Molecular basis and clinical application of targeted therapy in oncology

Abstract

Targeted therapy and precision oncology aim to improve efficacy and minimize side effects by targeting specific molecules involved in cancer growth and spread. With the advancements in genomics, proteomics, and transcriptomics with the accessible modalities such as next-generation sequencing, circulating tumor cells, and tumor Deoxyribonucleic Acid (DNA), more number of patients are being offered the targeted therapy in form of monoclonal antibodies and various intracellular targets, specific for their tumor. The harnessing of host immunity against the cancer cells by utilizing immune-oncology agents and chimeric antigen receptor T-cell therapy has further revolutionized the management of various cancers. These agents, however, have the challenge of managing the adverse effects that are peculiar to the class of drugs and very different from the conventional chemotherapy. This review article discusses the molecular basis, diagnostics, and use of targeted therapy in oncology.

Introduction

Targeted therapy is a type of cancer treatment that focuses on specific molecules or proteins involved in the growth and spread of cancer cells. Precision oncology uses genetic information to identify the best treatment options for each individual patient, leading to personalised therapy. This approach aims to improve the effectiveness of treatment while minimising side effects, as it targets specific pathways involved in cancer growth and progression.¹

The introduction of imatinib mesylate as a treatment for chronic myelogenous leukemia (CML) in 2001 was a breakthrough in the field of precision medicine. This drug, also known as Gleevec, is a tyrosine kinase inhibitor (TKI) that targets the Bcr-Abl protein produced by the Philadelphia chromosome (t(9,22)) present in CML cells. This targeted therapy effectively blocks the activity of the Bcr-Abl protein, which is responsible for the uncontrolled growth of CML cells. The use of imatinib mesylate resulted in a near-normal life expectancy for patients with CML, who previously had a very poor prognosis.^2,3

The sequencing of the human genome in 2001 also played a major role in the development of targeted therapy and precision medicine. This milestone provided a comprehensive understanding of the genetic makeup of the human body and allowed for the identification of specific genetic mutations and variations that contribute to the development and progression of various diseases. This knowledge has allowed researchers to develop targeted therapies that specifically target the genetic changes present in a particular disease.

The knowledge of the human genome, combined with advances in technology, has led to a proliferation of precision medicine trials in recent years. These trials include the use of protein markers to identify specific genetic changes, molecular profiling to identify potential targets for therapy, customised combinations of treatments to target multiple genetic changes.^{4, 5, 6}

Genomics, proteomics, transcriptomics

The advances in genomics and the understanding of the immune system's role in cancer have led to the development of targeted therapies that are specific to certain molecular changes or biological characteristics. However, genomics has also shown that cancer is a complex disease that requires a shift in the way treatment is approached, moving away from a focus on tumour types and towards gene-directed, individualised treatment based on biomarker analysis for each patient.^{7, 8, 9}

Genomics has been a key foundation for precision medicine studies. It has allowed for the identification of specific genetic mutations and variations that contribute to the development and progression of various diseases and has led to the development of targeted therapies that specifically target these genetic changes. However, beyond genomics, RNA and protein profiling have also been shown to be important in mediating the biological impact of diseases.¹⁰

Proteins are the effectors of signalling in cells and play a crucial role in many biological processes. Protein assays, which measure the levels and activity of specific proteins, can provide valuable information about the underlying biology of a disease. However, matching patients to drugs based on genomics has been more effective in improving outcomes than matching on the basis of protein assays, possibly due to technical limitations in protein assays.¹¹ Despite these limitations, protein and transcript assays may still provide essential information when integrated with genomics. Recently, panels that incorporate immune signatures, based on Deoxyribonucleic Acid (DNA) and Ribonucleic Acid (RNA) and/or proteins, have also gained clinical significance. This approach allows for a more comprehensive understanding of the disease and can provide insights into potential targets for therapy.¹²

