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COMPANY INFORMATION

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FACT SHEET
Pharmatest Services Ltd is a full-service bone biology contract research organization (CRO) specialized in preclinical in vitro, ex vivo and in vivo services for testing the efficacy of new drug candidates for osteoporosis, cancer and osteoarthritis. We provide rapid and cost-effective strategies with high quality. As the only source for commercial in vitro drug efficacy testing in the field of bone biology, we have developed unique in vitro assays designed specifically to provide fast and costeffective alternative for obtaining efficacy data for compounds still in discovery phase. These in vitro models are a perfect tool for selecting most potent osteoporosis or cancer bone disease drug candidates for further development. Our in vivo services include rat ovariectomy and orchidectomy models of osteoporosis, glucocorticoid-induced mouse osteoporosis model, osteoarthritis animal model and cancer tumor and metastasis models. In addition, Pharmatest provides many ex vivo bone analysis methods, such as BMD-measurement, histology services, histomorphometric analysis and mechanical testing, which are routinely used with our in vivo services, but can also be performed as stand-alone services. Our latest set of services for drug discovery include early ADMET assays and basic pharmacokinetics services for early stage drug discovery needs.

Pharmatest operates in 500 square meter state-of-the-art facilities in the city of Turku in the South-Western Finland. Pharmatest also has an office in Oulu, and international marketing office in Berlin, Germany.

All work at Pharmatest is performed using best possible equipment and procedures, and in accordance with The Pharmatest Services Quality Assurance Program. This ensures the quality of all services provided by Pharmatest.
Pharmatest Services Ltd Itinen Pitkkatu 4C 20520 Turku, FINLAND Tel: +358 2 278 4700 Email: info@pharmatest.fi

www.pharmatest.fi

1 IN VITRO BONE RESORPTION ASSAY


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FACT SHEET
Pharmatest's in vitro osteoclast assays can be used for preclinical testing of antiresorptive effects of drug candidates on human osteoclasts. In the assays, human osteoclast precursor cells are cultured on bone slices. All tests performed by Pharmatest include a group without treatment (baseline group) and a group treated with Pharmatest's reference inhibitor (control group).
Typical turnaround times are: Rapid Screening: up to 250 compounds / 2 months Preliminary: up to 50 compounds / 2 months Extensive: up to 20 compounds / 2 months

We have separate assays to test the effects of the test compounds on osteoclast differentiation and osteoclast activity. In the osteoclast differentiation assay, the test compounds are added into cultures of osteoclast precursor cells, and the cells are then allowed to differentiate into mature osteoclasts. After the culture period, the amount of tartrate-resistant acid phosphatase isoform 5b (TRACP 5b) secreted into the culture medium is determined as an index of the number of formed osteoclasts. In the osteoclast activity assay, the osteoclast precursor cells are first allowed to differentiate into mature osteoclasts, after which the test compounds are added, and the formed mature osteoclasts are then cultured for an additional two days, allowing them to resorb bone. After the culture period, the amount of Cterminal crosslinked telopeptides of bone collagen (CTX) released into the culture medium is determined as an index of bone resorption. With the optional toxicity testing you can also verify whether the possible effect of the drug is due to cytotoxicity or not. We offer various types of in vitro resorption projects; screening, preliminary and extensive. With a large number of compounds, we suggest starting with the screening, where 1 concentration is tested in 4 replicates, or the preliminary testing, where 3 concentrations of each compound are tested in 6 replicates. The results of the screening and preliminary testing are suggested to be used primarily for selecting the compounds for extensive testing, where 7 concentrations are tested in 8 replicates. The results of extensive testing can be used to select the most potent candidates for possible in vivo testing. The amount of concentrations and replicates selected for the two types of projects we offer have been chosen by meticulous validation. However, if your research requires different number of concentrations or replicates, please let us know. We will be happy to modify our assays accordingly.
Initiation of cell culture End of cell culture End of cell culture
- TRACP 5b-measurement from culture media - Morphological analysis from cells - CTx-measurement from culture media - Morphological analysis from cells

All work at Pharmatest is performed in accordance with The Pharmatest Services Quality Assurance Program to ensure the quality of all services performed at Pharmatest. As part of this Program, we guarantee that if the results of the control group are not statistically significantly different from the results of the baseline group, Pharmatest will repeat the testing without additional costs to the client.
140 Bone resorption (% from baseline) 120 100 * p<0.05 *** p<0.001

