5 Proliferation

The expression ‘tumour’ derives from the Latin tumere - describing a local swelling or lump. In the case of cancer such lumps are formed by an excessive growth of cells. Cells grow individually, i.e. in size, but growth in tissues is normally based on an increase in the number of cells; this growth happens through cell division, whereby each cell divides to produce two daughter cells, thus doubling the numbers with each round. The two daughter cells are identical and this process is also known as symetric cell division.

The cellular proliferation machinery is extremely powerful. This can be easily demonstrated by the fact that each individual starts with just one fertilised egg or ovum. During embryogenesis, cell numbers increase from the 16 cells of the morula stage (day 2-3) to hundreds of cells (blastocyst) only a week later, and further to form the embryo, fetus, child and eventually adult human being. A rough estimate suggests that it typically takes about 1,000,000,000 cells/gram of tissue, or roughly a 100,000 billion of cells in a person.

It is apparent that such a potent proliferation process needs to be well controlled and therefore the signals regulating this need to be controlled tightly. Maintaining the appropriate cell number in a tissue is an ongoing process as cells tend to need regular replacement; in fact an estimated 300 million cells are replaced in every minute.

How do cells normally proliferate and how is this regulated in the body?

5.1 Cell proliferation occurs as part of the cell’s life cycle

Cellular growth in tissues is based on repeated rounds of cells division.

To prepare for cell division the individual cell grows, producing more of its internal component parts, including various organelles, and increasing in volume. Once this process has been completed the cells can, if appropriate signals are received, engage the machinery for cell division. Multiple rounds of growth and cell division lead to proliferation and quick increase of cell numbers. The cell cycle can therefore be seen as the ‘motor’ driving proliferation. The cell cycle is characterised by a number of distinct stages with typical morphological changes associated with specific processes. Most importantly, this includes the

  • S-Phase (‘Synthesis’)
  • Gap Phases G1 & G2
  • M-Phase (Mitosis)

However, the default state is either quiescence (G0) or differentiated:

The life cycle of a cell.

Figure 5.1: The life cycle of a cell.

However, typically somatic cells only go through a limited number of rounds of division and therefore have limited proliferation potential. In fact, tissues are normally replenished from only a few, tissue-specific stem cells, also known as adult stem cells which are found in specific niches within a tissue. They only divide occasionally in a process known as asymmetric cell division. The process is termed ‘asymmetric’ because one of the resulting daughter cells remains at rest within the niche while the other daughter cell will move away from the niche to undergo a limited number of cell divisions. The cells resulting from these divisions will ultimately stop proliferation and will instead take on tissue specific and cell type-specific features (phenotypic commitment) leading to a state of terminal differentiation. In this process the cells typically loose the ability to proliferate again.

Therefore the cells in a tissue will typically not be undergoing cell cycling at any given time but rather be in a quiescent state (G0, from which they can re-enter the cell cycle) or exist as terminally differentiated specialist cells (Figure 5.1).

Finally, cells reach the end of their life span and then undergo senescence which an aging process linked to a number of morphological changes to the cell. Eventually cells that have become faulty should be eliminated by an orderly process of removal known as apoptosis so that they can be replaced with efficient new differentiated cells.

5.2 The restriction point R permits or prevents cell cycle entry

Mammalian cells adapted to growing in culture in the laboratory can divide continuously if sufficient space and nutrients are available. In 1974 American biochemist Arthur Pardee discovered that in order for this to happen the cells required the presence of certain factors, e.g. growth factors, during a specific time window in order after completion of the previous cycle to enter into the next cell cycle (5.2).

