Spring 2026 Biomedical Science Exam 2 material

agents that damage the DNA and cause mutations

- ionizing rays

- UV rays (*UVC rays)

- reactive oxygen species

- chemicals in the environment

- chemicals used in chemotherapy

UVC rays

UV-C from the sun does not reach the earths surface because it is absorbed by ozone layer

- Most of UV-A and some UV-B do reach earth's surface and can cause DNA damage

reactive oxygen species

chemical reactive molecules that contain oxygen; e.g., oxygen ions and peroxides

- produced during cellular metabolism

chemicals in the environment

hydrocarbons in cigarette smoke (tar, benzene)

- plant and microbial products such as aflatoxins

- (metabolites can intercalate DNA) produced by moldy peanuts

catagoeies of mutations

- substituiton

- transition

- transversion

- insertions

- deletions

- frame shift mutations

substitution mutation

a mutation that exchanges one base for another

transition mutation

One pyrimidine is changed to the other pyrimidine, or one purine is changed to the other purine

transversion mutation

Substitution of a purine with a pyrimidine, or substitution of a pyrimidine with a purine

insertions mutation

mutations in which extra base pairs are inserted into a new place in the DNA.  

deletions mutation

mutations in which a section of DNA is lost or deleted. 

frame shift mutations

Since protein coding DNA segment is divided into codons which is a triplet (3 letter code), insertions and deletions can alter the reading frame. 

serine structure

threonine structure

DNA damage: single base alteration (depurination)

It is a change in the DNA in which the purine (adenine or guanine) is removed from the deoxyribose sugar by hydrolysis of the b-N-glycosidic link between them. 

- After depurination, an apurinic site is formed where the sugar phosphate backbone remains, and the sugar ring has a OH group in the place of purine. 

- 5000 purines (a and g) can be lost per cell a day, but are efficiently repaired via BER pathway

single base alteration (depurination for cytosine and thymine)

Loss of pyrimidine base (cytosine and thymine) occurs by a similar mechanism, but at a substantially lower rate.  

cytosine deamination

Deamination of amino group attached to C-4 in cytosine results in the formation of uracil

- Can spontaneously occur at rate of 100 bases per cell per day

- Uracil in DNA can not base pair with guanine, and this disrupts hydrogen bonding

- Cells can correct this spontaneous deamination by the removal of uracil and repair by Base Excision Repair mechanisms. 

adenine deamination

Results in the formation of hypoxanthine

- Hypoxanthine can base pair with cytosine instead of thymine

- This results in a mutation where the original A-T base pair transforms into a G-C base pair.

oxidatatively damaged bases

Reactive oxygen species (superoxide  ion O2, hydroxyl radicals and H2O2) generated in cellular metabolism (normal products), can cause single base alteration. 

alkylation of base

- can cause apurnation (affects guanine)

- Occurs upon exposure to an alkylating agent.  E.g., anticancer agents [nitrogen mustard (cyclophosphamide) used to treat lymphomas]

- They add ethyl or methyl groups to DNA.  They are alkylating agents.  

- Results in miss-pairing, strand breakage or depurination.  Most susceptible site in DNA is N-7 of guanine which renders glycosidic bond susceptible to hydrolysis and loss of the base.

bifunctional alkylating agents

Alkylation of two bases by one alkylating agent (e.g., busulfan, anticancer drug used in CML=chronic myeloid leukemia).

- Results in interstrand or intrastrand cross-linkages.

UV induced DNA damage

UVA and UVB radiation that reaches the Earth’s surface contribute to the serious health effects

UV light induced dimers

UV induces covalent linkage between C-C of adjacent pyrimidine bases.     

- T-T dimers are formed when cells are exposed to UV rays (as in sunlight) (when there are adjacent thymine bases); they are the most common type.  They are the result of photochemical reactions.

- They interfere with base pairing during DNA replication.  

Uv rays from the sun can cause melanoma

80,000 T-T dimers per cell within one hour of sunlight exposure. 

Asymmetry

Borders (irregular)

Color (variegated)

Diameter (greater than 6 mm), about the size of a pencil eraser)

Evolving over time

miss match repair

Repairs miss-paired bases (well studied in bacteria)

base excision repair

Repairs  Non-Helix distorting damage

E.g., Repairing Deaminated base or

Depurinated base.

nucleotide excision repair

Repairs bulky helix distorting damage.

