1

Boards and Beyond: Hematology

1. Basic Pharmacology
2. Behavioral Science
3. Biochemistry
4. Biostatistics and Epidemiology
5. Cardiology
6. Cell Biology
7. Dermatology
8. Endocrinology
9. Gastroenterology
10. Genetics
11. Hematology
12. Immunology
13. Infectious Disease
14. Musculoskeletal
15. Neurology
16. Pathology
17. Psychiatry
18. Pulmonology
19. Renal
20. Reproductive
 2

- Coagulation
• Factor 10a converts prothrombin (2) thrombin (2a)
• Thrombin converts fibrinogen (1) fibrin (1a)
• Tissue Factor (Thromboplastin):
Constitutively expressed in subendothelium
No significant contact with circulating blood
Becomes exposed after endothelial damage
Converts factor 7 7a
Tissue factor/7a complex activates 10a
PT test: Tissue factor added to sample of plasma Time to clot is measured
• Thrombin (2a): Positive feedback Can activate cascade
• Factor 8:
Not produced by liver; synthesized by endothelial cells
Circulates bound to vWF (↑ plasma half-life of factor 8)
vWF releases factor 8 in response to vascular injury
• Calcium (factor 4): Required for clot formation
EDTA (blood collection tubes) chelates Ca2+ to prevent clotting
• Factor 13: Cross-links fibrin (stabilizes fibrin)
Breakdown of cross-linked fibrin D-dimer
• Factor 12: Hageman factor
Produces bradykinin (converts prekallikrein to kallikrein)
Kallikrein converts high-molecular weight kininogen (HMWK) to bradykinin
Kinins are the link between inflammation and coagulation
Factor 12 is activated by contact with negative charges; determines intrinsic pathway
PTT test: Plasma added to silica (negative charge) Time to form clot is measured
• Antithrombin 3 (AT3)
Serine protease inhibitor
Endothelium produces heparan sulfate Activates AT3
• Thrombomodulin
“Protein C V8 clot buster”
Activates protein C (protein S is a required cofactor)
Inactivates factors 5 and 8
• Plasminogen
Synthesized by liver; converted to plasmin by tPA/urokinase/streptokinase
Plasmin degrades fibrin into monomers Fibrin degradation products + D-dimer
• Vitamin-K Dependent Factors: 2, 7, 9, 10, Protein C/S
• Platelet Plug Formation
vWF is produced by endothelial cells and megakaryocytes; bind (carries) factor 8
vWF is stored in Weibel-Palade bodies (inside endothelial cells and α platelet granules)
vWF released in response to vascular injury (collagen exposure) Binds subendothelial collagen
vWF binds Gp1b on platelets in damaged endothelium (subendothelium) Adhesion
Gp2b/3a (platelet receptor) binds fibrinogen/vWF Platelets stick together Aggregation

α Granules Dense Granules

Fibrinogen, vWF, Platelet Factor 4 ADP, calcium, 5HT
HIT: IgG against Platelet factor 4-heparin complex ADP: Binds P2Y1/P2Y12; ↓ cAMP (platelet activation)
PDE inhibitors (↑ cAMP) block platelet activation
 Intrinsic (PTT) Extrinsic (PT)
Factors 12, 11, 9, 8 Factor 7 3
Hageman Factor (12) Tissue Factor (Thromboplastin)
Heparin Warfarin

Coagulation Tests

Common Pathway: Thrombin

Time (Bleeding Time)

Intrinsic: PTT
Factor 12 (Hageman)
Silica (negative charge) and
plasma Altered by Heparin

Extrinsic: PT
Tissue Factor (Activated factor
7) Altered by Warfarin

Patient PT
INR =
Control PT

Normal INR = 1
Warfarin ↑ PT (2-3)
 4

- Virchow’s Triad (“Virchow’s SHED”): The 3 Primary Influences of Thrombosis

1. Stasis
Post-op surgery, long drive/flight, atrial fibrillation, polycythemia, dilated cardiomyopathy
(CHF), sickle cell anemia
Atherosclerotic vessel narrowing, bruits, valve disease
2. Hypercoagulability
Factor V Leiden Mutation (factor V resistance to protein C inactivation)
G20210A Mutation of Prothrombin (↑ prothrombin levels)
Oral contraceptive usage Low clot risk
Heparin-Induced Thrombocytopenia
Syndrome
Antiphospholipid Antibody Syndrome: Patients with antibodies against anionic
phospholipids activate platelets and interfere with protein C activity High clot risk
3. Endothelial Damage
May result from LOW anticoagulant activity (PGI2, thrombomodulin, t-PA)
Hypercholesterolemia, Endotoxin, Endocarditis, HTN
Homocystinuria (deficiency of cystathione -synthetase)

- Factor V Leiden Mutation

• AD disorder; MCC of inherited hypercoagulability in Caucasians
• Point mutation of factor V gene
Missense mutation: Guanine Adenine
Change: Arginine Glutamine on position 506 (near cleavage site)
Factor V becomes resistant to degradation by activated protein C
Factor V becomes active longer; and is NOT inactivated by protein C
• Complications: DVT, cerebral vein thrombosis, recurrent pregnancy loss

- Prothrombin Gene Mutation

• Prothrombin 20210 gene mutation; guanine adenine
• Mutations results in ↑↑ prothrombin production
• Risk of venous clots

- AT3 Deficiency
• AD inherited deficiency of antithrombin; NO effect on PT, PTT, or thrombin time
• Diminishes increase in PTT following heparin administration
↓ AT3 levels available for heparin to activate) Heparin resistance
• Acquired AT3 Deficiency: Nephrotic syndrome (loss of AT3 in urine), DIC (consumption), cirrhosis

- Protein C/S Deficiency

• Loss of ability to inactivate factors Va and VIIIa (V8)
• Risk of venous clots
• Hallmark: Warfarin induced skin necrosis
 5

- Antiphospholipid Syndrome
• May occur as primary disorder or secondary to SLE
• Risk of thrombosis (both arterial/venous)
• Lab Findings:
Lupus anticoagulant antibodies Prolonged PTT
Anticardiolipin antibodies False positive Syphilis tests (VDRL/RPR)
Anti-ß2 glycoprotein antibodies

- Hemophilias
• X-linked recessive gene mutations; run in families but also occur de novo
• Findings:
↑ PTT (↓ factor 8 or 9), normal PT, bleeding time, platelet count
Recurrent joint bleeds
• Hemophilia A: Factor 8 deficiency
• Hemophilia B (Christmas Disease): Factor 9 deficiency
• Treatment:
Replacement factor 8/9 via IV
Desmopressin: Releases Weibel-Palade bodies; ↑ vWF, ↑ Factor 8
Aminocaproic Acid: Inhibits plasminogen activity (↓ plasmin, less breakdown of clots)
Cryoprecipitate: Precipitate of thawed FFP; contains factor 8, vWF, fibrinogen
 6

Glanzmann’s Thrombasthenia
AR deficiency of Gp2b/3a receptors
Defect in platelet aggregation Absent platelet aggregation
• Findings:
Prolonged bleeding time, normal platelet count
Blood smear: Isolated platelets (NO clumping)
↓ Gp2b/3a (↓ integrin α2bβ3)
Platelet-to-platelet aggregation and defective platelet plug formation

- Bernard-Soulier Syndrome
• AR deficiency of Gp1b receptors
Defect in adhesion Absent platelet adhesion
Loss of platelet-to-vWF adhesion
• Findings:
Prolonged bleeding time, ↓ platelet count (thrombocytopenia), LARGE platelets
Abnormal ristocetin test
7
 8

- Thrombus Formation
• Fibrin + Activated Platelets Thrombus (blood clot)

- Platelet Activation
1) Thromboxane A2
2) ADP
3) ↓ cAMP (inside platelets)
4) Activated platelets express glycoprotein 2b/3a

- Aspirin (Acetylsalicylic Acid, NSAID)

• NSAID MOA:
Irreversibly inhibits COX-1 and COX-2 Longer DOA than its own t1/2
Covalently acetylates serine residues in active site of COX
• Antiplatelet MOA:
Blocks formation of TXA2 (platelet activator)
Irreversible Antiplatelet effects last until new platelets are produced
• Uses:
Analgesic, antipyretic, anti-inflammatory (reduces pain, swelling, fever)
Antiplatelet drug (CAD, stroke, MI, angina)
Reduce polyp formation in patients w/ primary familial adenomatous polyposis
• Dosages:
Low Dose: Reduces platelet aggregation
Intermediate Dose: Antipyretic, analgesic
High Dose: Anti-inflammatory
• Toxicity:
GI mucosa damage (ulcers and bleeding due to COX-1 inhibition)
Reye’s syndrome: Aspirin use in child with viral infection Hepatic encephalopathy
Nephrotoxic: Interstitial nephritis
Hypersensitivity reaction (↑ LT production) High risk w/ nasal polyps or asthma
Increased bleeding time (no effect on PT/PTT)
Toxic Dose: Tinnitus (salicylate alters CN 8), vertigo, hyperventilation (respiratory alkalosis)
OD: Anion gap metabolic acidosis, hyperthermia, coma, death (antidote: NAHCO3)

- Thienopyridines
• Ticlopidine, Clopidogrel (Plavix), Prasugrel
• MOA:
Irreversible inhibitors of P2Y12 receptor
Block effects of ADP on platelets
• Uses:
Added to aspirin for prevention of MI/stroke
Backup drug if patient is allergic to aspirin
• Toxicity: Bleeding, TTP
 9

- Ticagrelor
• Cyclo-Pentyl-Triazolo-Pyrimidine (CPTP)
NOT a thienopyridine
• MOA:
Reversible inhibitor of P2Y12 receptor
Block effects of ADP on platelets
• Toxicity: Dyspnea

- PDE Inhibitors
• Dipyridamole, Cilostazol
• MOA:
Inhibit PDE3 in platelets to raise levels of cAMP (↑ cAMP) to inhibit platelet activation
Recall that LOW cAMP favors platelet activation
• Dipyridamole:
Also blocks adenosine uptake by cells Raises [adenosine] Vasodilation
Added to aspirin for prevention of MI/stroke
Chemical-inducer for cardiac stress test (for patients that cannot get on treadmill)
• Cilostazol:
Also raises cAMP in vascular smooth muscle Vasodilation
Use: PAD (vasodilates to increase distal blood flow, prevents DVT, reduces claudication)
• Toxicity: HA (meningeal vasodilation), flushing, hypotension
Reflects the vasodilating properties of these drugs

- Glycoprotein 2b/3a Receptor Blockers

• Abciximab, Eptifibatide, Tirofiban
Abciximab: Fab fragment of antibody to 2b/3a
• MOA:
Bind and block 2b/3a receptors
Block ability of activated platelets to adhere to other platelets
• Use:
IV drugs used in acute care settings (hospitals)
Acute coronary symptoms, cardiac stenting
• Toxicity: Acute thrombocytopenia (must monitor platelet count)
 1

- Anticoagulant (Antithrombotic) Drugs

• Acute: Help eliminate clots that are already formed
• Prevention: Reduce risk of clots in high risk patients

- PT/PTT
• PT (Extrinsic)
Test: Add tissue factor
Evaluates Factors: 1, 2, 5, 10, 7
Time: ~10sec
• INR = (Patient PT)/(Control PT)
Normal = 1
Therapeutic (ex. Warfarin) = 2-3
• PTT (Intrinsic)
Test: Add silica (negatively charged)
Evaluates Factors: 1, 2, 5, 10, 8, 9, 11, 12
Time: ~30sec

- Heparin
• Polymer of glycosaminoglycan
Can be administered as unfractionated or low molecular weight heparin
Natural mediator found in mast cells
Liquid medication (IV or SQ administration) Acute onset
• Unfractionated Heparin (UFH):
Widely varying polymer chain lengths
Activates antithrombin 3 (ATIII) Inactivates thrombin and intrinsic
pathway Increases PTT (intrinsic pathway) and thrombin timeMust monitor
levels High doses can affect PT (extrinsic pathway)
Binds to many plasma proteins and cells in body Variable responses in patients
• Low Molecular Weight Heparin (Enoxaparin/Dalteparin):
Small polymers only (SQ administration)
Primarily inhibit factor 10a Does not affect thrombin time
Fondaparinux: Only acts on factor 10a
PTT very slightly increased compared to UFH
Lower risk of binding plasma proteins More predictable dosage/effect than UFH
• Toxicity:
Bleeding, thrombocytopenia (HIT)
Osteoporosis (longterm use)
Mildly elevated LFTs
• Protamine: Antidote to reverse heparin effects

- Heparin-Induced Thrombocytopenia (HIT)

• Immune reaction to heparin Generation of anti-heparin antibodies
Type 2 HS IgG antibodies bind platelet factor 4-heparin complex
Widespread platelet activation, 5HT release, and thrombin activation
Macrophages remove IgG-Heparin-PF4 complex
Platelet number drops, but thrombus formation increases Hypercoagulable state
• Serotonin-Release Assay: Test to determine if patient has HIT syndrome
 1

- Factor 10a Inhibitors

• Rivaroxaban, Apixaban Factor 10a converts prothrombin to thrombin
• Use: Alternatives to warfarin
• Toxicity: Increases PT and PTT (factor 10a is involved in both pathways)

- Direct Thrombin Inhibitors

• Hirudin, Lepirudin, Bivalirudin, Desirudin, Argatroban, Dabigatran
All are IV/SQ administration except Dabigatran (oral)
• MOA:
Directly inhibit thrombin
Prolong PT, PTT, and thrombin time
• Uses:
HIT (Hirudin, Lepirudin, Bivalirudin, Desirudin, Argatroban)
Acute coronary interventions (Bivalirudin)
Atrial fibrillation (Dabigatran)

- Warfarin
• Oral, long-acting anticoagulant
• MOA:
Acts on liver Inhibits vitamin K epoxide reductase (VKOR)
Interferes synthesis (γ-carboxylation) of vit-K dependent clotting factors
Inhibits production of vit-K dependent factors 2, 7, 9, 10 and protein C/S
Affects extrinsic pathway (PT) Must monitor PT/INR (vit K levels differ in patients)
• Use: Chronic prophylaxis of coagulation (ex. A-Fib patient)
• Toxicity:
Initial risk of hypercoagulability (protein C deficiency is prothrombotic)
Skin necrosis (especially in protein C deficiency) Thrombosis of skin (dark purple lesions)
Teratogenic (heparin is safe for use in pregnancy)
OD Antidote: Vitamin K
 1

- Thrombolytics (Clot Busters)

• Tissue plasminogen activator (tPA), Streptokinase, Urokinase
• MOA: Activate plasminogen Convert plasminogen to plasmin (degrades fibrin)
• Uses: Acute MI, stroke
• Toxicity: High risk of bleeding

- Fresh Frozen Plasma (FFP)