If cells had been starved of such growth-factors or their protein synthesis inhibited for a few hours (3 hours after the last mitosis) the cells would not progress into the next cell cycle. However, if the same starvation was initiated a little bit later (after 4 h) it did not have the same effect and cells continued their progression through the cell cycle regardless. This suggests that there is a specific point during the progression of cells through the cell cycle at which the presence of such factors is critical in order not to ‘restrict’ further progression and for the cells to commit to entering into the cell cycle; otherwise cells would remain quiescent(G0). This point known as restriction point (R) sits between the early and late G1 phase. R represents a threshold of mitogenic (growth-inducing) signals that cells have to be exposed to before entering the cycle. Normally, cells enter into the cell cycle in the presence of the appropriate regulatory signals and sufficient supply of essential nutrients. In addition to requiring the presence of these growth promoting signals cells also check that there are no growth inhibitory signals or damage that would prevent progression through the cell cycle.

Entry at restriction point R depends on

  • Presence of

    • Nutrients

    • Growth signals

  • Absence of

    • Inhibitory signals

    • Damage/problems

These signals are integrated at the beginning of the G1 phase at the restriction point R. This represents a trigger point that determines whether the cell proceeds with the cell cycle or aborts/waits. Only if the balance of growth promoting and inhibitory signals reaches a certain threshold for the required period of time does the cell proceed to enter the cell cycle.

Natural progression of cancer (tumour)

Figure 5.2: Natural progression of cancer (tumour)

Events that lead to the lowering of this threshold reduce control of R over the cell cycle. This could allow premature entry into the cell cycle (even without appropriate mitogenic signalling) inducing unregulated proliferation and contribute to cancer development.11

5.2.1 Cell Cycle Progression

Once the cell has entered into the cell cycle (beyond R) it is committed to carry on with the whole sequence of cell cycle events wich once started follow a predetermined sequence. Each cell cycle phase is associated with a specific gene expression program in order to accomplish specific tasks. Typically, the tasks in one phase have to be successfully completed before the cell can progress to the next phase. Cell cycle check points exist that serve as control elements at the transition between different stages (i.e. G1, G2, M) to ensure that one stage of the cell cycle has been completed before the next one starts. For example, at G2 the cell makes sure that it is ready to undergo mitosis, for exmple checking that all DNA has been replicated and is damage free, or at the M-phase spindle check point that the microtubules have been assembled correctly.

In the event of problems or damage being detected at these check points the cell triggers repair mechanisms. If these attempts are unsuccessful and the problem can not be rectified signalling events lead to the triggering of apoptosis (‘suicide’) will be triggered.

The cell cycle follows a all-or-none principle: Once the cell has proceeded to enter into the cell cycle beyond R the cell cycle proceeds as a sequence of predefined events or stages. Progression through the cell cycle can only be paused but not stopped. If the cell cycle can not be completed (even after damage repair attempts) the consequence is that the cell will undergo apoptosis.

5.3 Cell cycle regulation

The progression through the different phases of the cell cycle depends on specific sets of cell machinery (e.g. for DNA replication in S-phase, for separation of chromosomes in M-phase). The presence of the proteins required for each phase depends on the timely expression of the corresponding set of genes by specific transcription factors. The level of gene expression and the activity of these genes therefore needs to be tightly controlled.

How does the cell ‘know’ when to express the different genes?

5.3.1 The progression through the cell cycle is controlled by cyclins

The cyclins and cyclin dependent kinases are the key proteins regulating the cell cycle. They were discovered in the ‘80s by British biochemist Tim Hunt and geneticist Paul Nurse, and American Leland Harrison.12 The cyclincs (A,B,D,and E) are a group of proteins which show a regular increase and decrease of concentration (oscillations) throughout the different stages of the cell cycle (Figure 5.3).

Cyclins are the regulators of the cell cycle.

Figure 5.3: Cyclins are the regulators of the cell cycle.

Cyclins exert their effects after binding to cyclin dependent kinases (CDK); each cyclin with a specific CDK partner. The binding in this complex begins activation of the kinase element of the CDK. Once fully activated the kinase functionality will then activate various target proteins by phosphorylation so that downstream expression of cell-cycle-stage specific genes can be triggered.

The progression through the different stages of the cell cycle is orchestrated as a sequence and the concentration of different cyclins increases at the transition between different stages of the cell cycle and then decreases again. These oscillations are regulated by feedback loops.

