From Bench to Bedside with Targeted Therapies

Published on 09/04/2015 by admin

Filed under Hematology, Oncology and Palliative Medicine

Last modified 22/04/2025

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Figure 45-1 Paul Ehrlich: birth of targeted therapy, 1911.

For the next 40 years, great progress was made in developing additional chemotherapeutic anticancer agents. Two approaches were used. The first approach involved synthesis of specific targeted agents—as with methotrexate. The next success was 6-mercaptopurine, another inhibitor of DNA synthesis, produced by George Hitchings and Gertrude Elion and shown by Joseph Burchenal to be effective in acute leukemia. ⁴ The synthesis of new therapeutic molecules has been greatly enhanced by the development of three-dimensional models of target molecules, enabling chemists to design small lead molecules and test them using in silico computer-based screening.

The second approach involved screening of large numbers of natural products and led to the discovery of taxanes and camptothecin. These successes were followed by discovery of the anticancer properties of vinca alkaloids, platinum-based agents, nitrosoureas, and anthracyclines, all of which remain in use today.

The pharmaceutical industry has used these two general approaches—synthesis of new compounds and broad screening of natural products—to develop many therapies against cancer and bring them to the clinic for investigation in therapeutic clinical trials ⁵ (Figure 45-2 ). The successful chemotherapeutic agents that have been developed are summarized in Chapter 46. Today new methods of targeting and screening are being used to find therapies that counteract the function, or loss of function, of the products of aberrant genes that cause cancer (see later discussion).

Two other major breakthroughs in the development of cancer therapies during this first phase deserve emphasis. The first is the development of combinations of therapies administered simultaneously. This was applied to leukemia by Emil Freireich, Emil Frei, and James Holland in 1956, soon after combinations of antibiotics were found to produce enhanced efficacy against bacterial infections such as tuberculosis. The principle is to use two or more agents that provide additive killing capacities against a target, but with different toxic side effects that are not additive. In the late 1950s and early 1960s, this principle was first applied successfully in a series of clinical trials by these investigators and others for the treatment of childhood leukemia, ⁶ and it was first used for the successful treatment of a solid tumor, testicular cancer, by M. C. Li and colleagues. ⁷

Leukemia researchers also pioneered the idea that treatment must continue beyond the time that the cancer is clinically detectable, in order to prevent recurrence due to the persistence of subclinical disease. Today this is standard practice for the care of many types of cancer. As with the combination therapy studies, the results of leukemia research in murine models provided the rationale for these studies.

A second major breakthrough in cancer therapy and the treatment of many diseases took advantage of a new technology invented by Kohler and Milstein, which enabled production of large quantities of a monoclonal antibody raised against a specific antigen. ⁸ This technique was rapidly applied to the production of antibodies targeting molecules on the surface of cancer cells. Major clinical responses in lymphoma patients were reported by Ronald Levy and colleagues in 1982, with an anti-idiotype antibody against the specific immunoglobulin molecule expressed on the surface of the patients’ malignant B cells. ⁹ Today, nearly half of the agents that are approved by the U.S. Food and Drug Administration (FDA) for the treatment of cancer are monoclonal antibodies, a revolution in targeted cancer therapy that has occurred in the past three decades.

Figure 45-2 New approaches and challenges for personalized cancer medicine Therapy development is shown as linear process, whereas in reality it is iterative in all its stages.

The Second Phase

Why not produce an anticancer therapy that targets a molecule that is known to cause cancer? A preliminary to this approach dates back to the 1940s and 1950s, when Charles Huggins discovered that depriving prostate cancer patients of androgen and breast cancer patients of estrogen can result in remission of their disease. ¹⁰ Both approaches are used in the clinic to this day, but the method has evolved from surgical removal of organs producing these hormones to targeted drugs that act on hormone receptors or hormone production.

The target for therapy switched from cytoplasmic hormone receptors to growth factor receptors on the cell surface in the early 1980s. Our group first hypothesized that inhibition of a receptor function might inhibit tumor cell growth (Table 45-1 ). We produced a monoclonal antibody, cetuximab, which binds to the EGF receptor, blocks activation of the receptor’s tyrosine kinase by its ligands, EGF and TGFα, and inhibits proliferation of cancer cells in culture and in human tumor xenografts.^11–13This was followed by reports on trastuzumab, a monoclonal antibody that binds to the closely related HER-2 receptor. ¹⁴ It was during this period that the term oncogene was first used, and these were the first experimental cancer therapies that targeted the products of an oncogene—in these cases a receptor tyrosine kinase. Since the mid-1980s, many oral chemotherapeutic agents have been developed that target these two tyrosine kinases and many other kinases, located both in receptors and free in the cytoplasm.