• Plasma after removal of RBC, WBC, platelets
Contains clotting factors
• Use: Reversal of anticoagulant drugs
Corrects deficiencies of clotting factors
 13

- Hemolysis
• Extrinsic (outside RBC):
Antibodies
Mechanical trauma (narrow vessels, mechanical heart valves)
RBC infection
• Intrinsic:
Failure of membrane (hereditary spherocytosis)
Hemoglobin (sickle cell anemia)
RBC enzymes (G6PD deficiency)

- Hemolysis
• Intravascular (RBCs lyse inside blood vessels)
Microangiopathic hemolytic anemia
Mechanical heart valves
• Extravascular (RBCs lyse in liver or spleen)
Spleen: Damaged RBCs accumulate in cords of Bilroth (red pulp)
Hypersplenism: ↑ Splenic removal of RBCs

- Haptoglobin
• Plasma protein; binds free hemoglobin; marker of hemolysis
• Intravascular Hemolysis:
↑ Free Hgb released directly into plasma Binds haptoglobin
↓↓ Haptoglobin
• Extravascular Hemolysis:
Some Hgb released from spleen; free Hgb may spill into plasma
↓ or Normal Haptoglobin

- Parvovirus B19
• DNA virus; infects RBC progenitor cells; ↓ EPO
• Dangerous for those with chronic hemolytic anemia (they cannot tolerate ↓ EPO)
Sickle cell anemia, hereditary spherocytosis, ß-thalassemia major
Aplastic Crisis: Severe pallor, weakness, lethargy with LOW reticulocyte count
 14

- Autoimmune Hemolytic Anemia (AHA)

• RBC destruction from autoantibodies developed against your own RBCs
Positive direct Coombs test
• Warm AHA:
Antibodies bind at body temp (98.6°F)
IgG antibodies develop against RBC surface antigens
RBC removed by spleen Extravascular hemolysis
Fatigue, pallor, tachycardia, jaundice, splenomegaly
Spherocytes: Small, round RBCs with less surface area and no central pallor (↑ MCHC)
Associated with SLE, NHL, CLL, and methyldopa, PCN (hapten)
• Cold AHA:
Antibodies bind only in cold (<86°F); in the limbs, fingertips, nose, ears
IgM antibodies develop against RBC surface antigens
IgM fixes complement; leaves C3 bound to RBCs Positive direct coombs for C3
Painful fingers/toes, purple discoloration (especially in cold weather)
Associated with CLL, Mycoplasma pneumoniae infections, infectious Mononucleosis (EBV)
• Direct Antiglobulin (Coombs) Test:
Use: Diagnosis of AHA
Tests for RBC antibodies; determines presence of IgG (warm) or C3 (cold) coated on RBCs
Patient RBCs added to anti-IgG serum Agglutination? Positive direct Coombs
• Treatment: Steroids, IVIG (throwing stick for dog), splenectomy (recall it is extravascular hemolysis)

- Indirect Antiglobulin (Coombs) Test

• Use: Screening for maternal anti-D antibodies (erythroblastosis fetalis)
• Tests for antibodies present in serum (but NOT coated on RBC)
• Patient serum added to sample RBCs Agglutination? Positive indirect Coombs
Intrinsic Hemolytic Anemias: Something is wrong with the RBC structure
1
itself

- Paroxysmal Nocturnal Hemoglobinuria

• Acquired deficiency of phosphatidylinositol-linked glycoproteins (CD55 + CD59)
• Mutation of PIGA; prevents formation of GPI anchors for complement inhibitors
Decay accelerating factor; DAF (CD55) Disrupts C3b attachment
MAC inhibitory protein (CD59) Disrupts MAC
These are required to protect RBCs by slowing down complement
Deficiency of complement inhibitors Complement running free Destroys host RBCs
• Findings: Hemolysis (intravascular) + Hypercoagulability + Pancytopenia
Fatigue, jaundice, anemia, abdominal pain, pink urine (especially at night)
Complement-mediated hemolysis at night (↑ complement activity due to ↓ pH during sleep)
Free plasma Hgb is thrombogenic Venous thrombosis risk (Budd-Chiari syndrome)
Free plasma Hgb binds NO Depletion of NO
↓ NO increases smooth muscle tone Abdominal pains, ED, dysphagia
Autoimmune attack against GPI-antigens Pancytopenia, aplastic
anemia Risk of developing acute leukemias (AML)
Hemoglobinuria ↓ Haptoglobin, dark urine, iron deficiency (loss in urine), renal failure
• Labs: ↑ LDH, ↓ Haptoglobin, ↑ Urine hemosiderin/hemoglobin
• Diagnosis: Flow cytometry (monoclonal antibodies against GPI-anchored proteins CD55/59)
• Treatment: Eculizumab (anti-C5 complement mAb; prevents C5a formation to ↓ hemolysis)

- Pyruvate Kinase Deficiency

• AR; pyruvate kinase defect
• ↓ ATP production Rigid RBCs Extravascular hemolysis
• ↑ 2,3-BPG causes ↓ hemoglobin affinity for O2
• Findings: Splenomegaly and hemolytic disease of neonate

- Glucose-6-Phosphate Dehydrogenase Deficiency (G6PD)

• X-linked recessive hemolytic disorder
MC human enzyme disorder Deficiency of glutathione
Extravascular and intravascular hemolysis
• G6PD is involved in HMP-shunt Responsible for reducing NAPD to NADPH
NADPH reduces RBC glutathione Protection against RBC oxidative injury (ROS)
• Oxidant stress: ROS from infection (macrophages), drugs (see below), fava beans
Protective against malaria infections; reason why G6PD deficiency is MC in Africa
• Hemolysis in G6PD Deficiency:
Sulfa drugs, dapsone, antimalarial drugs (primaquine, quinidine), nitrofurantoin, INH
• Hemoglobin sulfhydryl cross-linking Protein denaturation
Recurrent hemolysis (dark urine) after exposure to trigger
Heinz Bodies: Precipitation of oxidized hemoglobin
Bite Cells: Phagocytic removal of RBCs by splenic macrophages
• Diagnosis: Fluorescent spot test (detects NADP generation)
 1

- Hereditary Spherocytosis
• AD defect of proteins interacting with RBC membrane and cytoskeleton
Mutation of spectrin, ankyrin, band 3, protein 4.2 Intrinsic hemolytic
anemia Blood Smear: Small, round RBCs with ↓ surface area and NO central
pallor
• Spheroid shape Loss of deformability and propensity to be trapped by spleen
Splenic macrophages destroy spherocytes Extravascular hemolysis
Marked splenic congestion with prominent erythrophagocytosis in the cords of Billroth
Reticulocytosis, marrow erythroid hyperplasia Anemia, splenomegaly, jaundice
• Complications:
Aplastic Crisis: Parvovirus B19 induces transient suppression of erythropoiesis
Pigment gallstones (bilirubin stones)
• Labs: ↑ MCHC, ↑ RDW, ↓ haptoglobin, ↓ MCV, ↑ LDH
Normal O2 carrying capacity
Negative Coombs test
Abnormally low fluorescence of RBCs in the eosin 5-maleimide (EMA) binding test
Osmotic fragility on acidified glycerol lysis test (measure Hgb release in hypotonic solution)
• Treatment: Splenectomy, folate supplementation, blood transfusions
If a splenectomy is performed, Howell-Jolly bodies will appear
 1

- Lead Poisoning
• Etiology:
Children: Exposure to lead paint (old house construction)
Adults: Exposure via occupation (batteries, ammunition)
• Lead inhibits 2 enzymes required for heme synthesis:
1) Ferrochelatase
2) ALA dehydratase
• Findings:
Accumulation of ↑ lead, ↑ ALA, ↑ protoporphyrin, iron levels low or normal
Microcytic hypochromic anemia with basophilic stippling
Bone Marrow: Ringed sideroblasts (nucleated RBC precursors with Fe-loaded mitochondria)
Abdominal pain (lead colic), constipation
Teeth: Lead-lines (blue pigment at gum-tooth line due to reaction of lead with
plaque) Renal damage (PCT damage Fanconi-like syndrome)
CNS damage: HA, memory loss, demyelination (peripheral neuropathy with foot/wrist drop)
Children: Mental deterioration, behavioral problems, developmental delay, failed milestones
• Treatment: Chelators
Dimercaprol, calcium disodium EDTA, succimer, penicillamine
 1

Thalassemia
Low/absent globin chain gene production
α Thalassemia: ↓ α chains; can affect all types of Hgb
ß Thalassemia: ↓ ß chains
Major vs Minor
Major: Severely low globin production, transfusions, death Minor: Usually asymptomatic; identified on routine blood
All thalassemia’s result in a microcytic hypochromic anemia

α-Thalassemia
4 genes code for α-globin chains (αα/αα); located on chromosome 16

• 4 Genes = 4 different scenarios of deletions

Missing 1: α-/αα = α-Thalassemia Minima
No anemia; silent carrier
Missing 2: --/αα OR -α/-α = α-Thalassemia Minor
Mild anemia, cis deletion worsens outcome for carrier’s offspring
Missing 3: --/-α = Hemoglobin H (HbH) Disease
Excess ß-globin forms ß4 globin (HbH); moderate-severe anemia
HbH has excess affinity for O2 (useless for O2 delivery)
Abnormal RBC deformability Splenic sequestration
Splenomegaly, extravascular hemolysis (↑ bilirubin), ↑ LDH
Missing 4: --/-- = Hemoglobin Bart’s Disease (Hydrops Fetalis)
Excess γ-globin forms γ4 globin; incompatible with life
γ4 globin has extremely high affinity for O2 (useless for O2 delivery)
Massive total body edema (anasarca); high-output heart failure; fetal death
• Gene Deletions: ↓ α chains
Cis deletion (deletion on same chromosome); MC in Asians Worse Prognosis
Trans deletion (deletion on separate chromosomes); MC in Africans
 1

- ß-Thalassemia
• 2 genes code for ß-globin chains; located on chromosome 11
• Gene Point Mutations in Splice Sites and Promoter Sequences
NOT gene deletions (as seen in α-thalassemia)
MC in Mediterranean populations
• ß-Thalassemia Minor: Heterozygote, ß+
ß-chain is underproduced; asymptomatic
Diagnosis: ↑ HbA2 (>5%) on gel electrophoresis
• ß-Thalassemia Major: Homozygote, ß0
ß-chain is completely absent; severe anemia

- ß-Thalassemia Major
• Basophilic stippling (residual RNA in RBCs)
• Target cells (↑ SA:volume ratio)
• Anisopoikilocytosis: Abnormal RBC shapes and sizes
• Bone Marrow: Erythroid hyperplasia (↑ EPO)
Marrow expansion; osteoporosis
Crew-cut on skull x-ray
Skull deformities Chipmunk facies
• Extramedullary hematopoiesis Hepatosplenomegaly
Spleen/liver produce nucleated RBCs
• Risk of parvovirus B19-induced aplastic crisis
• Diagnosis:
↑ HbF (α2γ2), ↑ HbA2 (α2δ2)
HbF is protective in the infant; symptoms begin after 6 months, when HbF production wanes
• Treatment: Blood transfusions
Repeated blood transfusions Systemic iron overload Secondary hemochromatosis
 20

- Sickle Cell Anemia

• AR point (missense) mutation on position 6 of ß-globin chain
Glutamate Valine (Adenine Thymine)
Mutant HbA HbS; causes intravascular and extravascular hemolysis
Deoxygenated Hgb polymerizes HbS Deform (sickle) RBC Anemia, vaso-occlusions
Sickling triggered by: ↓ O2 (↑ altitude), acidosis (↓ pH), ↑ 2,3-BPG
Newborns initially asymptomatic due to remaining presence of HbF
• Heterozygotes (Sickle Cell Trait):
Resistance to symptomatic Malaria; 8% of Africans carry HbS allele (↑ malaria prevalence)
Risk of renal medullary carcinoma
• Complications:
Painful vascular-occlusive crises: MC in bones, penis, spleen, liver, lungs
Dactylitis (painful swelling of hands/feet), priapism
Acute chest syndrome (respiratory distress, new pulmonary infiltrates on CXR)
Splenic infarct/sequestration crisis Splenomegaly (in children/young adults)
By adulthood; progressive fibrosis and spleen shrinkage Autosplenectomy
Autosplenectomy Howell-Jolly bodies
Sickling in renal medulla Renal papillary necrosis Gross hematuria
Risk of infection by encapsulated organisms (S. pneumoniae, H. influenzae)
Risk of Salmonella osteomyelitis
Risk of aplastic crisis due to parvovirus B19 infection
↑ EPO; bone marrow expansion can cause crew-cut on skull x-ray (same as ß-thalassemia)
Avascular necrosis (bone collapse at femoral head)
↑ Free Hgb inactivates NO; ↑ vascular tone; ↑ platelet aggregation Risk of stroke
Hyperbilirubinemia Risk of pigment gallstones
• Diagnosis: HbS detection on gel electrophoresis
↓↓ HbA, ↑ HbF, ↑↑ HbS
↑ Reticulocytes, ↓ ESR (sickled RBCs lower the sedimentation rate)
• Treatment: Hydroxyurea (increases HbF transcription, ↓ WBC production)
 He moglobinopathy electrophoresis patt erns

Starting point of
elle ct rophoresis

,,

I I Normal
individual

IIII I I Sick le cell

rait

I I Sickle cell
disease

I I Hemoglobin
C trait

I Hemogllobin C disease

I I
Hemoglobin
SC d ise a se

0 Hemoglobin electrophoresis patterns

Cathode Diagnos is Hemoglobin A Hemoglobin S Hemoglobin F Anode

Normal - 99% 0% <1%

Sickle cell disease 0% 85-95% 5-15%

Sickle cell trait 50-60% 35-45% <2%

 2

- HbC Disease
• Mutation on ß-globin gene with different AA substitution than HbS
Glutamate Lysine
• HbSC Heterozygotes (1 HbC mutant gene, 1 HbS mutant gene)
Milder disease than HbS homozygotes
• HbC Disease causes extravascular hemolysis
• Diagnosis:
Blood Smear: HbC crystals inside RBCs
RBC dehydration (↑ MCHC), target cells

- EPO
• Produced by interstitial cells of peritubular capillaries; found in the renal cortex
• ↓ EPO in renal failure Normocytic anemia

- Aplastic Anemia
• Pancytopenia (↓ RBC, WBC, platelets) Anemia, bleeding, infections
• Bone marrow failure (acellular bone marrow)
• Etiology:
Idiopathic (T-cell mediated destruction of stem cells)
Radiation (Hiroshima)
Benzene (rubber factories, shoe repair shops)
Chemotherapy, chloramphenicol (grey baby syndrome), methimazole/PTU
Parvovirus B19 (MC in immunocompromised people)
Acute viral hepatitis, HIV, EBV
Fanconi anemia
• Treatment:
Blood transfusions
Epoetin alfa, G-CSF (filgrastim), GM-CSF (Sargramostim)
Romiplostim (TPO analog), eltrombopag (TPO receptor agonist)
Immunosuppressants: Antithymocyte globulin, cyclosporine