Key elements of cell cycle reglation:

Cyclins - Cyclin dedpendent kinase (CDK) - CDK Inhibitor

  • Increase in Cyclin
    • Complex and activates cyclin dependent kinases (CDKs)
    • Complex allows phosphorylation ⟶ triggers downstream transcription program
    • Initiates next phase of cell cycle
  • Increase in cyclin dependent kinase inhibitor (CKI)
    • Inactivates complex

This prinicple and the effect of feedback loops is discussed below in more detail for the transition from G0 to G1/S phase:

5.3.2 Cyclin D regulates the G0 to G1/S phase transition

The transition between cell cycle phases depends on feedback loops as illustrated in Figure 5.4.

Cyclin D regulates the transition of G~0~ to G~1~/S Phase

Figure 5.4: Cyclin D regulates the transition of G0 to G1/S Phase

In quiescent, G0 cells, the E2F–DP transcription factor is bound to p130, the principal pocket protein in these cells, which keeps it inactive (0). In G1, however, E2F–DP complexes to the protein Rb (retinoblastoma {#rb} protein) predominate. Early G1 is dependent on mitogenic signals (growth factors). Anti-mitogenic signals e.g. WAF1/KIP proteins are involved in regulation by binding to CDKs/complexes. Once passed the restriction point further cycling is mitogen-independent (late G1 phase).

Mitogenic signalling results in the synthesis of cyclin D (Cyc D) and the subsequent formation of active CDK4/6–cyclin-D complexes (1). This CDK4/6–cyclin-D complex carries out the initial, partial phosphorylation of RB. Partially phosphorylated RB binds to E2F–DP (2) and after binding to DNA transcribes specific genes, such as cyclin E (3). The increasing levels of Cyclin E to the formation of a complex with CDK2 which activates this kinase (4). CDK2 then completes phosphorylation of the remaining phosphorylation sites on RB which leads to the inactivation of RB-E2F-DP transcription factor (5). This inactivation of RB allows induction of a different set of E2F-responsive genes (6) which then continue to drive cells through the G1/S transition and to initiate DNA replication.

5.4 Therapeutic relevance

Our interest in understanding cancer is motivated by our desire to understand the mechanisms of actions, clinical application, and side effects of current cancer drugs so that their benefits and limitations are explained and potential avenues for novel therapies may be appreciated.

In the first instance, tumours are caused by excessive and unregulated proliferation. Cellular proliferation depends on mitosis and the presence of mitogens. The regulation of mitosis and the cell cycle itself has been are tightly regulated and once a cell has progressed beyond R an inability to complete the full cell cycle (e.g. because of DNA damage) triggers apoptosis.

Could we expose proliferating cancer cells to compounds that stop the cell cycle progression in order to kill cell by triggering apoptosis?

5.4.1 Cell cycle and proliferation as drug targets

In fact, the ability to interfere with the completion of he cell cycle and thus to potenitally trigger apoptosis is the mechanism that many commonly used cytotoxic chemotherapeutic anti-cacner drugs in share. Some drugs are active only in specific phases of the cell cycle because their mode of action others can target cells at any part of the cell cycel although the impact or damage cause by those drugs may only become apparent later when the cells need to complete specific cell cycle check points.

  • Cell cycle specific – active in specific phase
    • Mechanism based on causing ‘damage’

      • S phase - DNA replication
        • Requires unwinding of DNA - target via topoisomerase inhibitors
          • Requires DNA building blocks - target via anti-metabolites
      • M phase - Mitosis
        • Requires alignment of chromosomes/spindle - target via spindle toxins
    • Mechanism based on inhibition of cell cycle regulation - not damage (next generation drugs)

      • CDK4/6 inhibitors ⟶ palbocilib & ribocilib (2017)
  • Non-specific – no phase requirement
    • Mechanism based on interference with general proliferation
      • However, cell death can be triggered at specific cell cycle check points e.g. G1, G2

Drugs discussed above can kill cells by interfering with the cell cycle. However, we have established that tumours and, specificly the associated proliferation depend on mitosis but also on the presence of mitogens to enter into the cell cycle in the first place.

