L-13 Neoplasma II

• COM 5081 – Fundamentals of Pathology - Neoplasia II by R. Daniel Bonfil, Ph.D., Professor of Pathology.

Molecular Basis of Cancer – A Disease of Genes

• Biochemical and genetic markers suggest most cancers originate from a single cell with a mutation, having a monoclonal origin.
• The original progenitor cell doesn't acquire all cancer cell characteristics, becoming a "transformed" cell.
• Cancer develops through a multistep process, where cells become malignant via a progressive series of mutations affecting different genes in the founding and daughter cells which results in non-identical clones.
• Most cancers develop later in life, indicating the multistep nature of cancer development.
• Driver mutations aid cancer acquisition, whereas passenger mutations, which are more common, do not directly contribute.
• Tumors have a monoclonal origin but are genetically heterogeneous.

Molecular Basis of Cancer: Genes Involved

• Main targets of cancer-causing mutations (driver mutations) include:
• Growth-promoting proto-oncogenes
• Growth-inhibiting tumor suppressor genes
• Genes regulating programmed cell death (apoptosis)
• Genes involved in DNA repair

Oncogenes

• Proto-oncogenes (c-onc) encode cellular proteins regulating normal cell growth and are present in all nucleated body cells.
• Oncogenes are mutated proto-oncogenes which transform normal cells into cancerous cells and are counterparts of normal genes with "gain-of-function" mutations.
• Viral oncogenes (v-oncs) are in some oncogenic viruses (retroviruses) and contain exons of their homologous c-oncs only.

Proteins Encoded by Proto-oncogenes

• Growth Factors Include: sis, hst
• Transmembrane growth factor receptors (tyrosine kinase) Include: erbB, kit, fms, ret, ros, src, met, neu, trk
• Cytoplasmic kinases Include: pim-1, raf, mos, crk, vav
• Nuclear transcription factors (cell cycle regulators/apoptosis inhibitors) Include: myc, rel, fos, ets, jun, ski, myb
• ras guanosine triphosphatase Include: (GTPase), ras
• Membrane-associated kinases: Include: ick, fgr, fes, yes, fyn, abl, fps

Oncogenes in Different Types of Cancers

• c-Sis (codes for PDGF) is found in Astrocytoma
• HER2/neu (ERBB2) is a subset of breast carcinomas
• ERBB1 (codes for EGFR) is found in ~ 80% squamous lung ca and 50% glioblastomas
• Kit is found in GISTs and melanomas
• Abl is found in 95% of chronic myelogenous leukemia (CML) patients
• K-ras is found in 90% of pancreatic adenocarcinomas and 30% of all tumors
• N-mycs leads to Neuroblastomas
• L-myc leads to Small cell lung carcinomas
• с-тус found in ~90% Burkitt lymphomas t (8;14)

Proto-oncogenes Activation

• "Activation" in a single proto-oncogene allele can cause changes in cell and tumor growth, making oncogenes dominant.
• Activation can occur through:
• Translocation or transposition where a gene moves to a new locus under new controls.
• Gene amplification with multiple gene copies.
• Point mutations within a control element.

Oncogene Activation by Chromosomal Translocation

• Occurs in 90% of Burkitt Lymphomas
• Occurs in 95% of CMLs
• Part of Chromosome 8 with c-myc translocates to Chromosome 14 (t 8;14)
• Part of Chromosome 9 with c-abl translocates to Chromosome 22 (t 9;22)
• Philadelphia chromosome (Ph) result in an increased MYC protein and Tyrosine kinase

Oncogene Activation by Gene Amplification

• The N-myc gene is amplified in many neuroblastomas.
• Genes are amplified as extrachromosomal double minutes or as a chromosomally integrated homogeneous-staining region (HSR).

Single Point Mutations

• Mutations occur in the c-ras gene
• The proto-oncogene encodes for p21Ras protein, with GTPase activity, normally activated by external signals.
• A point mutation in a c-ras allele results in no GTPase activity which causes permanent activation of signaling pathways and stimulates proliferation.
• The most frequent oncogenic mutation is in human cancers, such as mutant K-ras.
• Mutant K-ras is present in 90% of pancreatic cancer, 50% of thyroid and bladder cancer, and 30% of lung cancer.
• Three c-ras genes exist; K-ras, H-ras, and N-ras.