Table 45-1

Rationale for Targeting EGF Receptors, 1980

Trastuzumab received regulatory approval for the treatment of breast cancer expressing high levels of HER-2 in 1998. ¹⁵ This was followed by imatinib against the product of the BCR-ABL gene translocation, another tyrosine kinase, in patients with chronic myelocytic leukemia in 2001. ¹⁶ Cetuximab entered clinical trials in 1990 and was approved for colorectal cancer in 2004^17,18and for head and neck cancer in 2006. These approaches to experimental therapeutics created a new paradigm for the development of therapies against cancer.

Three breakthroughs in knowledge and technology converged to greatly accelerate this change in therapeutic research on cancer:

(1) We have learned that accumulated genetic aberrations are the cause of cancer. This dates from observations by Bishop and Varmus in experiments exploring viral carcinogenesis with the SRC oncogene, published in 1976, showing that the genetic abnormality was intrinsic to human cells, not the Rous sarcoma virus. ¹⁹ Today, hundreds of genes are known to be altered in human cancers.

(2) There are more than 800 new drugs under development at pharmaceutical and biotechnology companies and universities, which are designed to target the products of the aberrantly functioning genes that cause cancer.

(3) New instruments and analytic techniques have reduced sequencing of a human genome from a 10-year, $3 billion project to a 10-day, $5000 project, which allows us to interrogate an individual patient’s cancer to detect genetic aberrations in the patient’s tumor. The speed and cost continue to improve dramatically.

As a result of this confluence of discoveries, there is pressure from all stakeholders in cancer care for development of new approaches to the treatment of cancer—using the genomic and molecular analysis of an individual patient’s cancer to select the targeted therapy that is most likely to provide clinical benefit. Although earlier approaches provided many successful chemotherapeutic agents and continue to be important, the new targeted approach to developing new cancer therapies dominates the field today. These new therapies are designed to target a particular gene product that has been shown to contribute to the malignant phenotype of cancer cells in correlative studies of molecular pathological data and clinical data and in laboratory experiments with cultured cells and animal models.^20,21

There have been some spectacular successes in clinical trials with experimental targeted therapies that enrolled only patients whose cancer was known to harbor an aberrancy in the target of that experimental drug or antibody. These successes have resulted in substantial prolongation of patients’ lives for many months or years.

The first example of this approach to reach the clinic involved trastuzumab, the monoclonal antibody against the HER-2 receptor. It is overexpressed on 20% to 25% of breast cancers and was found to be a biomarker predicting a worse prognosis in these patients. Clinical activity of trastuzumab was demonstrated in early studies only among patients whose cancers expressed high levels of HER-2. Genentech adopted a new paradigm for designing a randomized Phase III clinical trial, investigating standard chemotherapy with or without trastuzumab for treatment of patients with advanced, metastatic breast cancer. Only patients with high expression of HER-2 in their tumors were eligible to enroll. This enriched the trial with patients more likely to respond, and converted what would otherwise have been a negative study (too few responders) into a positive study, leading to FDA approval in 2002. ¹⁵ The optimal clinical situations for the use of trastuzumab have been refined continuously, as a result of dozens of clinical trials over the ensuing decade (see Chapters 36 and 50).

A second and most dramatic example involves the development of the ABL tyrosine kinase inhibitor imatinib for the treatment of patients with chronic myelocytic leukemia (CML), whose leukocytes nearly always carry a BCR/ABL gene rearrangement. Before this treatment, the median survival of patients with this disease was 3½ years. Today most patients on this and follow-up drugs live in remission more than 5 years, and some may reach a normal lifespan. ¹⁶ The incidence of CML is only 4000 cases per year. A drug against this disease was not considered to be a potential “blockbuster” compared with a treatment for lung cancer (over 200,000 cases/year). Movement from the research laboratory in a pharmaceutical company to clinical trials was encouraged by the persistent efforts of an academic clinician scientist, Brian Druker, who performed preclinical studies in his laboratory demonstrating efficacy against CML cells and then led the first clinical trials. The premise, which turned out to be prescient, was that blocking the abnormally active ABL tyrosine kinase—a product of the BCR-ABL rearrangement in patients with early CML—might produce especially effective responses, because this was the only known genetic abnormality in these patients. In contrast, most cancers have many genetic aberrations that contribute to their malignant activities at the time they are first detected.