E.g., repairing the T-T dimers.

In bacteria immediately after the replication, the parental strand is methylated whereas the daughter strand is not. 

Methylation does not occur until sometime after replication is completed.  

- Specific proteins (Mut proteins; Mut L, Mut S and Mut H) scan the newly synthesized DNA strand (not methylated strand) looking for mispaired bases`

-  The recognition of the new strand is based on methylation status.     

endonuclease

cuts the new strand at a site near the methylation point. 

exonuclease

removes the portion of the DNA containing the defect

DNA polymerase

adds the correct nucleotides and DNA ligase seals the nicks in the back-bone. 

base excision pair

Repairs damaged DNA throughout the cell cycle.  

- It is responsible primarily for removing small, non-helix distorting damage (e.g., Deamination of Cytosine to uracil). 

DNA glycosides (BER)

DNA glycosylases recognize the modified bases in the DNA

-  Each DNA glycosylase is specific for an abnormal base.  For example, in the diagram at right, cytosine is deaminated to give uracil.

- uracil DNA glycosylase recognizes the abnormal base and removes it.    

gene expression

Conversion of the information in a gene into a protein product

The edge of each base pair is exposed at the surface of the double helix, presenting a distinctive pattern of

- hydrogen bond donors

- hydrogen bond acceptors

- hydrophobic patches

- for proteins to recognize in both the major and minor groove.  

In most cases, the protein makes a series of contacts with the DNA, involving

- hydrogen bonds

- ionic bonds,

- hydrophobic interactions

gene control region

The entire section of the DNA involved in regulating and initiating gene transcription

- This includes promoter as well as regulatory sequences. 

In animals and plants, it is not unusual to have the gene regulatory region spread over

50,000 base pairs of DNA, much of these DNA serve as spacers

- In humans, average gene length is 3000 base pairs.  

It is the gene regulatory proteins that enable the genes of an organism to be turned on or off. 

enhancers

DNA sequences that increase transcription.

repressors

proteins that prevent transcription.  

translational control

Mechanism 1: Specific proteins interfere with binding to ribosomes. 

Mechanism 2:  Regulating the activity of translation factors via phosphorylation.    

Eukaryotic cells decrease their overall rate of protein synthesis in response to a variety of stressful situations, including

- deprivation of growth factors

- nutrients

- infection by viruses,

- sudden increase in temperature.  

This decrease is caused by phosphorylation of the translation eukaryotic initiation factor (eIF2) by specific protein kinases. 

Suppression of Gene Expression

Through RNA Interference (RNAi)

Recently discovered pathways (1998) that cause inhibition of translation and degradation of mRNAs

Small Interfering RNAs (siRNAs)

very similar to miRNAs in terms of functions. However, the way they are formed are different

miRNA is coded by the cellular genome.

siRNA are synthesized and introduced by transfection

or virally coded RNAs introduced into the cells by viral infection.

The RNAi technique

introduces into a cell or organism a double stranded RNA (dsRNA) molecule whose nucleotide sequence matches that of part of the gene to be inactivated. 

The ds RNA is processed by

Dicer enzyme ultimately to a 21 nts long double stranded RNA. 

The Argonaute associates with ds siRNA and converts it to

single stranded siRNA and forms the RISC.   

RISC directs the siRNA to hybridize with

mRNAs and causes its degradation

external factors of cancer

tobacco, infection (HPV), chemicals and radiation.

sunlight, diet, alcohol, carcinogens…..

internal factors of cancer

inherited mutations (e.g., BRCA1 and BRCA2), hormones, immune conditions and mutations that occur from metabolism, obesity, immunosuppression, chronic inflammation, age….. 

Cancers are Genetic Diseases

Loss of Control over Cell Cycle!

It is a collection of diseases that arise from the loss of control over cell division.

Because cell cycle is controlled by proteins, and proteins are encoded by genes, cancers are genetic diseases.

In cancer, cells proliferate abnormally and invade tissue locally or at locations distant from their site of origin. 

