation. When a BCOP test rules out GHS Category 1, and the EIT rules out GHS No Category, analysis of these results indicates the only other possible designation – Category 2. Per GHS, Category 2 classification defaults to Category 2A, because differentiation between Category 2A and 2B cannot be made. After testing 42 chemicals, we correctly identified 93% of the Category 2A/B chemicals as Category 2. The potential of the BCOP EIT dual-assay system, coupled with WoE evaluation, to correctly classify substances into GHS Category 1, Category 2A, and No Category is encouraging. We predict using this testing strategy would greatly reduce reliance on Draize Rabbit Eye Tests.

    Tues 9:15 AM - 4:30 PM. Abstract 2264, Poster Board P671
    A. Greminger1, P. Vij2, D. Sergeyev2, B. Varsho2, G. DeGeorge2
    1. ExxonMobil, Annandale, NJ
    2. MB Research Labs, Spinnerstown, PA

    The h-CLAT was developed as an in vitro method of identifying skin sensitizers and was promulgated by OECD as a testing guideline in the spirit of advancing the 3Rs (Replacement, Reduction and Refinement). The guideline test is limited by the sanctioning of only two currently validated vehicles (DMSO and saline), precluding in vitro sensitization testing of a wide range of insoluble chemicals. One such insoluble test chemical, Tetrakis (2-ethyl butyl) Orthosilicate, was evaluated in the h-CLAT using an alternative vehicle (i.e., ethanol) in an effort to broaden the utility of the test. This test measures the selective induction of the surface markers CD54 and CD86 in the human monocytic leukemia cell line THP-1, which functions as a dendritic (Langerhans) cell surrogate. Known reference chemicals such as mercaptobenzothiazole (MBT), used as a positive control, and isopropanol, used as a negative control, were also included to evaluate the utility of ethanol as an alternative h-CLAT vehicle. MBT induced positive responses for CD54 and CD86 expression as expected, whereas isopropanol resulted in negative responses for both cell-surface markers. The test chemical was found to be negative in this test.

    Thu 8:30 AM - 11:30 AM. Session: Late-Breaking 9: Toxicokinetics; Mixtures, Abstract 3488, Poster Board P284
    M. Cicchini, L. Pratt, P. Vij, B. Varsho, G. DeGeorge
    1. MB Research Labs, Spinnerstown, PA

    Predicting dermal sensitization is an important component of the acute toxicity testing battery. OECD test guideline assays have been established to measure the four key events of the dermal sensitization Adverse Outcome Pathway (AOP), allowing for screening of delayed-type 4 hypersensitivity potential. The second key event in the AOP is characterized by keratinocyte activation, for which the OECD 442D guideline details the in vitro ARE-Nrf2 Luciferase-based LuSens Test, herein referred to as the LuSens Test. This test has predictive power to identify skin sensitizers through their activation of cultured human keratinocytes. In order to explore the utility of the LuSens Test for predicting UN GHS sensitizers (Category 1) or non-sensitizers, we performed a series of assays using the LuSens Test protocol under the OECD 442D guideline. The LuSens Test correctly identified 5 of 6 sensitizers and 4 of 4 non-sensitizers from the validation chemical list, yielding an overall intra-laboratory accuracy of 90%. One limitation of this methodology for testing a wider array of chemicals and mixtures is limited solubility of potential test substances in the three guideline-approved vehicles, Dimethyl Sulfoxide (DMSO), Media, and Water. We sought to validate the use of additional vehicles applicable in the LuSens Test in order to expand its applicability domain. Propylene Glycol (PG), Polyethylene Glycol 400, 50-70% Ethanol, DMSO:Acetone:Ethanol (4:3:3), DMSO:Ethanol (1:1), Methanol, and 50% Isopropanol all demonstrated compatibility as vehicles in the LuSens Test, as determined by their ability to appropriately promote positive and negative control luciferase induction responses. Using DMSO or PG, when test articles were insoluble in the guideline-recommended vehicles, the LuSens Test was able to predict the dermal sensitization potential of nine commercially available mixtures. LuSens Test predictions agreed in 6 of 6 mixtures tested where a safety data sheet had a dermal sensitization prediction. For 2 of 2 lotion mixtures tested, which were not expected to induce a dermal sensitization response, no induction was seen. The last mixture was a shampoo that had a positive dermal sensitization prediction in both the LuSens Test and the Human Cell Line Activation Test (h-CLAT). Together, these data demonstrates that the LuSens Test can be performed on mixtures using an expanded list of intra-laboratory validated alternative vehicles.