Typical results of potential drug * compound X in 80 Pharmatests Bone Resorption (human) Assay. *** ( -10 60 M = 10 M test compound, etc.)
*** ***
40 20 0 Baseline Control -10M -9M -8M -7M -6M

- Human bone marrow cells - Cultured on bovine bone slices - 96-well format

Differentation assay Osteoclast progenitors

Activity assay

9 Days

Hematopoietic stem cells

Multinucleated osteoclast

Typical results of potential drug compound X in Pharmatests Bone Resorption (human) Assay. ( -10 M = 10 M test compound, etc.)
Pharmatest Services Ltd Itinen Pitkkatu 4C 20520 Turku, FINLAND Tel: +358 2 278 4700 Email: info@pharmatest.fi

Timeline of a Pharmatest testOSTEOCLAST in vitro bone resorption assay.

www.pharmatest.fi

RESORPTIONASSAY ASSAY 1 IN VITRO BONE FORMATION 2


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FACT SHEET
Pharmatest's in vitro osteoblast assays Pharmatest's in vitro osteoclast can be used for preclinical testing of assays can be used for preclinical anabolic bone effects of drug candidates testing of antiresorptive effects of on human osteoblasts. In the assays, drug candidates on cells human human osteoblast precursor are osteoclasts. In the assays, human cultured on plastic surface. All tests osteoclast precursor cells are performed by Pharmatest include a group without treatment (baseline group) and a cultured on bone slices. All tests group treated with Pharmatest's reference performed by Pharmatest include a activator (control treatment group). group without (baseline We have separate assays to test the effects of the test compounds on osteoblast differentiation and osteoblast activity. In the osteoblast differentiation assay, the test compounds are added into cultures of osteoblast precursor cells, and the cells are then allowed to differentiate into mature osteoblasts. After the culture period, the amount of intracellular alkaline phosphatase activity is determined as an index of the number of formed osteoblasts. In the osteoblast activity assay, the osteoblast precursor cells are first allowed to differentiate into mature osteoblasts, after which the test compounds are added, and the formed mature osteoblasts are then cultured for an additional time, allowing them to form bone nodules. After the culture period, the amount of calcium deposited into the formed bone nodules is determined as an index of bone formation. With the optional toxicity testing you can also verify whether the possible effect of the drug is due to cytotoxicity or not. We offer various types of projects; screening, preliminary and extensive. With a large number of compounds, we suggest starting with the screening, where 1 concentration is tested in 4 replicates, or the preliminary testing, where 3 concentrations of each compound are tested in 6 replicates. The results of the preliminary testing are suggested to be used primarily for selecting the compounds for extensive testing, where 7 concentrations are tested in 8 replicates. The results of extensive testing can be used to select the most potent candidates for possible in vivo testing. The amount of concentrations and replicates selected for the two types of projects we offer have been chosen by meticulous validation. However, if your research requires different number of concentrations or replicates, please let us know. We will be happy to modify our assays accordingly.
Initiation of cell culture
- Mouse bone marrow cells - 96-well format

group) and a group treated with Pharmatest's reference inhibitor (control group).
Typical turnaround times are: Rapid Screening: up to 500 250 compounds / 3 2 months Preliminary: up to 100 50 compounds compounds // 23 months months Extensive: up to 20 40 compounds / 3 2 months

All work at Pharmatest is performed in accordance with The Pharmatest Services Quality Assurance Program to ensure the quality of all services performed at Pharmatest. As part of this Program, we guarantee that if the results of the control group are not statistically significantly different from the results of the baseline group, Pharmatest will repeat the testing without additional costs to the client.
350 Bone formation (% from baseline) 300 250 * p<0.05 *** p<0.001

End of cell culture End of cell culture


- ALP-measurement from culture media - Morphological analysis from cells - Ca-measurement from culture media - Morphological analysis from cells

*** *

***

***

Typical results of potential drug compound X in 200 Pharmatests Bone Resorption (human) Assay. ( -10 M = 10 M test compound, etc.)
150 100 50 0 Baseline Control -12M -11M -10M -9M -8M

Differentation assay Osteoblast progenitors Osteoblast

14

Activity assay

21 Days

Mesenchymal stem cells

Typical results of potential drug compound X in Pharmatests Bone Formation Assay. ( -12 M = 10 -12 M test compound, etc.)
Pharmatest Services Ltd Itinen Pitkkatu 4C 20520 Turku, FINLAND Tel: +358 2 278 4700 Email: info@pharmatest.fi

