HCT=
(RBC/Bld Vol)x(RBC vol/RBC)=RBCxMCV
Blast Criteria
a. Big Cell
b. Increased nuclear/cytoplasm ratio
c. Immature Chromatin
d. Big nucleolus or two
e. A bunch of cell that look a like***
--Does not have to meet all the standards to be considered a blast
Myelophthisic/Leuko-erythroblastic
What hematologic malignancies look like
-Stuff that should be in the BM is in the peripheral blood
Immunophenotyping
-Differnent cell types express different antigens
-use mAbs to detect
-hint most markers for T-cells will be <10
-Malignancies can change surface expression so not as helpful as ppl thought it would be
-Access via flow cytometry (remember lyse RBC so will not detect poss RBC precursors)
B-cell and T-cell Antigens
B-cell: CD-20, IgG kappa/lambda, CD45(both)
T-cell: CD3, CD7, CD4 or CD8
CD45, CD33, CD34
Usually start with CD45: on almost all marrow cells and can help to ID a blast population (left ball of graft)
-Most blast tend to express CD34
-Maturing granulocytes tend to express CD33
-COMBO of CD33 & CD34 indicate AML if in large population (premature myeloid cell in peripheral)
Immunohistochemistry
-Works better for larger cells than flow cytometry
-use enzyme conjugated abs to desired cell surface protein and then use CHROMOGENIC substrate
Gilman's Hypothesis
Need to mutations for the AML to hurt the patients
Class I: increases proliferation
Class 2: decreases differentiation -together these will make the leukemia much more aggressive and dangerous
Summary of Malignancies Arising in the BM
AML
Rapidly proliferating clones: Blasts in marrow and often in blood stream
Can arise from 3 cell lineages:
(1) Megs
(2) Erythroid Lineage
(3) Myeloid lineage
Acute Lymphocytic Leukemias (ALL)
Arise from the lymphocyte lineage
-Rapidly proliferating clones: blasts in marrow and often blood stream
Typical Presentation of Acute Leukemias
-Clones can take over a variety of locations, usually blood stream and BM
-They can take over BM w/o raising the WBC count in peripheral
-Can sometimes present as cytopenias (decreased cellularity of the blood), would not expect a rapidly proliferating population to present as cytopenia
Blasts outside of Marrow (rare): Myeloid: Myeloid Sarcoma and Lymphoid: Lymphoblastic lymphoma
Acute Leukemia Diagnosis
-Use to simply be >20% blast in marrow, not very predictive of how the leukemia will progress
Other methods:Morphology (useless by itself), Immunophenotyping (blasts can change surface expression) and genotyping
-Use an many as possible, genotyping can predict AML even if blast count is low
Genotyping Acute Leukemias, Specific Mutations
t(15:17) PML-RARA (AML Subtype)
t(8;21) RUNX1-RUNXT1 (AML Subtype)
t(12;21) FLT3 mutation(+) (AML Subtype)
-These and others give increased prognostic value, predicts response to therapy, IDs molecular targets for therapy development
Gililand Hyposthesis AGAIN!!