- Fanconi Anemia
• Not to be confused with Fanconi syndrome (PCT disorder)
• AR DNA repair mutation; inherited aplastic anemia
Hypersensitivity to DNA damage (abnormal DNA strand cross-links)
Defect in DNA repair by homologous recombination
• Findings:
Short stature, café-au-lait spots, malformed thumbs
↑ Risk of malignancies
 2

- Pure Red Cell Aplasia

• Absence of erythroid precursors in bone marrow; markedly ↓ reticulocytes
• Normal granulocyte and platelet count in bone marrow
• Associated with thymoma

- Megaloblastic Anemia
• Hypersegmented neutrophils
• Etiology:
B12/folate deficiency
Orotic aciduria Most people (85-95%) are Rh positive
Methotrexate, 5-FU, hydroxyurea, zidovudine and possess the D-antigen
 2

Blood transfusion reactions

TYPE PATHOGENESIS TIMING CLINICAL PRESUTAflON IJONORBLOOD HOS1BLOOD

Allergic/ Type I hyperse nsitivity Within min utes Allergies: urt icaria, .,.-
A)
anaphylactic reaction agains t pla sma to 2-3 hr (due lo prn rit us
reaction proteins in transfused release of preformed Anaphylaxis:
i. (a,mtl ,

. ,
blo od inflam matory wheezing,
: ::: --·:·• '
• +

IgA-deficient in dividuals mediators in hypoteru io n,

should rece ive bloo d degran ulating mast respiratory arrest, Host mast llell
p rodu c ts without IgA cells) shock

Acute Type II hypersensiti vity During transfusion Fever, hypotension,

"
hemolytic reaction or with in 24 hr tachypnea,
transfusion Typically causes (due to preformed tachycard ia, =
h
reaction in travascular hemolys is antibodies) Hank pain ,
IDO'nll r llBC with A and} ll osu mli l -1\ antl-B lgG..
(ABO blood group hemoglobinuria
01 B gr,o p antlgem a
incom patib ility) (intravascular),
jaun dice
(ex:travascular)

.. .._,• ..
Febrile Cytokines created by Within 1-6 hr (due Fever, heada ches,
nonhemolytic donor WBCs accumulate to preform ed ch ills, flu sh ing
·.-·..
transfusion
reaction
dur ing storage of blood
produ cts
cytokines) More common in
ch ildren
: -
Domo, WBC: releases . .
IP'refolime cytol!ilrnes 11!1
Reactions prevented by
le ukoreduction of blood
products
Transfusion - Two-hit mec hanism: Within min utes to Respiratory dis tress,
related acute • Ne utrophils are 6 hr noncardiogenic
lung injury sequestered and pulmonary edema Hos

primed in pu lmona ry n utiop

vasculat ure due to

recipient risk factors Dom:ir a1mtll e1.1frocyt e,
l'gG
• N e utrophi ls are activated
by a product (eg,
antile u kocyte antibodies)
in the transfused blood
and release in flammatory
mediators ---. t capilla ry
pe rmeabi]ity
pulmonary edem a
----1-

Delayed
hemolytic
transfusion
Anam nestic response to a
foi:eign antigen on donor
RBC s (mos t com monly
Onset over 24 hr
Usually presents
within l-2 wk
Generally selfl imiied
and clin ically silent
Mild fever,
-
reaction Rh or other minor blood (due to slow hyperbilirubinemia Donor lllill.Cwl hi Host lgG

group antigens) previo usly destruct ion by lorelg11 amlg:e11s D

encountered by recipien t reticuloendotheliaI
Typically c.auses system)
extravascula r hemolysis
 2

Heme Synthesis
1. Succinyl CoA + Glycine; requires B6 (PLP)
Mitochondri

• Enzyme: -Aminolevulinic acid synthase (D-ALA)

RLS of heme synthesis; located in mitochondria
Heme and glucose inhibit D-ALA
• Product: -Aminolevulinic acid

2. -Aminolevulinic acid
• Enzyme: -Aminolevulinic acid dehydratase
Lead poisoning inhibits
Cytos

• Product: Porphobilinogen
• Porphyria: -Aminolevulinic acid porphyria
3. Porphobilinogen
• Enzyme: Porphobilinogen deaminase (PPG deaminase)
• Product: Hydroxymethylbilane
• Porphria: Acute intermittent porphyria
AD; ↓ PBG deaminase; random acute attacks
Exacerbated by CYP inducers; Tx is glucose
4. Hydroxymethylbilane
• Enzyme: Uroporphyrinogen III synthase
• Product: Uroporphyrinogen III
• Porphyria: Congenital erythropoietic porphyria
5. Uroporphyrinogen III
• Enzyme: Uroporphyrinogen decarboxylase (UROD)
• Product: Coproporphyrinogen III
• Porphyria: Porphyria cutanea tarda
Acquired UROD deficiency Accumulation of uroporphyrinogen (oxidized into uroporphyrin)
Uroporphyrin transported to skin Skin damage on exposure to
light Chronic blistering skin lesions, tea-colored urine (↑
uroporphyrin)
Liver: ↑ AST/ALT (uroporphyrin damages liver), ↑ iron storage (PCT is an iron overload)
Alcohol, HCV, HIV worsen disease
Treatment: Phlebotomy (removes excess iron to improve LFTS), hydroxychloroquine
6. Coproporphyrinogen III
• Enzyme: Coproporphyrinogen oxidase
• Product: Protoporphyrinogen IX
• Porphyria: Hereditary coporphyria
Cytos

7. Protoporphyrinogen IX
• Enzyme: Protoporphyrinogen oxidase
• Product: Protoporphyrin IX
• Porphyria: Variegate porphyria
Mitochondri

8. Protoporphyrin IX
• Enzyme: Ferrochelatase (located in mitochondria) + Fe2+
Located in mitochondria
Lead poisoning inhibits
• Product: Heme
 2
• Porphyria: Erythropoietic porphyria
 26

Porphobilinogen deaminase (PBG deaminase) deficiency: Acute intermittent porphyria

Uroporphyrinogen decarboxylase (UROD) deficiency: Porphyria cutanea tarda
 2

- Acute Lymphoblastic Leukemia (B-ALL)

• Pre-B cell malignancy; MC in children
• Blood Smear: Lymphoblasts (TdT+, CD10, CD19, CD20)
• Commonly metastasize to CNS and testes (need to inject chemotherapy prophylactically)
• 10-20x risk of trisomy 21 patients
• Prognosis: Good in children (cure rate
>80%) Poor prognosis in adults
• Etiologic Translocations:
t(12;21) TEL-AML1 fusion gene product Good prognosis
Philadelphia chromosome t(9;22) Poor prognosis (MC in adults)

- Acute Lymphoblastic Leukemia (T-ALL)

• Pre-T cell malignancy; MC in young adults (men in 20s)
• Presents as mediastinal mass (thymus enlargement)
May cause SVC syndrome or tracheal obstruction
• Blood Smear: Lymphoblasts (TdT+, CD1a, CD2-CD8)

- Acute Myelogenous Leukemia (AML)

• Malignancy of myeloblasts; MC in adult males age 65+
• BM suppression: Bleeding gums (thrombocytopenia)
• Blood Smear: Anemia, thrombocytopenia, myeloblasts
Myeloblasts: Myeloperoxidase (MPO) positive
Auer rods (accumulation of MPO)
↑ MPO can trigger DIC
DIC is a common presentation for AML
 2

- Acute Promyelocytic Leukemia (APL)

• t(15;17) PML-RARα fusion gene product Abnormal retinoic acid receptors
RARα = Retinoic Acid Receptor
• ↑ MPO DIC is a common presentation for APL
• Treatment:
All trans retinoic acid (vitamin A) Induce differentiation of promyelocytes
Arsenic trioxide Induce differentiation of promyelocytes

- Chronic Myeloid Leukemia (CML)

• Malignancy of myeloid progenitor cells Very high granulocytes and precursors
High basophils are a rare finding in normal people Warning sign for CML
• t(9;22) BCR-ABL fusion gene product
BCR gene (chromosome 22) fuses with ABL gene (chromosome 9)
BCR-ABL fusion gene creates non-receptor tyrosine kinase activity Transformation
Balanced reciprocal translocation of Philadelphia chromosome
Philadelphia chromosome (22) appears smaller than usual
• Findings: Peak incidence 65yo
Splenomegaly
LOW leukocyte alkaline phosphatase (LAP)
Blast Crisis: CML transforms to AML or ALL
• Treatment: Imatinib (BCR-ABL TYR kinase inhibitor)

- Chronic Lymphocytic Leukemia (CLL)

• Malignancy of naïve lymphocytes (NOT blasts; would be ALL)
• Small lymphocytic lymphoma has a very similar presentation
Same malignant cells, but presents with lower WBC
count
• Findings: Peak incidence 65yo
CLL is usually asymptomatic and found incidentally on CBC
Richter Transformation: CLL transforms into diffuse large B cell lymphoma (DLBCL; type of NHL)
Lymphadenopathy, splenomegaly, hepatomegaly can occur
Hypogammaglobulinemia (↓ IgG, IgA, IgM) Infections (MCC death)
Autoantibodies develop Autoimmune hemolytic anemia
• Blood Smear: CD5 B cells (co-expression of CD20/CD5)
CD5 (T cell marker) is found on B cells in CLL
Smudge Cells: Fragile lymphocytes get smooshed by blood smear prep
 29

- Hairy Cell Leukemia

• Rare chronic B cell malignancy; MC in adult males
• Associated with BRAF mutations
• Blood Smear: Mature B-cells with hair like projections
Express CD19, CD20, CD22 and CD103
• Findings:
Massive splenomegaly (red pulp engorgement)
Hairy cells cause BM fibrosis Dry tap on BM biopsy and pancytopenia
• Diagnosis:
Stain positive for tartrate-resistant acid phosphatase (TRAP)
Flow cytometry (CD103 expression on B cells)
• Treatment:
Cladribine (purine analog antimetabolite; adenosine deaminase inhibitor) Great efficacy
Pentostatin (adenosine deaminase inhibitor)

- Hodgkin Lymphoma
• Bimodal Distribution: Age 20s Age 60s
• Localized single group of LNs; orderly metastasis from LN to LN
• Reed Sternberg Cells: Malignant B cells with bilobular nucleus (owl-eyes)
Express CD15 and CD30; NOT positive for standard B cell markers (CD19, CD20, CD21, CD22)
Reed Sternberg cells only encompass a minority of enlarged LN
Reed Sternberg cells secrete mass amounts of cytokines Reactive lymphocytosis
B-symptoms are more common in Hodgkin lymphoma (↑ cytokines)
Macrophage 1α-hydroxylase activity Hypervitaminosis D Hypercalcemia
• Findings: Cervical lymphadenopathy, B symptoms, hypercalcemia
• Risk Factors: Prior EBV infection (infects B cells)
 30

- Non-Hodgkin Lymphoma (NHL)

• B and T cell malignancies; B cell cancers are MC
• Multiple malignant LNs with non-contiguous spread
• Extranodal involvement
GI (thickened bowel wall), skin changes

- DLBCL (NHL)
• MC NHL; common in HIV (AIDS defining malignancy)
• Treatment: Rituximab (CD20 mAb)

- Follicular Lymphoma (NHL)

• Indolent waxing and waning lymphadenopathy
• t(14;18) BCL-2 overexpression
Chromosome 14: IgG heavy chain
Chromosome 18: BCL2 Overexpression of BCL2 Anti-apoptosis

- Mantle Cell Lymphoma (NHL)

• Very aggressive B cell malignancy; arises from zone surrounding follicle or germinal center
• t(11;14) Cyclin D1 overexpression G1 to S
progression Chromosome 11: Cyclin D1
Chromosome 14: IgG heavy chain

- Marginal Zone Lymphoma (NHL)

• Lymphoma seen in chronic inflammatory disorders
Sjögren’s, Hashimoto’s, H. pylori (MALT lymphoma; induced by CagA/VacA genes)
• Translocation: t(11;18)

- Burkitt Lymphoma (NHL)

• Aggressive B cell malignancy; ↑ CD21 expression
Highly associated with EBV infection
• Starry sky morphology (clear spaces of white areas)
• t(8;14) translocation C-myc overexpression
• Endemic Form (Africa): Jaw mass
• Sporadic Form: Abdominal mass
3
 3

Non-Hodgkin lymph oma

TYl'E OCCUR S IN GENETICS COMMHIT5
Neoplasms of mature B cells
Bur kitt lymphoma Adolescents or young
t(8;14)- tra nsloca tion "S tarry sky" :ip pe,arance 1 shee ts of lymphocytes
::id ults with interspersed "tingible body" macrophages
of c - lU)'C (8) :md
heavy-c ha.in lg (a n ows in a ). Associated with EBY.
(14) Jaw les ion l]J in ,e ndemic form in Africa ; pelvis
or :ibdomen in spornd ic form.
Diffuse large B-cel
Usually older adults, M utatio ns in BCL-2, ost com mon type of no n - Hodgkin lymp homa
lymphoma
but 20% in children BCL-6 in ad ults.
Folllcular lymphoma
Adults t(l4 ;18 )- trnnslocat ion Indolent course with painless' waxing and
ofh eavy-cha i n lg (14) wan i ng" lymph ade nopathy . Bcl-2 norma
and .BC L-2 (18) lly inhibits. apoptosis.
Mantl ecell lymphoma
Adult males >> t(l1;14)- trnns location Very aggress ive, patients typica lly pre sent with
adult females of cyclin 01 (11) and late-stage dise:;is e .
heavy-chain lg (14),
CD5+
Marglnal zone Adnlts Associated with chronic inflammation (eg,
t(l l;18}
lymp homa Sjt:igren synd rome, chronic gastritis [tvlALT
lymphomal.
Primary central Considered an AIDS-defini n g illness. Variable
EBV related;
nervous system presentation: confusion, memory loss, seizures.
associated with HIV/
lymphoma CNS mass (often single, rin g enha ncing lesi on
AIDS
on MRI ) in immunocornpromised patien ts[i
needs lo be distinguished from toxoplasmosis
via CSF ana lysis or othe r lab tests.
Neoplasms of mature Tcells
Adult T-cell lymphoma Adults Ca used by HTLV Adults present with cutaneous lesions; common
(assoc iated with IV in Ja p an (T-ce ll in Tokyo), West Afr ica, and the
drug abuse) Caribbe:m.
Lytic bone le sions, hypercak e mia.
Mycosls fungoldes/ Adults Mycosis fu ngoide s: skin patches and plaques l!l
Se.zary syndrome (c uta neo us T-ce1l lym phoma), characterized by
atypica l CD4+ cells w ith ' l"l L: rifu 1 " 1 Ll
and intraepidermal neoplastic cell aggregates
( Paub ier micmabscess ). May progress to Sezar)'
syndrome (T-cell leu ikem ia).
 3

