The Complex Biology of Cancer Mutations

• The transformation of a normal cell to a cancer cell is a complex process resulting from the accumulation of phenotypic and genotypic alterations 
• Four major genetic alterations can contribute to cancer development: single-nucleotide variants (SNVs), copy number variants (CNVs), rearrangements (RE), and insertions and deletions (indels)
• Due to the constantly evolving nature of cancer cells and their mutations, cancer requires ongoing monitoring even after detection and during treatment

How does a normal cell transform into a cancerous cell? The transformation, driven by cancer mutations, is a complex biological process. The transition takes place over time with cells manifesting several signature phenotypic and genotypic alterations. In a normal cell, cell division is facilitated by several interacting genes through defined signal transduction pathways. In addition, mechanisms that control or regulate cell division also exist. The intricately maintained balance between cell proliferation and suppression sustains the normal functioning of cells.

Cancer cells, however, are typified by an imbalance between these two functions. In addition, over the course of cancer development, several characteristics are acquired by cancer cells. The typically understood characteristics, classified as major hallmarks of cancer, are: sustained proliferation, evading growth suppressors, activating invasion and metastasis, enabling immortality, inducing angiogenesis, resisting cell death, reprogramming of energy metabolism, and evading destruction by the immune system.¹ Such a display of complex characteristics involves accumulation of several alterations in genes (mutations). Understanding these mutated genes in cancer provide opportunities to develop novel ways to tackle the disease.

Genetic alterations that occur in germ cells (egg or sperm) are called germline mutations and are directly inherited by the progeny. Mutations occurring in any other cells of the body outside of germ cells are defined as somatic mutations and are not inherited by subsequent generations. Mutations occurring in developing somatic cells can give rise to populations of cells carrying the mutation. Depending on whether they occur in genes involved in growth and proliferation (oncogenes) or in growth suppression (tumor suppressor genes), they can exert diametrically opposite effects, but both favor unchecked development of cancer cells.

Cell growth is affected by the activation of cell surface receptors (such as receptor tyrosine kinases (RTK)), which in turn activate a series of phosphorylation events ultimately resulting in the expression of proteins, such as RAF, MEK, and ERK, which are responsible for growth, development and survival.² Mutations in these signal transduction genes can trigger development of different types of cancer.

Specifically, activating mutations in the RAS gene family—namely KRAS, NRAS, and HRAS—serve as key drivers in various malignancies; KRAS mutations are common in lung cancer, while NRAS and HRAS mutations are frequently identified in skin and bladder cancers, respectively.³ Mutations in these genes result in constitutive activation of the RAF-MEK-ERK signal transduction pathway, leading to uncontrolled cell division and growth. These are called activating mutations and they lead to a gain of function for the protein.

Conversely, other types of mutations, namely inactivating mutations, could result in loss of function of proteins, leading to an entirely different outcome. For example, the tumor suppressor gene TP53 has anti-proliferative functions and is involved in shutting down stressed or damaged cells. It is responsible for deciding the fate of damaged or stressed cells. It decides if a damaged DNA fragment has to be repaired or if the cell should be sent through a programmed cell death (apoptosis) pathway to prevent the damaged cells from posing a danger to normal cells, thus providing a cellular brake on cancer development. Deactivating mutations that result in a loss of function of TP53 render this brake nonfunctional, thus making cells vulnerable for cancer development and metastasis.⁴

From single base-pair alterations to complete chromosomal rearrangements, a range of genetic alterations result in cancer. The four major and significant ones are described below.

Mutations in a single nucleotide (SNVs) are point mutations that can cause missense, nonsense, or synonymous (silent) substitutions. While the majority of silent mutations may have no perceptible effect, missense or nonsense SNVs can function as activating mutations that drive oncogenesis.

