Hallmarks of Cancer

Cancer Background

Despite the many recent advances in our understanding and treatment of the disease, cancer is still among the leading causes of death worldwide. In 2018, there were 18.1 million new cases and 9.5 million cancer-related deaths worldwide, with cancer being the second leading cause of death in the United States of America.

• About one-third of all people in the US will develop cancer during their lifetimes.

• By 2040, the number of new cancer cases per year is expected to rise to 29.5 million and cancer-related deaths to 16.4 million.

Generally, cancer rates are highest in countries with the longest life expectancy, education level, and living standards, such as the US, Canada, and the UK. However, for some cancer types, such as cervical cancer, the reverse is observed.

Cancer: Causes and Treatments

Cancer is caused by a wide range of environmental factors, leading to DNA damage, hereditary and in utero mutations.

• Environmental factors, such as smoking, excessive alcohol consumption, and radiation exposure, contribute to an individual's lifetime risk of developing cancer.

• Inherited genetic mutations play a significant role in around 5 -10%of all cancers.

More than 50 mutations in specific genes have been linked to hereditary cancer syndromes.

Hallmarks of Cancer

The canonical hallmarks of cancer comprise six physical capabilities acquired during the multistep development of human tumors, built on a foundation of genomic instability and inflammation. Research and increased understanding of cancer in the last decade have provided two emerging hallmarks that contribute to establishing the core capabilities-reprogramming of energy metabolism and evading immune destruction. In addition to cancer cells, tumors exhibit another dimension of complexity: they contain a repertoire of recruited, ostensibly normal cells that contribute to the acquisition of hallmark traits by creating the "tumor microenvironment." Recognition of these concepts' widespread applicability increasingly affect the development of new means to treat human cancer.

Fig. 1. The Hallmarks of Cancer. Source: Hanahan and Weinberg, 2011

Sustaining Proliferative Signaling

A fundamental trait of cancer cells is their ability to support chronic proliferation. Healthy tissues carefully regulate the cell growth-and-division cycle, ensuring homeostasis and standard tissue architecture and function. In contrast, cancer cells deregulate these signaling pathways to maintain their survival. Growth factors (GF) that bind to cell-surface receptors containing intracellular tyrosine kinase domains are predominantly affected. This leads to the upregulation of intracellular signaling pathways that regulate cell cycle progression and growth.

Fig. 2. The deregulation of signaling pathways.

Cancer cells sustain proliferative signaling in several alternative ways:

Evading Growth Suppressors: Numerous tumor suppressors are involved in limiting cell growth and proliferation. They are often discovered through their characteristic inactivation in cancer cells. The two canonical tumor suppressors are the RB (retinoblastoma-associated) and TP53 proteins; they are essential regulators that govern apoptosis, proliferation, and senescence in cells.

• In-house production of GF ligands and cognate receptors, leading to autocrine proliferative stimulation.

• Cancer cells may stimulate normal cells within the surrounding stroma, subsequently supplying the cancer cells with various growth factors.

• Increased GF receptor expression, rendering cells hyperresponsive.

• Structural alterations in the receptor molecules that increase ligand-independent activation.

• GF independence may arise from constitutive down-stream activation of signalling pathways, negating the need to stimulate these pathways via ligand-mediated receptor activation.

• The RB protein integrates signals from diverse extracellular and intracellular sources, acting as a critical gatekeeper of cell-cycle progression, its absence can result in persistent cell proliferation.

• TP53 receives input from intracellular stressors DNA damage, suboptimal growth conditions and homeostatic imbalances can lead to TP53 halting cell-cycle progression until conditions are favourable, or initiate apoptosis if cellular damage is beyond repair.

• The nuanced effects of activated TP53 are highly context dependent, varying by cell type as well as the severity of cellular stress and DNA damage.