Instead of killing cells once they have entered into the cell cycel would it be possible to switch off proliferation before the start? This approach requires an understanding of what mitogens are and how they drive cells to enter into the cell cycle proliferate.

5.5 Growth factors are key regulators of cell cycle entry

Growth factors are signalling molecules, typically proteins or steroid hormones, that are involved in driving cellular functions including growth/proliferation and differentiation. While steroid hormones can enter the cell and interact with their respective receptors inside the cell, protein growth factors can not enter cells and need to relay their signal into the inside of the cell via specific surface receptor proteins.

Receptor tyrosine kinases (RTKs) represent a large family of these membrane bound growth factor receptor molecules (Figure 5.6. Their structure and architecture is linked to the key functions that they fulfil, i.e. as transmembrane receptors they have a large central part of the protein spanning the cell membrane (‘trans membrane domain’), while on the outside of the cell they have structures that allow binding of specific growth factors (‘ligand binding domain’) and facilitate interaction with other RTK proteins. Binding of a ligand will trypically induce changes which allow binding of a second RTK protein (dimerisation); this could involve the same RTK (homo-dimerisation), another RTK (it is called (hetereo-dimerisation), or even more than two RTKs (oligomerisation).

Human receptor tyrosine kinase families typically have an extracellular receptor domain, a transmembrane domain, and a intracellular signal transduction domain.

Figure 5.6: Human receptor tyrosine kinase families typically have an extracellular receptor domain, a transmembrane domain, and a intracellular signal transduction domain.

The part of the protein facing into the cell (‘cytoplasmic domain’) typically contains the tyrosine kinase functionality which gives this protein family its name. Its function is (auto)-phosphorylation which induces conformational changes which then allow binding of molecules for downstream signalling, typically the first step in the triggering of a complex signalling cascade leading to the expression of specific target genes to start the cell’s response to the signal, e.g. proliferation or differentiation.

There are over fifty transmembrane RTKs distributed which can be organised into about 20 subfamilies based on structural similarities. Many growth factor receptors are involved in signalling through proliferative pathways, i.e. pushing cells towards more rapid growth and cell division; consequently, such receptors also can potentially support uncontrolled cell growth and can thus be considered proto-oncogenes/oncogene (see below).

5.6 The ErbB / HER receptor family and Epidermal Growth Factor Receptor (EGFR)

The ErbB or HER RTK family groups together four proteins of similar structure which are involved with proliferation. One of them is the human Epidermal Growth Factor Receptor (EGFR) also known as Her1 or ErbB1 has a recognised role in a number of cancers. Similarly, the RTK known as Her2/Neu/ErbB2 is important as a driver of cancer cell proliferation.

HER Erb Common name
HER1 ErbB1 EGFR
HER2 ErbB2 Neu
HER3 ErbB3 -
HER4 ErbB4 -

5.6.1 The role of HER2 in cancer

HER2/neu, also known as ERBB2, is a growth factors and important proliferative signalling molecules and EGFR and HER2 therefore are potentially important oncogenes. Its involvement in cancer depends on increasing signalling driving downstream pathways in the cell. In this case, a typical mutation leading to its oncogene activity, i.e. sustained proliferative signalling, is a gene amplifications leading to the over-expression of the RTK proteins and therefore more proliferative signalling in the cell.

Specifically, HER2 is involved as a proliferative driver in 15-30% of breast cancers, in addition, it is also implicated in a number of other cancers such as ovarian and stomach cancer. Tumours expressing HER2 tend to proliferate faster due to the higher level of activity so that finding ways to suppress this higher Her2 signalling would make such cancers potentially amenable to HER2 directed therapy. Therefore, diagnosis of the HER2 status of a patient’s tumour is an integral part of the initial diagnosis, either when biopsies are taken and/or once the tumour lumps are examined by the pathologist after surgery.

