**3.1** *In vivo* **models**

52 Psoriasis

side effects from medication to treat psoriasis (54 %, 64 % and 69 % of psoriatic patients with 0–2 %, > 3% and > 10% BSA involvement, respectively). Patients also manifested that the reasons for treatment discontinuation were as following: lack of efficacy (60 %), inconvenience (23 %) and improvement of symptoms (22 %), side effects (20 %), cost (14 %)

Overall, results of worldwide surveys demonstrate that a substantial proportion of psoriatic patients are highly dissatisfied with current therapies, particularly those with greater psoriasis severity. A perceived lack of efficacy of available treatments suggests the importance of the development of more relevant treatments, in order to allow the

The most significant challenge for antipsoriatic drug development is to provide safe and effective long-term management of this disease. In general, a conventional vision of this process starts with the study of disease in relevant model systems, in order to determine cellular and molecular mechanisms involved in pathogenesis. Afterwards, new therapeutic approaches are developed in these models before clinical trials in humans (Guttman-Yassky & Krueger, 2007). The comprehension that psoriasis is an immune-mediated disease, which involves a complex interplay of T cells, natural killer cells, dendritic cells, macrophages and other leukocytes, has led to the development of new biological treatments. The positive results obtained with these agents have expanded our understanding on psoriasis pathogenesis. However, many questions remain regarding psoriasis pathogenesis, and other medications should be developed to offer individualized treatments able to improve patient's quality of life. Some of the challenges for this field include the improvement of efficacy and safety of new drugs, the solution of problems related to formulation/administration/costs of new agents,

Many psoriatic patients are unresponsive to current therapies or have aggressive disease that is not addressed by current approaches. The determination of relevant biomarkers directly related to psoriasis pathogenesis to be targeted with effective treatments could

The challenge of improving the safety of new antipsoriatic drugs is a very important aspect for long-term therapies, and can be overcome through the understanding of the toxicity mechanisms of new agents at early stages of drug development. Unfortunately, this is not always feasible during the drug development process, and the "safety question" should respond to what constitutes an acceptable risk. Thus, it is important to carefully analyse the

In the case of drugs approved for clinical use, their specific immunogenicity, costs, patient access and inconveniences for administration should be considerate. Other challenges

risk/benefit rate of new antipsoriatic agents, mainly in the case of severe disease.

and doctor's advice (14 %) (Poulin *et al.*, 2010).

establishment of more individualized therapies.

**2. Challenges for antipsoriatic drug development** 

and the development of more relevant psoriatic skin models.

allow quantitative assessment of treatment response (Rashmi *et al.*, 2009).

**2.1 Efficacy** 

**2.2 Safety** 

**2.3 Practical issues** 

#### **3.1.1 Spontaneous mutations**

Psoriasis is a typical human skin disease. Even if spontaneous mutation models do not exhibit every features found in psoriasis, various pathology-like characteristics can be observed, including hyperkeratosis and scaly formation (Mizutani *et al.*, 2003). Hundred of these spontaneous mutation models have been described in the literature (Sundberg *et al.*, 1990), but no one shows all the characteristics of psoriasis. However, these models can be really practical for studying individual characteristics such as hyperkeratosis (Schon, 2008). A comparison between the characteristics observed in the three major models of spontaneous mutations is presented in table 1.

#### **3.1.2 Xenotransplantation**

Animal models based on transgenic technology have been used extensively to study the pathogenesis of various skin diseases, including psoriasis (Raychaudhuri *et al.*, 2001, Jean & Pouliot, 2010). Xenotransplantation approach consists of grafting a piece of *in vivo* psoriatic skin (or an *in vitro* psoriatic substitute) on a genetically modified mouse. Currently, three major models are used: athymic nude mice (Fraki *et al.*, 1983), severe combined immunodeficient mice (SCID) (Raychaudhuri *et al.*, 2001), and spontaneous AGR129 model (Boyman *et al.*, 2004). The main difference between each model is the immunological potential of the immune system. Athymic nude mice have no thymus and therefore no T cells, whereas severe combined immunodeficient mice have no T and no B cells

Psoriatic Skin Models: A Need for the Pharmaceutical Industry 55

characteristics in rodents following the overexpression or underexpression of cytokines (or enzymes) (Bullard *et al.*, 1996, Danilenko, 2008, Keith *et al.*, 2005). It is important to note that psoriasis is a multisystemic skin disease, and that transgenic models consider only a single gene at the time. Thus, even if these models are interesting to observe isolated psoriasis-like features, they do not allow the study of all the characteristics of the pathology. There exist a broad variety of genetically modified *in vivo* models. An exhaustive list can be seen in table

