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Cell and Molecular Biology

Behrouz Mahmoudi

Molecular Biology of Cancer

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Cancer cells – The Basics

Neoplasia is an abnormal accumulation of cells that occurs because of an imbalance between cellular proliferation and cellular attrition. Cells proliferate as they pass through the cell cycle and undergo mitosis. Attrition, due to programmed cell death, removes cells from a tissue.

•Mutations leading to oncogenesis can be inherited or familial (1%) (single-gene or multifactorial disorders) or sporadic (99%).•Mutations can occur in coding DNA but also in non-coding DNA such as in microRNAs. •Cancer cells have lost their contact inhibition properties.

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Once it is initiated, a cancer progresses by accumulating additional genetic damage through mutations in caretaker genes encoding the cellular machinery that repairs damaged DNA and maintains cytogenetic normality

The original clone of neoplastic cells serves as a reservoir of genetically unstable cells, referred to as cancer stem cells.

Stages in the evolution of cancer

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1. Immortal (grow and divide indefinitely).2. Clonal origin from one ancestral cell that

becomes more mutated, deregulated over time, causing genetic instability such as chromosomal rearrangement and phenotypic mutations.

3. No longer subject to contact inhibition.4. Evade apoptosis (programmed cell death)

and senescence.5. Reduced requirement for growth factors.6. Insensitive to anti-growth signals.7. Ability to metastasize (invade other tissues).8. Sustained angiogenesis (formation blood

vessels).

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Cancer Cells -Types of Genes

The genes implicated in cancer development are classified into three types: • Oncogenes stimulate cell division

excessively. They are mutated from normal genes called proto-oncogenes that encode components of the cell's normal growth, such as growth factors.

• Tumor Suppressor Genes inhibit cell division and/or cause apoptosis. Mutations in tumor suppressor genes would promote cell division or allow genetically damaged cell to grow out of control.

• DNA Repair Genes correct mutation of a gene.

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Cancer Cells - Oncogenes and Proto-oncogenes

The mutation can be:• an activating gain-of-function mutation

in the coding sequence of the oncogene.• a mutation in the regulatory elements of

the oncogene.• an increase in the genomic copy number

of the oncogene.

Oncogenes can lead to hereditary cancer syndromes or sporadic cancers.

α-tubulin (violet), γ- tubulin (yellow) and DAPI (blue). Casimiro et al. show that in contrast to Cyclin D1-/- cells (left), those overexpressing Cyclin D (right) develop multiple centrosomes and abnormal spindle architecture, resulting in chromosomal instability.

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Cancer Cells – Tumor-suppressor GenesMutation or deletion of tumor suppressor genes is believed to initiate many forms of cancer. For tumors to develop, both alleles of the tumor suppressor gene must be inactivated (VERY DIFFERENT FROM ONCOGENES). In familial cancer syndromes, a mutant allele of a tumor suppressor gene is inherited and is present in every cell. However, tumorigenesis is not initiated until the second allele is inactivated in a somatic cell. In non-familial cases, inactivation of both alleles occurs via somatic mutation or deletion. The end result is the same in both cases, the lack of a functional tumor suppressor gene leads to tumor development.

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2-hit hypothesis of tumor suppression, in which the sequential inactivation of 2 gene alleles results in the development of retinoblastoma.

The first hit is usually a mutation in the DNA sequence of the gene (a small deletion or base substitution; star). This mutation can be transmitted through the germline, giving rise to an inherited form of cancer. The second hit ( loss of heterozygosity [LOH]) is often a gross chromosomal mechanism that occurs at higher rates in somatic cells and that leads to hemizygosity or homozygosity of the chromosome region containing the mutation. This includes nondisjunctional loss with reduplication of the chromosome carrying the mutated TSG, subchromosomal deletion, unbalanced translocation, and mitotic recombination.