Timeline of a Pharmatest testOSTEOBLAST in vitro bone formation assay.

www.pharmatest.fi

IN VITRO BONE OSTEOPOROSIS RESORPTION ASSAY MODEL (OVX/ORX) 1 EXPERIMENTAL 3


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FACT SHEET
Pharmatest offers Pharmatest's in both vitrorat osteoclast OVX and assays ORX models can be as used preclinical for preclinical testing testing services. ofWith antiresorptive these models effects we can of drug test how candidates your compounds on human can osteoclasts. prevent the In the development assays, human of osteoclast osteoporosis precursor or treat established cells are cultured osteoporosis. on bone We slices. offer Allthree tests performed different by project Pharmatest types, include rapid a group screening, without preliminary treatment and (baseline extensive group) projects. and All projects a group always treated contain with Pharmatest's a sham-operated reference control group inhibitor and (control an OVXgroup). group receiving vehicle.
Typical turnaround times are: Rapid OVX: Screening: up months to 250 compounds / 2 months Preliminary: up months to 50 compounds / 2 months Extensive: Regulatory: up months to 20 compounds / 2 months

Rat ovariectomy (OVX) and orchidectomy (ORX) models are the most commonly used experimental animal models of osteoporosis. The rat OVX model mimics postmenopausal bone loss and it is therefore recommended as a preclinical animal model for osteoporosis according to FDA Guidelines for Preclinical and Clinical Evaluation of Agents Used in the Treatment or Prevention of Postemenopausal Osteoporosis. The rat ORX model is a corresponding model for male osteoporosis. The rapid screening projects are 2-week studies that include only measurement of biochemical markers of bone turnover. These studies are designed for initial testing of large number of test compounds and for initial dose-response studies. Preliminary studies are 2-month studies that include measurement of biochemical markers of bone turnover, bone mineral density (BMD) measurements using peripheral quantitative computed tomography (pQCT), mechanical testing, ash weight analysis and static and dynamic histomorphometry. Extensive studies are 12-month regulatory studies that are performed according to FDA/EMEA guidelines, allowing the use of the results in the registration process of new drugs to obtain FDA and EMEA approval. Our imaging techniques and bone marker measurements can also be used as stand-alone services, where you just send your samples to us for analysis. We can for example process tissue samples for further histological analysis in our histomorphometry lab. The techniques available include tissue preparation and embedding to either paraffin or plastic to facilitate a wide array of histological analyses. We are experienced in a wide variety of staining techniques to make sure that we have the right tools for your needs. We can also perform BMD measurements, mechanical testing and ash weight analysis to your bone samples and measure

All work at Pharmatest is performed in accordance with The Pharmatest Services Quality Assurance Program to ensure the quality of all services performed at Pharmatest. Pharmatest. As part of this Program, we guarantee that if the results of the control group are not statistically significantly different from the results of the baseline group, Pharmatest will repeat the testing without additional costs to the client.

Typical results of potential drug compound X in Pharmatests Bone Resorption (human) Assay. ( -10 M = 10 M test compound, etc.)

Bone mineral density (BMD) readout from pQCT equipment which is used in our testOVX/ORX models (above) and typical bone resorption pit (below).

Pharmatest Services Ltd Itinen Pitkkatu 4C 20520 Turku, FINLAND Tel: +358 2 278 4700 Email: info@pharmatest.fi

www.pharmatest.fi

IN VITRO BONE BREAST RESORPTION CANCER BONE ASSAY METASTASIS MODEL 1 SYSTEMIC 4
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FACT SHEET
Pharmatestsin Breast Pharmatest's vitro osteoclast Cancer assays Metastasis canservice be used can forbe preclinical used for testing preclinical of antiresorptive testing of theeffects effect of drug drug candidates candidates on human on human breast osteoclasts. cancer boneIn metastases the assays,in human vivo. osteoclast This model closely precursor resembles cells are the cultured progression on bone of breast slices. cancer All tests in performed humans and by can Pharmatest be used to include test the a group effectiveness without of treatment drug compounds (baseline group) on bothand treatment a group of treated established with Pharmatest's disease and prevention referenceof inhibitor cancer (control metastases. group).
Typical turnaround turnaround times timesfrom are: the beginning of the in-life phase until the delivery of final report are: Rapid Screening: up to 250 compounds / 2 months Preliminary: Rapid screening project: up to 50 compounds 1 months / 2 months Extensive: project: Preliminary up to 20 compounds 2 months / 2 months Extensive project: 5 months

