Lecture 12: Oncogenes 1- start here tmr

* Understand viral oncogenes using the Rous Sarcoma Virus (RSV) model. * Explain how proto-oncogenes are converted into oncogenes. * Describe identification and activation of cellular oncogenes. * Explain Ras signalling and how Ras mutations contribute to cancer. * Understand KRAS-targeted therapies and mechanisms of resistance. * Describe mechanisms of Myc activation in cancer.

Why were tumour viruses initially considered important in cancer research?

Tumour viruses were initially suspected to cause many human cancers. Although only a minority of cancers are virally induced, studying tumour viruses revealed important mechanisms underlying cancer biology.

Why are viral genomes useful for understanding cancer mechanisms?

Viral genomes are small and contain only a few genes, meaning any transforming effects can often be traced to a limited number of highly potent genes.

Describe Peyton Rous' experiment and explain its significance.

Peyton Rous ground sarcoma tissue from chickens and passed it through a fine filter. The filtrate still caused sarcoma formation in healthy chickens, demonstrating that a virus rather than whole cells caused disease.

Why did RSV provide a unique opportunity for cancer research?

RSV allowed cancer to be induced predictably and repeatedly while producing large quantities of virus for experimental analysis.

How did RSV alter cultured chicken embryo fibroblasts?

RSV infection produced foci of transformed cells that displayed altered morphology and tumour-like metabolic behaviour.

What is cellular transformation?

Cellular transformation is the conversion of a normal cell into a cancer cell with altered growth and behavioural characteristics.

Why did formation of foci suggest transformation had occurred?

Foci indicated cells had lost normal growth constraints and were proliferating abnormally over neighbouring cells.

Describe the two hypotheses proposed regarding maintenance of the transformed phenotype.

The first proposed that virus remained continuously present and maintained transformation, while the second proposed a hit-and-run mechanism where transformation persisted without continued viral activity.

How was the hit-and-run hypothesis experimentally tested?

Temperature-sensitive RSV mutants were used to determine whether transformation persisted after viral protein activity was disrupted.

What happened at non-permissive temperatures in temperature-sensitive RSV experiments?

The transformed phenotype was lost because the defective viral protein could no longer function.

What conclusion was drawn from the temperature-sensitive RSV experiment?

The viral transforming protein was required both to initiate and maintain transformation.

Describe altered morphology in transformed cells.

Transformed cells became rounded and displayed abnormal organisation compared with normal cells.

What is loss of contact inhibition?

Loss of contact inhibition is the ability of cells to continue proliferating despite contact with neighbouring cells.

What is anchorage independence?

Anchorage independence is the ability of cells to grow and survive without attachment to a substrate.

Why is immortalisation important in cancer cells?

Immortalisation allows cells to continue dividing indefinitely rather than undergoing normal replicative limits.

Why do transformed cells show reduced growth-factor dependence?

Transformed cells become less dependent on external mitogenic signals because oncogenic pathways drive proliferation internally.

Why do transformed cells demonstrate increased glucose uptake?

Cancer cells have increased metabolic requirements and therefore increase glucose uptake to support rapid growth.

Which RSV genes were required for replication but not transformation?

Gag, pol and env were required for viral replication but were not responsible for transformation.

Which RSV gene was responsible for transformation?

The src gene was responsible for inducing transformation.

What is an oncogene?

An oncogene is a gene capable of transforming a normal cell into a tumour cell.

What is a proto-oncogene?

A proto-oncogene is a normal cellular gene that can become an oncogene after activation or mutation.

Why was discovery of c-src important?

Discovery of c-src showed that normal cells already contain genes capable of causing cancer under certain conditions.

What major conclusion emerged from studying src?

Normal vertebrate genomes contain proto-oncogenes that can be converted into oncogenic forms.

Why was most human cancer thought to involve mechanisms other than viruses?

The majority of cancers were not virally induced, suggesting other mechanisms such as mutagen exposure activate oncogenic pathways.

Give examples of mutagens discussed in the lecture.

Examples included UV light, X-rays, base analogues and DNA intercalating agents.

How were non-viral oncogenes identified experimentally?

DNA from tumour cells was transferred into recipient cells, and transformed recipient cells indicated the presence of oncogenic sequences.

Why was it concluded that only one gene often caused transformation during transfection experiments?

Only approximately 0.1% of donor DNA integrated into recipient genomes, making simultaneous transfer of multiple independent transforming genes unlikely.

What important conclusion arose from tumour DNA transfection experiments?

Activated oncogenes in tumour cells frequently originated from pre-existing normal proto-oncogenes.

How was H-ras identified?

H-ras was identified following transfection experiments using DNA from human bladder carcinoma cells.

Why was H-ras discovery important?

It represented the first example where a mutation in a specific gene was directly linked to human cancer.

Describe the normal Ras signalling cycle.

Ras cycles between an active GTP-bound form and an inactive GDP-bound form, regulated by GEFs and GAPs.

What roles do GEFs and GAPs play in Ras signalling?

GEFs activate Ras by promoting GTP binding while GAPs stimulate GTP hydrolysis and inactivate Ras.

Why do Ras G12V or G12C mutations promote cancer?

These mutations prevent efficient GTP hydrolysis, causing Ras to remain constitutively active.

Which Ras gene is most commonly associated with human cancers?

K-ras mutations are the most frequent drivers across human cancers.

Which KRAS mutation was highlighted as particularly common in lung cancer?

KRASG12C was highlighted as being particularly common in lung cancer.

How does AMG 510 inhibit KRASG12C?

AMG 510 forms a covalent bond with GDP-bound KRASG12C and inhibits downstream signalling and proliferation.

Why was AMG 510 more effective when combined with additional therapies?

Combination with chemotherapy, MEK inhibitors or anti-PD1 therapy improved tumour regression and immune responses.

Describe mechanisms responsible for resistance to KRAS inhibition.

Resistance mechanisms included mutations disrupting drug binding, additional KRAS mutations and downstream pathway alterations.

Describe the three mechanisms by which Myc oncogenes can arise.

Myc oncogenes can arise through gene amplification, chromosomal translocation or viral insertional mutagenesis.

Why does c-myc translocation promote Burkitt lymphoma?

The c-myc gene becomes controlled by a highly active immunoglobulin promoter, causing excessive expression and uncontrolled lymphocyte proliferation.