Oliveira AM, Ross JS, Fletcher JA. p53 and tumor suppressorgenes in breast cancer. In: Ross JS, Hortobagyi GH, eds. TheMolecular Oncology of Breast Cancer. Sudbury, MA: Jones andBartlett; 2005:358-372

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Hereditary Cancer Syndromes

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Retinoblastoma is a rare relatively tumor (1:20,000 incidence) that originates in the retina.•Sporadic (unilateral and 60% of cases)•Familial and autosomal dominant inheritance (most bilateral some unilateral and some asymptomatic and 40% of cases)•Induced by mutation in the tumor suppressor gene Rb (ch 13) encoding the Retinoblastoma protein (RB).

G1/S transition regulation

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In sporadic retinoblastoma, two somatic mutations occur in the same retinal cell, thus triggering tumorigenesis.

normal retina cell

precursor retina cell

Tumor retina cell

1st somatic hit (mutation)

2nd somatic hit (mutation)

The first mutation is recessive at the cellular level, although the clinical

condition follows an autosomal dominant mode of inheritance

A white pupillary reflex is the presenting manifestation of retinoblastoma in about

90% of patients in the United States.

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The presence of bilateral tumors indicates that the affected patient is a carrier of

familial retinoblastoma who can transmit the tumor to progeny.

precursor retina cell

Tumor retina cell

somatic hit (mutation)

In familial retinoblastoma, the 1st Rb mutation is inherited through the germline and is therefore present in all retinal cells.In this case, only one somatic hit is necessary to inactivate the tumor suppressor gene.

Here too, the Rb gene is recessive at the cellular level, although the

clinical condition follows an autosomal dominant mode of

inheritance.

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Heterozygosity should normally protect from mutant allele; however, frequently, there is loss of the normal Rb allele, uncovering the mutant allele.

Genetic mechanisms for the loss of heterozygosity (LOH) of wildtype RB1 uncovers the recessive allele.

Extra-ocular retinoblastoma

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Is RB sporadic, familial? What is the mode of

inheritance? Penetrance?

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Breast and ovarian cancers are the most common form of tumors in women (1:700 incidence in the US) that originate in breast and ovarian tissues.•Sporadic (90 to 95 of % of cases)•Familial and autosomal dominant inheritance (5-10% of cases)•Most hereditary cancers involves the tumor suppressor genes BRCA1 (Ch7) and BRCA2 (Ch13) encoding DNA repair proteins.

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A pedigree of Li-Fraumeni syndrome showing autosomal dominant inheritance

p53 therapy in a patient with Li-Fraumeni syndromeFusion PET/CT scan of the treated pelvic tumor (arrow) before (left) and after four intratumoral injections of Advexin depicting complete resolution of 2,3-FDG uptake at 2 mo (right) and progression of the untreated lesion (cross-hairs)

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Genetic of the Acute LeukemiasAcute leukemias often involve translocations: Cells of hematopoietic system rapidly divide: bone

marrow stem cells leading to red blood cells and white blood cells.

Ex. promoter fusions of an Immunoglobulin promoter to a proto-oncogene will overexpress the normal protein in B lymphocytes - promote excessive cell division.

Ex. gene fusions: CML = chronic myeloid leukemia: fusion of Bcr and abl genes by chromosome 9-22 translocation overactive tyrosine kinase.

Ex. PMLRAR = acute promyelocytic leukemia = fusion of PML and RAR (retinoic acid receptor); fusion protein defective.

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Translocations that aberrantly activate transcription factors in acute leukemias.

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Chronic Myelogenous Leukemia (CML) is an example of translocation leading to cancer.

The Philadelphia chromosome translocation, t(9;22)(q34;q11). The Philadelphia chromosome (Ph1) is the derivative chromosome 22, which has exchanged part of its long arm for a segment of material from chromosome 9q that contains the ABL oncogene. Formation of the chimeric BCR-ABL gene on the Ph1 chromosome is the critical genetic event in the development of chronic myelogenous leukemia.