Cancers arise due to mutations in

Proto-oncogenes and Tumor Suppressor genes

EGF-R, HER2, Src kinase (Proto-oncogenes)

P53, Rb, BRCA-1 and BRCA-2 (Tumor

suppressor genes)

CYCLINS and

CYCLIN DEPENDENT

KINASES

Regulate Cell Cycle

tumor / neoplasm

An abnormal cell that grows and proliferates out of control

benign

neoplastic cells do not become invasive, the tumor

cancerous tumor

its cells have acquired the ability to invade surrounding tissue.  

two heritable properties of cancer cells

reproduce in defiance of the normal restraints on cell growth and division

They invade and colonize territories normally reserved for other cells

adenoma

benign epithelial tumor with a glandular organization

adenocarcinoma

corresponding type of malignant tumor

- vast majority of the colorectal cancer are this type because colon has numerous glands within its tissue.     

A benign glandular tumor (an adenoma) remains inside

basal lamina

- marks the boundary of the normal structure whereas a malignant glandular tumor destroys duct integrity

cancer incidences

proto oncogene

a gene that is present in the cell that when mutated or expressed at very high levels can transform a normal cell to a cancer cell

c-Src (cellular Src in contrast to the viral Src); type of protooncogene

Regulated expression of proto-oncogenes is  important for normal cellular functions such as growth and differentiation.

When c-Src is mutated,

its expression is abnormal (very high) and it can cause cancer.

Mutated c-Src is an oncogene as it has the potential to cause cancer. 

examples of protooncogenes

Human Epidermal Growth Factor Receptor 1 (HER1) and 2 (HER2)-these are Receptor tyrosine kinases.

B-RAF- a cytoplasmic serine/threonine kinase.

C-Src- a cytoplasmic tyrosine kinase. 

c-Myc- a transcription factor

In normal epithelial cells, expression of -------- is tightly regulated

EGF-R (epidermal growth factor receptor)

In normal cells, when the EGF receptor is not phosphorylated, it is

inactive

In normal cells, binding of the EGF to EGFR causes

dimerization of the receptor and tyrosine phosphorylation of several residues

- This activates signal transduction and causes cell proliferation. 

- Receptor dephosphorylation and or internalization turns off signal transduction pathways.   

reason proto ongoene becomes an oncogene (point mutation / deletion)

translates into a changed amino acid that has different biochemical properties than the amino acid the DNA sequence originally encoded for

This change in amino acid can mimic phosphorylation, or inhibit binding of negative regulatory molecules, resulting in constitutively active protein.   

reason proto ongoene becomes an oncogene (gene aplification)

normal protein produced in much higher amounts

reason proto ongoene becomes an oncogene (chromosome rearrangement

- placement of strong enhancer causes overproduction of normal protein

- fusion to another actively transcribed gene results in either increased levels of fusion or fusion protein is hyperactive

How Point Mutations or Deletions Can Cause Cancer

There are 4 members in Epidermal Growth Factor Receptor (EGFR) family.  They are HER1, HER2, HER3 and HER4. 

- They are receptor tyrosine kinases localized to the plasma membrane.  They are important for cell growth, survival, adhesion, migration and differentiation.     

- Mutations and deletions in HER1 can lead to colon and lung cancer

HER-1

Mutations or NH2 terminal deletion in HER1 gene could lead to its constitutive  activation and results in cancer.  Observed in Lung and colon cancers. 

- Genzyme and Qiagen make kits to detect EGFR mutations in cancer tissue.

drugs that target HER-1 (EGFR1)

- Monoclonal antibodies (Intravenously given):

- Cetuximab (Erbitux) (Imclone Systems)

- Panitumumab (Amgen)

- Matuzumab (Merck Serono in collaboration with Takeda pharmaceuticals)

Tyrosine kinase Inhibitors

- Gefitinib also known as Iressa -1st selective tyrosine kinase inhibitor, marketed by AstraZeneka. 

- Erlotinib (Genentech, OSI pharmaceuticals, Roche)

MOA of cetiximab

It is a human monoclonal antibody.

Used to treat metastatic colorectal cancer and head and neck cancers.

Binds to the extracellular domain of the EGFR and inhibits signaling.

MMOA of gifitinib

It is  the first selective inhibitor that targets mutant EGFR

- Binds to the ATP binding pocket in EGFR1 cytoplasmic domain.

Used to treat lung cancers. 

HER-2

It is a member of Epidermal Growth Factor Receptor (EGFR) family. 

Approximately 12-40% of breast cancers have an amplification of the HER2 gene.

This leads to overstimulation of the receptor and abnormal cell proliferation.  

and colon cancer

interactions of Herceptin with HER2

Binding to HER2 interferes with the dimerization of the receptor.