Interestingly, chimeric mice populated with RB null cells do not demonstrate proliferative abnormalities- the only neoplasia observed were pituitary tumors later in life.¹ TP53 null mice also develop typically, showing normal cellular and tissue homeostasis, but again develop cancers later in life, such as leukemia's and sarcomas.²

Activating Invasion and Metastasis: A well-characterized alteration to aid invasion and metastasis is the loss of E-cadherin by carcinoma cells. E-cadherin aids the assembly of epithelial cell sheets and maintains the cells' quiescence within these sheets by forming adherent junctions with adjacent epithelial cells. The frequently observed downregulation and occasional mutational inactivation of E-cadherin in human carcinomas provided strong support for its role as a viral suppressor of this hallmark capability.³ Another important driver of cancer metastasis is the loss of Contact inhibition. Contact inhibition ensures healthy, noncancerous cells cease proliferation and growth when they come into contact with each other. This characteristic is lost when cells undergo malignant transformation, leading to uncontrolled proliferation and solid tumor formation.⁴

Contact inhibition can be achieved in a variety of ways:

• One mechanism involves the protein Merlin (coded by the NF2 gene), a tumour suppressor, which when absent results in the hereditary condition neurofibromatosis type 2.

• Merlin regulates contact inhibition via coupling cell-surface adhesion molecules (e.g., E-cadherin) to transmembrane receptor tyrosine kinases (e.g., the EGF receptor). In doing so, Merlin strengthens cadherin-mediated cell-to-cell attachments.

• Sequestering growth factor receptors, Merlin limits the capability of cancer cells to efficiently emit mitogenic signals.

• The second mechanism of contact inhibition requires the LKB1 epithelial polarity protein, involved in epithelial structure organisation and maintaining tissue integrity.

• LKB1 has been shown to negate the mitogenic effects of the Myc oncogene when it is upregulated in organized, quiescent epithelial structures.

• When LKB1 expression is reduced, epithelial integrity is compromised, making epithelial cells more susceptible to Myc-induced transformation.

Expression of other adhesion molecules is notably altered in many carcinomas, with those involved in cytostasis being downregulated. Adhesion molecules generally associated with the cellular movement during embryogenesis and inflammation are predominantly upregulated. For example, N-cadherin, typically expressed in migrating neurons and mesenchymal cells during organogenesis, is upregulated in many invasive cancer cells.⁵ Research shows that cell-to-cell contacts formed by dense populations of healthy cells propagated in 2D cell-culture seek to suppress further cell proliferation, presenting as confluent cell monolayers. Conversely, in vitro contact inhibition is absent in numerous cancer types, suggesting that contact inhibition is an in vitro analogous mechanism operating to maintain tissue homeostasis.

Enabling Replicative Immortality: Cancer cells require unlimited replicative potential to form macroscopic tumors, bypassing the Hayflick limit observed in healthy cells while avoiding programmed cell death. Telomeres and telomerase play a crucial role in this hallmark of cancer.

Fig. 4. Telomeres and Cancer.

Research suggests that cancer cells often experience telomere loss-induced crisis relatively early during multistep tumor progression due to their inability to express significant telomerase levels. Extensively eroded telomeres have been observed in premalignant growths, along with end-to-end chromosomal fusions, suggesting that cancer cells have passed through a substantial number of successive telomere-shortening cell divisions during their development from healthy cells before the acquisition of telomerase activity.⁶

Inducing Angiogenesis: Cancer cells, like healthy cells, need nutrients and oxygen and the capacity to remove metabolic wastes and carbon dioxide for survival; tumor-associated vasculature formed through angiogenesis caters to these requirements. During tumor development and progression, an "angiogenic switch" is almost always activated and remains on, causing normally quiescent vasculature to form new vessels that help sustain expanding neoplastic growth.⁷

Fig. 5. Cancer Cell Angiogenesis

The most prominent prototypes of angiogenesis induction and inhibition are vascular endothelial growth factor-A (VEGF-A) and thrombospondin-1 (TSP-1)

The past decade has seen significant focus on angiogenesis. Amid this wealth of new knowledge, other proangiogenic signals, such as members of the fibroblast growth factor (FGF) family, have been implicated in sustaining tumor angiogenesis when their expression is significantly upregulated.⁸ Historically, angiogenesis was thought only to be relevant for the formation of rapidly growing tumors. However, recent investigation indicates that angiogenesis also plays a fundamental role in the premalignant phase of neoplastic progression.