For example, a single nucleotide mutation resulting in substitution from valine to glutamic acid at codon 600 (V600E) in the BRAF gene accounts for about 90% of the BRAF mutation type in melanoma.⁵ Targeting melanoma with inhibitors of BRAF mutants and employing companion diagnostics approaches that include testing for BRAF mutations and determining the appropriate treatment plans are some of the options available for melanoma. Inhibitors of BRAF mutants, such as vemurafenib and debrafenib, have been developed for targeting melanoma.⁶

Another SNV of significance is an amino acid substitution in exon 21 of leucine to arginine (L858R) in the tyrosine kinase domain of the epidermal growth factor receptor (EGFR). This mutation significantly increases the efficacy of tyrosine kinase inhibitors (TKIs) like gefitinib, erlotinib, and afatinib in advanced non-small cell lung cancer (NSCLC).⁷

Copy number variants are quantitative alterations in the genome characterized by the gain (duplication/amplification) or loss (deletion) of DNA segments. These variations lead to a change in the number of copies of specific genes, which can result in their differential expression. In oncology, CNVs are significant drivers because they can lead to the overactivation of oncogenes or the functional silencing of tumor suppressor genes. For example, the amplification of the ERBB2 (commonly known as HER2) gene is a critical copy number gain observed in approximately 20% of breast cancers. This CNV serves as a primary biomarker for eligibility for HER2-targeted therapies.⁸

Chromosomal rearrangements occur when a segment of a chromosome is transferred to a different part of the same chromosome or to an entirely different chromosome. These structural events often lead to the shuffling of genetic material, which can result in the creation of gene fusions. Gene fusions occur when portions of two previously separate genes join together to create a new, chimeric gene that typically leads to uncontrolled cell signaling.

An example of this is the Philadelphia chromosome, made of a fusion between breakpoint cluster region gene (BCR) and Abelson murine leukemia viral oncogene homolog 1 (ABL1) gene, found in more than 95% of chronic myeloid leukemia.⁹ Similarly, fusion between the anaplastic lymphoma kinase (ALK) and the echinoderm microtubule-associated protein-like 4 (EML4) gene is frequently identified as a driver in NSCLC.¹⁰

Insertions and deletions (indels) involve the addition or loss of nucleotides within the DNA sequence. When the number of nucleotides involved is not a multiple of three, these result in frameshift mutations, which alter the entire downstream amino acid sequence and frequently introduce premature stop codons. This leads to the production of truncated proteins that are typically non-functional or rapidly degraded.

However, indels can also be in-frame, where the reading frame remains intact but the protein's structure and function are significantly altered. Along with SNVs, indels are among the most frequently observed somatic mutations in oncology, often occurring within critical domains of oncogenes and tumor suppressors.^11,12

During the course of establishing themselves, cancer cells exploit all resources in their environment to their advantage and acquire several new mutations. Of these, a few mutations, called the driver mutations, occur in oncogenes and tumor suppressor genes and facilitate tumor growth and proliferation. The adaptive significance provided by these driver mutations is crucial for cancer progression.¹³

An example is the T790M mutation in EGFR. During TKI therapy in NSCLC, cancer cells may acquire this specific driver mutation to develop resistance to first- and second-generation drugs.¹⁴ Identifying this mutation is critical for transitioning patients to alternative treatment options. In contrast, passenger mutations occur alongside drivers but do not confer a direct growth advantage or modify the tumor’s progression rate.¹⁵ They are essentially "byproducts" of the genomic instability inherent in cancer cells.

Cancer is a complex disease. Cancer cells acquire several mutations over the period of their existence. Cancer development and progression is conferred by processes akin to a Darwinian evolution. These involve continuous acquisition of genetic variation due to randomly occurring cancer mutations and selection of genetically advantageous variations by natural selection in the tumor environment. This selection eliminates those that are deleterious for cancer growth and selects those that have accumulated enough beneficial variations that enable the cancer cells to thrive.

This constantly evolving nature of cancer cells emphasizes the need for constant monitoring of cancer, even after detection and during treatment. Non-invasive approaches such as liquid biopsy, where cancer mutations can be detected from blood samples, enable residual disease detection, disease progression monitoring, and comprehensive genomic profiling.