A third example involves oral inhibitors of the EGF receptor tyrosine kinase, gefitinib and erlotinib (TKIs). In a series of very large and expensive randomized trials of chemotherapy plus or minus one or the other of these TKIs in patients with advanced lung cancer, the results showed no significant overall benefit from adding the TKI. However, a few patients had a substantial prolongation of survival. When it was discovered that mutations were present in the gene encoding the EGF receptor in the lung cancers of many of the patients who responded well,^22–24clinical trials were repeated with the requirement of an EGF receptor mutation for enrollment. The positive response rates for advanced lung cancer patients with a mutated EGF receptor were confirmed. ²⁵ This time, in the selected patient population, the results were positive in the majority of patients. Thus, a “failed” new targeted therapy was converted into a successful treatment for the 10% of lung cancer patients with mutated EGF receptor genes in their tumors, and the median duration of survival for those patients has doubled. In fact, for these patients the TKI therapy was more effective than chemotherapy, whereas for patients without the mutation, chemotherapy gave better outcomes than treatment with a TKI. ²⁵ This “resurrection of a cancer drug” brought clinical benefits to selected patients and economic benefits to the pharmaceutical companies, which were now able to market their new targeted drugs. This was a wake-up call to the industry, strongly suggesting the value of a new, selective approach to clinical trials with new targeted cancer treatments.

Among the lessons from these examples is the observation that for each case the circumstances were different, and an understanding of both the molecular biology and the clinical disease was critical in the development of a novel therapy that targeted a molecular abnormality in these patients’ cancers.

Von Hoff was the first to report on a clinical trial in which patient enrollment was governed by the result of a molecular analysis of tumors, using immunohistochemistry, fluorescence in situ hybridization, and, primarily, gene expression arrays. Of 66 patients treated with commercially available drugs that were felt to match the detected molecular markers, 27% had a progression-free survival at least 30% longer compared with the response to the previous drug(s) they had received. ²⁶ In this study, the patients served as their own controls, and gene mutations were not analyzed.

Recent examples have demonstrated the utility of identifying the targeted genetic aberration in the tumors of patients who enroll in clinical trials with experimental targeted therapies at the earliest stages.

The Biomarker-integrated Approaches of Targeted Therapy for Lung Cancer Elimination (BATTLE) trial for patients with advanced, heavily treated non–small-cell lung cancer was the first reported study with a panel of experimental drugs that (1) required a fresh tumor biopsy for genetic and molecular assays to detect particular aberrations that could serve as biomarkers for assigning therapy and (2) introduced an adaptive randomization design to assign a patient to receive one of the four experimental therapies that did, or did not, target an abnormality detected in that patient’s cancer. Essentially, this was a panel of four separate Phase II trials, testing the efficacy of “rationally” assigning patients to a particular trial. Importantly, nearly 100% of patients who were offered participation agreed to the biopsy procedure as a condition for receiving access to one of the experimental targeted therapies in a randomized study. The endpoint of the trial was improved duration of progression-free survival, which was subsequently confirmed by improved overall survival. The results showed that there were four situations where the results of the biomarker assays predicted a significant improvement in response, or in one case, a worse response, to an experimental drug targeting that biomarker. ²⁷ This suggested that preselection of patients for a panel of Phase II studies of new therapeutic agents based on biomarkers showing abnormalities in the targets of these agents is useful and may improve clinical outcomes.

Another trial at MD Anderson Cancer Center retrospectively analyzed response rates and survival of patients treated with experimental drugs that matched genetic abnormalities in their cancers. Over a 4-year period, more than 1000 patients with advanced, heavily treated cancers were tested for mutations in hot spots in 11 genes and for loss of expression of PTEN. An experimental drug was often available targeting the detected genetic aberration for most of their genes in ongoing Phase I/II trials. A total of 291 patients with genetic aberrations could be evaluated for response to an experimental targeted therapy. Of these, 175 received a targeted therapy that matched an aberrant gene target in their cancer, and 116 were treated with nonmatched therapy (based on nonavailability of matched therapy at the time). The overall response rate of the first group was 27% (including two complete responses), significantly greater than the second group, which was 5%. Median overall survival was 13.4 months compared to 9.0 months, a 49% increase. ²⁸ In most reports analyzing the results of Phase I trials, the main objective is determination of maximal tolerated dose and toxicities, and the objective response rate to new therapies is less than 10%. This trial, which achieved a much higher response rate, was a nonrandomized study and involved many types of cancer and many separate clinical trials of new drugs at varying doses. Although the results support the benefit of matching experimental therapy to genetic aberrations, this conclusion now must be confirmed in a randomized study. Today a number of cancer centers are carrying out prospective clinical trials using next-generation sequencing technology to assess cancers and using the results for assigning patients to trials of experimental targeted therapies.