- Multiple Myeloma
• Malignancy of plasma cells; dependent on IL-6
Overproduction of certain type of immunoglobulin (paraprotein)
↑ IgG (50%), ↑ IgA (20%), ↑ Ig light chains (15%)
Overproduced Ig lacks antigenic diversity (NOT good at detecting antigens)
• AL Amyloidosis: Primary systemic
amyloidosis Light chain cast nephropathy
Ig light chains combine with Tamm-Horsfall mucoprotein Form casts
Large, waxy, eosinophilic casts made of Bence Jones proteins
Ig light chains (lambda/kappa) Bence Jones (BJ) proteins are light chains in urine
• Serum Protein Electrophoresis (SPEP):
Electrical current separates serum proteins based on size and charge
Gamma fraction contains Ig Multiple myeloma is a monoclonal gammopathy
Presents with ↑ Gamma Gap M-spike (monoclonal paraproteinemia)
• Findings:
Recurrent bacterial infections (loss of humoral immunity); MCC of death
Risk of primary light chain AL amyloidosis
Myeloma cell cytokine secretion activates osteoclasts
Osteolytic (punched-out) bone lesions (bone pain, fractures, osteopenia)
Hypercalcemia (metastatic calcifications, constipation, weak muscles)
BM replacement by plasma cells Normocytic anemia (fatigue)
Risk of Fanconi syndrome or Type 2 (proximal) RTA
LOW anion gap (IgG is cationic; pushes Na+ ions out of plasma)
• Blood Smear: Rouleaux formation (RBCs stacked in straight lines like poker chips)
• BM Biopsy: ↑ Monoclonal plasma cells; clock-face chromatin; intracytoplasmic inclusions of IgG
• Treatment: Bortezomib, Carfilzomib (proteasome inhibitors)
 3

- Myeloproliferative Disorders
• Malignant hematopoietic neoplasms
• JAK2: Gene codes TYR kinase
JAK2 Mutation: ↑ TYR phosphorylation

- Polycythemia Vera
• High blood viscosity (↑ RBC mass, ↑ blood volume, ↑ blood flow resistance)
• Findings:
HTN, flushing, red puffy skin (facial plethora)
Erythromelalgia (burning painful red/blue skin)
Thrombosis risk; episodic clots in extremities, DVTs, Budd Chiari syndrome
Aquagenic pruritis (itchy skin after taking a shower)
• Complications:
Progression to myelofibrosis
Leukemia (AML or CML)
Gout (↑ DNA turnover, ↑ purine metabolism)
• Treatment: Phlebotomy
Hydroxyurea (inhibits ribonucleotide reductase)
Ruxolitinib (JAK2 inhibitor)

- Essential Thrombocytosis (Thrombocythemia)

• Malignant proliferation of megakaryocytes/platelets (↑↑ platelet count)
• Findings:
Bleeding and thrombosis
Erythromelalgia may occur
• Blood Smear: Markedly increased platelets (abnormally large and irregular shapes)
• Treatment: Aspirin, hydroxyurea

- Myelofibrosis
• Obliteration of bone marrow with fibrosis due to ↑ fibroblast activity
“Bone marrow cries teardrops because it’s fibrosed and is a dry tap.”
• Findings:
Myelophthisic Anemia: Leukoerythroblastosis with teardrop-shaped RBCs; severely ↓ RBCs
Extramedullary Hematopoiesis: Massive splenomegaly (early satiety, abdominal pain)
3
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Chrom osomal translocations

TRANSLOCATION ASSOCI ATED DISORD ER NOTES
t(8 ;14)
Burkitt (Burk-8) lymphoma (c-my c act ivation The lg heavy chain genes on chromosome 14
t(ll ;l4 ) are constitutively expressed. When other
) Mantle cell lymphoma (cyclin DI activation )
t(ll ;l8 ) genes (eg, c-myc and BCL-2) are translocated
Marginal zone lymphoma
next to this heavy chain gene region, they are
t(l4;18) Follicular lymphoma (BCL-2 act ivatio n) overexpressed.
t(l 5;17) APL (M3 type of AM L; responds to all-trans
retinoic acid)
t(9; 22) (Ph il adelph CML (BCR-ABL hybrid), ALL (less common,
ia chromosome) poor prognostic factor); Philadelphia Crea M L
cheese

Acute lymphoblastic
Most frequentl y occurs in chi]dren; less common in adu]ts (worse prognosis). T-cel1 ALL can
leukemia /lymphoma
present as med iasti nal mass (presen ting as SVC -like syndrome ). Associa ted with Down syndrome.
Peripheral blood and bone marrow have t t t lymphoblasts IJ.
TdT+ (marker of pre-T and pre-B ce lls), CDlO+ (marker of pre-B cells).
Most respon sive to the rap y:
M ngnosis.
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 3

- Amyloidosis
• Extracellular deposition of fibrillar proteins Tissue damage Functional compromise of organ
Aggregation of misfolded proteins Formation of insoluble fibrils
Amyloid is a fibrillar protein that deposits in interstitial tissue
Pressure atrophy of adjacent cells at deposition site Systemic or local organ dysfunction
Linear, nonbranching filaments form a β-pleated sheet
• Amyloid is derived from (3) major precursor proteins:
1. AL Ig light chains (lambda/kappa) Bence Jones (BJ) proteins are light chains in urine
Primary systemic amyloidosis
AL (light chain amyloidosis) Plasma cell dyscrasias (multiple myeloma)
2. SAA (serum amyloid A) Acute phase reactant (released by liver during inflammation)
Secondary systemic amyloidosis
SAA is released during malignancy, autoimmunity, and familial Mediterranean fever (FMF)
FMF = Heredofamilial Amyloidosis (pyrin mutation leads to↑ IL-1); FMF treatment is colchicine
Chronic inflammatory conditions (IBD, RA, etc.)
3. APP (amyloid precursor protein) Generates Aβ (β-amyloid)
β-Amyloid forms cerebral plaques Deposits in cerebral vessels in AZD
• Other precursor proteins:
1) Transthyretin (carrier protein for thyroxine and vitamin A) Age-related
2) β2-Microglobulin (light chain component of MHC)
3) Prion proteins (PrPC, α-helical PrPSC, β-pleated)

- Amyloidosis (Histological Features)

• H&E Stain: Amyloid is eosinophilic (pink)
• Congo Red Stain: Amyloid shows apple green birefringence under polarized light

- Amyloidosis (Etiologies)
1) Excessive production of proteins prone to misfolding and aggregation
2) Mutations that produce proteins cannot fold properly
3) Defective/incomplete proteolytic degradation of extracellular proteins
Normally, proteasomes degrade misfolded proteins intracellularly

- Amyloidosis Diagnosis
• Abdominal fat pad biopsy
 3

- Amyloidosis (Clinical Organ-Based Findings)

• General Symptoms: Fatigue, dyspnea, edema, paresthesias, weight loss
• Renal Symptoms:
Kidney is MC organ affected Amyloid in glomeruli
Deposits in glomerular mesangium and tubular basement
membrane Proteinuria Nephrotic syndrome
Generalized pitting edema
• GI Symptoms:
Macroglossia (tongue enlargement)
Malabsorptive diarrhea
• Cardiac Symptoms:
Restrictive cardiomyopathy (amyloid infiltrates myocardium)
Diastolic heart failure, ↑ wall thickness (infiltration of amyloid; NOT hypertrophied
myocytes) Senile Cardiac Amyloidosis Transthyretin deposits in heart; found
in 25% of people >80yo Isolated Atrial Amyloidosis ANP deposits in atria Risk of atrial
fibrillation
• Neurologic Symptoms:
Aβ (derived from APP) deposits in brain Amyloid plaques Alzheimer disease
Gene for APP is found on chromosome 21 Down syndrome
Familial Amyloid Polyneuropathy Transthyretin deposits in nerves Peripheral neuropathy
• Spleen
Symptoms:
Splenomegaly
Sago Spleen: Deposition of tapioca-like granules (‘grains of sand’) in splenic follicles
Lardaceous Spleen: Large, map-like deposition in walls of splenic venous sinuses and red pulp
• Liver Symptoms:
Hepatomegaly
Normal liver function, despite pressure atrophy of adjacent hepatocytes
• Pancreas Symptoms:
T2DM Amylin deposition in pancreatic islets
• Thyroid Symptoms:
Medullary carcinoma of thyroid Calcitonin deposits in tumor
• Hematologic Symptoms:
Factor X deficiency in the AL (light chain) type
Pinch Purpura: Skin hemorrhages around orbit
• Long-Term Hemodialysis Patients:
Patients with ESRD
Β2-microglobulin deposition in carpal ligament (tunnel) of wrist Median nerve compression
Β2-microglobulin deposition in joints Destructive joint disease
 3

- Malignant Cancer Spread

1) Seeding within body cavities
Ex: Ovarian cancer seeding into peritoneal cavity
Ex: Medulloblastoma seeding from brain to CSF in spinal cord
2) Lymphatic spread
Carcinoma: Lymphatic spread 1st Hematogenous spread
Skip Metastasis: Cancer cells spread to most proximal LNs Continue to spread to nearest LN
Sentinel LN: 1st regional LN that receives lymphatic flow from a primary tumor
3) Hematogenous spread
Sarcoma

- Cancer Genes
1) Oncogenes: Over-expressed (mutated) versions of proto-oncogenes
2) Tumor Suppressor Genes
Normally prevent uncontrolled growth
Loss of tumor suppressor gene (point mutation) Transformation
Usually requires mutation of both (2) alleles “Two-Hit”
Hypothesis
3) Apoptosis Regulatory Genes
Underexpression (inactivation) of pro-apoptotic genes
Overexpression of anti-apoptotic genes
4) Tumor-Host Regulatory Genes: Genes that enhance or inhibit immune recognition of tumor cells

- Darwinian Selection in Cancer Cells

• Even though malignant tumors are monoclonal in origin they are typically genetically
heterogeneous by the time of their clinical presentation due to subclonal tumor cells
acquiring mutations (Darwinian selection Selective mutations give advantage for growth)
• Explains why over time cancer becomes more aggressive and less responsive to therapy
 4

- Gene Amplifications
• Double minutes (extrachromosomal structures)
• Homogenously staining regions (insertion of amplified genes into new chromosomal regions)
HSR = Diagnostic for gene amplification
• Examples:
1) Amplification of ERBB2 (HER2, an EGF receptor) or estrogen receptors 20% of Breast CA
2) Amplification of N-MYC (Neuroblastoma) or L-MYC (Small Cell Carcinoma of Lung)

- Overexpression of miRNAs
• Overexpression of miRNAs Reduce expression of tumor suppressors genes
• Deletion of miRNAs Overexpression of proto-oncogenes
Leukemia/Lymphoma Overexpression of BCL2 (anti-apoptosis)

- Epigenetics
• Reversible, heritable changes in gene expression that occur without mutation
Posttranslational modifications of histones
DNA methylation
• Oncogenesis: Tumor suppressor genes can be silenced via methylation of the promoter region

- Hallmarks of Cancer Enabling Factors for Cancer

1. Self-sufficient growth signals
2. Insensitivity to growth inhibitory signals
3. Altered cell metabolism
4. Evasion of apoptosis
5. Limitless replicative potential (immortality)
6. Sustained angiogenesis
7. Invasion and metastasis
8. Evasion of immune surveillance
1. Cancer-promoting inflammation
2. Genomic instability
Mutations in genes that regulate some or all
of these 8 cellular traits form the basis of the
molecular origin of cancer
 4

- Self-Sufficiency in Growth Signals

• Proto-oncogene gain-of-function mutation Oncogene activation
• Cancers secrete their own growth factors or induce stromal cells to produce growth factors
Autocrine activity
Ex: Glioblastomas secrete PDGF and express PDGF receptor
Ex: Sarcomas secrete TGF-α and express TGF-α receptor
• Oncoproteins promote cell proliferation:
Mutations in signaling molecule genes
Overproduction or unregulated activity of transcription factor
Mutations that activate cyclins (CDKs) or inactivate cyclin inhibitors CDKIs
Stimulus-independent expression of growth factor and its receptor
Mutation in genes encoding growth factor or its receptor (RTK mutant) Constitutive signaling
Amplification of ERBB2 (EGF receptor) gene Ex. HER2 amplification in breast CA
Amplification of ERBB1 (EGF receptor) gene Squamous cell carcinoma of lung, glioblastoma,
epithelial tumors of head and neck
RAS mutation Blocks hydrolysis of GTP to GDP Constantly active
Translocation of MYC Burkitt lymphoma
Fusion of ABL-BCR genes Constitutively active tyrosine kinase activity in CML

- Self-Sufficiency in Growth Signals: RAS Oncogene

• RAS is the most commonly mutated oncogene in human tumors (~30% of cancers)
Usually a point mutation within the GTP-binding pocket or GTPase enzyme
Interferes w/ breakdown of GTP Unable to revert back to inactive conformation
• Growth factor binds Exchange of GDP for GTP Activation of RAS Hydrolysis of GTP to GDP
Normally inactive when bound to GDP, but stuck bound to GTP in RAS mutations
NF1 = GTPase activating protein (GAP) tumor suppressor gene
Loss of function of GAP (NF1) = Gain of function of RAS
• RAS signals RAF/MAP kinase pathway and PI3 kinase/AKT/mTOR pathway MYC transcription factor
PTEN = Negative regulator of PI3 kinase/AKT pathway
Cowden Syndrome: Germline loss of PTEN
BRAF mutation 60% of melanomas

MYC is the most important transcription factor

upregulated by RAS mutations or ABL-BCR fusion.

Other transcription factors include FOS, JUN, MYB, and REL.

