HC13: Heterogeneity in cancer

Types of heterogeneity

In cancer, there are 3 types of heterogeneity:

• Inter-patient: between patients
• Inter-tumor: between tumors
• Intra-tumor: within the tumor

Examples are:

• Well/moderately differentiated versus poorly differentiated
• Tumors with a strong T-cell infiltration versus with a poor T-cell infiltration
• Cancer cells are different when they are at the core than when they are at the invasive margin
  • Because the environment is different
    • For example the amount of immune cells or the access to oxygen
• The more clones there are in a tumor, the more likely the chance that 1 is able to metastasize

Heterogeneity forms a major issue for the treatment of cancer.

Intra-tumor and inter-tumor heterogeneity

Intra-tumor heterogeneity:

There are 2 origins of intra-tumor heterogeneity:

• Every time a tumor cell divides, it is an opportunity for a new clone to arise
• DNA repair deficient tumors are likely to be more clonal heterogenous

The fact that a new clone evolves, doesn’t mean it will survive:

• It has to compete with other clones
  • Competition for nutrients/oxygen
  • Proliferation
• It has to evade the immune system

Inter-tumor heterogeneity:

Inter-tumor heterogeneity is a morphological heterogeneity between tumors → one tumor is well/moderately differentiated while the other is poorly differentiated.

Heterogeneity becomes clear when the protein expression is observed. For instance, 1 tumor can have high levels of PD-L1 expression while the other doesn’t. Tumor cells which are negative for PD-L1 respond negative to the treatment.

Not only the tumor cells present in tumors can differ, but also other cells, for instance immune cells.

Origins of cancer cell heterogeneity:

Heterogenic tumors arise as follows:

1. Epigenetic changes occur during cell division
2. A clone that is able to evade the immune system survives and keeps dividing
3. The clone keeps getting better at evading the immune system

Of all the clones, 1 clone can remain resistant to therapy.

Clonal cooperation:

Clonal cooperation refers to characteristics of cancer provided by distinct cancer cell clones. Different clones, which all are present in the same tumor, have different functions:

• Clone A: tumor proliferation
• Clone B: angiogenesis
• Clone C: invasion

Clonal competition:

Aside clonal cooperation, there is competition between cancer cells. Cells compete for nutrients, space and oxygen. Many clones that are generated during tumor progression do not survive. This competition is driven by selective pressure, which is driven by either external factors or intrinsic features of cancer cells:

• Immune recognition
• Competition of nutrients/oxygen
• Inter-clonal competition

Inter-patient heterogeneity

Different genes are mutated in different cancers. Only TP53 mutations occur in nearly every type of cancer. The large majority of mutated genes differs considerably from one tumor to the other tumor type. They play different roles in healthy tissues. This inter-patient heterogeneity has consequences for the treatment.

An example is HER2, which is mainly expressed in breast and urothelial cancer. These cancer types can be treated with trastuzumab.

Different mutation profiles:

Inter-patient heterogeneity can also occur in the same cancer type. In case of colorectal cancer, patients with a KRAS mutation cannot have a BRAF mutation and vice versa. They are mutually exclusive. This is caused by KRAS and BRAF both being oncogenes which are in the same pathway. In case of lung cancer, mutual exclusivity is visible between KRAS, BRAF and EGFR. Activation of more than 1 of these genes would lead to an excess of signaling, which no longer forms an advantage for cancer cells.

Identification

Intra-tumor heterogeneity can be identified by cutting a section of the tumor into 5 different regions (region A-E). With these biopsies, multiple things can be done:

• Next generation sequencing
• Histology
• Cell culture
• Xenotransplantation

An example of possible results is:

• TP53 mutation in region D
• EGFR amplification in region C
• PTEN mutations in both regions

In this case, PTEN was mutated first → the mutation that is present in all the clones is probably mutated first. Different clones in the primary tumor may leave and metastasize at different sites.

Driver and passenger genes

There are several differences in driver and passenger genes:

• Driver genes
  • High mutation frequency
  • Have important characteristics → are selected for their mutations and divide
  • Likely important and form the initiative of cancer formation
  • Only a small fraction of all genes
• Passenger genes
  • Low mutation frequency
    • Exceptions: some very big genes
  • The larger the gene, the higher the chance of acquiring mutations
  • Occur randomly

The same passenger genes usually aren’t found in different tumors because they mutate randomly and aren’t significant. However, there are some exceptions → in some cases, passenger genes can be found in relatively high sequences. This can be explained by their size:

• TP53: 2500 nucleotides
  • Normal mutation frequency: 54/2500 = 0,0216
• FAT4: 16000 nucleotides
  • Normal mutation frequency: 17/16000 = 0,001

In conclusion, TP53 mutates more than 21x as much as FAT4. Furthermore, mutations in TP53 are often truncating while mutations in FAT4 are often missense. Even though passenger mutations dominate in number, they usually do not mean anything.

Prognosis

Tumors with more mutations correlate with improved survival in colorectal cancer. Depending on which type of alterations are present in tumor cells, survival rates differ:

• Patients with POLE and POLD1 mutants have the best survival chances
  • These are polymerase mutants
• Mismatch repair deficient patients have a reasonably good prognosis
• DNA replication repair proficient patients have the worst prognosis

The same counts for breast cancer. Because POLE and POLD1 mutants are more often mutated, it is more likely that new antigens are presented → a strong immune response is more easily triggered, making the tumor easier to be removed.

Treatment

It is important to assess different features of different patients in order to prescribe them with the best treatment:

• 5-FU treatment
  • MMR proficient cancer patients are sensitive to 5-FU treatment → 5-FU is inserted in DNA, causing the tumors to be destroyed
  • MMR deficient patients don’t respond very well to 5-FU
• Cituximab
  • Cituximab is an example of a EGFR blocker
  • If the EGFR receptor is amplified it can be targeted with antibodies and inhibitors to activate the pathway
  • KRAS doesn’t need EGFR for its development → there is no use in putting a EGFR blocker at the start of the pathway

Inter-tumor heterogeneity:

In case of inter-tumor heterogeneity, treatment is difficult. A certain drug may only be effective for 1 tumor → the other tumor(s) can still metastasize.

Intra-tumor heterogeneity:

There is also selection induced by treatment → treatment can be a very strong form of selective pressure. For instance, EGFR mutant lung cancer which carriers an EGFR exon 19 deletion is treated with a drug which targets this type of mutation. However, if clones with different EGFR mutations which are not sensitive to the treatment are also present, these clones start to spread. EGFR exon 19 deletion activates the other clone → deletion leads to activation.A clone that isn’t good at competition can grow when another clone is killed by therapy.

Ideally, the primary mutation should be targeted in therapy → is present in all the different clones. This almost never is the case in real life.