Two recent highly successful Phase II trials of experimental targeted drugs took an approach to testing a new experimental therapy similar to that used in the registration trial of trastuzumab, the trials of TKIs in patients with mutated EGF receptors, and in most of the patients with CML treated with imatinib. Each trial studied a single drug. Large numbers of patients were screened for enrollment, based on the presence of the genetic aberration that the experimental drug was designed to target.

In the first clinical trial, crizotinib, which already was in trials targeting the EML4-ALK rearrangement in lymphoma, was tested for activity against patients with this rearrangement in their lung cancer. This was found to occur in less than 5% of lung cancers. More than 1700 patients were screened at multiple institutions to populate a Phase II trial with 82 preselected patients. The drug was successful, with a response rate of 57%, an additional 35% achieving stable disease, and a median time to progression of greater than 6 months at the time of publication. ²⁹ These results led to successful follow-up trials and FDA approval, within only 5 years of discovering this genetic aberration in small numbers of lung cancer patients (the drug was already available).

Another clinical trial explored the efficacy of vemurafenib which was designed to block the activity of BRAF, a gene that was reported to be mutated in many patients with melanoma. The complete plus partial response rate with advanced melanoma patients whose cancer had the targeted BRAF mutation was over 70%, with a median duration of 7 months. ³⁰ FDA approval followed soon afterward.

These were remarkable results in patients with advanced cancers that were well known to be minimally responsive or resistant to chemotherapy. It is clear that genomic screening can improve response rates with targeted therapies, benefiting patients and shortening the time required for testing an experimental therapy. However, we will not know how generalizable this conclusion is until the results of all such targeted trials—positive and negative—are reported and shared. Negative trials of experimental drugs are not typically reported, although they inform decision making by the pharmaceutical company that is developing them. Meanwhile, the results just described support pursuing this approach to testing targeted experimental therapies in a screened subset of cancer patients.

Next Steps in Drug Development

Progress will be hastened if a number of challenges are addressed—some involving gaps in scientific knowledge and technology, and others involving changes in the clinical trials process.

Scientific Knowledge and Technology

A major scientific challenge is the need to balance the tremendous power of next-generation genomic sequencing and other technologies for interrogating molecular aberrations in a patient’s cancer with the mandate to keep down the costs of clinical care. The most sophisticated genomic tests (see Chapter 24) add substantial costs for instrumentation and data analysis, although they may reduce the cost of care in the long run. An added factor is the requirement in the United States that all tests performed to guide clinical care must be carried out in laboratories with Clinical Laboratory Improvement Amendment (CLIA) certification, which ensures high standards, but adds costs. Similar standards are used in Europe. Finally, sophisticated, deep-level next-generation sequencing often identifies hundreds of genetic aberrations in human cancer specimens, ³¹ and most of these are not actionable because their significance in the tumor’s biology and the patient’s disease is not known, or drugs that target them are not yet available.

For these reasons, clinical testing for genomic aberrations in most medical centers currently is limited to assays that measure mutations (insertions, deletions, or substitutions of nucleotides) in selected genes, alterations in gene copy number, and structural rearrangements in selected genes ³² (Figure 45-3 ). Available experimental or approved drugs that target the products of aberrant genes are currently limited to a few dozen genes. At the author’s institution, we currently are sequencing 740 hot spots in 46 genes. This detects only the first of the three categories of genomic analysis listed above. To capture all three categories and expand the gene coverage, we are planning to move to the targeted deep sequencing of the exons (open reading frames) of a few hundred genes. Both of these approaches to sequencing enable multiplexed assays on many genes to be run on a single extracted sample of tumor DNA. This is a huge advance over the technology just a few years ago, when selected regions of genes were sequenced in a separate assay for each individual mutation, and the amount of tumor tissue available rapidly became a limiting constraint. A number of cancer centers have started programs assaying DNA in patient’s cancers to guide therapy (e.g., Refs. 33, 34).

Figure 45-3 The major classes of genomic alterations that give rise to cancer. CML, Chronic myelogenous leukemia; TS, tumor suppressor. ³²

With 800 experimental targeted drugs in the clinical pipeline and a few dozen already approved, current second-generation sequencing of advanced solid tumors yields a match between a genetic aberration and a targeted drug about 40% of the time. The yield is higher in reported cases of deep sequencing with more complete genomic analysis. The ingenuity of companies developing new technologies for detecting genetic aberrations guarantees that advancements will continue to enable more rapid and complete analysis for less cost. The limiting factors have become the need for bioinformatics and computational tools and for trained personnel to use these tools to interpret the sequencing data.

