extra reading

toxicity through through extensive target molecule modulation

toxicity through through extensive target molecule modulation

too much of anticoagulant coumadin → uncontrollable bleeding and death

toxicity through modulation of similar proteins

antiviral drug against protease may also inhibit proteases needed for normal proteases eg those regulating blood pressure

toxicity through modulation of unrelated protein eg ion channels

many compounds block ion channels eg K+ channel hERG → potentially life threatening cardiac rhythm changes

drugs discovered by serendipity

discovered by chance eg penicillin, chlorpromazine and sildenafil

chlorpromazine (no longer really used)

psychosis drug originally discovered through investigation of drugs directed towards treating shock in surgical patients

how does taxol/paclitaxel work?

enters cancer cell and binds microtubule beta subunit preventing MT depolymerisation and alignment of chromosomes in centre of cell (metaphase) → mitotic arrest

what do functional assays measure

effect or response of a molecule eg enzyme inhibition or gene expression

how is potency of a molecule for a functional assay usually reported?

IC50 or EC50 - conc required to cause 50% maximal response

what do binding assays measure

extent to which a molecule binds its target

Kd

conc required to reach 50% of target binding - how result of binding assays are expressed

differences between biochemical and cellular assays

Cellular assays use intact cells whereas biochemical don’t. Biochemical assays used for both functional and binding assays whereas cellular mainly used for functional assays

counter assay

detects interference with primary assay to help eliminate false positives

orthogonal assay

measures activity differently from primary assay to confirm activity seen in primary assay

secondary assay

confirms activity in primary assay by testing functional activity downstream of target

-cillin family structure activity relationship

contain beta lactam ring which is important for inhibiting cell wall biosynthesis

hit ranking and clustering

confirmed hit compounds ranked according to effectiveness

freedom to operate evaluation

hit structures checked to determine if patentable

genetic differences causing a drug to not be as effective

slight encoded differences in drug target molecule or proteins in drug transport and metabolism

most common allele for beta1 adrenergic receptor and how does this differ in others

serine at p49 and arginine at p389. In some people glycine replaces one of these residues

How do the effects of metoprolol differ based on beta 1 adrenergic alleles present?

Two copies of common allele = daytime diastolic BP reduced by 14.7+/ - 2.9mmHg. One variant allele = smaller BP reduction, two variants = no significant effect

what are thiopurine drugs eg azothiprine used to treat?

leukaemia, Crohn’s, rheumatoid arthritis and IBD and organ transplant

what are thiopurines?

purine antimetabolites

why do some patients treated with thiopurines show signs of toxicity at typically well tolerated doses?

rare variations in gene encoding xenobiotic metabolising enzyme thiopurine methyltransferase which methylates sulphur - less stable in toxic patients

when is a drug having more side effects accepted?

if drug for severely ill patients eg with late stage cancer where there’s greater consequences for not having effective treatment

how does cancer develop multiple drug resistance

proliferation of cancer cells overexpressing ABC transporter proteins which pump the drugs out od the cell

bcr-abl formation

chromosome 9 ABL1 gene fuses BCR gene on chromosome 22. Changed chromosome 22 = Philadelphia chromosome

when is imatinib used?

to treat adult and paediatric CML in blast crisis, accelerated phase or chronic phase after IFN alpha therapy fails

blast crisis characterisation

high blast cells in blood or bone marrow and enlarged spleen

imatinib resistance in imatinib resistant blast crisis patients (T315l)

SNP in Abl gene causing Thr → Ile at amino acid 315 which still allows ATP to bind but creates steric hindrance and preventing imatinib binding

what cells usually express yamanaka factors

embryonic stem cells

Sox2 and Oct4 function

activate transcription factor Nanog and others, activating pluripotency and blocking differentiation

cMyc function

demethylates chromatin allowing Oct4, Sox2 and Nanog to be accessed and transcribed

Klf4 function

prevents apoptosis allowing cell to complete transition

what causes FOP?

dysregulated activity of bone morphogenetic protein receptor kinase ALK2, 97% of patients have gain of function mutation R206H which alters receptor intracellular domain

saracatanib (AZD0530)

dual SRC/ABL inhibitor which is both potent and selective for BMP and not TGF beta

saracatanib scaffold

quinazoline - yielded many approved drugs inc gefinitinib and vandetanib

FOP ALK2 function

abnormally transduces BMP signaling in response to activin A, a molecule that normally transduces transforming growth factor β (TGF-β) signaling but not BMP signaling

mouse model of FOP

FOP iPSC derived ectopic bone formed in mice

what are organoids

3D multicellular aggregates derived from stem cells that differentiate and self-organize to create the structural features and cell–cell interactions of mature tissues

neural organoids derived from autistic patient iPSCs

showed normal early neuronal differentiation followed by relative increase in inhibiory GABAergic over glutamatergic fate due to FOXG1 overproduction

benefits of using human stem cells to study chlamydia over genetically manipulated mice cells

different macrophage response to bacteria between humans and mice and immortalised cell lines are very far from the normal genetic state. Also monocytes and macrophages difficult to genetically manipulate

how were iPSC models of chlamydia pathogenesis made

human iPSC derived macrophages and infected them with chlamydia, observing genes dysregulated in chlamydia infection to find potential drug targets IFRF5 and IL10RA