Week 7 Flashcards
Monoclonal antibodies
i. Abs derived from single B cell that has been fused with a multiple myeloma tumor cell → grows forever in culture
- B cell has info on what Ab to make, and myeloma cell is the machine
ii. Very expensive to produce, but promising use as anti-inflammatory agents
- Now trying to grow Ab in plants (cheaper)
Murine monoclonal antibodies
monoclonal made using B cells directly derived from immunized mice
i. (-omab)
ii. Can result in immune complex formation
1. HAMA - human anti mouse antibodies → serum sickness
2. Solved this problem with Chimeric and other forms of mAB
Chimeric monoclonal antibodies
mABs engineered at the DNA level to have mouse VL and VH domains but human C domains
i. (-ximab)
ii. Less likely to be recognized by patient’s own immune system - can be given several times, but will eventually develop HACA (human anti-chimeric antibody)
Humanized monoclonal antibodies
only CDRs of V domains are from the mouse
i.(-zumab)
Human monoclonal antibodies
all human, use a humanized mouse with SCID
i.(-umab)
NK cells
large granular lymphocytes (LGL), killers (similar mech to CTL)
i. No rearranged V(D)J genes, NOT thymic derived
ii. Have NK receptors which recognize DAMPs from stressed or dysregulated cells
iii. Part of the innate immune system
Antibody-dependent cell mediated cytotoxicity (ADCC)
- Requires LGL and antibody
- Not MHC-restricted the way CTL-mediated killing is
- NK cells have receptors for Fc of IgG → ab-dependent way to bind target cell
- IgG ab binds target cell → NK cell binds Fc IgG → NK cell triggered, delivers lethal signal to target → target dies via apoptosis
- Many mABs work by triggering ADCC
mABs in tumor therapy
i. mABs activate complement → tumor lysed or phagocytosed
ii. OR mABs activate ADCC
mABs tagged with radioisotope
used in imaging and as highly targeted therapeutic drug depending on which radioisotope is attached
BiTE: Bispecific T cell engager
i. Used specifically in lymphoma and non-philadelphia chromosome ALL
ii. CD-19 and CD3 targets
Hemostasis
ability of the body to stop bleeding from a damaged blood vessel and repair the defect in the vessel wall so that normal blood flow to and from the involved area can be maintained
Hemostasis components (5)
1) Coagulation cascade
2) Anticoagulation regulatory pathways
3) Fibrinolytic system (breaks down formed clots)
4) Endothelial cell lining of blood vessels (prevents clots in resting state, promotes clot formation following injury)
5) Platelets
Coagulation factors are synthesized in the ____
Name 3 exceptions
Liver
- Tissue Factor: NOT made in liver, expressed on surface of many cell types outside of vasculature
- VWF: NOT made in liver, produced by endothelial cells/megakaryocytes
- Factor VIII: can be made in liver, but also spleen, lung, and kidney
Most coagulation factors are ____ proteases
serine
Serine proteases
i. Includes majority of enzymes in coagulation cascade
ii. Serine proteases exist as Zymogens (inactive precursor proteins) → cleaved at arginine residues → active serine proteases
1. Exception: Factor XIII
iii. Factor XII, XI, IX, X, VII, II (prothrombin), and Prekallikrein
Coagulation cofactors (name 4)
initiate or accelerate enzymatic reactions (lack enzyme activity themselves)
i.Factor III (Tissue Factor), Factor V, Factor VIII, high molecular weight kininogen (HMWK)
Intrinsic tenase (4 components and what it activates)
Factor IXa (serine protease) + Factor VIIIa (cofactor) + phospholipid surface + calcium → activate factor X
Extrinsic tenase (4 components and what it activates)
TF (cofactor) + factor VIIa (serine protease) + phospholipid surface + calcium → activate factor IX or X
Prothrombinase complex (4 components and what it activates)
Factor Xa (serine protease) + Factor Va (cofactor) + phospholipid surface + Calcium → activate prothrombin (factor II) to Thrombin (IIa)
