12 - Non-clinical tox

Question 1

Sulfanilamide

Answer 1

Marketed as cough medicine
* Changed from solid to liquid form for kids
* Kills >100 people
* There was NO tox testing

Question 2

Poly and diethylene glycol

Answer 2

Polyethylene glycol (PEG) is a common excipient in meds (despite tox)
* Contain repeats of ethyl ester
* DIethylene (DEG) glycol used in Sulfanailamide

Question 3

DEG toxic metabolites

Answer 3

Metabolised to HEAA and DGA
* Accumulate in kidney
* Renal toxicity, metabolic acidosis, neuropathies

Question 4

How do reduce HEAA

Answer 4

1. Supportive care
2. Fomepizole (inhibitory of alcohol dehydrogenase)
3. Ethanol (STOP metabolism of DEG)

Question 5

Some regulatory bodies in medicine

Answer 5

FDA, TGA (Aus), EMA (Eu), MHLW/PMDA (Japan)

Question 6

AMA

Answer 6

Approved Market Authorisation
* Drug companies need to have to market a drug in each jurisdiction

Question 7

ICH

Answer 7

International Council for Harmonisation
* Guidelines for scientific studies to establish safety, quality and efficacy
* Standard format for reg
* Common Technical Doc (CTD)

Question 8

Common Technical Document (CTD)

Answer 8

Internationally agreed set of specifications for a submission
* 5 modules
* Non-clinical and clinical data

Question 9

TGN1412 (2006)

Answer 9

Phase 1 clin. trial
* Given TGN1412 ➔ CD28 superagonist
* Expansion of T cells well tolerated in animals
* Humans ➔ cytokine storm

Question 10

MABEL

Answer 10

Minimum Anticipated Biological Effect Level
* Do the calc. for agonist receptor occupancy
* Determine the Kd experimentally

Question 11

Non-clinical Toxicology Studies to Support Safety in Humans

Answer 11

1. Single Dose Toxi
2. Repeat Dose Tox
3. Genotoxicity
4. Carcinogenicity
5. Reproductive Tox
6. Safety Pharmacology
7. Pharmacokinetics Studies

Question 12

GLP

Answer 12

Good Laboratory Practice
1. Resources
2. Characterisation (test items and systems)
3. Rules
4. Results (record all changes)
5. Quality Assurance

Question 13

Single Dose (Acute) Tox Studies

Answer 13

• 2 mammalian, one non-rodent species
• 2 routes: clinical (oral) + parenteral (IV)
• Dose should exceed clinical exposure (overdose pharma)
• NOT MANDATORY

Question 14

Repeat-Dose Toxicity (RDT) Studies

Answer 14

Detect longer term effects
* Species: 2 mammalian (1 non-rodent)
* Route: usually clinical
* Dose: up to max. tolerated dose (MTD), may be limited by solubility (MFD used)

Question 15

Repeat-Dose Study Duration

Answer 15

Will be more than below values if clinical treatment duration is longer
1. 14 or 28 day repeat-dose (both species)
2. 6 month repeat-dose (rodent)
3. 9 month repeat-dose (non-rodent)
4. 2 year carcinogenicity (rodent only)

Question 16

RTD Data Collection

Answer 16

Provides more insight into the possible response of the drug
* e.g. toxicokinetics

Question 17

Genotoxicity

Answer 17

DNA damage ➔ mutations
* Reactive electrophiles attack nucleophilic sites on DNA bases
* Guanine contains multiple sites where adduction can occur (important positions N7 and O6)

Question 18

ICH Genotoxicity Testing Requirements

Answer 18

1. (all of) Ames test, cytogenetic test for chromosomal damage, in vivo test for genotox in rodent haemopoietic cells
2. (both of) Ames test, in vivo test for genotox with 2 diff. tissues

Question 19

Ames test

Answer 19

Salmonella mutated to be histidine dependent
* Put on histadine-deficient plate with drug and liver extract
* If histadine-independent revertants grow back ➔ mutated back to functional gene