Epidermal

thickness

Hypomorphic CD18 + + + + (Bullard *et al.*, 1996,

E (CD103) + + ? + (Schon *et al.*, 2000) K14/p40 + ? ? + (Kopp *et al.*, 2001)

pTek-*tTA*/Tie2 + + + + (Voskas *et al.*, 2005) K14/VEGF + + + + (Xia *et al.*, 2003)

K5/Stat3C + + + + (Sano *et al.*, 2005) IKK2 + + ? - (Pasparakis *et al.*, 2002) c-Jun/JunB + + + + (Zenz *et al.*, 2005) K14/KGF + + + - (Guo *et al.*, 1993)

K14/IL-20 + + - - (Blumberg *et al.*, 2001) K14/amphiregulin + + + + (Cook *et al.*, 1997) K14/IL-1 + + - ? (Groves *et al.*, 1995) K14/IL-6 + - - - (Turksen *et al.*, 1992) K10/BMP-6 + + + + (Blessing *et al.*, 1996) Involucrin/integrins + + + + (Carroll *et al.*, 1995) Involucrin/MEK1 + + ? + (Hobbs *et al.*, 2004) Involucrin/amphiregulin + + + + (Cook *et al.*, 2004) Involucrin/IFN- + + + - (Carroll *et al.*, 1997) Chymotryptic enzyme + + ? + (Hansson *et al.*, 2002)

Reproduced and modified from Jean *et al.*, 2010 according to the copyright policy of the

K14/TGF- + + ? Some

Table 2. *In vivo* genetically modified models of psoriasis

Abnormal

differentiatio

Increased

vascularizatio

Epidermal T cell

infiltratio

+ + + + (Keith *et al.*, 2005,

References

Breban *et al.*, 1996)

Kess *et al.*, 2003)

animals (Vassar & Fuchs, 1991)

n

n

n

2 (Jean & Pouliot, 2010).

*Targeting the immune system* HLA-B27/β2 microglobulin rat

*Targeting vascular endothelium*

*Targeting epidermal proteins*

publisher. 2010 InTech.

Model


Table 1. Examples of spontaneous mutation models and their characteristics

(Raychaudhuri *et al.*, 2001). As for AGR129 model, it is characterized by the absence of T and B cells and by the presence of immature natural killer (NK) cells, less cytotoxic than mature NK cells (Boyman *et al.*, 2004). A weaker system is potent to dwell skin transplants for a longer time on a compromised mouse upon rejection. Thus, the amount of transplant rejection is reduced in the AGR129 model compared to the others. Boyman *et al.* demonstrated that human uninvolved psoriatic skin grafted onto AGR129 mice spontaneously developed psoriatic plaques without the injection of any activated immune cells or any other exogenous factor, suggesting that uninvolved psoriatic skin is not exactly comparable to the normal human skin of healthy patients (Boyman *et al.*, 2004, Gudjonsson *et al.*, 2007, Jean & Pouliot, 2010). However, the absence of an inflammatory system could be a significant weakness of these models, since the importance of the immunology has been described by many research groups.

#### **3.1.3 Genetically modified models**

Development of rat and mouse transgenic models was an important step in the field of *in vivo* models. These genetically modified animals allow the observation of psoriasis-like

Epidermal acanthosis

Dermal infiltrate (mast cells and macrophages)

Best spontaneous model of psoriasis described

Positive Koebner reaction after tape-stripping

Hyperproliferative skin

inflammatory cells in the

Dilation of blood vessels

(Raychaudhuri *et al.*, 2001). As for AGR129 model, it is characterized by the absence of T and B cells and by the presence of immature natural killer (NK) cells, less cytotoxic than mature NK cells (Boyman *et al.*, 2004). A weaker system is potent to dwell skin transplants for a longer time on a compromised mouse upon rejection. Thus, the amount of transplant rejection is reduced in the AGR129 model compared to the others. Boyman *et al.* demonstrated that human uninvolved psoriatic skin grafted onto AGR129 mice spontaneously developed psoriatic plaques without the injection of any activated immune cells or any other exogenous factor, suggesting that uninvolved psoriatic skin is not exactly comparable to the normal human skin of healthy patients (Boyman *et al.*, 2004, Gudjonsson *et al.*, 2007, Jean & Pouliot, 2010). However, the absence of an inflammatory system could be a significant weakness of these models, since the importance of the immunology has been

Development of rat and mouse transgenic models was an important step in the field of *in vivo* models. These genetically modified animals allow the observation of psoriasis-like

Table 1. Examples of spontaneous mutation models and their characteristics

Infiltration of

in the dermis

skin

Increased dermal vascularization

Proliferation and hyperkeratosis of stratified squamous

epithelia

Homozygous *asebia* (Scd1ab/Scd1ab)

Flaky skin mice (Ttcfsn/Ttcfsn)

Spontaneous chronic proliferative dermatitis

(Sharpincpdm/Sharpincpdm)

described by many research groups.