The diagnosis is typically carried out using either immunohistochemistry (IHC) or fluorescence in-situ hybridisation (FISH). For IHC tumour levels of HER2 are graded from 0-3 and levels of 3+ are considered positive, while 0-1 are considered negative.

Monoclonal anti-body based therapy based on Herceptin/Trastuzumab has shown effects in the (neo-)adjuvant therapy of HER2-positive breast cancer, i.e. used before (neo) or after primary surgery (typically lumpectomy) to improve long term outcome of therapy, but also for the treatment of advanced stages of breast cancer in combination with paclitaxel.13

Trastuzumab is also used in the form trastuzumab emtansine (Kadcyla), an antibody drug conjugate, a molecule that combines the targeting from the anti-body with the inhibition of tubulin by emtansine (also see chapter on tubulin targeting agents) which is indicated for pre-treated metastatic BC patients in the US. Other therapies that target the HER2-driven proliferation include Lapatinib (Tyverb), Pertuzumab (Perjeta).

5.6.2 HER2 vs. other HER/EGFR family members

Receptor tyrosine kinases (RTKs) EGFR (HER1), HER3, and HER4 typically have a “closed” conformation with the dimeriation sub-domain inaccessible; the ligand binding induces conformational changes enabling dimerisation. The Her family members bind to a set of 11 different growth factor ligands. However, HER2 is considered an orphan receptor, i.e. it has no known ligand that binds to it. Furthermore, HER2 is in an ‘always open’ conformation and can dimerise without the need for ligand binding. This means dimersiation could occur without the requirement of growth factor binding. HER2 prefers to undergo hetero-dimerisation, i.e. it will preferentially bind to another EGFR family member such as EGFR but can at high HER2 receptor concentrations undergo homo-dimerisation. Consequently, an increase of the number of HER2 receptors on a cell will lead to a higher chance of HER2 dimers triggering downstream proliferative signalling. Once either homo-dimerisation or hetero-dimerisation have occurred the cytoplasmic parts of the RTK undergo autophosphorylation and binding to the sites for SH2 (src homolog 2) domains and PTB (phoshotyrosine binding) can occur – allowing the activation of the next step. In the case of HER2 downstream intracellular pathways depend on the specific dimerisation partner i.e. signalling occurs through either the MAP-kinase or PI3 kinase pathway (Figure 5.7). From a clinical perspective, such homodimers are predictive of the likley therapeutic response to Herceptin/trastuzumab.

HER2 dimersiation with different partners signals through the MAP kinase (MAPK) and PI3K pathways.

Figure 5.7: HER2 dimersiation with different partners signals through the MAP kinase (MAPK) and PI3K pathways.

5.6.3 Signalling via the MAP kinase pathway

This signalling can be divied into three main elements - activation of the RTK, relay of the signal into the cytoplasm, ant the MAP activation cascade.

5.6.3.1 RTK activation

After the binding of the respective ligand to the extracellular receptor domain, or in the case of HER2 even in absence of a binding, RTK monomers are able to bind to one another forming homodimers or heterodimers, respectively (Figure 5.8). The subsequent auto-phosphorylation leads to conformational changes allowing access to binding sites thus initiating the intracellular part of the signalling pathway. RTK receptors typically are controlled by some common signalling mechanism in which an initial extracellular binding event occurs involving the respective ligand. The cytoplasmic tails (intracellular domains) of such receptors are now in close proximity, allowing one kinase domain on one receptor to phosphorylate a matching residue on the other receptor (receptor auto-phosphorylation). The phosphorylation now leads to conformational changes in the cytoplasmic tails fo the receptors that expose previously inaccessible binding sites that allows binding of intracellular adaptor or signalling proteins. Specifically, they expose the SH2 (src homolog 2) binding sites which in turn allow docking of other proteins containing the matching SH2 domains. SH2 (Src homology 2) domains, originally recognised in the Src oncoprotein, form part of a number of different adaptor proteins involved in signal transduction, specifically in RTK pathways.14

Dimersiation and activation of human receptor tyrosine kinases.

