H&N tumor biology and carcinogenesis

Question 1

What are the five basic phases of the cell cycle,

and what occurs during each phase?

Answer 1

● Quiescent phase (G0): Resting state
● Gap 1 (G1): Preparation for cell division; increase in transcription/translation and ~ doubling of macromolecules
● Synthesis phase (S): Replication of chromosomes
● Gap 2 (G2): Continued cellular growth
● Mitosis phase (M): Chromosomes are separated, and two
daughter cells result

Question 2

What is the term given to cells in permanent cell-

cycle arrest?

Answer 2

Senescent

Question 3

What are the key checkpoints within the cell cycle?

Answer 3

● G1/S checkpoint (“restriction point”): Prevent entry into
S-phase, rate-limiting step
● Intra-S phase checkpoint: Halt progression of S-phase if
damaged DNA is detected
● G2/M checkpoint: Prevent entry into M phase
● M checkpoint: Ensure correct replication of DNA and avoid
mitotic exit if errors exist

Question 4

If a lesion (i.e., partially replicated DNA, strand
breaks, or other errors) is identified at a check-
point, what processes can be activated?

Answer 4

● Recruitment of DNA repair effector complexes
● Temporary cell-cycle arrest, which can lead to senescence
or apoptosis, depending on the cell and the lesion

Question 5

What key tumor suppressor protein controls pro-

gression through the G1/S checkpoint (restriction point) and the G2/M checkpoint?

Answer 5

p53 (activates p21 → inhibits cyclin and cyclin-dependent

kinase (Cdk) complexes)

Question 6

What two key classes of molecules regulate a cell’s

progress through the cell cycle?

Answer 6

● Cdks: Catalytic subunit; require cyclin for activation; result
in phosphorylation (activates/inactivates molecules nec-
essary for progression through the cell cycle)
● Cyclins: Regulatory subunit that activates Cdk molecules
when bound to form a heterodimer

Question 7

Although more than 15 Cdks have been identified,
four have key biologic functions within the cell
cycle. What are they, and what do they do?

Answer 7

● Cdk 1: Controls G2 phase and M phase (prophase) →
Cyclin A dependent
● Cdk 2: Controls G1 to S transition and S phase → cyclin E
(and A) dependent
● Cdk 4: G0 to G1 transition → cyclin D dependent
● Cdk 6: G0 to G1 transition → cyclin D dependent

Question 8

Specific families of activators and inhibitors regulate functional activities of the Cdk com-
plexes. Identify the primary activators and inhibitors.

Answer 8

● Activators: Cdk-activating kinase (CAK) and Cdc-25
● Inhibitors: Cdk inhibitors (CKI) → Inhibitor of kinase 4
(INK4a), Cdk Interacting protein/kinase inhibitory protein
(Cip/Kip)

Question 9

What type of gene helps to control cell growth or

progression through the cell cycle?

Answer 9

Tumor suppressor gene (also called antioncogenes)

Question 10

What type of gene promotes cell growth and

progression through the cell cycle?

Answer 10

Proto-oncogenes. Once a proto-oncogene is mutated, it is

known as an oncogene.

Question 11

The classic retinoblastoma tumor suppressor protein (pRb) functions to inhibit what key transcription factors, effectively preventing formation of cell cycle–related proteins and arresting the cell
in G0 phase?

Answer 11

E2F factors

Question 12

Phosphorylation of pRb by what important cyclin-
Cdk complex(es) results in dissociation of pRb from E2F and subsequent entrance into the cell cycle?

Answer 12

Cyclin D-Cdk4 and cyclin D-Cdk6

Question 13

Release of E2F from pRb inhibition results in the
transcription of multiple genes necessary for the
function of the cell cycle. Transcription of what key cyclin protein results in activation of Cdk2, progression from G1 to S phase, additional pRb inactivation, and p27 degradation?

Answer 13

Cyclin E

Question 14

What genes are considered members of the Cip/
Kip family of tumor suppressor genes, and in what
phase of the cell cycle do they inhibit cyclin-Cdk
complexes?