Direct dysregulation of MYC occurs when there is

a translocation t(8;14) of MYC in Burkitt
lymphoma (aggressive B cell tumor).
 4

- Self-Sufficiency in Growth Signals: ABL Oncogene

• Non-receptor tyrosine kinase
• Chronic Myeloid Leukemia (CML):
Part of ABL gene is translocated from chromosome 9 chromosome 22
(Philadelphia) Fuses w/ BCR gene ABL-BCR generates
constitutive tyrosine kinase activity
ABL-BCR activates all signals downstream of RAS

- Insensitivity to Growth Inhibitory Signals

• Tumor suppressor genes (RB, TP53, APC, BRCA, MEN1, NF1, NF2, PTEN, VHL, WT1, etc.)
• Products of tumor suppressor genes apply brakes to cell proliferation (anti-proliferative)
1) Signal dividing cells to enter G0 (quiescence dormancy)
2) Signal cells to enter post-mitotic differentiated pool Loss of replicative ability
3) Signal cells to enter non-replicative senescence (permanent cell cycle arrest)
• Loss of tumor suppressor gene Cells unable to slow down proliferation
“2-Hit”: Both alleles of tumor suppressor gene must be lost for expression of disease

- RB Tumor Suppressor Gene

• RB = “Master Brake” and “Governor of Cell Cycle”
Inactivated (lost) in most human cancers
• Anti-proliferative effect: Regulates G1 S phase
• Rb Active: Hypophosphorylated state
Complexed with E2F Prevents E2F from translocating to the nucleus and causing
transcription of proteins involved in proliferation
Rb-E2F complex = Anti-proliferative (inhibits progression into S-phase)
• Rb Deactivated: Phosphorylated state
Growth factors (EGF, PDGF) Cyclin D complexes with CDK4/6
Cyclin D-CDK4/6 phosphorylates (deactivates) Rb
Deactivation of Rb allows E2F to translocate Initiates progression of cell cycle into S phase

- TP53 Tumor Suppressor Gene

• TP53 is the most commonly mutated tumor suppressor gene in human tumors (~70% of cancers)
• TP53 = Guardian of the Genome
Controls expression of genes involved in cell cycle arrest, DNA repair, and apoptosis
Activated by cellular stress: DNA damage, anoxia, faulty oncogene signaling
Short half-life when cell is not under any stress (destroyed by MDM2)
If stressed ATM phosphorylates p53 Allows p53 to escape MDM2 Increases t1/2
• TP53 Mechanism of DNA Repair:
1) DNA damage ATM = Ataxia telangiectasia mutated
2) Phosphorylation (activation) of MDM2 = Potent inhibitor of p53
p53
3) Transcription of p21
4) p21 prevents Rb phosphorylation Keeps Rb active
Apoptosis
5) G1-S blockade Keeps cell in G1; inhibits progression into S-phase
6) Allows time to pause for DNA repair before proliferation continues
7) GADD45 activation DNA repair
8) If DNA damage is beyond the ability to repair p53 will induce apoptosis via BAX and PUMA
 4

- Cervical Carcinoma
• Begins with cervical dysplasia at basal layer of squamocolumnar junction (transformation zone)
Classified as CIN 1, CIN 2, or CIN 3 (severe, irreversible dysplasia or carcinoma in situ)
• HPV-16 and HPV-18
E6 gene Inhibits p53
E7 gene Inhibits Rb

- Insensitivity to Growth Inhibitory Signals: TGF-ß

• Growth factor; however, it is a potent inhibitor of proliferation of many types of cells
Activates CDKI’s and repression of CDK4 and MYC Growth inhibition
May become pro-oncogenic in late cancers
• TGF-β mutations
TGF-β receptor mutation Colon, stomach, endometrium cancers
Inactivation of SMAD (transduces TGF-β) Pancreatic cancer
• In late cancers, TGF-β signaling can activate epithelial-to-mesenchymal transition (EMT),
which promotes migration, invasion, and metastasis

- Contact-Mediated Growth Inhibition

• Normal cells STOP proliferation once they form confluent monolayers
• Cancer cells continue to proliferate after they come in contact with each other
• E-cadherins mediate cell-cell contact in epithelial layers Maintains contact inhibition
NF2 acts downstream of E-cadherin to help maintain contact inhibition
E-caderin binds ß-catenin (E-cadherin is lost in APC) Proliferation
Germline loss-of-function of E-cadherin Gastric adenocarcinoma

- Altered Cell Metabolism

• Warburg Effect: Shift of glucose metabolism away from mitochondrial oxidative phosphorylation
toward aerobic glycolysis (fermentation in the presence of oxygen) in tumors
Fermentation: Glucose converted to lactose (↑ lactic acid)
Aerobic glycolysis provides cancer cells w/ metabolic intermediates needed for synthesis
of cellular components (carbon for macromolecule synthesis, etc.)
• PET Scan:
Injection w/ 18F-flurodeoxyglucose (glucose analog) Preferentially taken up by tumors
• Autophagy
Sever cell nutrient deficiency Cells arrest growth Cannibalize own organelles for energy
Cancer cells may mutate to avoid autophagy (dormancy) or undergo autophagy (survival)
• Oncometabolite Production
Isocitrate dehydrogenase (IDH) mutations in the Krebs cycle
IDH acquires ability to produce 2-hydroxyglutarate (2-HG)
2-HG (oncometabolite) inhibits TET2 (DNA methylation regulator) Epigenetic changes
Apoptosis Pathways
Intrinsic (Mitochondrial) Extrinsic
Anti-Apoptosis Pro-Apoptosis • Death receptors, FAS, FAS-ligand
• BCL2, BCL-XL, MCL-1 • BAX/BAK, BAD, BID, PUMA

Chemotherapy and radiation activate intrinsic pathway

 4

- Intrinsic Apoptosis Pathway

• P53 senses DNA damage beyond repair Induces apoptosis via upregulation of PUMA
• Cancer:
1) Loss of P53 Prevents upregulation of PUMA Anti-apoptosis
2) Amplification of MDM2 (P53 inhibitor) Anti-apoptosis
3) Overexpression of BCL2 Anti-apoptosis
• Activation:
BAX and BAK (pro-apoptotic): Increase permeability of mitochondrial outer membrane
BH3-Only Proteins: BAD, BID, PUMA Neutralize effects of BCL2 Pro-apoptosis
Form pores in mitochondrial membrane Release of cytochrome c
Cytochrome c binds APAF-1 in cytosol Activates caspase-9 Activates caspase-3
• Inhibition:
BCL2 (anti-apoptotic): Keeps mitochondria impermeable Prevents release of cytochrome c
Antagonizes BAX and BAK
BCL2 Overexpression: Follicular B cell lymphoma t[14;18]

- Limitless Replicative Potential (Immortality)

• Hayflick Limit: Normal human cells have a maximum capacity of 70
replications Progressive shortening of telomeres
Shortened telomeres are recognized as dsDNA breaks P53/RB cause cell cycle arrest
• Nonhomologous End-Joining (NHEJ)
Last ditch effort to save cells that have P53 or RB mutations
• Tumor cells must lose growth restraints, avoid cellular senescence, and avoid NHEJ
Malignant cancers upregulate telomerase (normal cells lack expression of telomerase)

- Sustained Angiogenesis
• Solid tumors cannot enlarge >1-2mm until angiogenesis is established
• P53 normally stimulates expression of antiangiogenic molecules
Thrombospondin-1 Represses expression of VEGF
• Gain of function mutation of RAS-MAP Kinase pathway or MYC Upregulates VEGF
• Hypoxia triggers angiogenesis via hypoxia-inducible factor HIF-1α Activates VEGF

- Invasion and Metastasis

• Begins with clonal proliferation of transformed cell (capacity to metastasize)
1. Loss of cell-cell adhesion (down-regulation of E-cadherins)
E-cadherin sequesters ß-catenin Loss of E-cadherin ß-catenin translocation (proliferation)
SNAIL and TWIST transcription factors promote EMT Suppresses E-cadherin expression
2. Local degradation of basement membrane and interstitial connective tissue
Release of proteases (MMPs, cathepsin D, urokinase plasminogen activator) Degrade BM
Cleavage of type IV collagen, release of VEGF from ECM
3. Attachment to fibronectin in ECM Degradation of ECM
4. Release of cytokines to stimulate cell motility through matrix and CT
Hepatocyte growth factor (HGF) and scatter factor (SCF) Motility
5. Invasion of capillary (intravasation) Enter circulation
*Note: Some malignant cells are killed by T cells in circulation
6. Tumor cells that evade host immunity form tumor cell emboli (coated w/ fibrin/platelets)
7. Attachment to target organ capillary Extravasation Invade new metastatic site
CXCR4 Extravasation of circulating tumor cells to new tissue site (metastasis)
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 4

- Evasion of Immune Surveillance

• Antitumor activity is primarily mediated by T cells (CMI)
Tumor antigens are presented on the cell surface by MHC class I (recognized by CD8 CTLs)
• Cancer Immunoediting (how cancer avoids host immunity)
Acquire mutations in ß2-microglobulin Prevents formation of functional MHC I
Expression of proteins that activate immune checkpoints Inhibits CTL function
• Immunocompromised (↓ T cells) = ↑ Risk of oncogenic DNA viruses (EBV, HPV)

- Hereditary Nonpolyposis Colon Cancer (Lynch Syndrome)

• Autosomal dominant
• Inheritance of one defective DNA mismatch repair (MMR) allele
Each affected individual inherits one defective copy of MMR
Two-hit necessary (bi-allelic loss) Loss of heterozygosity
Microsatellite instability Repeated sequences predispose for replication errors
• High risk of colorectal cancer WITHOUT previous polyps

- Xeroderma Pigmentosum (XP)

• Autosomal recessive
• Defect in nucleotide excision repair (NER)
• Inability to repair pyrimidine dimers created by UV light
↑ Risk of skin cancer

- Defects in DNA Repair by Homologous Recombination

• Autosomal recessive cancer syndromes
• BRCA1 and BRCA2 are involved in homologous DNA repair
• Chromosomes are susceptible to damage by ionizing radiation
1. Ataxia-Telangiectasia (ATM mutation Neural symptoms)
2. Fanconi Anemia (DNA-cross link repair mutation Bone marrow aplasia)
3. Bloom Syndrome (Helicase mutation Developmental defects)

- Lymphoid Cancers
• Mutation in lymphocyte expressing genes
RAG1/RAG2, AID mutations

- Direct-Acting Carcinogens
• Requires no metabolic conversion to become carcinogenic
Alkylating agents (chemotherapy) May invoke
leukemia Acylating agents
 4

- Indirect Acting Procarcinogens

• Require metabolic conversion to become carcinogenic
Cytochrome P450 mediated metabolism
Induce mutations of P53
• Polycyclic hydrocarbons (combustion product)
Benzo[a]pyrene Scrotal cancer in chimney pipes, tobacco smoke
Benzo[a]pyrene is metabolized into epoxides Covalent adducts w/ cell DNA
• Aromatic amines and Azo dyes
ß-naphthylamine (aniline dye) Bladder cancer
Aflatoxin B1 (Aspergillus - moldy peanuts) HCC

- Activation of Pro-Carcinogens to Carcinogens

• Exogenous procarcinogens are activated into carcinogens in the liver via microsomal
cytochrome P-450–dependent mono-oxidases found in the smooth ER of hepatocytes

- Radiation-Induced Cancer
• Ionizing radiation Formation of dsDNA breaks
• Leukemia: MC radiation-induced cancer
• Papillary thyroid carcinoma: 2nd MCC

- Basal Cell Carcinoma (BCC)

• Most common skin cancer
Most common cancer due to UVB sunlight exposure
• Most common malignancy (of all cancers) in humans
• Locally invasive, but never metastasizes
 4

- Human T-Cell Leukemia Virus 1 (HTLV-1)

• Oncogenic RNA retrovirus
Adult T-Cell Leukemia/Lymphoma (ATLL) develops after long latency period (~50 yrs)
Virus exhibits tropism for CD4 T Cells Rash, lymphadenopathy, lytic bone lesions (↑ Ca2+)
Contains Tax protein Proliferation (upregulates cyclin D), activates NF-κB
Common in Japan, Africa, Caribbean

- Oncogenic Viruses
1. HPV
2. EBV
3. HHV-8 (Kaposi Sarcoma)
4. Merkel Cell Virus (Polyoma Virus)
5. Hepatitis B Virus (HBV) and Hepatitis C Virus (HCV)

- Human Papillomavirus (HPV)

• Benign squamous papilloma (warts): HPV 1, 2, 4, 7
• Benign genital papilloma (warts): HPV 6, 11
Low malignant risk
• Squamous cell carcinoma of cervix or anogenital region: HPV 16,
18 High malignant risk
Includes risk for oropharyngeal cancers
• E6 Oncoprotein
Binds and degrades P53
Stimulates expression of TERT (telomerase)
• E7 Oncoprotein
Binds E2F binding site Degrades RB
Inhibits CDK inhibitors p21 and p27
Activates cyclin A and cyclin E

- Epstein Barr Virus (EBV)

• Herpesvirus (HHV-4)
• Oncogenesis
CD21 EBV receptor attaches and infects B cells
LMP1 (CD40 mimicry) NF-κB and JAK/STAT pathway Polyclonal B cell proliferation
Generation of immortal B lymphoblastoid cell lines
EBNA2 Activates cyclin D and SRC (oncogenes)
vIL-10 Viral cytokine pirated from host genome Prevents MΦ-T cell activation
• Cancers:
Burkitt lymphoma from t(8;14) c-MYC gain of function
Hodgkin lymphoma
Nasopharyngeal carcinoma (China)
Uncommon: T-cell and NK-cell tumors, gastric carcinoma, sarcomas

- HBV and HCV

• 80% of HCCs worldwide are due to infection w/ HBV or HCV
• Immunologically mediated chronic inflammation Hepatocellular injury w/ ROS and NF-κB
• HBx protein (HBV) and core protein (HCV) activate carcinogenic signaling pathways
 4

- Helicobacter pylori
• 1st bacterium to be classified as carcinogenic
• H. pylori infections are associated w/ gastric adenocarcinoma and MALT
lymphoma Carcinogenic strains contain the CagA gene
MALT lymphoma = Monoclonal B-cell tumor of the stomach (marginal zone lymphoma)
• Chronic gastritis Gastric atrophy Metaplasia Dysplasia Neoplasia
Sequence takes decades, and only occurs in 3% of patients

- Cancer Cachexia
• Progressive loss of body fat and lean body mass, weakness, anorexia, anemia
• NOT caused by nutritional demands of the tumor
• Cachexia results from action of cytokines produced by the tumor and the host
TNF (MΦ’s, tumor cells) Suppresses appetite and inhibits LPL

- Paraneoplastic Syndromes
• Set of signs and symptoms that is a consequence of cancer, but unexplained by local mass
effects, metastasis, or elaboration of hormones
• Commonly caused by small cell lung CA, breast, RCC, and hematologic cancers
Hypercalcemia (Ectopic PTH, TGF-α, vitamin D)
Cushing syndrome (hypercortisolism from ectopic ACTH) Hypercalcemia secondary to
Nonbacterial thrombotic endocarditis (hypercoagulability) osteolytic bone metastasis is NOT
a paraneoplastic symptom
Small Cell Lung CA = ACTH
 5

- Grading and Staging of Cancer

• Grade = Level of differentiation
• Stage = Degree of spread
TNM Staging System Greater clinical value than grading

- TNM Staging of Cancer

• T: Tumor (size/depth of invasion)
T0: In situ (tumor has not spread beyond its original starting position)
T1-T4 (levels increase with increasing tumor size)
• N: Spread to regional lymph nodes
N0: No LN involvement
N1-N3: Increasing number of LNs involved
• M: Metastasis
M0: No distant metastasis
M1-M2: Increasing number of metastasis sites