Apoptosis

Apoptosis is the process of programmed cell death characterized by distinct morphological characteristics and energy-dependent biochemical mechanisms. Apoptosis is considered a vital component of processes including normal cell turnover, proper development and functioning of the immune system, hormone-dependent atrophy, embryonic development and chemical-induced cell death.

Fig. 6. VEGF Signaling Pathway

Resisting Cell Death: It is well established that programmed cell death via apoptosis serves as a genetic defence mechanism against cancer development. The apoptotic machinery is composed of both upstream regulators and downstream effector components. Regulators are subdivided into two primary circuits:

1. The extrinsic program is involved in receiving and processing extracellular death-inducing signals.

2. The intrinsic program, responsible for sensing and integrating intracellular signals.

Each program culminates in the activation of a predominantly latent protease (caspases 8 or 9), which proceeds to initiate a cascade of proteolysis involving effector caspases responsible for the execution phase of apoptosis, in which the cell is progressively disassembled and then consumed, both by its neighbors and by phagocytic cells. Currently, the intrinsic apoptotic program is more widely implicated as a barrier to cancer pathogenesis.⁹

Fig. 7. Apoptosis Pathway

Apoptotic cells undergo morphological changes involving extensive plasma membrane blebbing followed by karyorrhexis. Apoptotic bodies are formed by separation of cell fragments during a process called “budding.” They consist of cytoplasm with tightly packed organelles with or without a nuclear fragment. The organelle integrity remains enclosed within an intact plasma membrane. These bodies are subsequently phagocytosed by macrophages, parenchymal cells, or neoplastic cells and degraded within phagolysosomes. There are three main types of biochemical changes observed in apoptosis:

• Activation of caspases.

• Breakdown of DNA and protein.

• Membrane changes and recognition by phagocytic cells.

Each program culminates in the activation of a predominantly latent protease (caspases 8 or 9), which proceeds to initiate a cascade of proteolysis involving effector caspases responsible for the execution phase of apoptosis, in which the cell is progressively disassembled and then consumed, both by its neighbors and by phagocytic cells. Currently, the intrinsic apoptotic program is more widely implicated as a barrier to cancer pathogenesis.¹⁰

Cancer cells have evolved a variety of strategies to limit or circumvent apoptosis:

• The most common mechanism involves losing the TP53 tumor suppressor function, eliminating a critical damage sensor from the apoptosis-inducing circuitry.

• Cancer cells may increase anti-apoptotic regulator expression, such as Bcl-2, Bcl-xL, or the survival signals Igf1/2 via downregulating pro-apoptotic factors (Bax, Bim, Puma).

• Alternatively, tumors can also short-circuit the extrinsic ligand-induced death pathway to avoid apoptosis.

The variety of apoptosis-avoiding mechanisms presumably reflects the diversity of apoptosis-inducing signals that cancer cell populations encounter during their evolution to the malignant state. In addition to apoptosis, necrosis also plays a role in resisting cell death; cell death by necrosis appears to be under genetic control in some situations, rather than being a random and uncoordinated process.^{11 12}

Necrotic cell death releases proinflammatory signals into the surrounding tissue microenvironment, allowing necrotic cells to recruit inflammatory cells. Evidence suggests that immune-inflammatory cells can be actively tumor-promoting in the context of cancer, as these cells can foster angiogenesis, cancer cell proliferation, and invasiveness. Furthermore, necrotic cells can release bioactive regulatory factors, such as IL-1α, which can directly stimulate neighboring viable cells to proliferate, with the potential to facilitate cancer progression.¹³

Conclusion

The hallmarks of cancer comprise six biological capabilities acquired during the multistep development of human tumors. These hallmarks constitute an organizing principle for rationalizing the complexities of cancer development and progression. From sustaining proliferative signaling to avoiding cell death, genome instability generates the genetic abnormalities required for multiple hallmark functions. The Conceptual progress of the hallmarks of cancer in the last decade has presented two further hallmarks, the reprogramming of energy metabolism and evading immune destruction. In addition to cancer cells, tumors exhibit another dimension of complexity: they contain a repertoire of recruited, ostensibly normal cells that contribute to the acquisition of hallmark traits by creating the "tumor microenvironment." Recognition of the widespread applicability of these concepts will increasingly affect the development of new means to treat human cancer.