In the Breast Cancer Metastasis model the animals are inoculated with a human breast cancer cell line, which will cause the animals to develop metastases to bone and soft tissue. The drug compound is then administered for 3 to 4 weeks and the progress of the disease is monitored by xray radiography. At the end of the experiment the animals are radiographed and tissue samples are processed for histomorphometric analysis. The radiographs can be analyzed for lesion number and areas and the histology samples can be analyzed for tumor area, bone area and osteoclast number at tumor-bone interface. The experiment can also be modified to include a fluorescent reporter gene for faster, non-invasive in vivo imaging which also enables earlier detection of cancer metastases. This imaging can be used either alone or in conjunction with radiographic imaging. The use of fluorescent reporting will also facilitate non-invasive detection of soft tissue tumors and partially replace histomorphometry, thus significantly reducing the duration and costs of the experiment. All experiments consist of a control group with no treatment and experiment groups treated with drug compounds. Data from the experiment groups are compared against the control group. The model is suitable for testing preclinical in vivo efficacy of any new drug compound targeting mechanisms of cancer metastases. This model is fast and highly cost-effective. The radiography data is available for the client in only 4 weeks from the inoculation. Furthermore, the amount of test compounds needed for the experiment is extremely low due to the use of mice as test animals and the short duration of the animal experiment.

All work at Pharmatest is performed in accordance with The Pharmatest All work at Pharmatest is performed in Services Quality Assurance Program to accordance with The Pharmatest ensure quality of allProgram services Servicesthe Quality Assurance to performed at Pharmatest. As part of this ensure the quality of all services Program, we that if the results performed at guarantee Pharmatest. of the control group are not statistically significantly different from the results of the baseline group, Pharmatest will repeat the testing without additional costs to the client.

Cancer cell inoculation


- Human breast cancer cells - Athymic female nude mice - Intra-cardiac inoculation

Monitoring and drug administration


- Weight and clinical signs - Osteolytic lesion are by x-ray, if necessary - All administration routes

Sacrifice and end measurements

- Osteolytic lesion area - Macroscopical findings - Blood sampling, if necessary - Histology, if necessary

Typical results of potential drug compound X in Pharmatests Bone Resorption (human) Assay. ( -10 M = 10 M test compound, etc.)

Figure: Osteolytic bone destruction caused by breast cancer metastases can be seen in the radiograph of mouse tibia and femur (left). The same destruction can be seen on a cellular level in histology of right mouse proximal tibia with tumor cells stained purple (H&E staining).

0
Prevention model; drug administration started

10
Therapeutic model; re-randomization according to lesion size and drug administration started

24
In-life phase, days

Timeline of a testMETASTASIS in vivo systemic bone metastasis model.

Pharmatest Services Ltd Itinen Pitkkatu 4C 20520 Turku, FINLAND Tel: +358 2 278 4700 Email: info@pharmatest.fi

www.pharmatest.fi

IN VITROPRIMARY BONE RESORPTION TUMOR MODEL ASSAY 1 CANCER 5


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FACT SHEET
Pharmatest's Pharmatests Breast in vitro Cancer osteoclast primary assays tumor model can be used can be for preclinical used for testing preclinical of antiresorptive testing of theeffects effect of drug drug candidates candidates on human on human breast osteoclasts. cancer primary In the soft assays, tissue tumors human osteoclast e i t h e r s u precursor b c u t a n e o cells u s l y are or cultured orthotopically on bone in mice. slices. All tests performed by Pharmatest include a group This model without is useful treatment for testing (baseline the group) capacity and of a drug group compounds treated with to Pharmatest's either prevent the reference growth of inhibitor primary (control tumors, or group). treat established tumors, depending on the research approach.
Typical turnaround times are: Rapid Screening: up to 250 compounds / 2 months Preliminary: up to 50 compounds / 2 months Extensive: up time to 20is: compounds / 2 months Typical turnaround 2 months