- Lab Diagnosis of Cancer

• Sampling: Excision (biopsy), fine-needle aspiration (FNA), cytologic smears
Frozen Section: Permits histologic evaluation within minutes
• Flow cytometry (fluorescent-labeled antibodies) Leukemia and lymphoma diagnosis
• PSA Prostate CA diagnosis
• Carcinoembryonic Antigen (CEA) Colon, pancreas, stomach, breast CA diagnosis
• Alpha Fetoprotein (AFP) HCC, yolk sac tumors, teratocarcinoma diagnosis
• Molecular Analysis:
FISH or PCR Ewing sarcoma, leukemia, lymphoma
Point Mutation Analysis JAK2 mutation in polycythemia vera
 5

Goljan Pathology Neoplasia

- Tumors
• All tumors are clonal expansions of neoplastic cells with supporting stroma composed
of non-neoplastic connective tissue and blood vessels
Desmoplasia: Infiltrating fibrous stroma (seen in malignant tumors)
• Tumor type is based on characteristics of its parenchyma
Benign: Well-differentiated (resemble normal surrounding tissue); localized lesion
(usually epithelial or CT origin) without spread; amenable to surgical resection
Malignant: Undifferentiated; cancer; invade and metastasize to other sites

- Malignant Tumors
• Desmoplasia: Infiltrating fibrous stroma (makes them hard “scirrhous tumor”)
• Carcinoma: Epithelial cell origin
Squamous Cell Carcinoma (SCC): Oropharynx, larynx, upper/middle esophagus, lung,
cervix, skin
Adenocarcinoma: Lung, distal esophagus to rectum, pancreas, breast, liver,
endometrium, ovaries, kidneys, prostate
Transitional Cell Carcinoma: Bladder, ureter, renal pelvis
• Sarcoma: Mesenchymal (connective tissue) cell origin

Cell Type Benign Malignant

Epithelium -carcinoma suffix
Ectoderm Squamous/Transitional/Basal Papilloma Squamous/Transitional/Basal* Carcinoma

Ectoderm (Neural) Meningioma, Craniopharyngioma, Neuroma Glioma, Blastoma

Endoderm (Glandular) Adenoma (colon polyp, pituitary adenoma) Adenocarcinoma

*BCC invades but does NOT metastasize

Mesenchyme (CT) -oma suffix -sarcoma suffix
Mesoderm Blood Cells: N/A Blood Cells: Leukemia, Lymphoma
Blood Vessels: Hemangioma Blood Vessels: Angiosarcoma
Smooth Muscle: Leiomyoma Smooth Muscle: Leiomyosarcoma
Skeletal Muscle: Rhabdomyoma Skeletal Muscle: Rhabdomyosarcoma
CT: Fibroma, Chondroma CT: Fibrosarcoma, Chondrosarcoma
Bone: Osteoma Bone: Osteosarcoma
Fat: Lipoma Fat: Liposarcoma
Melanocyte: Nevus (mole) Melanocyte: Melanoma

ALL Layers (Germ Cell) Mature Teratoma Immature Teratoma

(Ectoderm, Endoderm, Dermoid Cyst Teratocarcinoma
Mesoderm) Placenta: Hydatidiform Mole Placenta: Choriocarcinoma (totipotent)
 5

- Tumor-Like (Non-Neoplastic) Conditions

• Hamartoma: Nonneoplastic, disorganized overgrowth of tissues in their native location
Ex. Bronchial hamartoma, Peutz-Jeghers polyps
• Choristoma: Heterotopic rest; mass of non-neoplastic cells in a foreign location
Ex. Pancreatic tissue growing in stomach wall, brain tissue in nasal cavity

- Tumor Differentiation
• Benign or Low-Grade Malignancy
Well-differentiated (resembles parent tissue)
Keratin pearls (squamous tissue)
Glands w/ lumens
• High-Grade Anaplastic Malignancy
No glands, no keratin

Properties of Malignant Tumors

Glycolysis (Anaerobic Metabolism is Primary Source of Energy)
↑ Lactate production
PET scan (radiolabeled glucose w/ CT scan) can detect ↑ glucose uptake
Angiogenesis (Required for O2)
TNF released by surrounding macrophages stimulate tumor cells to secrete VEGF
VEGF acts on endothelial cells to develop new capillary sprouts
Organelle Properties
Large, irregular nucleus
↑ Chromatin
↓ Mitochondria
↓ ER, ↑ Free ribosomes
↓ Cadherins (loss of cell adhesion)
Protein Activity
↑ Telomerase (allows increased cell longevity, inhibits apoptosis to allow survival)
↑ Decay-Accelerating Factor (prevents formation of MAC (complement)

- Invasion and Metastasis of Primary Tumor

• Clonal proliferation of transformed cell (capacity to metastasize)
1. Loss of cell-cell adhesion (down-regulation of cadherins)
2. Attachment to laminin in basement membrane Release of MMPs to degrade BM
3. Attachment to fibronectin in ECM Degradation of ECM
4. Release of cytokines to stimulate cell motility through matrix and CT
5. Invasion of capillary (intravasation) Enter circulation
*Note: Some malignant cells are killed by T cells in circulation
6. Tumor cells that evade host immunity form tumor cell emboli (coated w/ fibrin/platelets)
7. Attachment to target organ capillary Extravasation Invade new metastatic site
 5

- Pathways of Dissemination
• Carcinoma: Lymphatic spread to LNs
Afferent lymphatic vessel LN Efferent lymphatic vessel Thoracic Duct Systemic
Travels to LN first, then enters systemic circulation Hematogenous spread
Exception: Follicular carcinoma of thyroid only participates in hematogenous spread (no LNs)
• Sarcoma: Hematogenous spread
Invades capillary or venule Directly spreads to distant site via systemic circulation
Does NOT travel to LNs
• Portal Spread
Malignant cells in portal vein Metastasize to liver
• Vena Cava Spread
Malignant cells in vena cava Metastasize to lungs

- Seeding of Malignant Cells

• Malignant cells exfoliate from serosal surface Implant and invade tissue in a body cavity
• Peripheral lung cancers (adenocarcinoma) can seed parietal/visceral pleurae Malignant
pleural effusion
• Glioblastoma multiforme (GBM): High-grade cancer arising in the brain that commonly
exfoliates malignant cells into the CSF Seeds brainstem and spinal cord

- Vertebral Metastasis
• Vertebrae: Most common site of metastasis to bone
• Breast Cancer: Most common cancer that metastasizes to bone
• Prostate Cancer: 2nd most common
• Batson’s Plexus
Conduit for transport of metastasis to bone (vertebral column)
Has connections with vena cava and vertebral bodies
Ex: Breast tumor Intercostal vein Vena cava Batson’s Plexus Vertebrae

- Osteoblastic Metastasis
• Prostate Cancer: Most common cancer producing osteoblastic metastases
• Breast Cancer: 2nd most common
• Malignant cells in metastatic sites secrete cytokines
Activates osteoblasts
Initiates bone formation
• ↑ Serum alkaline phosphatase (ALP)
Osteoblasts use this enzyme in bone formation
• Osteoblastic metastases produce radiodensities that are identified in radiographs

- Osteolytic Metastases
• Osteolytic metastatic cancers: Lung cancer, renal cell carcinoma, breast cancer
• Malignant cells in metastatic sites produce chemicals (prostaglandin E2, IL-1)
Activates osteoclasts
Initiates bone destruction
• Osteolytic metastases produce radiolucencies that are identified in radiographs
• Pathologic fractures and hypercalcemia may be present
 5

- Sites Where Metastasis is More Common than Primary Tumors

• Lymph Nodes: Metastatic breast and lung cancer most common
• Lungs: Metastatic breast cancer most common
• Liver: Metastatic lung cancer most common
• Bone: Metastatic breast cancer most common
• Brain: Metastatic lung cancer most common

Cancer Epidemiology
nd
Cancer is the 2 most common cause of death for men/women in the USA
Men (Incidence) Women (Incidence)
1) Prostate 1) Breast
2) Lung 2) Lung
3) Colon 3) Colon

Men (Death) Women (Death)

1) Lung 1) Lung
2) Prostate 2) Breast
3) Colon 3) Colon
Children Cancers
nd
Cancer is the 2 most common cause of death in children 1-14 years old (#1: Accidents)
1) Acute lymphoblastic leukemia
2) CNS tumors (medulloblastoma)
3) Neuroblastoma
4) Others: Rhabdomyosarcoma, Wilms (kidney) tumor, retinoblastoma, osteosarcoma, Ewing sarcoma

- Cancer Geography
• Worldwide: Malignant melanoma increasing at most rapid rate
• China:
EBV Nasopharyngeal carcinoma
Alcohol, smoking SCC of esophagus
• Japan:
Smoked foods Stomach adenocarcinoma
• Southeast Asia:
Hepatitis B Virus (HBV) Hepatocellular carcinoma (HCC)
Aflatoxins (Aspergillus) in food (moldy peanuts) Postnecrotic cirrhosis
• Sub-Saharan Africa:
EBV Burkitt lymphoma
HHV-8 Kaposi sarcoma

- Screening Procedures to Detect Cancer

• Pap Smear
Detects cervical squamous dysplasia (precursor for SCC of cervix)
HPV-16 and HPV-18 can cause cervical cancer
• Mammogram
Detects non-palpable breast masses
• Colonoscopy
Detects colon polyps (precursor for colon cancer)
 5

- Cancer Biomarkers
• Prostate Specific Antigen (PSA): Prostatic adenocarcinoma, BPH
Serum PSA test has LOW sensitivity and specificity
• Carcinoembryonic Antigen (CEA): Colon, pancreas, stomach, breast carcinomas
• Alpha-Fetoprotein (AFP): HCC, yolk sac tumors, teratocarcinomas, neural tube defects (NTDs)
• Human Chorionic Gonadotropin (HcG): Testicular tumors
• CA-125: Ovarian tumors
• Assays of shed DNA from dying tumor cells can be collected in blood, stool, sputum,
or urine can look for mutated APC, P53, and RAS sequences

- Carcinogenic Gene Mutations

• Point mutation (most common), balanced translocation, viral genome insertion,
deletion, amplification (multiple copies of same gene), overexpression (↑ transcription)

Proto-Oncogene Mutations
Gain-of-function (translocation, amplification, overexpression, point mutation)
ABL Chronic myelogenous leukemia, Acute lymphoblastic leukemia (ALL)
ERBB2 (HER2) Breast carcinoma
C-MYC Burkitt lymphoma
N-MYC Neuroblastoma, Small cell carcinoma of lung
RAS 15-20% of all cancers
RET MEN2A, MEN2B, Leukemia
SIS (PBGFB) Osteosarcoma, Astrocytoma

Tumor Suppressor Gene Mutations

Loss-of-function (deletion, inactivation) AWAYS POINT MUTATIONS
APC Familial adenomatous polyposis (FAP), colon/stomach cancers (somatic)
BRCA1/2 Breast, ovary carcinoma
NF1 Neurofibromatosis Type 1: Pheochromocytoma + Wilms tumor + Neurofibrosarcomas
Somatic: Neuroblastoma
NF2 Neurofibromatosis Type 2: Bilateral acoustic neuromas (schwannoma) + Meningioma
Somatic: Schwannoma, meningioma
P53 Li-Fraumeni Syndrome: Breast carcinoma, brain tumors, leukemia, sarcomas
(17p) Somatic: P53 gene is the most common gene producing cancer
TGF-ß Retinoblastoma, osteogenic sarcoma, breast, lung, colon carcinomas
VHL von Hippel Lindau Syndrome: Cerebellar hemangioblastoma, retinal angioma, renal cell
carcinoma (bilateral), pheochromocytoma (bilateral)
WT1 Wilms tumor

- Von Hippel-Lindau Syndrome

• Autosomal dominant
• Loss of VHL gene on chromosome 3p
• Angiomas of the retina and hemangioblastomas of the cerebellum
• Risk of developing renal cell carcinoma and pheochromocytoma
 5

- Retinoblastoma
• Malignancy of the eye in children <5 years old
• Clinical Findings:
Leukocoria (abnormal white reflection of light on eye), loss of red reflex, strabismus
• 2-Hit Hypothesis:
Requires mutation in both Rb (tumor suppressor) gene alleles Loss of heterozygosity
• 60%: Sporadic (Nonheritable) Unilateral
Two somatic mutations of the RB1 suppressor gene on chromosome 13q occur
• 40%: Autosomal Dominant Bilateral
One of the RB1 genes on chromosome 13 is mutated in germline cells (before birth)
All somatic cells inherit one copy of mutated Rb gene from carrier parent
Subsequent 2nd mutation of RB1 gene on remaining chromosome 13 after birth Tumor
Risk for developing second malignancies Osteosarcoma, soft tissue sarcoma

- Rb (Tumor Suppressor) Gene

• RB1 is the “master brake” and governor of the cell cycle (Inhibits G1 S phase)
• Rb Active: Hypophosphorylated state
Complexed with E2F Prevents E2F from translocating to the nucleus and causing
transcription of proteins involved in proliferation
Rb-E2F complex = Anti-proliferative (inhibits progression into S-phase)
• Rb Deactivated: Phosphorylated state
Growth factors (EGF, PDGF) Cyclin D complexes with CDK4/6
Cyclin D-CDK4/6 phosphorylates (deactivates) Rb
Deactivation of Rb allows E2F to translocate Initiates progression of cell cycle into S phase

- Methods of Rb Dysfunction
1. Loss of function of Rb
2. Gene amplification of CDK4 or Cyclin D
3. Loss of function of CDK inhibitors (p16/INK4a)
4. Viral oncoproteins that bind and inhibit Rb *Binds to same site where E2F binds!
E7 protein of HPV (binds hypophosphorylated form of RB Prevents binding to
E2F) Polyoma virus, Hepatitis B
- Familial Adenomatous Polyposis (APC)
• Autosomal dominant colorectal cancer from malignant transformation of polyps
Germline loss of APC tumor suppressor gene on chromosome 5 (75% of colon cancers)
Thousands of adenomatous polyps develop in colon by the 20s
2nd Hit Both alleles of APC gene lost Adenoma develops
 5

- Canonical Wnt/APC/ß-Catenin Signaling Pathway

• WNT signaling (frizzled receptor) plays a major role in controlling cell fate, adhesion,
and cell polarity during embryonic development

Wnt Signal ON Wnt Signal OFF

Wnt binds frizzled receptor Activates Dsh
Dsh inhibits Gsk3 (and destruction complex)
Prevents phosphorylation of ß-catenin
NO degradation of ß-catenin (↑[ß-catenin])
ß-catenin (cytosol) translocates into nucleus and
binds transcription factor
Wnt does NOT bind frizzled receptor
APC (negative regulator of ß-catenin) holds it
Gsk3 phosphorylates ß-catenin
ß-catenin is ubiquitinated
Degradation of ß-catenin (↓[ß-catenin])
Failure to migrate into nucleus