    Thu 8:30 AM - 11:30 AM. Session: Late-Breaking 3: Model Systems; Nanotoxicology, Abstract 3383, Poster Board P173
    P. Vij1, S. Lebrun2, G. DeGeorge1
    1. MB Research Labs, Spinnerstown, PA
    2. Lebrun Labs, Anaheim, CA

    The EPA eye irritation classification system is routinely used to categorize ocular toxicity. The EPA system classifies chemicals that damage the eye after 24 hours (Category III, II, or I) and those that do not cause damage (Category IV). This latter EPA classification is aligned with the standard definition of an “ocular non-irritant” and is appropriate for test substances routinely applied to the eye area. The OptiSafeTM (“Optimized for Safety”) test is a novel, shelf-stable, test-tube based method that can be used to discriminate ocular irritants/corrosives from non-irritants and does not use animal tissues or cells. The OptiSafeTM test determines whether a substance is an ocular non-irritant by measuring damage via a proxy for the corneal stroma (water-soluble molecules), damage to phospholipid bilayers (water-insoluble molecules), and the potential to induce pH extremes in a system (pH buffering system of the eye). Chemicals in this study were selected based on a wide range of EPA classifications, chemical and physical properties, high quality in vivo reference data, and chemical stability. Selected chemicals (38) including surfactants not previously tested were aliquoted into coded vials and tested blind in triplicate. The coded vials were tested, and results were reported as either EPA Category IV (nonirritant) or not (EPA Category III, II, or I). The OptiSafeTM test method applied to these 38 test chemicals achieved a sensitivity of 100% (27/27), specificity was 81.8% (9/11), and overall accuracy was 94.7% (36/38). The better accuracy of OptiSafeTM (~ 95%) versus the OECD Test Guideline 492 EIT method (~ 86%) is best attributed to its higher sensitivity. These results suggest that OptiSafeTM may be an important tool in the complete classification of hazards, especially for surfactants, as well as cosmetics and other substances applied to or around the eye.

    Thu 8:30 AM - 11:30 AM. Session: Late-Breaking 3: Model Systems; Nanotoxicology, Abstract 3385, Poster Board P175
    N. Guan1, P. Vij2, G. Adamson3, A. Nordone4, D. Sergeyev2, M. Cicchini2, B. Varsho2, G. DeGeorge2
    1. Givaudan Fragrances SH Ltd., Shanghai, China
    2. MB Research Labs, Spinnerstown, PA
    3. Givaudan Fragrances Corp., East Hanover, NJ
    4. Givaudan UK Ltd., Ashford, United Kingdom

    Proper identification and classification of the skin sensitization potential for new consumer products, chemicals, and pharmaceuticals are important for hazard communication and subsequent risk management, upon which in vitro toxicology methodologies are heavily relied. The h-CLAT method, as defined by the OECD 442E test guideline, uses the dendritic cell proxy THP-1 monocytic cell line, to addresses the third key event in the sensitization Adverse Outcome Pathway (AOP). The third key event in the AOP is marked by dendritic cell activation, during which time the cell surface markers CD86 and CD54 are induced. In this assay, if a test article (a volatile liquid fragrance) induces CD54 and/or CD86 expression to a level of at least 200 and 150 RFI, respectively, it is predicted to be a skin sensitizer. A major limitation to the versatility of the h-CLAT is the number of recommended vehicles: Saline, media, or dimethyl sulfoxide (DMSO). However, the OECD 442E test guideline states that another vehicle may be used if sufficient scientific rationale is provided. In this study, the test article was not soluble in the recommended vehicles. Therefore, we sought to determine a suitable alternative vehicle and provide a sufficient scientific rationale for using such a vehicle. A previous in-house study suggested ethanol to be a potential alternative vehicle in this test (Abstract Number 2264; SOT 2019), so the test article’s solubility was tested in ethanol. When it was determined the test article was indeed soluble in ethanol up to the recommended 100 mg/ml test concentration, the h-CLAT was performed as otherwise recommended by the guideline. Additionally, 2-mercaptobenthiazole and isopropanol; positive and negative controls, respectively, were concurrently tested using ethanol as their vehicle. The h-CLAT correctly predicted the isopropanol negative control not to be a skin sensitizer, while correctly predicting the positive control 2-mercaptobenthiazole to be a skin sensitizer. Using ethanol as the vehicle, the test article was predicted to be a skin sensitizer, with the RFI levels of CD54 and CD86 being induced above 200 and 150, respectively. Here we provide scientific evidence and rationale for ethanol to be used as a suitable vehicle with the h-CLAT method.