Answer 14

● p21, p27, and p57

● G1 phase

Question 15

DNA damage results in activation of what key tumor-
suppressor gene and proapoptotic factor, which in turn activates p21 (a Cip/Kip Cdk complex inhibitor)
and results in arrest of the cell cycle in G1 phase?

Answer 15

p53

Question 16

Transforming growth factor-β is a growth inhibitor,
which results in activation of which INK4 tumor
suppressor gene causing subsequent cell-cycle
arrest in G1 phase?

Answer 16

p27

Question 17

What genes are considered members of the INK4a
family of tumor-suppressor genes, and in what
phase of the cell cycle do they inhibit cyclin-Cdk
complexes?

Answer 17

● p16 and p19

● G1 phase

Question 18

Name the tumor suppressor gene that is a
member of the INK4a family that prevents p53
degradation and therefore results in cell-cycle
arrest at the G1-checkpoint.

Answer 18

p19

Question 19

Name the tumor suppressor gene that is a member of the INK4a family that inhibits cyclin D-Cdk4/6 complexes and therefore results cell cycle arrest during the G1 phase.

Answer 19

p16

Question 20

What important enzymes function to dephos-

phorylate the targets of cyclin-Cdk complexes, such as pRb?

Answer 20

PP1 and PP2A (phosphatases)

Question 21

To promote an orderly progression through the
cell cycle, cyclin-Cdk complexes must be degraded
to allow the next phase of the cell cycle to
progress uninterrupted. For example, S-phase
complexes cannot be active during M phase and so forth. What important enzymatic process selectively targets these complexes for degradation?

Answer 21

Ubiquitin-dependent protein degradation (ubiquitylation)

Question 22

The phases G1, S, and G2 are collectively referred

to as what?

Answer 22

Interphase

Question 23

What are the five stages of mitosis (M phase)?

Answer 23

● Prophase: Preparatory; cyclin A-Cdk1 active; condensation
of chromatin, polarization of centrosomes, and initiation
of mitotic spindle formation
● Prometaphase: Nuclear envelope breaks down, mitotic
spindle microtubules attach to chromosomes
● Metaphase: Alignment of chromosomes at metaphase
plate
● Anaphase: Separation of sister chromatids
● Telophase: Cytoplasmic division (cytokinesis) into two
daughter cells; chromatid decondensation (expansion)

Question 24

Describe the “two-hit” hypothesis (Knudson,

1971) for carcinogenesis.

Answer 24

Loss of two alleles for a tumor-suppressor gene is necessary
to result in loss of function and tumorigenesis. This
hypothesis is not applicable to proto-oncogenes and
dominant negative tumor suppressor genes (e.g., p53).

Question 25

In an effort to understand carcinogenesis, Fearon
and Vogelstein (Cell, 1990) presented a model of
tumor progression that involved which three
hypotheses?

Answer 25

● Inactivation of tumor suppressor genes or activation of
proto-oncogenes results in the formation of cancer.
● A series of defined genetic events lead to the develop-
ment of cancer.
● This linear progression may vary, but ultimately it is the
accumulation of genetic events that results in the
development of a malignant phenotype.

Question 26

What is the estimated average of time required for
the accumulation of enough genetic alterations to
produce traditional head and neck squamous cell
carcinoma (Renan, 1993)?

Answer 26

20 to 25 years

Question 27

Carcinogenesis models describe a linear progres-
sion from precancerous lesions to overtly malignant tumors. Describe the steps involved for each
of the following models:
1. Genetic progression model for head and neck
cancer
2. Multistep carcinogenesis

Answer 27

1. Hyperplasia → dysplasia (mild, moderate, severe) →
   carcinoma in situ → invasive carcinoma
2. Initiation → promotion → progression

Question 28

In upper aerodigestive tract tumors, what term
describes the histopathologic changes seen in
mucosa surrounding invasive carcinoma and result
in an increased incidence of second primary tumors?

Answer 28

Field cancerization (Slaughter, 1953)

Question 29

What hypotheses focusing on the clonal abnor-

mality required for tumor growth have been put forth to explain field cancerization?