Normal Ras Protein Signaling

• Normal Ras protein causes GTPase activity
• Inactive RAS protein is activated through external Growth Factor
• Inactivation of RAS protein by hydrolysis of GTP
• Active RAS enables cell growth Signaling by Mutant Ras Protein
• Mutant Ras protein has Inactivation of GTPase activity
• Active Ras protein is uncontrolled resulting in Increased Cell division leading to Cancer

Tumor Suppressor Genes (TSGs)

• Normal genes act as negative cell growth regulators and suppress tumor formation.
• Both alleles of a TSG must be inactivated through "loss-of-function" mutations to disrupt normal cell growth regulation and promote tumor formation; mutated TSGs act as recessive genes.
• "Loss of heterozygosity" (LOH) in a TSG only predisposes for tumor formation, it is a common genetic event in cancer where one allele is lost.

Pathogenesis of Retinoblastoma

• In Chromosomes 13 normal RB are tumor suppressor genes
• Sporadic Form is unilateral.
• Familial Form is bilateral

Many Cancers Have a Disabled G₁ Checkpoint

• Mutations in Rb or other Genes that Affect its Function and cause a disabled Checkpoint (M = mitosis, S= DNA)
• Normal Cell Cycle
• The start of the cell cycle begins with Growth Inhibitors stimulating the cell division
• RB protein becomes Hypophosphorylated
• Forms a complex with the transcriptional activator, E2F, and cannot active transcription of genes
• Results in cells arrested at the G₁ stage
• DNA replication is initiated → Mitosis
• Abnormal/Cancer Cell Cycle
• Inactivating mutations in Rb or CDK inhibitors' genes
• Activating mutations in cyclin D, CDK4,6 genes
• Oncogenic viruses (e.g., HPV) encode proteins (e.g., HPV E7) that bind to and degrade RB
• There is now an Insensitivity to Growth Inhibitors and the Loss/inactivation of RB protein
• E2F freely activates genes that promote cell division with Cell cycle progression in absence of GFs

p53: The Guardian of the Genome

• Wild-type p53 expression increases in response to DNA damage.
• P53 (aka TP53) inhibits cell division through quiescence or senescence, or induces apoptosis.
• Over 50% of cancers have somatic mutations affecting both p53 alleles.
• Patients with Li Fraumeni syndrome (rare) inherit one mutant p53 allele in the germline and present a 25-fold increased risk for many cancer types.

BRCA1 and BRCA2 and Familial Cancers

• BRCA1 (Ch17) and BRCA2 (Ch13) tumor suppressor genes participate in genomic maintenance, including DNA DSB repair by homologous recombination.
• Germline mutations in these genes cause genomic instability in breast and ovary cells.
• Females with a germline mutation in BRCA1 or BRCA2 have:
• A 50-70% risk of developing breast cancer (BrCa) by age 70.
• A 40-50% and 10-20% risk for ovarian cancer, respectively.
• Higher occurrence of germline BRCA1/2 gene mutations in Jewish females of Eastern European (Ashkenazi) descent increases risk for BrCa.
• Males with a mutant BRCA2 germline allele have a >7-fold increased risk of developing prostate cancer by age 65.

Autosomal Dominant Syndromes

• Familial retinoblastoma has the gene RB and main cancers
• Li-Fraumeni syndrome has the gene TP53 and main cancers of, Sarcomas, breast cancer
• The Familial adenomatous polyposis (FAP) has the gene APC and it's main cancel is Colorectal cancer
• Neurofibromatosis 1 and 2 has the gene NF1, NF2 and main cancers of Neurofibromas, acoustic neuromas, meningiomas
• Familial breast cancer has the gene BRCA1, BRCA2 (DNA repair genes) and the main cancers Breast, ovarian, prostate cancer
• Von Hippel-Lindau has the VHL gene with the main cancer Renal cancer
• Wilm's tumor has the WT1 gene and the main cancer Pediatric renal cancer

Autosomal Recessive Syndromes of Defective DNA Repair

• Xeroderma pigmentosum has the gene Nucleotide excision repair and main cancer Skin cancers in early childhood
• Bloom's syndrome has the BLM gene and it is a greatly increased risk of cancer
• Fanconi anemia has the gene DNA cross-links repair and its main cancer is Acute myeloid leukemia (AML)

Apoptosis-regulating genes

• Code for apoptosis-controlling proteins (programmed cell death).
• Overexpression of BCL-2 genes, which encode anti-apoptotic proteins, causes cells to ignore apoptosis signals, increasing abnormal cell survival. Overexpression of the anti-apoptotic gene Bcl-2 in diffuse large B-cell lymphomas makes malignant B cells resistant to apoptosis and causes tumore growth.