Two key rate-limiting gaps in scientific knowledge were identified recently in a report from the President’s Council of Advisors on Science and Technology. ³⁵ These were (1) identifying and validating the protein targets that are critical to the survival of cancer cells and are “druggable,” and (2) predicting more effectively the efficacy and toxicity of candidate drugs before high investments of funds and time are made in clinical trials.

The new fields of systems biology and computational biology have emerged to address the first of these questions. Through extensive computer-based analysis of data from biological studies on the biochemical steps that activate or suppress molecular processes in cells, investigators have been able to draw complex maps of interconnected pathways that regulate the activities of proteins in those cells. Candidate “drivers” that form pivotal controlling nodes in these interconnected pathways are identified in a number of ways: (1) They may connect multiple signaling pathways; (2) they may be recurrently mutated in human cancer specimens; (3) introduction of specific RNAi’s can block a signaling pathway that changes major phenotypic functions in the cell (such as maintaining viability); and (4) knockin or knockout of the gene’s activity in genetically engineered mouse models (GEMMs) results in molecular alterations, changing the phenotype in significant ways that affect malignant behavior.^36,37It is important to note that up to now, most targeted therapies discussed in this chapter were developed with earlier approaches that involved identification of abnormal functioning of a target in biological experiments with cancer cells, not because of detection of aberrant genes in broad screens of human cancers.

The Human Cancer Genome Atlas and additional data collected from genomic testing of cancers in multiple centers will identify more genetic aberrations that are cancer drivers and potential targets for new drugs. These new approaches will also enable identification of driver genes that are present at low frequencies—for example, less than 5%—in few types or only one type of cancer. It must be cautioned that a mutation in a known oncogene does not necessarily mean that it is a driver in a particular cancer, and a gene could have “driver” status in a cancer without a genetic aberration, working instead through amplification or an epigenetic mechanism, or through changes in expression of message or posttranslational modification of the protein it encodes. It also must be emphasized that a driver molecule may be activated by a variety of mechanisms related to its situation in one or more activated regulatory pathways ³⁸ (Figure 45-4 ).

The ultimate driver gene is one which is so critical to a cancer cell’s survival that when its function is blocked, the cell dies, typically by apoptosis. The observation of this phenomenon led to the “addiction hypothesis” of Weinstein, who postulated that in cancer cells that are driven by a mutation in a critical regulatory pathway, alternative pathways that could promote or bypass the driving gene’s activity may be downregulated to the point where, unlike the situation in normal cells, they may be irretrievably damaged when the driving, addicting gene is suddenly blocked by a targeted therapy. ³⁹

The identification of a driver gene does not invariably lead to creation of a drug effective against the gene’s product. The tyrosine kinase oncogenes have turned out to be “druggable” targets, and a few dozen new agents against these targets are in clinical use or in clinical trials—both antibodies against receptors on the cell surface and their ligands, and low molecular-weight drugs that act intracellularly. Suppressor genes such as p53, genes controlling transcription factors such as MYC, and genes whose products have more challenging molecular characteristics such as RAS have turned out to be less “druggable.” However, by knowing the upstream molecules that interact with these gene products, or the downstream molecules that they activate, targets may be identified that counteract the activity of these genes and are more “druggable.” ⁴⁰

Figure 45-4 Activation of EGFR signaling in non–small-cell lung cancer (NSCLC) can occur via disruption of several different components at multiple levels of the pathway. (A) Different proteins in the EGFR pathway can be activated (red) or inactivated (blue) by underlying genetic or epigenetic changes at the DNA level, leading to aberrant pathway activity and oncogenic signaling in NSCLC. Examples of key oncogenes affected include EGFR, RAS, PIK3CA, and AKT. Conversely, examples of tumor suppressors that are inactivated include PTEN and RASSF1. (B) Genetic and epigenetic mechanisms responsible for the disruption of genes in the EGFR signaling pathway in NSCLC include DNA copy number alterations (amplification or deletion), point mutations, and DNA methylation changes. Thus, it is important to consider multiple aspects of the genome and epigenome simultaneously to elucidate the mechanisms driving pathway deregulation. (This illustration was generated using Ingenuity Pathway Analysis software). ³⁸

