Lecture 5: Disorders caused by Chromosome Rearrangements (focus on cancer)

Balanced vs. Unbalanced Rearrangements

True or false: BALANCED rearrangements are OFTEN Benign?

TRUE

• but it is NOT ALWAYS

4 ways balanced rearrangements cause disease

****Of the 4 Ways Balanced rearrangements cause disease Which mostly result in CONSTITUTIONAL vs. CANCER diseases?

****What is an Example of a disease caused by GENE disruption?

Hemophilia A

• X-linked condition causing severe bleeding (loss of clotting factors)

****What is the most common CAUSE of Hemophilia A + What is the mechanisms + outcome?

****What are CIS-REGULATORY ELEMENTS in genetic disease?

• contrast them to Trans-regulatory elements

Cis-regulatory elements are non-coding DNA sequences on the same molecule of DNA that regulate nearby genes

• trans-regulatory elements are diffusible molecules like proteins or RNA that can regulate target genes anywhere in the genome

******Balanced Rearrangements POSITION EFFECTS. What are the 3 Cis-regulatory elements in genetic disease?

1. Promoters: DNA sequence immediately upstream of a gene where RNA poly and transcription factors bind; required to initiate transcription

2. Enhancers: Noncoding DNA elements that boost gene transcription. Can act at long distances, even if upstream, downstream or intronic.

3. Locus Control Regions (LCR): “super enhancers” coordinates expression of an entire gene cluster

****If a rearrangement separates a gene from its enhancer/LCR what will happen?

The gene will become silent despite being intact

****Position effect in Genetic diseases: What is an example of a Separation from LCR

• What is the Mechanism

• What is the result?

B-thalassemia

B-globin genes (hemoglobin) are arranged in a cluster on chromosome 11

• expression is developmentally regulates (different protein in embryo, fetus and adult)

LCR upstream of cluster is essential for switching them ON in a RBC precursor

• If balanced translocation SEPERATES the LCR form the cluster the B-globin genes are intact but they CANNOT BE EXPRESSED

******Position effects in Cancer: ENHANCER HIJACKING

• What is an example

• the mechanism

• OUTCOME?

Proto-oncogene activation in B-cell Lymphomas

IgH heavy chain locus is on chr 14q32. IgH has VERY STRONG ENHANCERS + PROMOTERS that norm drive antibody gene expression in a B-cell

In B-cell malignancies, chromosomal translocations bring PROTO-ONCOGENES under Igh control

• This “enhancer high jacking” leads to ONCOGENE OVEREXPRESSION + lymphoma development

*****What is BURKITT’S LYMPHOMA associated with?

• Hint: Think Position effects

• What is the mechanism + result of this disease

Burkitt’s lymphoma = aggressive B-cell Lymphoma

Typically associated with t(8;14) translocation putting MYC under control of the IGH enhancer

• Proto-oncogenes brough into proximity with new cis-regulatory elements

MYC = Proto-oncogene

• Normally regulates cell growth + proliferation

• When overexpressed drives uncontrolled division

MECHANISM: IgH enhancer drives MYC overexpression → lymphoma

*****What are FUSION GENES? They are a KEY MECHANISM to what?

Rearrangement joins 2 genes together to form a new fusion transcript/protein

Key mechanism in CANCER

******What are 2 examples of FUSION GENES

1. BCR-ABL (t(9;22)) - Philadelphia chromosome → CML

2. PML-RARA (t(15;17)) - Acute promyelocytic leukemia

******ONCOGENIC FUSION PROTEINS: Philadelphia Chromosome (Ph)

• What is it a hallmark of

• What is the mechanism

• What is the outcome?

Hallmark of CML (chronic myeloid leukemia)

• First cancer marker chr. discovered

Reciprocal translocation t(9;22)(q34;q11)

• Fuses BCR gene (chr 22) with ABL1 gene (Chr 9)

Creates BCR-ABL fusion protein (CONSTITUTIVELY ACTIVE TYROSINE KINASE)

• Always on growth signal

CONTINUOS SIGNALLING → UNCONTROLLED PROLIFERATION

******ONCOGENIC FUSION PROTEINS: PML-RARA

• What disease/cancer does it cause

• What is the mechanism

• What is the outcome

Acute promyelocytic Leukemia (APL)

t(15;17) resulting in Fusion of PML (tumor suppressor) + RARA

• NORMAL RARA: responds to retinoic acid (Vit A) → Switches ON GENES that drive Myeloid differentialtion

Fusion protein (PML-RARA) stays in OFF MODE → PML affinity for repressors acts as a Super repressor → causes DIFFERENTIATION BLOCK

RESULT: IMMATURE PROMYELOCYTES ACCUMULATE

***What is the HALLMARK of APL (Acute promyelotic leukemia)?

Accumulation of immature promyelocytes

****What is the importance of discovering these cancer-causing translocations?

potential to target cells with these genetic mutations. TARGETED THERAPY

• Find a therapy that seeks and destroys a specific translocation or cells that have learned to depend on a translocation for survival and/or growth, and you can kill cancer without killing surrounding, healthy cells

***CML before 2000

• survival

• Disease progression

• Treatment options available?