**3.1.3 Genetically modified models** 

mutation

**Model Characteristics References Psoriasis-like Psoriasis-unlike**

Lack of the

Alterations of the cutaneous lipid metabolism different from psoriasis

(Schon, 2008, Zheng *et al.*, 1999)

(Sundberg *et al.*, 1990, Danilenko, 2008, Stratis *et al.*, 2006, Sundberg *et al.*, 1994, Schon, 1999)

1999)

Lack of T cells and neutrophils

Comprises aspects not find in psoriasis

immunological side

Lack of T cells (Schon,

characteristics in rodents following the overexpression or underexpression of cytokines (or enzymes) (Bullard *et al.*, 1996, Danilenko, 2008, Keith *et al.*, 2005). It is important to note that psoriasis is a multisystemic skin disease, and that transgenic models consider only a single gene at the time. Thus, even if these models are interesting to observe isolated psoriasis-like features, they do not allow the study of all the characteristics of the pathology. There exist a broad variety of genetically modified *in vivo* models. An exhaustive list can be seen in table 2 (Jean & Pouliot, 2010).


Table 2. *In vivo* genetically modified models of psoriasis

Reproduced and modified from Jean *et al.*, 2010 according to the copyright policy of the publisher. 2010 InTech.

Psoriatic Skin Models: A Need for the Pharmaceutical Industry 57

Facing the absence of exogenous material-free models, our group developed a new pathological skin model to study psoriasis *in vitro* by using the self-assembly approach (Michel *et al.*, 1999) (Fig. 3). Briefly, normal and pathological fibroblasts are thawed and cultured with ascorbic acid for a period of time of four weeks. Then, dermal sheets are produced and removed from flasks. Two fibroblast sheets are superimposed to form a new dermal equivalent. Seven days later, normal or pathological keratinocytes are seeded on the dermal equivalent to obtain a new epidermal equivalent. After another 7 days of culture, the substitutes are raised to the air–liquid interface to favour cell differentiation and stratification. Finally, biopsies are taken after 21 days of culture at the air–liquid interface, and samples are analyzed using histological, immunohistochemical, physico-chemical or

**3.2.3 Self-assembly approach** 

permeability techniques (Jean *et al.*, 2009).

the publisher. 2010 InTech.

**Seeding of fibroblasts** 

**4. Conclusion**

Fig. 3. The self-assembly approach for the production of skin substitutes

**Day 0 Day 35** 

**Superposition of sheets Day 28** 

Schematic representation of the various steps of skin substitutes production in function of time. Reproduced and modified from Jean *et al.*, 2010 according to the copyright policy of

**Seeding of keratinocytes**  **Keratinocytes**

**Air-liquid interface** 

**Day 42 Day 63** 

**Biopsies** 

In 2009, Jean *et al.* showed that self-assembled skin substitutes partially maintained psoriasis-like features such as a thick epidermis, hyperproliferation as well as abnormal cell differentiation of epidermal cells (Jean *et al.*, 2009). In 2011, they demonstrated for the first time that pathological substitutes produced by the self-assembly approach can be treated with an anti-psoriatic molecule and react positively to the treatment such as observed in psoriatic skin *in vivo*. This functional study suggests that the self-assembled skin substitutes could be useful to better understand the mechanisms through which retinoic acid regulates cellular physiology in psoriatic skin, and could become an effective and innovative

Psoriasis is characterized by the presence of physical and psychological pains, which can severely affect the quality of life of psoriatic patients. Currently, a broad spectrum of antipsoriatic treatments, both topical and systemic, is available for the management of psoriasis. These treatments only allow to control psoriasis without curing it. Challenges for antipsoriatic-drugs development are numerous, and the pharmaceutical industry strongly

dermopharmaceutical tool for the screening of new treatments (Jean *et al.*, 2011).