It was initially hoped that a drug which successfully targeted the product of an aberrant gene in one type of cancer would be effective in other types of cancer with the same genetic abnormality. However, this has not always turned out to be the case, because different types of cells can express pathways that may bypass the drug’s target. An example is found with vemurafenib, which, as noted, is effective against the majority of melanomas expressing a mutated BRAF gene at V600E, but is not effective against colon cancers with the same genetic aberration. ⁴¹ This has been investigated by screening a panel of colon carcinoma cell lines bearing V600E RAF mutations with a short-hairpin RNA (shRNA) library representing the full complement of 518 human kinases, in the presence or absence of the BRAF inhibitor PLX4032. It was found that shRNA vectors targeting and eliminating EGF receptors converted resistant cells into becoming sensitive to inhibition by the BRAF inhibitor. ⁴² This effect was duplicated by treating cells with a drug or antibody against the EGF receptor. Thus inhibition of the EGF receptor tyrosine kinase appears to be required to permit sensitivity to this BRAF inhibitor in colorectal cancer cells. Most melanoma cells do not have the active EGF receptors that are seen in typical epithelial cells, and therefore are sensitive to the drug inhibiting BRAF. Clinical trials are testing this important experimental observation in patients with colorectal cancer bearing the V600E BRAF mutation, by adding an EGF receptor inhibitor to a BRAF inhibitor.

The above experiment is one example of many biological studies with cell lines and tumor biopsies that provide explanations for resistance to targeted drugs, involving activation of bypass pathways. For example, MET amplification can lead to resistance against the EGF receptor inhibitor gefitinib by activating ERB-B3. ⁴³ Likewise, the efficacy of MEK inhibitors in basal-subtype breast cancer cells is limited by a feedback loop involving activation of the PI3K pathway by activation of the EGF receptor tyrosine kinase. ⁴⁴ This suggests that a combination of inhibitors of MEK plus inhibitors of EGF receptors or the PI3K pathway may be effective in this situation.

In summary, combination treatments with agents against two different targets, in order to attack a driver gene and a bypass pathway concurrently, may provide a way to overcome resistance. Many clinical trials are testing this hypothesis.

With the technologies available today, an attractive way to investigate the mechanisms of failure (resistance) after initial response to an experimental targeted agent in a clinical trial is to biopsy the recurrent tumor and use genomic and expression array analysis—in comparison with the original tumor—to determine what new event or events have conferred resistance.

The bypass mechanisms that explain resistance to a targeted drug in a primary tumor may be discovered by direct comparison of genomic aberrations in primary tumor specimens from patients who are sensitive, or resistant, to a drug targeting a genetic aberration that they share. For example, clinical responses occur only in a minority of the colon cancer patients treated with the EGF receptor inhibitors cetuximab or panitumumab. It was found that colon cancers with a mutation in K-RAS were not responsive to these EGF receptor inhibitors. ⁴⁵ Furthermore, when resistance to these anti-EGF receptor antibodies developed in colon cancer patients who were initially responsive, biopsy of the recurrence showed that a mutation in RAS (either from outgrowth of a minor subpopulation below the limits of detection in the primary tumor, or occurring de novo) accounted for resistance in the majority of patients.^46,47Unfortunately, an effective inhibitor of K-RAS is not available to administer with the EGF receptor inhibitor. Because nearly 50% of colon cancers have mutations in K-RAS, screening for these mutations provides benefits to the patient by avoiding a therapy that will not be beneficial, and to controlling the costs of medical care by avoiding the use of these expensive agents in this clinical situation. Testing for this biomarker is now standard of care for treatment of colon cancer patients.

The complexity of the challenge in identifying genes that can modulate sensitivity and resistance to targeted therapies is exemplified by the many mechanisms of resistance that have been discovered in patients with malignant melanoma who were initially responsive to therapy with vemurafenib against V600E B-RAF, and subsequently relapsed. Six different mechanisms identified in tumor biopsies from these patients are listed in Table 45-2 .^48–54In all cases, confirmatory studies were carried out with cell lines containing the V600E B-RAF mutation, to demonstrate that the proposed mechanism was able to confer resistance to the inhibitor. The common theme in this series of observations and in studies of resistance to other targeted therapies is that there are two predominant resistance mechanisms—new aberrations of the targeted gene and activation of bypass pathways.

Table 45-2

Mechanisms of Resistance to Vemurafenib Treatment in Patients with V600E B-RAF Malignant Melanoma

These data naturally stimulate the formation of hypotheses on how to overcome resistance to drugs that target B-RAF. One approach that was tested recently was the addition of an inhibitor of MEK, administered concurrently with an inhibitor of mutated B-RAF. The rate of complete or partial response with the combination was 76%, as compared with 54% with monotherapy (P = 0.03). ⁵⁵ Of course the hope is that these responses will be more durable, resulting in substantial prolongation of life—but the results will not be available for a few years.