Genes Involved in DNA Repair

• Fix mistakes when DNA is copied.
• TSGs acts wild-type as DNA repair genes like BRCA1, BRCA2, and p53
• "Loss-of function” mutations will affect DNA repair genes
• Impair the ability of the cell to recognize and repair genetic damage will cause genomic instability which will cause mutations
• Lynch syndrome Mutations in MLHL, MSH2, MSH6, PMS2, and/or EPCAM genes which cause an issue involved in DNA mismatch repair

Driver versus Passenger Mutations

Driver mutations

• Directly cause oncogenesis.
• Occur in oncogenes (gain-of-function mutations).
• Occur in Tumor suppressor genes (loss-of-function mutations).

Passenger mutations

• Are neutral mutations
• May occur anywhere in the genome
• Selective pressure by certain oncologic therapies may favor tumor progression.

Epigenetic Changes

• Affect gene expression without DNA sequence alteration.
• Histone modifications alter gene transcription through methylation or acetylation.
• MicroRNAs interfere with complementary mRNAs, preventing translation into proteins.
• Hyper/hypomethylation alter covalent addition/removal of –CH3 at C5 cytosine bases, causing genes activation/inactivation.

Carcinogenesis Multistep Process

• Most malignant is not due to mutation of a single gene
• The number of mutations found varies
• Accumulation of modest number of mutations in proto-oncogenes and tumor suppressor genes
• The genes that cause cancer are diverse and complex

Multistep Model of Colon Carcinogenesis

• Starts with a Normal Colon to Mucosa at Risk
• Loss of function mutations of TSGs, which can both be acquired, or inherited
• DNA methylation abnormalities in inactivation of normal alleles
• Gain-of-function mutations in proto-oncogenes
• Followed by homozygous loss of TSGs
• TP53 at 17p13 LOH at 18q21
• telomerase and other mutations

Hallmarks of Cancer

• Sustaining proliferative signaling
• Evading growth suppressors
• Resisting cell death
• Enabling replicative immortality
• Inducing angiogenesis
• Activating invasion and metastasis
• Avoiding immune destruction
• Tumor-promoting Inflammation
• Genome Instability and Mutation
• Deregulating Energetics

Sustaining Proliferative Signaling

• Employing activated of oncogenes, production of growth factors (GFs), constitutive activation of GF receptors, can have an increased number of GF receptors.
• Constitutive stimulation of transcription factors

Evading Tumor Suppressors

• Cancer cells bypass mechanisms that normally inhibit cell proliferation.
• Inactivating of RB/CDK inihibitors
• activating mutations in cyclins D/E and CDKs 4,6 or 2
• causing an unaffected Growth Inhibitors

Resisting Cell Death

• Mutations in genes can result in cell death
• Loss of function mutations in p53
• Overexpression of MDM2
• Overexpression in anti-apoptotic members

Enabling Replicative Immortality

• Upregulation of the telomerase enzyme leading to maintenance in cell aging
• Cancer has Loss of -function mutations in p53 cause senescence

Inducing Angiogenesis

• Formation of new blood vessels
• Active around tumors and adequate blood supply
• Creation and spread to other tissues (metastasize)

Activating Invasion and Metastasis

• Spreading the cancer cause secondary tumors and invade neighboring tissues to cause a primary tumor
• The matrix will invaded distant issues and cause more tumors

Avoiding Immune Destruction

• Immune responses can inhibit or eliminate tumors
• Strategies to avoid Immune Destruction includes inhibition of immune response and lead to antigens

Tumor Promoting Inflammation

• Cancer cells display high inflammation towards the tumor mediated by immune cells
• Tumor effects will cause cells to have more infections, GFs, anti-proteases, and metastases

Genome Instability and Mutation

• Requires Successive Accumulation of mutations leading to accelerator rates
• Mutations that have less of functionality during DNA damage will cause multistep
• Genome instability occurs through therapies

Deregulating Energetics

• Normal cells create 36-38 molecules that are completely broken down
• Cancer molecules don’t require tradition molecules
• The metabolic shift takes some time, but intermediates can contribute to more elements
• High levels of glucose will increase with high uptake in cells