Vitamin K dependent factors (name 6 and their properties)
Factor II, VII, IX, X, anticoagulant protein C and S
- All contain Gla domain (9-13 glutamic acid residues) → post-translational modification to y-carboxyglutamic acid residues (Gla Residues)
a. y-carboxylation carried out by y-glutamyl carboxylase (enzyme)
b. Vitamin K required to generate precursor
i. No Vitamin K (reduced form)→ can’t make Gla residues - Gla residues (negatively charged) bind calcium → change shape of protein, allows binding to anionic phospholipid surface
Enzyme that gamma carboxylates the vitamin K dependent factors
y-glutamyl carboxylase
Fibrinogen to fibrin (explain the process)
Fibrinogen → Fibrin (by Thrombin) → formation of insoluble fibrin network
i.Factor XIII: stabilizes the forming clot
- Covalently links fibrin molecules together to form a stable clot
- Transglutaminase that forms amide bonds between specific lysine and glutamine residues of fibrin
ii. Fibrinogen: soluble plasma protein, cleaved by thrombin to Fibrin
iii. Fibrin: participates in forming actual blood clot
Structure of fibrinogen
Made of 6 polypeptides (2 pairs of 3) - A-alpha, B-beta, gamma
Von Willebrand Factor
i. Carrier protein for factor VIII in the plasma
ii. Stored in endothelial cells and platelets
iii. Binds factor VIII in circulating plasma → prolongs half life
iv. Deficiency → bleeding disorder similar to hemophilia A (factor VIII deficiency)
Extrinsic coagulation pathway
Tissue damage → Tissue Factor (TF) binds Factor VIIa → activate factor X
ii.Xa + Va → activate factor II → IIa → fibrinogen converted to fibrin
Intrinsic coagulation pathway
all necessary components contained in plasma
i. Surface contact → contact factors (XII, prekallikrein, HMWK) → activate factor XI → activate factor IX → IXa + VIIIa → activate factor X
ii. Xa + Va → activate factor II → IIa → fibrinogen converted to fibrin
iii. TENET = twelve, eleven, nine, eight, ten
Initiation phase of coagulation cascade
Vascular disruption → expose plasma TF → encounter free factor VIIa on cell surface (in presence of calcium) → TF-VIIa
- → binds Factor IXa, converts IX to IXa
- → converts VII to VIIa
- → activates Xa
Xa begins process of clot generation → activate V, bind Va → form prothrombinase complex → activate thrombin
Amplification phase of coagulation cascade
occurs on surface of activated platelets adhered to exposed subendothelium
i.Thrombin:
- Amplifies procoagulant signal
- Activates Factor V and VIII → Va and VIIIa
- Activates XI → XIa
ii.Platelets activated → Va and VIIIa bind platelet surface → Xa forms prothrombinase complex with Va → thrombin formation from prothrombin
Propagation phase of coagulation cascade
i. Assembly of procoagulant complexes on cell surface
ii. Intrinsic tenase complex (VIIIa+IXa) → activate more factor X on platelet phospholipid surface
1. 50-100x faster than extrinsic tenase
2. → Xa binds Va → prothrombinase complex → converts prothrombin to thrombin
3. → thrombin cleaves fibrinogen to fibrin → form fibrin clot → activate XIII to XIIIa → cross-link/stabilize clot
iii. Platelets: surface for coagulation reactions to occur, generate more thrombin
1. Secrete contents of granules containing components of coagulation
Thrombin as the central enzyme (5)
i. Prothrombin converted to Thrombin by tissue thromboplastin in the presence of calcium → fibrinogen to fibrin by thrombin
ii. Cleaves and activates V and VIII to Va and VIIIa
iii. Activates XI to XIa which then generates for IXa
iv. Amplifies coagulation response
v. Most potent activator of platelets
4 steps of primary hemostasis
i. Platelet Adhesion
ii. Platelet Activation
iii. Platelet Aggregation
iv. Fibrin formation and support of local coagulation
1. Locally thrombin converts fibrinogen to fibrin stabilizing clot
End goal of primary hemostasis
formation of platelet plug (weak, unstable)