Receptors are down- regulated and disappear from the membrane.

herceptin

monoclonal antibody

-  It was the 1st pharmacogenomics based targeted treatment for breast cancer.

- It interferes with HER2 receptor and blocks cell signaling

- Monoclonal antibodies (written as mAb) are identical antibodies produced by one type of B cells and are obtained from clones of a single cell

chronic myelogenous leukemia

hematologic cancer resulting from malignant transformation of pluripotent stem cells

- A specific chromosomal abnormality called Philadelphia chromosome (Ph) is observed in ~95 % of patients

philidelphia chromosome

- This chromosomal abnormality results from the translocation of parts of chromosome 9 and 22

A proto-oncogene called Abelson or c-abl located on chromosome 9 is translocated to chromosome 22 near a gene called Bcr (break point cluster region protein), and a part of chromosome 22 breaks off and moves to chromosome 9.    

Abl is a tyrosine kinase that requires cytokine to be activated.  In its monomeric form, Abl is inactive.   When cytokines are present, Abl proteins are dimerized, phosphorylated and activated.

treatment of CML

Gleevec (Imatinib mesylate)

- It is a p210 tyrosine kinase (BCR/Abl) inhibitor. 

- It inhibits proliferation and induces apoptosis in bcr-abl positive cells. 

Dasatinib- from Bristol- Meyers Squibb

- for treatment of adults with ALL (Acute Lymphoblastic Leukemia) and it is also used to treat CML patients.  (Dasatinib is a Abl-Bcr kinase inhibitor, also inhibits Src kinase).

tumor supressor genes

inhibit multiple steps in signal transduction pathways or modulate transcription; thus, contribute to inhibition of cell proliferation.     

They regulate and inhibit inappropriate cell growth and cell proliferation.  

examples of tumor supressor proteins

are RB-1 (retinoblastoma gene) and p53.

RB-1 MOA

Progression from G1 to S requires E2F transcription factors.  RB-1 can bind to E2F and inhibit its activity.

RB-1 Retinoblastoma Tumor Suppressor Gene

is inactivated by

HPV encoded protein

belongs to the pocket protein family, whose members have a pocket for the functional binding of other proteins

E6 protein can inactivate another tumor suppressor protein p53 and can lead to

cervical cancers

- Vaccine called Gardasil (Merck) and Cervarix (Glaxo).  The vaccine contains virus like particle but no genome, so it is non-infectious

-  Vaccination can prevent cervical HPV-induced cancers

Main Functions of p53the Guardian of the Genome

apoptosis

programmed cell death

- In multicellular organisms, cells that are no longer needed or are a threat to the organism are destroyed by a tightly regulated cell suicide process known as programmed cell death

- This happens naturally during embryonic development

also occurs in cells that contain extensive DNA damage.  So, it is a normal process of eliminating the unwanted cells

Many anti-cancer drugs kill cancer cells by causing

DNA damage and through induction of apoptosis

- Most of the cytotoxic anticancer drugs in current use have been shown to induce apoptosis in susceptible cells

- Topoisomerase I and II inhibitors, alkylating agents etc.,

Bax

pro-apoptotic

Increases the permeability

Bcl2

anti-apoptotic

Decreases the permeability

In normal mitochondria Bcl2 binds to

bax

- Quenches the action of Bax

In DNA damaged cells p53 accumulates in the

cytoplasm

- it interacts with Bcl2 and activates Bax

apoptotic caspade cascade

p53 functions as a

tetrameter

People who inherit only one functional copy of the P53 gene will most likely develop

tumors in early adulthood

present in 15 of the 16 tetramers.  Mutant p53 exerts dominant negative effect. This may cause loss of p53 function.

BRCA 1 and 2

Breast Cancer type 1 and 2 Susceptibility Proteins

- belongs to the tumor suppressor gene family

- involved in the repair of DNA double strand breaks

where does BRCA 1 and 2 bind to

a protein called RAD51 to repair the breaks in the DNA

These breaks occur during:

Naturally during:

- chromatid exchange

- Medical radiation

- Environmental exposure

BRCA1 and BRCA2 helps to repair double stranded DNA breaks during

S and G2 phase of cell cycle.

In cancer, protooncogenes are

activated

- EGFR2 (HER2), c-Src, c-Myc, B-Raf

in cancer, tumor suppressor genes are

deactivated

- P53, RB-1, BRCA1 and 2