Class I (proliferation/survival advantage):
-FLT3-ITD, FLT3-TKD, Kit, Ras, PTPN11 Jak2 (usually dectected via sequencing)
Class II (impaired differentiation/apoptosis):
-t(15;17) PML-RARA, t(8;21) Runx-Runx1T1, (inv(16) or t(16;16)))CBFB-Myh11, MLL fusions (usually detected via cytogenetics)
BOLDED= AML diagnosed with presence even w/o blast count
AML with AUER RODs
t(8;21) Runx-1-Run1T1 Mutation
Clinical Presentation: younger patients/kids
Auer rods: crystalization of granule contents (excess myeloperoxidase production)
Immunophenotype: CD34+, CD13+, CD33weak
Prognosis: good response to chemo
APL: PML-RARA
t(15;17) fusion of PML (trans factor) with RARA (retinoic acid receptor)
-Class II mutation: Inhibits granulocyte differentiation
-TREATMENT: All Trans Retinoic Acid (ATRA), induces cell differentiation of the blasts to granulocytes (CLINICAL REMISSION)
-CLINICAL PRESENTATION: DIC, SEVERE THROMBOCYTOPENIA***
-Morphology: AUER RODS in stacks (faggot cells)*** sometimes see Bi-nuclear appearance
Immunophenotype: Weak/absent: CD34, HLA-DR, CD13+, CD33+
Prognosis: good if diagnosis is made
AML: CBFB-MYH11
inv(16) or t(16;16)
-Class II mutant, clinical presentation: younger patients/kids
-Morphology: Myelomonocytic* (mixed granulocyte-monocyte feats), Increased Eosinophils in blood and marrow (not known why)
-Immunophenotype: CD34+, CD117+ (blasts); CD13+, CD33+ (granulocytes); CD14+, CD11b+ (monocytes)
Prognosis: GOOD; optimal with high dose CYTARABINE
AML w/ Normal Cytogenetics
40-50% of AML cases
-Any age group
Morphology: undifferentiated or variable granulocytic or monocytic/monoblastic
-Immunophenotype: blast markers (CD34, CD117)+, can show any lineage markers
-Prognosis: have to look at molecular genetics, pairing of mutation will determine
-For some BM transplant doesnt help (NPMN! or CEBPA+, FLT3-ITD-, 60% 4-yr survival)
-for other it will (FLT3-ITD+ or NPM1-, CEBPA-, FLT3-ITD-, ~29% 4-yr survival)
ALL (Acute Lymphoblastic Leukemias)
TdT: important for VDJ recombination and marker of B and T cells
-CD19,CD10 and uCD20 are markers of B cells, can also express myeloid markers (CD13, CD33)
-Can also express CD34 (GENERAL BLAST MARKER*****)
ALL: BCR-ABL1
-t(9;22), proliferation activator (class I), diff from the mutation found in CML which is a class II mutation and prevents maturation of the cell, and have similar cytogenetics
-Clinical Presentation: Older adults (25% of all cases)
Morphology: Big agranular blasts, hard to tell ir myeloid or lymphoid
Immunophenotype CD10+, CD19+, TdT+
Prognosis: Poor
ALL: MLL rearranged
t(v;11q23), or t(11;19) Class II mutation, also found in AML
Clinical Presentation: most common leukemia in kids
Morphology: big agranular blasts
Immunophenotype: CD10-, CD19+, TdT+
Prognosis: Poor
ALL, Mutations that made St. Jude Famous
-t(12;21); TEL-AML1 (ETV6-RUNX1), class II (inhibits differentition)
-Clinical Presentation: Kids, 25% of pediatric B-ALL
Morphology: big agranular blasts
Immunophenotype: TdT+, CD34+, CD20-
Prognosis: 90% cure rate
ALL: T-ALL
-Common motif for lymphoid malignancies: oncogene>>>>Ig or TCR promoter
-Clinical Presentation: Kids 25% of pediatric T-ALL, often wih thymic mass**
Morphology: Big agranular blasts
Immunophenotype: TdT+, CD3+, CD5+, can express myeloid or B-cell antigens as well
Prognosis: High Risk: more intense chemotherapy regime, pretty good survival rate
Myeloproliferative Disease
Definition: Chronically proliferating clones which differentiate to circulating blood cells
Chronic Myelogenous Leukemia (CML): shows blast profile
CMML (myelodysplastic/myelomonocytic): hybrid b/t myelocytes and granulocytes
-Can also see Chronic: eosinophilic, neutrophilic or mastocytosis leukemia. Need to differentiate from a Reactive condition
--Best indicator if prominence of mature to immature cells is out of order, "Myeloid Bulge" (more blasts than more mature cells)
Myeloproliferative Diseases of Erythroid and Megs Lineages
Polycythemia Vera, elevated RBC count (erythroid lineage)
-Essential thromboythaemia/Primary myelofibrosis: elevated platelet count (meg lineage)
-Can progress to AML
CML that is Undectable
Cryptic Translocation, Cannot see with Routine Cytogenetics
-Will progress to AML (can be myeloid or lymphoid)