Although driver genes and genes to which cancer cells are addicted are obvious targets for drug development, there are other genes and pathways that do not cause cancer but are important for the survival and growth of cancer cells. This has generated the concept of “non-oncogene addiction.” ⁵⁶ Examples include gene products involved in DNA damage repair, protein chaperones and heat shock proteins, histone modification, and buffering of reactive oxygen species. Attacking these pathways that support the malignant phenotype, in addition to targeting critical oncogenes, may be a fruitful approach to combination therapy of cancer.

The series of experiments just discussed demonstrate the appropriateness of the term precision medicine. This term describes the situation in which a more precise understanding of the various genetic aberrations in a particular patient’s cancer, and their interactions with each other and with the surrounding microenvironment, can provide decision support to the treating oncologist—enabling selection of a targeted drug or drugs that are appropriate for that patient’s illness. In this chapter, the emphasis has been on genomics as the basis of precision medicine. This is because at the present time, genomic biomarkers are the most useful for drug development and for making clinical decisions. In the future, it is anticipated that biomarkers from at least four other sources will contribute more and more to precision medicine:

• Expression of genes. Data are being reported suggesting that a considerable fraction of mutated genes are not expressed. In the case of suppression genes, this is expected, because this is the basis for their impact on the cell’s phenotype. For potential oncogenes, however, this prevents the genetic aberrancy from having an impact. Furthermore, the level of expression of a gene—unrelated to the presence of a mutation—may alter cell behavior and phenotype. Again, the EGF receptor provides an example. This receptor is overexpressed (10- to 1000-fold) on a large fraction of epithelial cancers and is likely to create a “non-oncogene addiction” in these cells. ⁵⁶ Thus an important area of cancer research will involve closely examining the expression of RNA and proteins coded for by aberrantly functioning genes, to assess the importance of their expression levels in identifying a biomarker signature useful for precision cancer care.

• Epigenetics. Current technologies and computational methods in this area are not adequate to inform clinical decisions. Many investigators believe that epigenetic changes play a major role in oncogenesis and the malignant phenotype. ⁵⁷ This is a topic that will likely need to be visited more thoroughly in the future, as advancements are made in knowledge and assay techniques.

• Immunologic profile. As discussed in Chapters 50-52, immunological interventions are beginning to affect cancer therapy, leveraging recent gains in understanding the functioning of the human immune system. The patient’s immune system is likely to play an important role in controlling malignancy, as was postulated five decades ago. With increased understanding of the role of cytokines derived from the patient’s lymphocytes and leukocytes in promoting or inhibiting cancer cell growth, a profile of the patient’s immune functions is likely to become a component of the biomarker profile informing precision cancer care. ⁵⁸

• Proteomics. Evaluation of the functional status of proteins in a patient’s cancer (e.g., phosphorylation status) and detection of circulating protein markers derived from cancers (e.g., HCG levels) are promising areas for the clinic and should expand in value as biomarkers, as mass spectrometry techniques and the use of reverse phase protein arrays are developed into more useful clinical tests. ⁵⁹

Predicting Drug Efficacy and Toxicity

The other major scientific gap identified in the Report from the President’s Council of Advisors on Science and Technology underscored the need for improved ways of predicting at the earliest possible stage of development the efficacy and toxicity of candidate drugs. This requires continued efforts at developing improved models of human cancer in cell culture and animal models.

At the level of cell culture, screening on arrays containing hundreds of human cancer cell lines is providing new data on mechanisms of drug action, on potential synthetic-lethal targets which predict pairs of targeted drugs that may be effective when either alone is not, ⁶⁰ and on patterns of gene aberrations and gene expression that predict sensitivity or resistance to specific targeted experimental drugs.

At the level of drug-testing systems in animal models, progress has been made on two fronts:

• Growth of primary human tumor specimens in direct xenografts, without an intermediate step of culture on plastic surfaces, has produced models of human cancer in mice that are more closely comparable to primary human cancers. ⁶¹

• GEMMs with inducible promoters on mutated genes permit activation of the aberrant gene in specific organ sites and at specific time points, creating murine models that can more closely mimic human cancers. ²¹

Mouse models are discussed in Chapters 8 and 9.