1. Lipinski MM, Jacks T. The retinoblastoma gene family in differentiation and development. Oncogene. 1999 Dec 20;18(55):7873-82. doi: 10.1038/sj.onc.1203244. PMID: 10630640.

2. Ghebranious N, Donehower LA. Mouse models in tumor suppression. Oncogene. 1998 Dec 24;17(25):3385-400. doi: 10.1038/sj.onc.1202573. PMID: 9917000.

3. Berx G, van Roy F. Involvement of members of the cadherin superfamily in cancer. Cold Spring Harb Perspect Biol. 2009 Dec;1(6):a003129. doi: 10.1101/cshperspect.a003129. Epub 2009 Sep 23. PMID: 20457567; PMCID: PMC2882122.

4. Pavel, M., Renna, M., Park, S.J. et al. Contact inhibition controls cell survival and proliferation via YAP/TAZ-autophagy axis. Nat Commun 9, 2961 (2018). https://doi.org/10.1038/s41467-018-05388-x

5. Cavallaro U, Christofori G. Cell adhesion and signalling by cadherins and Ig-CAMs in cancer. Nat Rev Cancer. 2004 Feb;4(2):118-32. doi: 10.1038/nrc1276. PMID: 14964308.

6. Cleal K, Norris K, Baird D. Telomere Length Dynamics and the Evolution of Cancer Genome Architecture. Int J Mol Sci. 2018;19(2):482. Published 2018 Feb 6. doi:10.3390/ijms19020482.

7. Hanahan D, Folkman J. Patterns and emerging mechanisms of the angiogenic switch during tumorigenesis. Cell. 1996 Aug 9;86(3):353-64. doi: 10.1016/s0092-8674(00)80108-7. PMID: 8756718.

8. Baeriswyl V, Christofori G. The angiogenic switch in carcinogenesis. Semin Cancer Biol. 2009 Oct;19(5):329-37. doi: 10.1016/j.semcancer.2009.05.003. Epub 2009 May 29. PMID: 19482086.

9. Adams JM, Cory S. The Bcl-2 apoptotic switch in cancer development and therapy. Oncogene. 2007 Feb 26;26(9):1324-37. doi: 10.1038/sj.onc.1210220. PMID: 17322918; PMCID: PMC2930981.

10. Adams JM, Cory S. The Bcl-2 apoptotic switch in cancer development and therapy. Oncogene. 2007 Feb 26;26(9):1324-37. doi: 10.1038/sj.onc.1210220. PMID: 17322918; PMCID: PMC2930981.

11. Galluzzi L, Kroemer G. Necroptosis: a specialized pathway of programmed necrosis. Cell. 2008 Dec 26;135(7):1161-3. doi: 10.1016/j.cell.2008.12.004. PMID: 19109884.

12. Zong WX, Thompson CB. Necrotic death as a cell fate. Genes Dev. 2006 Jan 1;20(1):1-15. doi: 10.1101/gad.1376506. PMID: 16391229.

13. Grivennikov SI, Greten FR, Karin M. Immunity, inflammation, and cancer. Cell. 2010 Mar 19;140(6):883-99. doi: 10.1016/j.cell.2010.01.025. PMID: 20303878; PMCID: PMC2866629.

• International Agency for Research on Cancer- https://www.iarc.who.int/

• Cancer Research UK- https://www.cancerresearchuk.org/

• Hallmarks of Cancer- Hanahan D, Weinberg RA. Hallmarks of cancer: the next generation. Cell. 2011 Mar 4;144(5):646-74. doi: 10.1016/j.cell.2011.02.013. PMID: 21376230.

• Innovative Approaches for Cancer Treatment- Pucci C, Martinelli C, Ciofani G. Innovative approaches for cancer treatment: current perspectives and new challenges. Ecancermedicalscience. 2019;13:961. doi: 10.3332/ecancer.2019.961. PMID: 31537986; PMCID: PMC6753017.