In the primary tumor model, human breast cancer cells are injected to animals either subcutaneously or orthotopically into the mammary fat pad. This will cause the animals to develop primary tumor to the site of injection. The drug compound is then administered and the progress of the tumor development is monitored by measuring the size of the tumor at desired intervals during the experiment. At the end of the experiment the animals are radiographed, euthanized and tissue samples are processed for possible future histomorphometric analysis. The experiment can also be modified to include a fluorescent reporter gene for faster, non-invasive in vivo imaging which also enables earlier detection of cancer metastases. The use of fluorescent reporting will also facilitate non-invasive detection of soft tissue tumors and partially replace histomorphometry, thus significantly reducing the time needed for the experiment. All experiments consist of a control group with no treatment and experiment groups treated with drug compounds. Data from the experiment groups are compared against the control group. This model is fast and highly cost-effective. The preliminary tumor size data is available for the client in only 4 weeks from the cancer cell injection. Furthermore, the amount of test compounds needed for the experiment is extremely low due to the use of mice as test animals and the short duration of the animal experiment.

All All work work at at Pharmatest Pharmatest is is performed performed in in accordance with The accordance with The Pharmatest Pharmatest Services Services Quality Quality Assurance Assurance Program Program to to ensure ensure the the quality quality of of all all services services performed performed at at Pharmatest. Pharmatest. As part of this Program, we guarantee that if the results of the control group are not statistically significantly different from the results of the baseline group, Pharmatest will repeat the testing without additional costs to the client.

Cancer cell inoculation


- Human breast cancer cells - Athymic female nude mice - Mammary fat pad (orthotopic) or subcutaneous

Monitoring and drug administration


- Weight and clinical signs - Tumor volume

Sacrifice and end measurements

- Tumor volume and weight - Macroscopical findings - Blood sampling, if necessary - Histology, if necessary

Typical results of potential drug compound X in Pharmatests Bone Resorption (human) Assay. ( -10 M = 10 M test compound, etc.)

0
Prevention model; drug administration started

6
Therapeutic model; re-randomization according to tumor volume and drug administration started

30
In-life phase, days

Figure: Representive results of primary tumor experiment can be seen above, whereas the lower image shows a typical GFP-expressing soft tissue tumor.

Timeline of a Pharmatest testTUMOR in vivo primary tumor model.

Pharmatest Services Ltd Itinen Pitkkatu 4C 20520 Turku, FINLAND Tel: +358 2 278 4700 Email: info@pharmatest.fi

www.pharmatest.fi

IN VITRO BONE OSTEOARTHRITIS RESORPTION ASSAY MODEL 1 EXPERIMENTAL 6


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FACT SHEET
Pharmatest's in vitro osteoclast assays can be used preclinical Pharmatest's in vivo for osteoarthritis testing of antiresorptive effects of model can be used for preclinical drug candidates on human testing ofosteoarthritis drug osteoclasts. candidates. In the assays, human osteoclast precursor cells are cultured on bone slices. All tests performed by Pharmatest include a group without treatment (baseline group) and a group treated with Pharmatest's reference inhibitor (control group).
Typical turnaround times are: Typical turnaround is: 2 months Rapid Screening: up time to 250 compounds / 2 months Preliminary: up to 50 compounds / 2 months Extensive: up to 20 compounds / 2 months

Pharmatest uses rabbit experimental osteoarthritis model. Using these models, we can test your compounds efficacy in the treatment of osteoarthritis. In this study, the animals are first operated and intervention is started immediately . A normal osteoarthritis study is composed of a partial medial meniscectomy operation, followed by an in-life phase during which the drug compound is adminstered. After the in-life phase the test subjects are euthanized and histology samples are collected. In the final phase of the study histomorphometric analysis is performed to assess the effect of your drug compound on osteoarthritis. Turnaround time of this model depends largely on the pharmacokinetic characteristics of the drug compound, i.e, the duration of the experiments in-life phase.

All work at Pharmatest is performed in accordance with The Pharmatest Services Quality Assurance Program to ensure the quality of all services performed at Pharmatest. Pharmatest. As part of this Program, we guarantee that if the results of the control group are not statistically significantly different from the results of the baseline group, Pharmatest will repeat the testing without additional costs to the client.

Figure: Representive results of osteoarthritis model. Image on the right clearly shows how the drug compound X has inhibited the destruction to femoral boneof surface caused osteoarthritis Typical results potential drug by compound X in (sites A, B and C) when compared to the control Pharmatests Bone Resorption (human) Assay. group (left). ( -10 M = 10 M test compound, etc.)