ß-catenin unphosphorylated ß-catenin phosphorylated Ubiquitination

Transcription factor binding Cell Growth/Proliferation NO expression of growth factors

*Destruction Complex: Dsh, CKI, axin, GSK3, APC

Loss of APC is carcinogenic because APC is part of the destruction complex that stabilizes ß-catenin.
Loss of APC results in destabilization of ß-catenin Allows ß-catenin to thrive separated from the complex.
No degradation of ß-catenin Unregulated cell proliferation via Wnt growth proteins Colon cancer

- Lynch Syndrome (Hereditary Nonpolyposis Colon Cancer)

• Autosomal dominant
• Inheritance of one defective DNA mismatch repair (MMR) allele
Two-hit necessary (bi-allelic loss) Loss of heterozygosity
Microsatellite instability Repeated sequences predispose for replication errors
• High risk of colorectal cancer WITHOUT previous polyps

- Li-Fraumeni Syndrome
• Autosomal dominant
• Inheritance of one defective allele of P53 on chromosome 17p
Two-hit necessary (bi-allelic loss) Loss of heterozygosity
• 25-fold greater risk of developing malignant tumor by age 50
Sarcomas, breast cancer, brain tumors, leukemias, adrenocortical carcinoma

- BRCA1/BRCA2 Tumor Suppressor Genes

• DNA repair protein
Loss of BRCA genes result in cells that develop chromosomal breaks and aneuploidy
Weak correlation w/ sporadic cancers, only associated w/ familial cancers
• Mutations associated with breast, ovarian, and pancreatic cancers
BRCA1/BRCA2 mutations = 25% of familial breast cancers
BRCA1 = ↑ Risk of ovarian cancer (men have slightly ↑ risk of prostate CA)
- Defects in DNA Repair by Homologous Recombination
• Autosomal recessive cancer syndromes
• Chromosomes are susceptible to damage by ionizing radiation
1. Ataxia-Telangiectasia (ATM mutation Neural symptoms)
2. Fanconi Anemia (DNA -cross link repair mutation Bone marrow aplasia)
3. Bloom Syndrome (Helicase mutation Developmental defects)
 5

- Bcl-2
• Anti-apoptotic protein involved in the intrinsic (mitochondrial) pathway
• Function: Keeps mitochondria impermeable Prevents release of cytochrome c
• Bcl-2 Overexpression:
Follicular B cell lymphoma
85% have translocation of chromosomes 14 and 18 t[14;18]

- BAX and BAK

• Pro-apoptotic proteins
• Function: Form pores in the mitochondrial membrane Release of cytochrome C
• Results in activation of caspases Apoptosis

Virus Mechanism Associated Cancer

HCV (RNA) Produces postnecrotic cirrhosis
HTLV-1 (RNA) Activates TAX gene, stimulates polyclonal T-cell
proliferation, inhibits p53 suppressor gene
EBV (DNA) Promotes polyclonal B-cell proliferation, which
increases risk for t(8;14) translocation and
formation of follicular B cell lymphoma
HBV (DNA) Activates proto-oncogenes,
inactivates p53 suppressor gene
HHV-8 (DNA) Acts via cytokines released from HIV and HSV
HPV 16 and HPV 18 Type 16 (~50% of cancers): E6 gene product
(DNA) inhibits p53 suppressor gene
Type 18 (~10% of cancers): E7 gene product
inhibits RB1 suppressor gene
Hepatocellular carcinoma
T-cell leukemia and lymphoma
Burkitt lymphoma, CNS lymphoma in
AIDS, mixed cellularity Hodgkin
lymphoma, nasopharyngeal carcinoma
Hepatocellular carcinoma
Kaposi sarcoma in AIDS
Squamous cell carcinoma of vulva,
vagina, cervix, anus (associated with
anal sex), larynx, oropharynx

- Carcinogenic Agents
• Direct Acting:
Contain unstable electron deficient atoms that react w/ electron-rich atoms in DNA
Ex: Alkylating agents, acylating agents, nickel
• Indirect Acting:
Require metabolic conversion to carcinogen
Ex: Polycyclic hydrocarbons (tobacco, smoked meats or meats cooked over flame)
are metabolized by CYP450 in the liver Converted to DNA-binding epoxides
 5

Carcinogen Exposure/Sources Associated Cancer

Aflatoxin Ingestion of maize and peanuts grown in hot/humid Hepatocellular carcinoma
(Aspergillus) climates
Alkylating agents Oncology chemotherapy Malignant lymphoma
Arsenic Herbicides (common in vineyard workers), Squamous cell carcinoma of skin
fungicides, animal dips; metal smelting; Lung cancer
intentional/accidental poisoning Liver angiosarcoma
Azo dyes Used in paints, printing inks, varnishes, leather Hepatocellular carcinoma
products, carpets, food products
Asbestos Roofing material (roofers with over 20 years of Bronchogenic carcinoma
experience have had contact with this); insulation Pleural mesothelioma
for pipes in ships in shipyards, old homes; old cars
with brake liners
Benzene Component of light oil; used in printing industry, Acute leukemia
dry cleaning, paint, adhesives and coatings Hodgkin lymphoma
Beryllium Used in the space industry (missile fuel and space Bronchogenic carcinoma
vehicles; metal alloys in aerospace appliances and
nuclear reactors)
Cadmium Industrial industries where ore is being smelted; Prostate cancer
electroplating; welders who have welded on Lung cancer
cadmium-containing alloys or worked with
silver
solders; found in some batteries
Cyclophosphamide Chemotherapy agent Transitional cell carcinoma of bladder
Diethylstilbestrol Once used to treat women with threatened Daughters exposed to mothers who
(DES) abortions took DES may develop clear cell
carcinoma of vagina/cervix
β-Naphthylamine Workers in the rubber, chemical, leather, textile, Transitional cell carcinoma of bladder
(aniline dyes) and metal, and printing industries
aromatic amines
Nickel Nickel plating, by-product of stainless-steel Bronchogenic carcinoma
welding, ceramics, batteries, spark plugs Nasal cavity cancer
Oral contraceptives Birth control pill Breast and cervical cancer
Hepatic adenoma (may rupture)
Polycyclic Most common carcinogen in US (tobacco smoking) Squamous cell carcinoma: Skin
hydrocarbons (scrotum cancer from soot in chimney
Formed when coal, soot (chimney sweeper), wood, cleaners), oral cavity, midesophagus,
gasoline, oil, tobacco, or other organic materials are larynx, lung
burned; also formed in food when fish or meats are Adenocarcinoma: Distal esophagus,
charbroiled on an open flame pancreas, kidney
Transitional cell carcinoma: Bladder,
renal pelvis
Polyvinyl chloride PVC pipes, adhesive plastics, refrigerant Liver angiosarcoma
Radon and decay By-product of decay of uranium, hazard in quarries Bronchogenic carcinoma
products and underground mines
Silica Chemical of silicon dioxide, rock quarries, Bronchogenic carcinoma
sandblasting
 6

Pathoma: Neoplasia

- Neoplasia
• Benign or malignant
• Unregulated, irreversible, monoclonal (neoplastic cells derive from single mother cell)
• Cancer is detectable (clinical signs/symptoms) when 30 or more cell divisions occur
Mass = 1 gram (109 cells)

- G6PD Isoforms
• G6PD is an X-linked isoform enzyme (ex. G6PD-A, G6PD-B, etc.)
X-chromosomes should display a normal 1:1 ratio of G6PD isoforms
The ratio will still be 1:1 in hyperplasia (polyclonal)
• During neoplasia, the ratio will NOT be 1:1 because unregulated proliferation derives
from a single cell
Ex: ALL tumor cells would express the SAME isoform of G6PD

- Hyperplasia
• Regulated, reversible, and polyclonal (multiple cells produce daughter cells)
Ex: Uterus smooth muscle growth during pregnancy

- B Cell Clonality
• Determined by light chain phenotype (kappa or lambda)
Some B cells have kappa light chain, some have lambda light chain (ex. 3:1 ratio)
The ratio will still be 3:1 in hyperplasia (polyclonal)
The ratio will NOT be the same during lymphoma (monoclonal)
6
 62

- Tumor Suppressor Genes

• Regulate cell growth and suppress (decrease) risk of tumor formation
• p53:
Regulates G1 S
Slows cell cycle in response to DNA damage (upregulated DNA repair enzymes)
Induces apoptosis if DNA repair is not possible (↑BAX, ↓Bcl2 Cytochrome c leakage)
Loss of p53 is seen in >50% of cancers
• Rb:
Regulates G1 S
Rb holds E2F transcription factor in place (CDK4 phosphorylation E2F translocation S-phase)

Benign Tumors Malignant Tumors

• Mobile; slow growing • Fixed; rapid growing
• Well circumscribed; encapsulated • Poorly circumscribed
• Well differentiated • Poorly differentiated (anaplastic)
• Low mitotic activity • High mitotic activity
6
 6

- Cell Cycle-Nonspecific (CCNS) Drug

• Acts on tumor stem cells when:
1) They are traversing the cell cycle
2) They are in the resting phase (G0)

- Cell Cycle-Specific (CCS) Drug

• Acts selectively on tumor stem cells when:
1) They are traversing the cell cycle
2) Does NOT act on them when they are in the G0 phase

- Log-Kill Hypothesis
• Describes action of CCS drugs Follows first-order kinetics
• Applies to hematologic malignancies
• Given dose kills a constant proportion of a tumor cell population
Does NOT kill a constant number of tumor cells
• Capacity of a chemotherapy drug to kill tumor cells is logarithmic
Ex: 3-log-kill dose of a drug reduces a cancer cell population of 10 12 cells to 109
The same dose would reduce a starting population of 106 cells to 103 cells
In both cases, the dose reduces the numbers of cells by 3 orders of magnitude (3 logs)

Growth Fraction = The proportion of cells in

a tumor population that are actively dividing
 6

- Resistance to Chemotherapy Drugs

1. Increased DNA Repair
Alkylating agents, cisplatin
2. Formation of Trapping Agents
Some tumor cells increase their production of thiol trapping agents (glutathione)
Bleomycin (alkylating agent), cisplatin, anthracyclines
3. Changes in Target Enzymes
Changes in the drug sensitivity of dihydrofolate reductase
Increased synthesis of dihydrofolate reductase
Mechanisms of resistance of tumor cells to methotrexate
4. Decreased Activation of Prodrugs
Resistance to the purine antimetabolites (mercaptopurine, thioguanine) and the
pyrimidine antimetabolites (cytarabine, fluorouracil)
Decrease in the activity of the tumor cell enzymes needed to convert these prodrugs
to their cytotoxic metabolites
5. Inactivation of Anticancer Drugs
Increased activity of enzymes capable of inactivating anticancer drugs
Purine and pyrimidine antimetabolites
6. Decreased Drug Accumulation
Increased expression of a normal gene (MDR1) for a cell surface glycoprotein (P-glycoprotein)
This transport molecule is involved in the accelerated efflux of many anticancer drugs

- Strategies in Cancer Chemotherapy

• Primary Induction
First-treatment modality, or as a sole-therapy because there are no alternative options
Hematologic cancers, advanced solid tumors
Curative in a small fraction (mainly palliative to extend life)
• Neoadjuvant Chemotherapy
Administration of chemotherapy before local surgery (makes local therapy more effective)
• Adjuvant Chemotherapy
Adjuvant to local treatment procedures (surgery/radiation)
Goal is to reduce the risk of local and systemic recurrence, and extend overall survival

- Goals of Combination (Chemotherapy/Chemotherapy) Therapy

1. Each drug should be active when used alone against the particular
cancer
2. The drugs should have different mechanisms of action
3. Cross-resistance between drugs should be minimal Rescue Therapy
4. The drugs should have different toxic effects
Methods to alleviate toxicity
- Rescue Therapy of chemotherapy drugs
• Methotrexate Toxic to GI tract and BM
Leucovorin (Folinic acid): Form of tetrahydrofolate accumulated more readily by normal
cells Rescue of normal cells because leucovorin bypasses DHFR step in folate
synthesis
• Mercaptoethanesulfonate (Mesna)
Traps acrolein released from cyclophosphamide Reduces risk of hemorrhagic cystitis
• Dexrazoxane
 6
Inhibits free radical formation Protection against cardiotoxicity of anthracyclines (doxorubicin)
 6

- Alkylating Agents
• CCNS Drugs
• MOA:
Form reactive molecular species Alkylate nuclear DNA Cell death
Major site of alkylation within DNA is the N7 position of guanine
Cross-link DNA bases (guanine) Abnormal base-pairing DNA strand breakage
• Tumor Resistance:
Increased DNA repair
Formation of trapping agents (thiols)
Decreased drug permeability
Alkylating Agents
Nitrogen Mustards Cyclophosphamide, Ifosfamide, Chlorambucil, Mechlorethamine
Nitrosoureas Carmustine, Lomustine
Alkyl Sulfonate Busulfan
Alkylating-Like Drugs Platinum Analogs (Cisplatin/Carboplatin/Oxaliplatin)
Dacarbazine, Procarbazine

- Cyclophosphamide (Alkylating Agent)

• Nitrogen mustard
• MOA: Prodrug; bioactivation at liver P450 Acrolein (toxic breakdown product)
• Toxicity:
GI distress, myelosuppression, alopecia Hemorrhagic cystitis (bleeding from
Interactions with drugs metabolized by P450 bladder) from cyclophosphamide can
Hemorrhagic cystitis (effect of acrolein) be prevented by (1) Mesna (thiol binds
SIADH (↑ ADH) toxic acrolein) and (2) Adequate
hydration
- Ifosfamide (Alkylating Agent)
• Nitrogen mustard
• Isomer of cyclophosphamide
• Toxicity:
Fanconi syndrome (polyuria, electrolyte loss, metabolic acidosis)
Encephalopathy

- Chlorambucil (Alkylating Agent)

Nitrogen mustard

- Mechlorethamine (Alkylating Agent)

• Nitrogen mustard
• Toxicity: Severe vesicant (blister-forming) actions, sterility

- Carmustine/Lomustine (Alkylating Agents)

• Nitrosoureas
• MOA: Prodrug; bioactivation at liver
• Use: Brain CA (highly lipid-soluble)
• Toxicity: Myelosuppression, CNS toxicity (encephalopathy, seizures)
 6

- Busulfan (Alkylating Agent)

• Alkyl sulfonate
• Use: Myeloablation to prepare for BM transplant (single high dose), CML
• Toxicity: Skin pigmentation, seizures (high doses), pulmonary fibrosis (ground glass opacities)

