Renske de Veer (rdeveer)
Cancer development and immune defense
Lecture 1. Introduction course
Lecture 2. Stem cell biology & acute myeloid leukemia
Haematopoiesis = making blood, occurs in bone marrow.
- Stem cells can differentiate to mature blood cells (neutrophil, monocytes, megakaryocyte
(produce platelets). Cells leave bone marrow and enter blood circulation.
- Granulocytes: 1-2 days
- Platelets: 1 week. Erythrocytes don’t have nucleus, can survive 80 days.
- B/T cells are silent when there is no infection.
- Nucleated cells have short life span
Lymphoid progenitor: proliferate and differentiate to B/T cells.
Myeloid progenitor: Proliferate and differentiate to red and white blood cells.
Stem cells: self-renewal. Give rise to daughter cell (will never return to stem cell) and self-renewal of
itself.
“Quiescent”: divide very few times to protect against xenobiotics and stress.
CD34 and CD38 are markers on cell membrane of stem cells that can be detected with flow
cytometry. Most immature stem cells are always CD34+.
Recognition cell/transplantation models
Cannot prospectively identify a stem cells. Select cells that look like stem cells or fall into the
population of stem cells, but don’t know if these cells will behave like stem cells. SCID (severe
combined immunodeficient mice) are used to haematopoiesis. Key to defining what markers on stem
cells and what cells are present on progenitor cells.
Development and hierarchy leukemia
Leukemia: disbalance proliferation/differentiation/apoptosis
- >20% immature cells in bone marrow: acute leukemia.
- Types: myeloid vs. lymphoid, acute vs. chronic.
o Myeloid origin: granulocytes or monocytes
o Immature lymphocytes: lymphoid origin.
o Chronic myeloid is discovered by chance, not as dramatic as acute leukemia. Chronic
myeloid cells can still differentiate.
8 different maturation stages of acute myeloid leukemia. Cells that are arrested in a certain stage of
differentiation and expanded.
Treatment: 3 heavy chemotherapeutic treatments in isolation (prone to infections), then autologous
or allogeneic stem cell transplantation.
Every step of differentiation stem cells lose sell-renewal potential. Moreover, which each step the
chance of genetic mutation is higher (more cells).
- Increased cell division: > chance genetic mutations
- More cells: > chance something goes awry
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- First mutation in limited-renewing cell (late stage): mutation disappears due to limited self-
renewal and differentiation. More mutations are necessary in one cell for leukemia to arise.
- First mutation in earlier stage: re-activation of self-renewal machine. When additional
mutations occur (blocking apoptosis etc) leukemia may arise (leukemic progenitors →
leukemia).
- First mutation in stem cells: stays present due to self-renewal. Additional mutations will
result in leukemia (long-term leukemia stem cells → short-term leukemia stem cells →
leukemic progenitors → leukemia).
Limiting dilution assay: it is not possible to give mice leukemia by injecting 1 leukemic cells, because
many cells are required for leukemia to arise.
- Some types of leukemia are still not possible to transplant to mice to generate leukemia.
In patients with leukemia there is a very small populations which is called leukemia stem cells. Look
like normal haemopoietic stem cells and behave like normal haemopoietic stem cells, but have
leukemic properties. These cells are responsible for leukemic blast cells.
- Fast dividing cells will be attacked by conventional chemo (very old fashioned). Slow dividing
cells won’t be affected by chemo and will survive. Survival of these “quiescent” cells are
responsible for recurrence of leukemia.
Type of mutation can direct cells to lymphoid leukemia or myeloid leukemia.
T(8;21): present in 10% of leukemia patients. Selected CD34+CD38- stem cell population: see this
mutations.
First mutation cannot transform a cell to a leukemia stem cell: pre-malignant cell. Can still
differentiate.
MLL-AF9 is a very aggressive oncogene. When transplanting myeloid progenitor leukemic cells with
this mutation in mice, all mice will develop leukemia. These myeloid progenitor leukemic cells look
like normal granulocty-macrophage progenitors cells, not like stem cells. However, a small set of
genes identical to stem cells is expressed on these cells. This indicated that MLL-AF9 mutations
reactivate self-renewal program in GMP.