The study of RNA transcripts, called transcriptomics is the study and utility in understanding the causation of disease. High-throughput technologies such as microarrays and RNA sequencing are utilised in transcriptomic analysis and can be a valuable tool in identifying the right therapy for patients. The first solid tumour precision medicine trial that used transcriptomics in the clinic, the WINTHER trial, demonstrated that transcriptomics optimally increased the likelihood of patients to be matched to precision therapy. However, the use of transcriptomic biomarkers is currently limited by various challenges, such as degradation of RNA in formalin-fixed, paraffin-embedded samples, complexity of bioinformatic analysis, and reproducibility of the results.¹³

Next-generation sequencing (NGS) has shown that genomic changes in advanced cancers do not conform to the traditional categories defined by the organ where the tumour originated. Additionally, NGS has revealed that metastatic tumours have unique and complex genomic and immune profiles, which further emphasises the need for personalised treatment based on genomic analysis.^14,15 The cost and time required for sequencing have decreased significantly in recent years thanks to the technological advancements, making it more accessible for researchers to conduct these types of studies and for clinicians to use the information in the treatment of their patients.

Circulating tumour DNA and cells

Circulating tumour DNA testing, which stands for circulating tumour DNA, is a non-invasive method that is increasingly being used to select anti-cancer therapy and monitor treatment response. ctDNA testing is based on the detection of DNA that has been leaked into the bloodstream from a patient's tumour. Because ctDNA reflects the genetic makeup of the entire tumour and not just a single biopsy sample, it can provide a more comprehensive understanding of the tumour's genetic heterogeneity, which can be particularly useful in cases where the tumour has multiple subclones.¹⁶ ctDNA testing is also useful for monitoring treatment response, as it can detect changes in the levels of ctDNA in the bloodstream as the treatment progresses. This can provide important information about the dynamics of subclones within the tumour and help guide therapy decisions.¹⁷

Circulating tumour cells (CTCs) are cells that have detached from a primary tumour and are present in the bloodstream. The presence of CTCs has been independently associated with poorer survival outcomes in cancers like breast cancer and hormone-resistant prostate cancer (HRPC). For example, in a study of patients with untreated metastatic breast cancer, the number of CTCs in the bloodstream correlated with shorter progression-free survival and overall survival.^18,19 CTCs may also be utilised as a predictive biomarker for chemotherapy and immunotherapy. Studies have shown that the presence of CTCs in the bloodstream can predict whether a patient will respond to treatment or not. However, the use of CTCs in clinical practice has not yet been fully established, and more research is needed to understand the clinical utility of CTCs.²⁰ Serial CTC analyses, or the repeated measurement of CTCs over time, may also enable real-time surveillance of the disease.²¹

Clinically applicable targeted therapy in oncology

Currently, targeted therapy can be utilised in myriad ways in cancer. Some of the most common types of targeted therapies include monoclonal antibodies, antibody-drug conjugates (ADC) and TKI. The ability to harness host immunity against cancer cells has led to the development of newer modalities like immunotherapy and chimeric antigen receptor T cells (CAR-T) therapy in various cancers with no specificity for the cell or organ of origin of cancer.

Monoclonal antibodies

Monoclonal antibodies are laboratory-made proteins that mimic the immune system's ability to fight off harmful invaders. These antibodies can be designed to specifically bind to certain molecules, such as receptors on the surface of cancer cells or extracellular ligands to which these antibodies bind. By binding to these receptors or ligands, monoclonal antibodies can block the signals that convey growth and division, ultimately leading to the death of the cancer cells.

The extracellular ligands bind the receptors on the cell surface and guide cellular processes. Bevacizumab, a monoclonal antibody which binds the soluble vascular endothelial growth factor A (VEGF-A), preventing the interaction of VEGF-A with VEGF receptor and neo-vascularisation pathways. This drug has shown benefit in combination with chemotherapy or as single-agent maintenance in many cancers like colorectal, lung, renal, ovarian cancers, and glioblastoma multiforme.^{22, 23, 24, 25, 26}

Cell surface receptor targeting, one of the most widely used mechanism in precision oncology, has transformed the way many malignancies are treated. The cell survival is assisted by the cell surface receptors, which activate intracellular downstream signalling pathways upon ligand binding on the surface. Over expression or abnormal expression of receptors on the cancer cells can be targeted by using monoclonal antibodies against the receptors. This minimises the toxicity to normal cells. A common example is epidermal growth factor receptor [EGFR] family of receptors. Among the four members of the EGFR family, ErbB1 (HER1), ErbB2 (HER2/neu), ErbB3 (HER3) and ErbB4 (HER4), the ErbB2 [Her-2/neu] over expression in many cancers drives tumourigenesis. The role of overexpression of ErB3 (HER3) as a prognostic marker and as a potential resistance to anti-Her-2 therapy has been explored in trials.²⁷