Answer 29

● Abnormal, and genetically unique, clones form inde-
pendently at multiple sites due to exposure to similar environmental carcinogens.
● A single tumoral clone forms and subsequently migrates
via lateral movement through the mucosa (shown in
several studies to be accurate).

Question 30

What are the two predominant environmental
carcinogens that have been associated with head
and neck cancer, show a dose response, and can
function synergistically?

Answer 30

● Tobacco

● Alcohol

Question 31

Why might alcohol, in particular, function as a
synergistic carcinogen with other environmental
carcinogens?

Answer 31

It may decrease the effectiveness of both local and systemic
detoxification enzymes (e.g., cytochrome P450 system).

Question 32

Does cessation of smoking and alcohol consumption reduce the risk of head and neck cancer development? If so, how long does it take for the
risk to reach the level of never-smokers?

Answer 32

Yes. Some studies have shown 20 years or longer.

Question 33

How does diet impact the development of cancer

of the upper aerodigestive tract?

Answer 33

A high intake of fruits and vegetables and low intake of red
meat or processed meats has been associated with a
decreased risk of head and neck cancer (INHANCE
Consortium, 2012).

Question 34

Describe the cell-cycle dysregulation that is
commonly seen in patients with environmentally
related head and neck cancer.

Answer 34

(Posner, Goldman’s Cecil Medicine, 24th ed., Chp196)
● Loss of p16 (normally inhibits cyclin D)
● Upregulation of cyclin D
● Loss of p53 (normally inhibits cell cycle progression and
promotes apoptosis)
● Upregulation of EGFR (enhances mitogenic signaling)
● Upregulation of COX-2 (increased angiogenesis, de-
creased apoptosis)
● Increased chromosomal instability (increased aneuploidy)

Question 35

What genetic conditions are related to an in-

creased risk of head and neck cancer?

Answer 35

● Fanconi anemia (AR; DNA repair gene mutation)
● Cowden syndrome (AD: PTEN hematoma tumor syndro-
me; PTEN is a tumor suppressor gene)
● Mutations in the cytochrome P450 enzymes (CYP1A1
mutations in Asian populations)

Question 36

What circular, double-stranded DNA virus com-
monly infects the basal layer of cutaneous of
mucosal squamous epithelium and is spread by
sexual contact?

Answer 36

Human papilloma virus (HPV)

Question 37

What two HPV strains are considered “low-risk” for
the development of cancer and are frequently
associated with papillomas and warts?

Answer 37

HPV 6 and 11

Question 38

What two HPV strains are considered “high-risk”
for the development of cancer and have been
associated with cervical, anogenital, and head and
neck (predominantly oropharyngeal) carcinomas?

Answer 38

HPV 16* and 18

*90% of HPV(+) head and neck carcinomas

Question 39

The HPV DNA encodes nine open reading frames
(genes) on a single strand of its double-stranded
circular DNA. Seven of these are considered early phase genes (E), and two are considered late-
phase genes (L). What are the general functions of the E and L genes, respectively?

Answer 39

● E genes: Regulate the transcription viral DNA

● L genes: Encode capsid proteins involved in viral spread

Question 40

Name the two viral onco-proteins in HPV-related
tumorigenesis, and identify the two genes, that
control the transcription of these viral proteins.

Answer 40

● E6/E7: Onco-genes

● E1/E2: Transcriptional regulators

Question 41

When HPV DNA integrates into host DNA, the
process can result in deletion or loss of function of
the E1 and E2 viral genes. This in turn results in
what?

Answer 41

Loss of regulation of E6 and E7and subsequent increased

transcription of these viral genes

Question 42

What HPV protein functions to inhibit the function
of p53 by targeting it for ubiquitin-dependent
degradation? What is the result?

Answer 42

● E6
● Progression through G1 checkpoint and inhibition of
apoptosis

Question 43

What HPV protein phosphorylates pRb and thus
targets it for ubiquitin-dependent degradation?
What is the primary result?

Answer 43

● E7
● Release of pRb inhibition of E2F, activation of cell cycle-
related transcription, progression through the G1 check-
point

Question 44

In addition to E2F-related transcription, the
degradation of p53 results in over expression of
what important protein? What impact does this
have on cell-cycle progression?