PHASES: chronic, accelerated, acute
-Can also confuse with sepsis, get a BM
Mastocytosis
--Usually presents as benign cutaneous lesions in kids (itchy) that goes away by itself
-Release of histamine can cause systemic symptoms that confuse clinician
Morphology: Aggregates of blank looking cells, round or spindle shapes, s/t with eosinophils
Immunophenotype: Tryptase+, CD117+, CD25+
Treatment: Imatinib
Prognosis: highly variable
Polycythemia Vera
-Jak2 mutation in >95% of cases-detectable in PB leukocytes
-Clinical presentation: Thromboisis?hypertension/stroke or MI. increased RBC #
Morphology: hypercellular marrow, erythroid hyperplasia, increased Megs
-Immunophenotype: no known features
Prognosis: >10yr survival is common, can progress to myelofibrosis MDS, acute leukemia
Epo will usually be low, one way to differ between neoplastic and reactive
Essential Thrombocythemia
Jak2 mutation in ~50% of cases-detectable in PB
-Clinical Presentation: Thrombosis, increased platelets, can also be due to Fe deficiancy, infecion or chronic inflammation, can also have bleeding and splenomegaly
Morphology: Increased Megs (very abnormal and tend to cluster)
Immunopheno: not known, can not put Meg through flow cytometry (too fragile)
-Prognosis: >10yr survival is common, can progress to myelofibrosis MDS, acute leukema
Primary Myelofibrosis
Jak2 mutation ~50%, disputed diagnostic criteria
-Morphology: increased Megs, bizarre shapes, clusting and fibrosis (TEAR DROPS?)
No known immunopheno
Prognosis: usually shorter survival than ET, can progress to marrow failure, acute leukemia
-Cells of bone marrow will relocate to spleen and liver: Extra-medullary hematopoiesis
Treatment: supportive care, blood transfusions, splenectomy are dangerous (pts too old, will bleed) and HSCT is curative but high mortality rate b/c pts are old as hell
Myelodyisplasias
-Usually present as unexplained cytopenia, bicytopenia or pancytopenia
-see poorly functioning clones, mostly in elderly (>70)
5 major adult forms (best to worst prognosis): (1) Refractory cytopenia with unilineage dysplasia (2) refractory anemia with ring sideroblasts (3) myelodysplastic syndrome with isolated del(5q) (4) refractory cytopenia with multilineage dysplasia (5) refractory anemia with excess blasts
-Accumulation of several mutations over time in different mutations
Refractory Cytopenia with Unilineage Dysplasia
Clinical pres: unexplanined cytopenai
Morphology: weird looking precursors: binucleation or irregular nuclei, can show fibrosism high or low cellularity, megaloblastoid features
Prognosis: surival unclear, rarely progresses to AML
Refractory Anemia with Ring Sideroblasts
-Nonspecific cytogenetic abnormalities (trisomes, monosomes) may be present
-Morphology: ring sideroblasts usually with dyspoietic feats (in red cell series only)
--Iron granules form ring around nucleus
-Prognosis: rarely progresses to AML
MDS with Isolated del(5q)
-All the Megakaryocytes are MONOCUCLEAR
-Clinical pres: Anemia often severe, usually elderly
-Morph: megs are mononuclear
-Prognosis: good median survival; treatable with LENALIDOMIDE, 10% progress to AML
Refractory Cytopenia with multilineage dysplasia
-Two or more lineages show dysplastic changes
-Clinical pres: Anemia often severe: usually elderly
-Morph: Granulocytes dont granulate normally (if affected), nuclei dont lobulate normally
Prognosis: Median survival 30 months, 10% progress to AML in 2 years
Myelodysplastic Case Study
77y.o. man with newly diagnosed pulmonary adenocarcinoma
-anemia was not worked up completely, has had for past 2 years (b/f diagnosis): Fe studies, folate, B12 levels are all normal
-Gave chemo for initial diagnosis
-Really had Refractory Anemia with trilineage dysplasia
BM biopsy: Myeloid hyperplasia
Poor prognosis, gave chemo that made him sick and was not going to help
ALL: T-ALL
-Common motif for lymphoid malignancies: oncogene>>>>Ig or TCR promoter
-Clinical Presentation: Kids 25% of pediatric T-ALL, often wih thymic mass**
Morphology: Big agranular blasts
Immunophenotype: TdT+, CD3+, CD5+, can express myeloid or B-cell antigens as well
Prognosis: High Risk: more intense chemotherapy regime, pretty good survival rate