In most patients first mutations occurs at stem cell level or even more differentiated cells that
generated leukemia initiating cells. These cells will differentiated further and expand, resulting in
bulk myeloid leukemia.
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- Normal stem cell: generation all mature blood cell types, life long
- Cancer stem cell: continuous generation more differentiated immature leukemia blasts. Does
not imply that cancer stem cell is directly derived from normal stem cell. It may re-activate
self-renewal process however. Since stem-cells are very “quiescent” it is not very likely for
this to happen.
Consequences for therapy
- One patient had mutations in transcription factor A and a mutation in growth factor receptor
F, no mutation in signalling molecule N. Patient was successfully treated, but relapsed after 2
years. There was still a mutation in transcription factor A, but not in growth factor receptor F,
and now a mutation in signalling molecule N. This indicates mutation A was the first one, and
then F. All cells with mutation A and F were killed, but not cells with only mutation A.
Therefore, these cells could undergo mutation N and develop leukemia again.
Targeted therapy is more specific, with less side effects.
- There is chance on resistance.
- Should be a relevant target: not very useful when only present in 1% of the patients.
- Expensive drug
- Not for all patients: patients have different mutations, choose a large group with same
mutations.
Summary
- AML is composed of different cell populations depending during which step of differentiation
the mutation occurs .
- Pre-leukemic stem cells
- Leukemic stem cells produce blast
- Blasts cause complaints/disease
o Differences in maturity
o Differences in chemo-sensitivity
- Precision medicine: target all malignant cells
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Lecture 3. Clonal evolution in cancer
Leukemia; pre-malignant, used as example.
- Very good access to tumour, tumour circulates through body. Access to stem cells.
Hematopoiesis is regulated by growth factors and cytokines outside of the cell, feedback
mechanisms.
- Kidney → Oxygen tension → not enough erythrocytes → cells recognize → will promote
production of signalling molecule that stimulates production of erythrocytes → molecule
travels to bone marrow → erythrocytes are produced.
- When ligand binds to external receptors, internal phosphorylation activates domains inside
the cell → signals can be transferred to other proteins (second messengers) → that can
physically move to other parts of cell and physically interact with proteins that regulate cell
cycle and whole machinery that determines which genes are transcribed inside that cell.
Long-tail of cancer: a lot of mutations are recurrent in some patients, only in low percentages. Only a
few genes are mutated in many patients.
Genes that are mutated in leukemia can be functionally classified: splicing factor, epigenetic
regulator, transcription factor, apoptosis, signal transduction.
Some mutations never occur together in the same patient. You can have one driver mutation in a
specific process, but having another mutation in this process does not give an advantage, because
the process is already disrupted.
The more mutation → worse prognosis (→ more aggressive the tumour → more complex tumour →
more difficult to treat).
Clonal evolution in cancer = accumulation of mutations in the cells of a body during a lifetime, and
the effects of those mutations on the fitness of those cells.
- Natural selection in cancer: cells in pre-malignant and malignant neoplasms evolve by natural
selection. There must be variation in the population (genetic and epigenetic changes in
different mutant cells that distinguish them from normal cells), the variable traits must be
heritable (both daughter cells inherit the genetic and epigenetic abnormalities of the parent
cell, and may also acquire new genetic and epigenetic abnormalities in the process of cellular
reproduction), the variation must affect survival or reproduction. → A cell that acquires a
mutation that increases its fitness will generate more daughter cells than competitor cells
that lack that mutation. In this way, a population of mutant cells (clone) can expand in the
neoplasm → clonal expansion.
Driver- and passenger mutations: tumours typically have more passenger mutations than driver
mutations.
- Driver: a mutation that gives a selective advantage to a clone in its microenvironment,
through either increasing its survival or reproduction. Driver mutations tend to cause clonal
expansions. genes of which the mutations are significantly higher than the background
mutations rate
- Passenger mutations: a mutation that has no effect on the fitness of a clone but may be
associated with a clonal expansion because it occurs in the same genome with a driver
mutation. This is known as a hitchhiker in evolutionary biology.
Acquisition of genetic mutations
- Random mutations occur in genome: 2-10 mutations per cell division → in adult, cells may
harbour up to 1000-2000 acquired mutations in genome