Question 20

Emergency “A Rule” of DNA Polymerisation

Answer 20

If DNA polymerase encounters lesion in DNA template strand (due to adducts) ➔ use this rule
* ADD deoxyadenosine and pass over lesion ➔ allows replication to continue and cell survival
* Often OK (most mutations occur in non-critical codons)

Question 21

In Vivo Carcinogenicity

Answer 21

1 long-term rodent (generally rat – 2y)
* Long–med in second rodent species
* Organs examined for tumours
* If tumour findings, follow up with mechanistic studies (to determine relevance to humans)

Question 22

Reproductive Studies

Answer 22

Allow exposure of mature adults and all stages of development from conception to sexual maturity
* Divided into subcategories

Question 23

ICH S5 (r3) Requires observations through one complete life cycle

Answer 23

1. Premating ➔ conception
2. ➔ Implantation
3. ➔ Closure of hard palate
4. ➔ END of pregnancy
5. Birth ➔ Weaning
6. ➔ Sexual maturity

Question 24

Types of Safety Pharmacology studies

Answer 24

1. Primary Pharmacodynamic - Studies on MOA and effects in relation to target
2. Secondary Pharmacodynamic - Studies on off-target effects etc.
3. Safety Pharmacology (S7A) - Potential undesirable pharmacodynamic effects in therapeutic range or above

Question 25

What systems does safety pharma usually look at?

Answer 25

1. CV 2. CNS 3. Respiratory

Question 26

Safety Pharmacology: CV

Answer 26

* ***BP***, ***HR***, ***ECG*** changes (esp. **QT** interval) * Also **in vitro** and **ex vivo** studies - detect ***repolaristion*** and ***conductance*** abnormalities

Question 27

Safety Pharmacology: CNS

Answer 27

* ***Motor***, ***sensory***, ***behaviour*** * **Reflex** response * Body **temp** * For behaviour ➔ ***Irwin's*** test

Question 28

Irwin’s Test (CNS)

Answer 28

1. Observations in home ***cage*** (e.g. **bizarre** behaviour) 2. ***Open field*** observation (e.g. **alterness**) 3. ***Manipulation*** observations (e.g. *visual* **placing**) 4. ***Invasive*** manipulation (e.g. **pain** response)

Question 29

Safety Pharmacology: Respiratory

Answer 29

*Whole* body **plethysmography** of rodents determine: * ***Respiratory*** rate * ***Tidal*** vol. * *Peak* ***inspiration*** / *expiration* AND **O2** *saturation*

Question 30

What questions can be asked to determine one's chances of detecting toxicity?

Answer 30

1. Are the **numbers** *sufficient*? (rule of ***3***) 2. Are the **dose** levels *relevant*? (***allometric*** scaling)

Question 31

Event rate

Answer 31

How many **cases** (***Y***) did you see when (***X***) number of people were **exposed**? * ***Y/X*** * What if Y = **0**? But X was *too* **small** * Need ***SUFFICIENT*** numbers

Question 32

If trial finds occurrence of NO EVENTS does this still contribute to risk estimation?

Answer 32

**Yes** it does * If ***no*** event occurs in **n** trials we can be **95%** confident that ***longrun*** risk **≤ 3/n** (rule of three)

Question 33

Rule of Three

Answer 33

Provides excellent ***estimate*** of probabilities for **n≥30** * No. of **subjects** needed = ***3x*** *estimated* **frequency** of event

Question 34

Allometric Scaling

Answer 34

***Direct*** comparisons of dose/unit body weight are **NOT** always ***suitable*** (e.g. **metabolic** processes /rates are *proportional* to body **SA**) * ***Conversion*** tables are available for common species (**Km** factors) * 𝑯𝒖𝒎𝒂𝒏 𝐸𝑞𝑢𝑖𝑣𝑎𝑙𝑒𝑛𝑡 𝐷𝑜𝑠𝑒 (𝑚𝑔/𝑘𝑔) = 𝐴𝑛𝑖𝑚𝑎𝑙 𝑑𝑜𝑠𝑒(𝑚𝑔/𝑘𝑔 ) × (𝑎𝑛𝑖𝑚𝑎𝑙 𝑘𝑚 / 𝒉𝒖𝒎𝒂𝒏 𝑘𝑚)