Pharmatest Services Ltd Itinen Pitkkatu 4C 20520 Turku, FINLAND Tel: +358 2 278 4700 Email: info@pharmatest.fi

www.pharmatest.fi

IN VITRO BONE RESORPTIONMODEL ASSAY OF OSTEOPOROSIS 1 GLUCOCORTICOID-INDUCED 7


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FACT SHEET
Pharmatest's cause in vitro osteoclast Glucocorticoids osteoporosis by affecting can both be bone resorption and bone assays used for preclinical formatio n .antiresorptive T h e b o n e eeffects ff e c ts of of testing of glucocorticoids caused by dramatic drug candidates on human decrease in bone formation due to osteoclasts. In theof assays, human extensive apoptosis osteoblasts and osteoclast precursor cells are osteocytes. Pharmatest offers a mouse cultured bone slices. All tests model ofon glucocorticoid-induced osteoporosisby where 3-6 months old intact performed Pharmatest include a male mice are administered with group without treatment (baseline glucocorticoids 3-8 weeks. With this group) and afor group treated with model, we can test how your new Pharmatest's inhibitor osteoporosis drugreference candidates can protect (control group). against glucocorticoid-induced
osteoporosis. The model can also be used to test whether or not your new glucocorticoid analogs have detrimental Typical turnaround times are: side-effects on bone.
Rapid Screening: up to 250 compounds / 2 months Preliminary: up to 50 compounds / 2 months Typical turnaround 3 months Extensive: up time to 20is: compounds / 2 months

We offer an extensive range of imaging techniques and ex vivo bone analysis techniques that can be utilized to help determine the effects of new drug candidates on bone during and after the in-life phase. These techniques include: - BMD measurements performed by pQCT analysis - Static and dynamic histomorphometric analysis - Ash weight analysis - Mechanical testing - Biochemical markers of bone turnover Our pQCT equipment allows in vivo imaging of living animals, making it a perfect tool for monitoring changes in cortical and trabecular bone parameters during a long study period. The ash weigh analysis shows the effects of drug candidates on the amount of mineralized bone. Our histomorphometric analysis includes determination of both static and dynamic parameters that will give detailed information about bone architecture and bone quality, uncovering the effects of drug candidates at cellular level. Biochemical markers of bone turnover enable rapid detection of the effects of drug candidates on bone turnover, allowing the prediction of longterm bone effects of the drug candidates. A group without treatment and a group with our reference compound prednisolone are included in all projects performed with the model.

All All work work at at Pharmatest Pharmatest is is performed performed in in accordance with The accordance with The Pharmatest Pharmatest Services Quality Quality Assurance Assurance Program Program to to Services ensure ensure the the quality quality of of all all services services performed performed at at Pharmatest. Pharmatest. As part of this Program, we guarantee that if the results of the control group are not statistically significantly different from the results of the baseline group, Pharmatest will repeat the testing without additional costs to the client.

Glucocorticoid administration
- Baseline blood samples - BMD-measurements

Monitoring and drug administration


- Weight and clinical signs

Sacrifice and end measurements

- BMD-measurements - Blood sampling - Histology

Typical results of potential drug compound X in Pharmatests Bone Resorption (human) Assay. ( -10 M = 10 M test compound, etc.)

0
Prevention model; drug administration started

6
Therapeutic model; drug administration started

28
In-life phase, days

Figure: Representive histomorphometry section from in vivo osteoporosis model.

Timeline of a Pharmatest testGIO in vivo osteoporosis model.

Pharmatest Services Ltd Itinen Pitkkatu 4C 20520 Turku, FINLAND Tel: +358 2 278 4700 Email: info@pharmatest.fi

www.pharmatest.fi www.pharmatest.fi www.pharmatest.fi

IN VITRO BONE MODELS RESORPTION ASSAY 1 ADME/Tox 8


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FACT SHEET
In addition to our efficacy testing Pharmatest's in vitro osteoclast services, Pharmatest is now able to assays can be used for preclinical provide ADME/Tox studies to speed testing of antiresorptive effects of up your drug discovery. These drug candidates on human studies are essential for selection osteoclasts. In the assays, human of safe compounds with cells favorable osteoclast precursor are pharmacokinetic properties for cultured on bone slices. All tests further drug development. performed by Pharmatest include a group without treatment (baseline group) and a group treated with Pharmatest's reference inhibitor Pharmatest can help you select the best (control group). drug candidates from a large set of
compounds. We can accomplish this by designing the most cost-effective and time-saving strategy for performing Typical turnaround times are: ADME/Tox assays and efficacy tests for your drug candidates in an optimal order. Rapid Screening: up to 250 compounds / 2 months
Preliminary: up to 50 compounds / 2 months