Figure 5.8: Dimersiation and activation of human receptor tyrosine kinases.

5.6.3.2 Relay of signal

The activation of the RTK has conveyed the signal (the binding of the ligand) from the outside of the cell membrane to the inside; the next step is to relay the signal from the cyoplasmic side of the membrane into the cytoplasm. The docking of proteins containing SH2 domains allows adaptor proteins that contain these SH2 domains to recognise phosphorylated (i.e., activated) tyrosine residues and subsequently bind to sites containing these residues (SH2 binding site) (Figure 5.9 - (1)) .

GRB2 (growth factor receptor-bound protein 2) is one example of a adaptor protein containing the SH2 domain. GRB2 functions as adaptor protein in the HER2 triggered RTK pathway that on the one hand binds the activated tyrosine kinase and on the other hand then allows subsequent binding of a protein named SOS1 via further binding domain, named SH3 domain.

SOS1 is a GNEF (Guanine nucleotide exchange factor). Once bound to GRB2 SOS1’s role is to bind to the inactive form of the Ras protein which contains guanosine-diphosphate, GDP. Once the inactive Ras-GDP has bound SOS1 catalyses releases (‘exchanges’) of the the GDP. Release of the GDP makes the Ras binding sites available again for the binding of GTP to create the active form of Ras, Ras-GTP.

5.6.3.3 The Ras cycle and its helpers

Ras proteins are members of a family of proteins known as ‘small GTPases’ with the H-Ras, K-Ras, and N-Ras forms being most relevant in cancer with mutations being found in 20-30% of human cancers.

Ras acts like a cellular ‘on/off’ switch for a number of different signalling pathways, with the MAPK pathway being particularly important for proliferation (Figure 5.9) .

In its inactive form Ras is bound to guanosine diphosphate (GDP). After release of GDP with the help of the GNEF SOS1 Ras is able to bind GTP (5.9 - (2)), which carries and additional phosphate group. Binding triggers a conformational change of Ras making it active and thus allowing it to bind and thus activate Raf (3) and thus engage the MAPK pathway that ultimately leads to target gene expression (see below).

Ras function depends on two type of ‘helpers’: In order to terminate this signal the active form of Ras needs to be ‘switch off’ by hydrolysis of GTP to GDP. Catalysis of this hydrolysis step by Ras would be too slow and additional helper proteins known as GTPase activating proteins (GAP) accelerate this process (4), thus inactivating Ras-GTP to Ras-GDP (5). Loss of function of GAPs can lead to continued proliferation signalling to Raf and is thus relevant in cancer. The GNEF (Guanine nucloside exchange factor) SOS1 releases releases (‘exchanges’) GDP to make the binding sites available again for the binding of GTP (1).

The Ras cycle relays the signal from the membrane to the MAPK pathway.

Figure 5.9: The Ras cycle relays the signal from the membrane to the MAPK pathway.

5.6.3.4 Activation of mitogen activated protein kinases (MAPK)

Principle: Mitogen activated protein kinases (MAP kinases or MAPK) phosphorylate proteins specifically on threonin, tyrosin, and serine residues. This family of kinases typically depend on activation by two phosphorylation events relayed from an two upstream tiers of kinases. The kinase that activates MAPK is known as a MAP2 kinase (MAP kinase kinase) and in turn has to be activated by a further upstream kinase, MAP3 kinase (or MAP kinase kinase kinase). Once activated, MAPK typically will activated specific gene expression programs via activation of transcription factors (Figure 5.10). In the case of the HER2 linked pathway ‘Raf’ is the ‘gatekeeper’ for this specific signalling cascade and the binding of RAS-GTP opens a binding pocket with a kinase domain. Raf acts as a MAP3K (MAP kinase kinase kinase). Once activated Raf can then activate the next downstream element, a MAP2 kinase called MEK. MEK in turn can then activate the actual MAPK Erk. Erk activation leads to phosphorylation of transcription factors such as AP1 family members jun and fos which bind to DNA and trigger transcription of their target genes and subsequent proliferation.