While hormone receptor-positive breast cancer is often treated with hormone therapy, HER2-positive breast cancer is often treated with targeted therapies such as trastuzumab. About 20% of women with breast cancer have an amplified ERBB2/neu oncogene or overexpression of the HER2 receptor. Trastuzumab has been granted multiple approvals for use in breast cancer treatment (Fig. 1). However, resistance to the treatment, both intrinsic and acquired, has limited its effectiveness. Another HER2 antibody, pertuzumab, has been developed as a neoadjuvant treatment in combination with trastuzumab to reduce the risk of cancer recurrence.

Fig. 1.

The various anti-Her-2 therapies in clinical use and research. Shown are the monoclonal antibodies (A) tyrosine kinase inhibitors (B), –drug conjugates, (C) and other drugs like bi-specific monoclonal antibodies etc. TCR- T cell receptor.

EGFR directed therapy is also employed in treatment of colorectal cancers. A concept based on mutated or wild-type receptor, can be predictor of the use of the monoclonal antibodies. Constitutive activation of the downstream signalling pathway in the case of mutated EGFR receptor [RAS mutations negates the effect of surface receptor blockade. Hence, anti-EGFR therapy [cetuximab, panitumumab] is employed in clinical practice only in patients with wild-type status of EGFR.²⁸

Tyrosine kinase inhibitors

Tyrosine kinase inhibitors, also known as TKIs, are drugs that target enzymes called tyrosine kinases that are found in cancer cells. These enzymes play a critical role in the growth and spread of cancer cells, and TKIs work by binding to these enzymes and blocking their activity. This can lead to the cancer cells dying or being unable to grow and spread.

The landscape of management of lung cancer changed around the turning of the century when actionable mutations were discovered in patients of metastatic lung cancer. These patients were usually elderly women, never smokers and had adenocarcinoma as the histology. The presence of mutations in the intracellular domain of the EGFR receptor, in the exons 18, 19, 20, and 21, led to development of EGFR and TKI. These mutations were seen commonly in patients with Asian descent. In the landmark trial, IPASS, gefitinib (iressa) was shown to be more effective than chemotherapy in patients harbouring the mutations.²⁹ Since then numerous TKI have been developed against mutations or amplifications/over expression of many receptors in multiple cancers (ALK, ROS-1, BRAF, VEGF, MET, RET), which are in regular clinical use (Fig. 2).^{30, 31, 32, 33, 34}

Fig. 2.

Targets of various tyrosine kinase inhibitors and their mechanism of action in various malignancies. VEGF- Vascular endothelial growth factor; VEGFR- VEGF receptor; PDGF- Platelet-derived growth factor; PDGFR- PDGF Receptor; FGF- Fibroblast growth factor; FGFR-FGF Receptor; SCF- Stem cell factor; KIT- A type of proto-oncogene; GFL- Glial cell line-derived neurotrophic family ligands; HER-2- a member of epidermal growth factor receptor [ErbB2]; ALK- Anaplastic lymphoma kinase; EGFR- Epidermal growth factor receptor; Ras/Raf/MEK/MAPK and PI3K/AKT/mTOR- Downstream signalling pathways.