Pharmatest offers three types of in vitro ADME/Tox projects: rapid screening, preliminary and extensive. Rapid screening in vitro ADME/Tox projects include rapid screening of physicochemical properties, permeability and metabolism of a large number of drug candidates. The physicochemical property tests include studying ionization characteristics, solubility, lipophilicity, plasma protein binding and non-specific binding. The permeability tests are performed using human enterocyte-like colon carcinoma cells (Caco-2). The metabolic screening consists of determination of metabolic stability of the drug candidates and identification of their metabolites produced in phase I and II metabolic pathways. Based on the results of the rapid screening ADME/Tox projects, most promising drug candidates can be selected for further testing in preliminary and extensive projects. Preliminary in vitro ADME/Tox projects include preliminary tests for permeability, metabolism, drug-drug interactions and hepato/cytotoxicity. Permeability is tested both from apical to basolateral and basolateral to apical surface using Caco-2 cells, including a separate substrate test for a general efflux pump, P-glycoprotein (P-gp). The metabolic tests include determination of the metabolic stability of the drug candidates and their enzyme kinetic values for intrinsic clearance and prediction of in vivo metabolism. The drug-drug interaction tests include characterization of the capability of the drug candidates to inhibit cytochrome P450 enzymes (CYPs) using specific substrates and liver microsomes / hepatocytes. Based on the results of the preliminary projects, most promising drug candidates can be selected for further testing in extensive projects. Extensive in vitro ADME/Tox projects include detailed permeability, metabolism and drug-drug interaction tests. Permeability is tested from apical to basolateral surface using Caco-2 cells, including a P-gp substrate test, and describing the contribution of transporters, intracellular metabolism and cell accumulation. The metabolic testing consists of determination of metabolic stability of the drug candidates and their enzyme kinetic values for prediction of in vivo metabolism as well as detailed characterization of their metabolites. The drug-drug interaction tests include identification of CYPs and uridine diphosphateglucuronosyltransferases (UGTs) that contribute to metabolism of the drug candidates using specific inhibitors and liver microsomes/hepatocytes or recombinant CYPs and UGTs. After the in vitro ADME/Tox properties of the drug candidates are known, they can be tested further with our in vivo ADME/Tox models. Pharmatest offers two types of in vivo ADME/Tox projects: rapid screening and preliminary. Rapid screening in vivo ADME/Tox projects include simplified and animal-saving determination of bioavailability, maximum concentration in plasma and half-life of the drug candidates as well as identification of their metabolites and elucidation of their preliminary structures. Preliminary in vivo ADME/Tox projects include a comprehensive characterization of oral bioavailability, pharmacokinetics and metabolism of the drug candidates. In vivo toxicological properties of the drug candidates can be characterized both with acute and repeated dose toxicity tests by monitoring clinical signs of the animals and using histopathology evaluation, clinical chemistry and hematology parameters. In vivo toxicological characterization is also available for all in vivo efficacy models offered by Pharmatest.

Extensive: up to 20perform compounds / 2required months Pharmatest will then the efficacy tests in-house and outsource the ADME/Tox assays to our highly experienced network of partner CROs. All work at we Pharmatest is to performed in This way are able offer our accordance with The Pharmatest customers large packages of projects, Services Assurance Program to including Quality a complete set of ADME/Tox ensure the efficacy qualityprojects of all needed services projects and in performed at Pharmatest. As part of this preclinical phases of drug discovery, from Program, we guarantee that if toxicology the results basic pharmacokinetics and of theto control groupefficacy are not studies, statistically tests regulatory all significantly from If the results of managed bydifferent Pharmatest. necessary, the group, Pharmatest will repeat we baseline can also put you directly in contact the without additional costs to the withtesting the member of our partner network client. that is best equipped to perform your ADME/Tox studies.

The ADME/Tox assays offered by our partner network include comprehensive in vitro assays for prediction of ADME/Tox properties of your drug candidates, as Typical results of potential drug compound X in well as their further characterization in Pharmatests Bone Resorption (human) Assay. vivo. ( -10 M = 10 M test compound, etc.)

Pharmatest Services Ltd Itinen Pitkkatu 4C 20520 Turku, FINLAND Tel: +358 2 278 4700 Email: info@pharmatest.fi

www.pharmatest.fi