Human receptor tyrosine kinase families typically have an extracellular receptor domain, a transmembrane domain, and a intracellular signal transduction domain.

Figure 5.10: Human receptor tyrosine kinase families typically have an extracellular receptor domain, a transmembrane domain, and a intracellular signal transduction domain.

5.6.4 RTK signalling via the PI3K/Akt pathway

An alternative way of signalling by RTKs in response to receptor binding (not required by HER2) and dimerisation is via the PI3K (Phosphoinositol3 kinase) pathway (Figure 5.11). PI3K binds to the (auto)phosphorylated RTK and then phosphorylates the membrane bound phospholipid PIP2 (phospho-4,5 inositol) to PIP3 (phospho-3,4,5 inositol). The PIP3 form of phosphoinositol allows proteins that possess a PH domain to bind leading to their co-localisation at the plasma membrane. The PIP3 thus allows binding and co-localisation of the proteins Akt and PDK1.

Binding of PIP3 to the PH domain of Akt also exposes a phosphorylation site which can be first activated by PDK1 (also bound to PIP3), followed by a second phosphorylation by a protein called PDK2 (also known as mTORC2). In this form Akt is fully active and able to phosphorylate downstream cytosolic and nuclear effectors such as e.g. mTOR. The active (PIP3) form of phosphinositol that allows binding is inactivated with the help of a phosphatase, PTEN(phosphatase and tensin homolog), that hydrolyses PIP3 to the inactive from, PIP2, and thus ends the initial signalling{#PTEN}. Thus PTEN is a important tumour suppressor for this pathway with clinical relevance in e.g. glioblastoma, breast cancer and in up to 70% of prostate cancers PTEN is only partially active.

Human receptor tyrosine kinase families typically have an extracellular receptor domain, a transmembrane domain, and a intracellular signal transduction domain.

Figure 5.11: Human receptor tyrosine kinase families typically have an extracellular receptor domain, a transmembrane domain, and a intracellular signal transduction domain.

5.7 Summary

Growth factors drive cell proliferation. Cell will not enter into the cell signal if there is not an appropriate balance of growth activating signals (+) and growth inhibiting (-) signals up to the moving beyond the restriction point R.

Growth factor pathway signalling is known to occur via either a cell surface receptor (RTKs) or by binding to an intracellular receptor l (steroid hormones).

RTKs come in many form (~ 20 subfamilies) and their function is transmit a signal (extracellular ligand concentration) to the inside of the cell where these signals eventually will be responsible for driving various gene expression programs. Some of these are linked to growth but other signalling pathways link to differentiation, angiogenesis, or apoptosis.

Even where a RTK is involved in growth / proliferation it is important to realise that there are a range of factor/receptors; this diversity and detailed regulation is important so that provide fine grained control over proliferation and make it possible for only specific cell types to grow in specific tissue at the desired time - rather than have all cells in the body grow at any time.

Typically RTKs are activated by binding of a ligand - the exception is HER2 which is an orphan receptor that can form various homo- and heterodimers. After ligand binding (apart from HER2) and dimerisation RTKs relay the signal to the cytosolic side of the membrane. Within the cell HER2 is involved with MAPK signalling (dimers of HER2-EGFR, HER2-HER2) and PI3K signalling (HER2-HER3) to eventually target the appropriate transcription factor for activation of a specific gene expression program.

As for other similar pathways there a number of steps (signalling nodes) involved in each case and mis-function (e.g. constant “on” or “off” signalling) of any node could lead to inappropriate and sustained activation of proliferation.

Signalling pathways are not typically working as a simple chain but rather form part of a wider network which extensive cross-talk.