Answer 44

● p16
● Normally inhibits Cdk4/6, but with the los of p53 and
pRb, does not meaningfully result in cell cycle control.
Can be used as a biomarker of HPV activity.

Question 45

In addition to HPV, what viruses have been

associated with head and neck cancer?

Answer 45

● Epstein-barr virus (EBV): Nasopharyngeal carcinoma
● Human immunodeficiency virus (HIV): Increased risk of
head and neck cancers
● Merkel cell polyomavirus: Merkel cell carcinoma
● Human T-lymphotrophic virus (HTLV-1): Human T-Cell
lymphoma/leukemia
● Kaposi sarcoma associated herpesvirus (KSHV): Kaposi
sarcoma

Question 46

Whereas the epidermal growth factor receptor
(EGFR) is normally expressed in the epithelium of
several organ systems (dermis, gastrointestinal
tract, kidney), it has been found to be dysfunc-
tional in what percentage of head and neck
(squamous cell) cancers?

Answer 46

80 to 90%

Question 47

EGFR is a transmembrane glycoprotein that, when
activated by binding an extracellular ligand, results
in dimerization, tyrosine kinase activation, and a
complex downstream pathway that ultimately
results in what major outcomes?

Answer 47

● Cellular growth and proliferation
● Apoptosis
● Angiogenesis
● Invasion
● Metastasis

Question 48

How does EGFR expression relate to prognosis in

head and neck squamous cell carcinoma?

Answer 48

Increased expression and amplification are related to
decreased recurrence-free survival and cancer-specific
survival rates.

Question 49

What strategies have been used to target aberrant signaling in head and neck squamous cell carcinoma based on a better understanding of EGFR signaling?

Answer 49

● Tyrosine kinase inhibitors (geftinib)
● Monoclonal antibodies inhibiting dimerization (cetuxi-
mab)
● Antisense oligodeoxynucleotide or small interfering
mRNA inhibition of mRNA expression

Question 50

What immune cell is primarily responsible for
identifying normal cells that have been altered by
viral or tumor activity? What is the general
function of these cells in patients with head and
neck cancer?

Answer 50

T cells
Decreased: T cells demonstrate increased apoptosis, decreased recruitment from the thymus, and poor proliferation. Overall immunosuppression

Question 51

What important normal immune system function is
designed to prevent the recognition of “self” through processes such as anergy, suppression, and ignorance, which makes stimulating the immune system
to target self-derived tumor cells challenging?

Answer 51

Tolerance

Question 52

What cell-based therapeutic model has been
studied in head and neck cancer with promising
(although early) results that focuses on using the
immune system to target cancer cells specifically?

Answer 52

Immunotherapy focused on manipulation of the following:
● Cell signaling (i.e., cytokines such as interleukin-2)
● Vaccination (i.e., Gardasil, whole-cell vaccines, dendritic-
cell vaccines, etc.)
● T cells, dendritic cells, or antibodies

Question 53

What type of therapy has the potential to prevent
head and neck cancer from occurring, recurring,
or progressing?

Answer 53

Chemoprevention

Question 54

What prominent chemopreventive agents have

been studied for use in patients with head and neck cancer?

Answer 54

● Retinoids and vitamin A (betacarotene)
● COX-2 inhibitors
● Aspirin
● Selenium
● Vitamin D
● Interpheron-α2a
● α-tocopherol (Vitamin E)
Many others are being studied. There is great controversy
surrounding most of these agents as studies have
demonstrated conflicting results.

Question 55

What chemopreventive effects are retinoids purported to have on head and neck cancer?

Answer 55

They can retard progression of premalignant oral lesions
(leukoplakia and erythroplakia) and have also been asso-
ciated with lower rates of second primary tumors.
Subsequent studies have called these results into question.

Question 56

True or False. The risk of head and neck cancer

increases in the setting of vitamin A toxicity.

Answer 56

False. The risk increases with vitamin A deficiency.

Question 57

What inflammatory mediator is elevated in head

and neck cancer cells?

Answer 57

Prostaglandins as a result of the upregulation of COX-1 and -2