*****True or false: The FIRST successful TARGETED cancer treatment was to treat CML?

TRUE

*******What is the mechanism behind how the First TARGETED Cancer therapy to treat CML WORKS?

• Impact>

• Proof- of-concept

Small-molecule tyrosine Kinase inhibitor (TKI)

• Binds to the ATP-binding pocket of BCR-ABL = prevents phosphorylation of downstream targets (halts the abnormal “always on” growth signal)

Impact: Blood counts normalize within weeks. Lon term survival dramatically improved CML = now a manageable chronic condition

Proof-of-concept: drugging the molecular driver can transform cancer care

Nxt gen TKIs are being developed to overcome resistance

**APL before targeted therapy

Used to be the MOST LEATHAL LEUKEMIA

• characterized by promyelocyte accumulation and severe Coagulopathy (DIC). Patients often died within weeks of diagnosis from bleeding complications

Standard chemotherapy was Ineffective - cure rates very low

Discovery: t(15;17) → PML-RARA fusion blocks differentiation of myeloid precursors

******What is the modern targeted therapy to cure APL?

• what are the components

• How does it work

• Impact?

ATRA + Arsenic - Differentiation therapy

• All-trans retinoic acid (ATRA): binds the RARA portion of the fusion = releases repressor complexes → allows promyelocytes to differentiate into neutrophils

• Arsenic Trioxide (ATO): directly degrade the PML-RARA fusion protein + restores normal nuclear body function

Impact: cure rates >90. APL no longer the most lethal but one of the most curable cancers in adults

• Cancer therapy can sometimes mean restoring normal maturation not just killing cells

*****What are Chromosomal Instability syndromes?

Rare INHERITED DISORDERS caused by defects in DNA repair or Cell cycle checkpoints, leading to an increased rate of chr. breakage and rearrangement

****What are CHROMOSOMAL INSTABILITY syndromes Clinically characterized by?

Growth + developmental problems

Immunodeficiency

strong predisposition to cancer

*****What do cells from Individuals affected by Chromosomal instability syndrome often show HYPERSENSITIVITY TO?

DNA damaging agents → thus why they are predisposed to Cancer

• this may be used in diagnostic workup

*****What are 4 of the more common chromosomal instability syndromes?

****Info dump on BLOOM SYNDROME

• Cause?

• Mechanism

• CLINICAL FEATURES?

• HALLMARK?

Autosomal recessive condition cause by Defects in the BLM gene (RecQ helicase family)

• defects in DNA helicase leads to impaired homologous recombination repair(error prone) resulting in increased sister chromatid exchange + genomic instability

CLINICAL FEATURES: growth deficiency, recognizable facial features, photosensitivity (Malar rash of face), Immunodeficiency, predisposition to cancer esp. leukemias, lymphomas + GI (areas of high cell renewal)

Hallmark = INCREASED SISTER CHROMATIC EXCHANGE

• chromosomes are stained so that sister chromatid genetic material can be distinguished

• sister chromatid exchange is rare in a lymphocyte from a normal individual but occurs at a high frequency in a lymphocyte from a patient with bloom syndrome

****Info dump on FANCONI ANEMIA SYNDROME

• Cause?

• Mechanism

• CLINICAL FEATURES?

• HALLMARK?

Caused by defects in the FANC pathway - DNA damage response network (mostly autosomal recessive, rare x-linked)

Result = defect in DNA interstream crosslink repair ==> chromosomal breakage

CLINICAL FEATURES: progressive bone marrow failure (Pancytopenia - decrease in levels of all cell types), Congenital anomalies, hyperpigmentation, increased cancer risk esp. AML, head/neck SCC

Chromosome breakage assay: increase in chromosome aberrations induced by DNA cross-linking agents (DEB or MMC) = increase in spontaneous chromosome breaks

HALLMARK = increase in spontaneous chromosome nonsymmetric TRIRADIAL + QUADRIRADIALS

****What is CHROMOTHRIPSIS?

• cause + result?

Chromosome shattering

• catastrophic event where a chromosome breaks into many pieces in a single cellular crisis + gets stitched back together in a haphazard order

Result: Massive complex rearrangement, loss and gain of DNA segments (deletions, duplications, inversions).

• Tens to hundreds of rearrangements all at once (instead of gradual accumulation)

*****CHROMOTHRIPSIS MECHANISM?

• Hint: 3 potential models

Not fully understood

1. micronuclei hypothesis: A chromosome or fragment gets trapped outside the nucleus during mitosis → undergoes faulty replication + shatters → reintegrated with errors

2. Telomere crisis/ dicentric chromosomes: breakage-fusion-bridge cycles lead to pulverization

3. Ionizing radiation: Could also cause shattering

****CHROMOTHRIPSIS SIGNIFICANCE (aka. what can chromothripsis result in/ what disease is it associated with)

• in both CANCER + CONSTITUTIONAL/GERMLINE disease

CANCER:

• Associated with poor prognosis + aggressive biology

• Can create oncogene amplifications or disrupt tumor suppressors in a single step

• Extrachromosomal DNA (ecDNA) Formation: circular DNA fragments carrying oncogenes. They can amplify to very high copy numbers, segregate unevenly and act as mobile enhancers driving tumor heterogeneity, aggressiveness and therapy resistance

CONSTITUTIONAL/GERMLINE

• RARE but can occur. Patients often present with a unique complex rearrangement not fitting of typical CNV syndromes

***Prevalence of Chromothripsis across cancer types

SARCOMA = most common

Blastoma

then CARCINOMA

****Group the Structural rearrangements talked about today into one of the 2 following categories