- Platinum Analogs (Alkylating-Like Agents)

• Cisplatin, Carboplatin, Oxaliplatin
• MOA:
Form reactive molecular species Alkylate nuclear DNA Cell death
Major site of alkylation within DNA is the N7 position of guanine
Cross-link DNA bases (guanine) Abnormal base-pairing DNA strand breakage
• Toxicity:
Peripheral neuropathy Rescue therapy for cisplatin nephrotoxicity:
Ototoxicity (hearing loss) (1) Forced hydration
Cisplatin: Nephrotoxicity (acute kidney injury) (2) Amifostine
(3) Mannitol
- Dacarbazine (Alkylating Agent)
• MOA: Methylates DNA (methyl carbonium) Inhibits DNA synthesis
• Use: Hodgkin’s Lymphoma
• Toxicity: Phototoxicity, flu-like syndrome

- Procarbazine (Alkylating Agent)

• Oral drug
• MOA: Methylates DNA (also forms H2O2) dsDNA breaks
• Uses: Brain CA (penetrates CNS well)
• Toxicity:
Disulfiram-like reaction Antimetabolites
MAO inhibitor (tyramine cheese effect) Structurally similar to endogenous metabolites
Leukemogenic (can create leukemia) CCS DrugsAct on S-phase of cycle
Cytotoxic and immunosuppressant properties
- Methotrexate Methotrexate, 6-MP/thioguanine, azathioprine, Hydroxyurea, 5-
• Antimetabolite
• MOA: Antagonist of folic acid production
Competitively inhibits dihydrofolate reductase
Indirectly inhibits function of thymidylate synthase by depleting N5, N10-THF (cofactor)
Inhibits de nova purine synthesis
Polyglutamate derivative production Cytotoxic
• Resistance:
Decreased drug accumulation
Changes in the drug sensitivity or activity of dihydrofolate reductase
Decreased formation of polyglutamates
• Uses:
Many malignancies, ectopic pregnancy, RA, psoriasis, IBD, vasculitis
Wean patients off steroids in chronic disease
• Toxicity:
Myelosuppression; reversible with leucovorin (folinic acid) rescue
Hepatotoxicity, megaloblastic anemia, mucositis (mouth ulcers), pulmonary fibrosis
Teratogenic (folate deficiency NTD)
 6

- Mercaptopurine (6-MP), Thioguanine

• Antimetabolite; purine (thiol) analogs (mimics hypoxanthine/guanine)
• MOA:
Added to PRPP by hypoxanthine-guanine phosphoribosyltransferase (HGPRTase)
Converted into toxic nucleotides Inhibit enzymes for purine synthesis
Inhibits triphosphate incorporation into both RNA/DNA
• Uses:
Prevent organ rejection (solid organ transplant)
Wean patients off steroids in chronic disease
• Toxicity:
Myelosuppression, hepatotoxicity (↑ LFTs)
6-MP is metabolized by xanthine oxidase (inhibited by allopurinol/febuxostat)
Xanthine oxidase inhibitors boost effect (use caution as it increases toxicity)

- Azathioprine
• Prodrug; converts into 6-MP
• MOA: Same as 6-MP
• Use: IBD (UC/Crohn’s), RA
• Toxicity: Same as 6-MP

- Hydroxyurea
• Antimetabolite drug; inhibitor of BOTH purine and pyrimidine synthesis
• MOA: Inhibits ribonucleotide reductase (↓ DNA synthesis)
Increases HbF transcription
• Use:
Myeloproliferative disorders (CML, polycythemia vera)
Sickle cell anemia (↑ HbF)

- Fluorouracil (5-FU)
• Antimetabolite; pyrimidine analog (mimics uracil)
• MOA:
Activated (converted) to 5-FdUMP
Covalently complexes with thymidylate synthase and folate
Covalent complex inhibits thymidylate synthase “Thymineless death of
cells” Incorporation of FdUMP metabolite into DNA Inhibits DNA synthesis
Incorporation of FUTP metabolite into RNA Inhibits RNA processing
Leucovorin (folinic acid) enhance effects of 5-FU (extends half-life)
• Uses:
Colon, pancreatic, breast CA
Skin (topical): Actinic Keratosis, Basal Cell Skin Cancer
• Toxicity:
Myelosuppression, GI distress, alopecia, coronary vasospasm
Neurotoxic (rare); cerebellar ataxia, encephalopathy
Palmar-plantar erythrodysesthesia (hand-foot syndrome)

- Cladribine
• Antimetabolite; purine analog (mimics adenosine)
• Use: Hairy cell leukemia
 7

- Cytarabine
• Antimetabolite; cytosine arabinoside (pyrimidine analog)
• MOA:
Activated (converted) to AraCTP
AraCTP inhibits DNA pol Inhibits DNA chain elongation DNA chain termination
• Use: Leukemias/lymphomas
• Toxicity: Myelosuppression, megaloblastic anemia, neurotoxicity (cerebellar ataxia)

- Gemcitabine, Capecitabine
• Antimetabolite; pyrimidine analog
• MOA:
Gemcitabine diphosphate Inhibits ribonucleotide reductase Inhibits DNA synthesis
Gemcitabine triphosphate Incorporated into DNA Chain termination
• Use:
Gemcitabine: Colon CA
Capecitabine: Pancreatic CA
Toxicity: Myelosuppression, pulmonary toxicity

- Natural Product Anticancer Drugs

1. Vinca Alkaloids (Vinblastine, Vincristine, and Vinorelbine)
2. Podophyllotoxins (Etoposide, Teniposide)
3. Camptothecins (Topotecan, Irinotecan)
4. Taxanes (Paclitaxel, Docetaxel)

- Vinca Alkaloids
• Vinblastine, Vincristine, Vinorelbine
• MOA:
Bind to ß-tubulin Inhibit microtubule polymerization (contrast to taxanes)
Prevents formation of mitotic spindle
Act in the M-phase of cell cycle
• Toxicity: Neurotoxic (areflexia, peripheral neuropathy, paralytic ileus)

- Podophyllotoxins
• Etoposide, Teniposide
• MOA:
Inhibits topoisomerase 2 DNA breaks with no resealing
Act in the S/G2 phase of cell cycle
• Toxicity: Myelosuppression, GI distress, alopecia

- Camptothecins
• Topotecan, Irinotecan
• MOA:
Inhibits topoisomerase 1 DNA breaks with no resealing
Act in the S/G2 phase of cell cycle
• Uses:
Topotecan: Small Cell Lung Cancer, Ovarian Cancer (Advanced)
Irinotecan: Colon Cancer (Metastatic)
• Toxicity: Myelosuppression, extremely severe diarrhea
 7

- Taxanes
• Paclitaxel, Docetaxel
• MOA:
Inhibit disassembly of microtubules into tubulin monomers ENHANCE tubulin
polymerization Microtubules cannot breakdown Inhibition of mitosis
Act in the M-phase of cell cycle
• Toxicity:
Myelosuppression, hypersensitivity reactions (wheezing, urticaria, hypotension)
Nab-paclitaxel is an albumin-bound taxane that lowers risk of hypersensitivity reactions
Peripheral neuropathy (nerves are heavily dependent on microtubule function)

- Antitumor Antibiotics
• Naturally occurring microbial products derived from Streptomyces
1. Anthracyclines
2. Dactinomycin (Actinomycin D)
3. Bleomycin
4. Mitomycin

- Anthracyclines
• Doxorubicin, Daunorubicin, Idarubicin, Epirubicin, Mitoxantrone
Antitumor antibiotic; CCNS
• MOA:
Intercalate between DNA base pairs
Inhibit topoisomerase 2
Generate free radicals
Block synthesis of both DNA/RNA
• Toxicity: Cardiotoxicity: EKG abnormalities, arrhythmias, cardiomyopathy, systolic HF (↓ LVEF)
Dexrazoxane: Iron chelator; cardioprotective rescue for anthracyclines (doxorubicin)

- Dactinomycin (Actinomycin D)
• Antitumor antibiotic; CCNS
• MOA:
Intercalate between DNA base pairs
Inhibits RNA pol (both eukaryotic/prokaryotic) Blocks RNA transcription
Induce dsDNA breaks
• Use: Childhood cancers (neuroblastoma, Ewing’s sarcoma, osteosarcoma)
• Toxicity: Myelosuppression

- Bleomycin
• Antitumor antibiotic; CCS
• MOA:
Generates free radicals dsDNA and ssDNA breaks
Acts in the G2 phase of cell cycle
• Uses: Lymphomas, testicular CA, head/neck CA
• Toxicity: Bleomycin is inactivated by an enzyme densely located in lungs/skin
Pulmonary fibrosis (pneumonitis)
Skin Toxicity: Blister formation, hyperkeratosis, flagellate erythema (red streaks on skin)
 7

- Mitomycin
• Antitumor antibiotic CCNS drug
• MOA:
Metabolized by liver enzymes to form an alkylating agent that cross-links DNA
Targets hypoxic tumor cells

- Trastuzumab
• Monoclonal antibody against HER-2 (HER2/neu, ERB2, CD340)
• MOA: Binds surface protein that overexpress HER-2/neu for EGF
Triggers antibody-dependent cell-mediated cytotoxicity (ADCC)
• Use: HER2-positive Breast CA
• Toxicity: Cardiomyopathy (↓ LVEF)

- Cetuximab, Panitumumab
• MOA: Inhibits EGFR
• Uses: Metastatic colon cancer (wild-type KRAS)
EGFR is overexpressed in >80% of colon
CA’s
• Toxicity: Acne rash on face, diarrhea, ↑ LFTs

- Erlotinib
• EGFR TYR kinase inhibitor
• Use: NSCLC
• Toxicity: Acne rash on face, diarrhea

- Imatinib, Dasatinib, Nilotinib

• TYR kinase inhibitor
• MOA: Inhibits non-receptor tyrosine kinase activity of BCR-ABL fusion gene
• Use:
CML (Philadelphia translocation; BCR-ABL)
Kit-positive GI stromal tumor
• Toxicity: Fluid retention (peripheral or periorbital edema)

- Rituximab
• MOA: Monoclonal CD20 antibody
Binds to surface protein (CD20) Depletion of B cells
• B Cell Depletion Mechanisms:
1) Induces complement-mediated lysis
2) Antibody cytotoxicity
3) Induction of apoptosis of B lymphocytes
• Use: B cell malignancies (NHL, CLL), autoimmune diseases (RA, MS, immune thrombocytopenia)
• Toxicity: High risk of opportunistic infections
PML, PCP, cryptococcosis, CMV colitis, HBV reactivation

- Bevacizumab, Ziv-Aflibercept, Ranibizumab

• MOA: Angiogenesis inhibitor (inhibit VEGF binding to endothelial cells) NO O2 for tumor
• Ranibizumab: Used for retinopathy (↓ VEGF in the eye); age-related macular degeneration
• Toxicity: HTN, arterial thrombosis, impaired wound healing, bleeding
 7

- Bortezomib, Carfilzomib
• Proteasome inhibitors
• MOA: Inhibits 26S proteasome
26S proteasome is a large protein complex that degrades ubiquitinated CDKs
Induce arrest at G2-M phase and apoptosis
• Use: Multiple Myeloma, Mantle Cell Lymphoma
• Toxicity: Peripheral neuropathy, herpes zoster (VZV) reactivation

- Tamoxifen, Raloxifene, Toremifene

• Selective estrogen receptor modulator (SERM)
• MOA:
Competitive inhibitor of estrogen receptors (ER) in breast tissue
Blocks the binding of estrogen to receptors of estrogen-sensitive breast tumor cells
• Use: ER-Positive Breast CA
Raloxifene: Prevention of osteoporosis (ER agonist activity in bone)
• Toxicity: Risk of thrombosis (DVT, PE), hot flashes (menopausal type symptoms)
Tamoxifen: Risk of endometrial hyperplasia/cancer (ER agonist activity in uterus)
Raloxifene: ER antagonist activity in uterus NO risk of endometrial CA

- Flutamide, Bicalutamide, Apalutamide, Enzalutamide

• Nonsteroidal androgen receptor antagonist
• Use: Prostate Cancer
• Toxicity: Gynecomastia, hot flushes, hepatic dysfunction

- GnRH Receptor Agonists

• Leuprolide, Goserelin, Nafarelin
• Action: Decreases sex hormone production (physiologic antagonists)
Sustained NON-PULSATILE administration of GnRH analogs INHIBITS release of FSH/LH
Target: GnRH receptors (located on gonadotrophs in the anterior pituitary)
Inhibits FSH/LH release via downregulation of GnRH receptors
Induces gonadal suppression Hypogonadism
• Use: Prostate Cancer
• Toxicity: Bone pain, hematuria, gynecomastia, impotence, testicular atrophy

- Aromatase Inhibitors
• Anastrozole, Letrozole, Exemestane
• MOA: Inhibit aromatase (blocks testosterone to estrogen conversion); ↓ estrogen
• Use: ER-Positive Breast CA (only in post-menopausal women)
• Toxicity: Osteoporosis (↓ estrogen)
7
 Common hereditary cancer syndromes

Sy,nd rome Gene Pathoge" ,esl

ne oplasms,

Coloreclal eanoor
MSH2. MLH1.
l:ynch syndrome Endometrial cancer
MSH5, PMS2
• Ovarian cancer

,. Co1011ec tal
Famllal
canoer D molds &
ad nom tou AP<C
osteomas
polyposls

H ,mangiobla tomas
von Hlppel-Llndau • Ole ar ,oell renal
VHl.
syndr.om.e caroiooma • Autos.omal dom in ant
P ocnromoeytomai Caused by In· ct1,v· Ing mu!a ·on in oorrespo dlng t mor

• Sarcomas .uppr.. ssor g n .

• Breast cance • Deletion or rem aining normal allele ,(second hit) I ads lo loss or
Ll• Fraum I'll ·• Brain tumors t\elarozy·gosity & malignant tr n:sformatlon
TP53
synd.rome • Adre rioaortlcal
caminoma
leu emi•a

P r a:thyroid
adenomas
Mul tlple endocrine
MEN1 • Piluilary adenomas
neopl· i · typ: 1
Pancreatic
adenomas

Medul lary thyroid

ca. o r Auto om 'I ,domln nt
Multiple endocrine • Pheochromocytoma • A.c Uvatln g (gain-of-function) mulalio n {n prolo-oncogene
,RET
naopl asl · type 2. • Para:lh yroid • Con tililuou s stlmu fa ion of ee I diVision predisposes lo lumor
hyperpl Sia growth
(Mi=N2A )

* R,STOL,e.Hr'\. ;.-..t!l.vtesI/ WF t-0b,t'\ol Gp 1-b

,11.lo l"\ C.r" "'"'-a...\r,st-1:><.. -\-""-. \eJi- iv-..e, f'V\a,,rcl,.-Sc:,\Jl, r vWF
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