The paradigm of treatment of HER-2 positive breast cancer has been transformed with the discovery of TKI. Lapatinib, the first approved TKI to treat HER2-positive metastatic breast cancer was used in combination with capecitabine, a chemotherapeutic agent. Neratinib with capecitabine has also been approved for the treatment of advanced or metastatic HER2-positive breast cancer following two or more prior anti-HER2 therapies.³⁵ These inhibitors possess the ability penetrate the blood–brain barrier, providing clinical benefit in patients with brain metastasis.³⁶ In patients who have received prior anti-HER-2 therapy and have metastatic breast cancer with brain metastases, tucatinib, the new generation anti-HER-2 TKI, can be utilised in combination with trastuzumab and capecitabine.³⁷

There are numerous such TKIs in use in lung cancer, renal cell carcinoma, thyroid cancer, hepatocellular cancer, melanoma and in many other cancers.^38,39

Antibody–drug conjugates

Resistance to monoclonal antibodies is common in clinical practice. To overcome this, antibody–drug conjugates (ADCs) have been developed and certainly they are more effective.⁴⁰ These ADCs have the properties of cytotoxic chemotherapy and targeted therapy. ADC has these both molecules joined by a linker.

Brentuximab vedotin is the anti CD30 ADC, used for relapsed refractory Hodgkin lymphoma and anaplastic large cell lymphoma. It has also been used for other CD30 positive lymphomas. The structure of ADC is Fig. 1 and approved for clinical use ADCs are depicted in Table 1.⁴⁰ There are multiple ADCs under development for lung cancer.⁴¹

Table 1.

Antibody–drug conjugates.

ADC	Year approval	Antigen	MAb	Payload	Indications
Brentuximab vedotin	2011	CD30	Chimeric IgG1	Monomethyl auristatin F	Hodgkin lymphoma, ALCL
Trastuzumab emtansine	2013	Her2	Humanised IgG1	Derivative of maytansine	Breast cancer
Trastuzumab deruxtecan	2019	Her2	Humanised IgG1	Exatecan derivative	Breast cancer, gastric cancer
Sacituzumab govitecan	2020	TROP2	Humanised IgG1κ	Metabolite of topoisomerase 1 inhibitor irinotecan (SN38)	Triple negative breast cancer
Polatuzumab vedotin	2019	CD79b	Humanised IgG1	Monomethyl auristatin F	Diffuse large B cell lymphoma

Targeting intracellular pathways

Understanding of the numerous intracellular pathways have paved way for newer targeted therapies. These are often added on to other targeted therapies or are utilised stand alone. Examples of these include the mTOR, cyclic-dependent kinase (CDK) 4/6, and PIK3CA pathways. A drug named everolimus was approved in breast cancer patients who are hormone receptor positive but have developed resistance to it.⁴² Addition of this drug showed advantageous progression free survival benefit. Another mechanism of overcoming hormone receptor therapy resistance is inhibition of Rb phosphorylation. This leads to cell cycle arrest and reverses endocrine resistance. This class of drugs, CDK 4/6 inhibitors, block phosphorylation of retinoblastoma protein. Palbociclib, ribociclib, and abemaciclib are the approved CDK 4/6 inhibitors. They used in combination with endocrine therapy in the first-line in post-menopausal women or whose disease progressed even while on hormonal therapy.⁴³ PIK3CA mutations often occur in patients who progress while on endocrine therapy. Alpelisib, is a PIK3CA inhibitor which has shown improvement in progression frees survival of nearly 11 months when added to hormonal therapy.⁴⁴ The new drug showing a lot of promise in this class is capivasertib.⁴⁵

Immunotherapy

The Nobel Prize winning work of James Allison and Tasuko Honjo, has paved way for immunotherapy to be utilised in oncology. The class of drugs called immune checkpoint inhibitors (ICIs) or immuno-oncology agents (I–O) direct the host immunity towards the cancer cell. The cancer cells express neoantigens on their surface following various genetic and epigenetic changes in the cell, effectively triggering a T cell response. The host T cell immune response is normally regulated by immune checkpoints as a protective mechanism to prevent overactivity of the T cells to cause autoimmunity and an inflammatory state. The cancer cell expresses the ligands to these immune checkpoints, thereby preventing the T cells from activation. The immune checkpoint inhibitors blunt this ability of the cancer cell to evade host immunity and direct the T cells towards the tumour. However, the milieu and microenvironment of the tumour and the native tissue decides the efficacy of the ICIs in cancers. As a result, the balance between markers of response and resistance define the efficacy of the ICIs in various cancers (Fig. 3). High expression of the programmed death ligand [PD-L1], high tumour mutational burden and deficiency of mismatch repair genes are commonly used as biomarkers for use of ICIs in clinical practice. The use of ICIs is, as a result, tumour agnostic and has seen extensive research and development of molecules (Fig. 4).

Fig. 3.

The various biomarkers for response and resistance to immunotherapy. INF: Interfron; MDSC: Myeloid-derived suppressor cell; TAM: Tumour-associated macrophages; T-Reg: T regulatory.

Fig. 4.

The timeline of development of various immuno-oncology drugs in clinical practice.

Currently there are more than a dozen approved anti-PD1/PD-L1 directed drugs which are used extensively to treat patients with advanced non-small cell lung cancer (NSCLC), colorectal cancers, triple negative breast cancer, head and neck cancers and many others.

CAR-T cell therapy

Chimeric antigen receptor T cells [CAR-T Cells] are bio-engineered cells which have a tumour-associated antigen (TAA) binding domain, which is an extracellular hinge domain. In addition, a transmembrane domain, an intracellular domain combined with a with a costimulatory molecule expressed on a T cell complete the structure of the CAR-T cells.⁴⁶ The effect is specific. These tumour antigen directed T cells achieve precision killing of the cells expressing the antigen in a non-MHC restricted manner. The production, bio-engineering, expansion and transfusion of CAR-T cells is depicted in Fig. 5.

Fig. 5.

Figure depicting the process of CAR-T cell therapy. CAR- Chimeric antigen receptor.

The CAR-T cell therapy indications are rapidly expanding. At present, there are six approved CAR-T cell products by US-FDA. These are namely; axicabtagene ciloleucel, tisagenlecleucel, brexucabtagene autoleucel, lisocabtagene maraleucel, idecabtagene vicleucel, ciltacabtagene autoleucel. CAR-T cell therapy has potential to invoke the immune system against the cancer cells. Till date, CAR-T cells have received approval for treatment for relaped and refractory B cell lymphomas and acute leukemias; diffuse large B cell lymphoma, follicular lymphoma and mantle cell lymphoma. The response CAR-T cell therapy (idecabtagene vicleucel) studied in multiple myeloma has shown the benefits.⁴⁷ The field of CAR-T cell therapy is investigated in India by Narula G and Purwar R et al. for B cell malignancies and trials in India have entered in phase 2 state. As these CAR-T are very costly, the indigenously developed CAR-cell therapies will be boon for Indian patients as well as the other low- and middle-income countries. The initial success of CAR-T cells is largely limited to B cell lymphoid malignancies.

These newer therapies do come with the peculiar side effects apart from other transfusion related side effects. The cytokine release syndrome (CRS) and neurotoxicity are the two important and life threatening adverse effects. CRS manifests are fever, hypoxia, hypotension and invariably any other systemic involvement can be seen. Neurotoxicity presents are confusion, seizures and might lead to coma. Early diagnosis and treatment with steroids remains the cornerstone of treatment of these adverse effects.⁴⁸

Clinical trials of CAR-T cell therapies are being done in solid tumours like breast cancer, renal cell cancer, high grade glioma, prostate and lung cancer.⁴⁷ The CAR-T cell is being evaluated in T cell acute lymphoblastic leukemia in phase 1 clinical trial.⁴⁹ Human immunodeficiency virus (HIV) infection can be targeted by CAR-T cells on the ‘kick and kill’ strategy wherein the viral reservoir can be targeted for long term cure. Similarly, the other viral diseases can be the target of these smart CAR-T cells.⁴⁷ Despite all such advances, cancer cell remains so far immortal in many refractory diseases and permanent cure remains a challenge. Understanding the resistance mechanisms are important and on such basis newer and effective therapies are being developed. Strategies of overcoming the T cell exhaustion are being explored.⁵⁰

Conclusion

Targeted therapy is an exciting and promising area of cancer treatment that is showing great potential in the fight against cancer. By focusing on specific molecules or genes that are involved in the growth and spread of cancer cells, targeted therapy can be more specific and less toxic than traditional chemotherapy. The current challenge lies in optimal utilisation of the available modalities of targeted therapy for each patient—a concept of personalised precision medicine. The unique adverse effects of these drugs and their management remains a clinical challenge.

Disclosure of competing interest

The authors have none to declare.