Clinical Trials

The sequencing of clinical trials that is currently used to study new cancer drugs is presented in Chapter 48. There are two advances that may improve the efficiency and effectiveness of trials with experimental cancer therapies. The first is the increased use of adaptive randomization of patients to receive one of two drugs, or one from a panel of drugs. Initially, patients are assigned randomly to receive one or another of the drugs. As results accumulate, patients are assigned preferentially (but not invariably) to drugs that are producing more benefit. Eventually, with accrual of data on additional patients, poorly performing drugs are eliminated and new drugs can be added. ⁶² This method depends on powerful preset biostatistical tools that track and calculate probabilities of benefit sequentially, as results on each patient are added continuously to the database. The BATTLE Trial in advanced lung cancer, discussed earlier, was one of the first with targeted experimental therapies to use adaptive randomization, and the I-Spy 2 Trial for breast cancer is one of many current examples that is ongoing. This approach ⁶³ is quite different from the standard randomization protocol, in which the number of patients to be studied is predetermined by a calculation of the number needed for statistical significance of the results. Patients are then assigned randomly to one of two or more arms of the trial, and results remain undisclosed until the trial reaches a predetermined point for evaluation. The advantage of adaptive randomization is the capacity to study more experimental agents and reach yes-or-no decisions with fewer patients, which speeds up the process and reduces the number of patients who are receiving a drug that is not going to be beneficial. However, there is concern about the possibility of moving too swiftly and discarding an experimental drug inappropriately because decisions are made on a small number of patients, which bypasses the value of randomizing large numbers in order to stratify for unknown variables that might affect responses.

The other major advance in clinical trial design involves more stringent criteria for selecting patients in the earliest Phase II setting, once the dose and toxicity issues are settled in Phase I. A number of the successful trials described in this chapter enrolled only patients whose cancers had been shown to bear the genetic aberration that the experimental drug was designed to target. It is possible that off-target effects of an experimental drug may make an important contribution to its efficacy. However, the logic of this approach to testing drugs that target the products of a specific aberrant gene is obvious, provided that the experimental drug does indeed work primarily by acting on that target. With this approach, fewer patients will have to be studied to reach statistically valid conclusions, drug approval by the FDA will be achieved in shorter time frames, and patients will receive treatment with a drug that is more likely to be effective. A positive result in a clinical trial performed in this manner validates the genetic aberration tested for in patients’ cancers as a useful biomarker. The successes with this approach have led to Phase I/II trials, where biomarker-based enrollment in Phase II on a fixed dose and schedule is a seamless transition from Phase I.

As noted, most cancers are likely to have a number of driver genetic aberrations—a typical estimate is five—and it is not clear which or how many need to be targeted to eliminate the malignant cells. Because most drugs given at maximal tolerated doses only partially block their targets, and because of the redundancies in most signaling and regulatory pathways, it is likely that there are few occasions when a cancer cell is so totally addicted to a particular aberrant gene that a single drug targeting the product of that gene will cause death of the cancer cell. This reasoning, combined with knowledge of reported clinical outcomes data, suggest that optimal anticancer treatment is likely to require administration of drugs in combinations. In the past, combinations were selected based on the efficacy of individual drugs with nonoverlapping toxicities, whereas today a more rational selection of combinations of targeted drugs is based on knowledge of intersecting signaling and regulatory pathways and data on the genomic aberrations in more than one driver gene in a particular cancer.^64,65

Bringing two or more investigational drugs into clinical trials for use in combinations has been a challenging task, because the FDA required demonstration of the efficacy and safety of each drug alone before a combination could be tested. The FDA has released a new Guidance Document that provides guidelines for exploring combinations more efficiently, in situations where one or both new drugs are unlikely to provide benefit to patients when administered alone. ⁶⁶

With the proliferation of data on the genomic and molecular aberrations in many thousands of patients’ cancers and data on their responses to both targeted and conventional therapies, we will have new opportunities to learn by retrospectively correlating genomic and clinical data and comparing effectiveness of all available treatment options. The growing fields of health information technology and medical informatics should make this possible.^67,68Examples of insights that can be gained include discovery of a rare toxicity of a cancer therapy and discovery of an unanticipated association between responsiveness to a drug and a particular signature of genomic aberrations. However, many issues remain to be solved. Perhaps the most daunting is the need for standardized nomenclature and formats for electronic reporting of data, and interoperability between multiple clinical databases from multiple sources. Large health care providers such as Kaiser Permanente have solved many of these problems with sophisticated information systems that process and analyze large amounts of clinical data. There should be a federated system of access to data that protects patient confidentiality, but allows wide access for data mining and analysis as well as hypothesis testing. Another factor that prevents data sharing is the desire of academic institutions, clinical researchers, and pharmaceutical companies to sequester data that they can control because of concerns about protection of intellectual property.

Once analysis of databases and results of research projects on genomic-based cancer treatments have produced results, this information must be gathered and organized in a way that supports clinical decision making. Doctors and patients need analytical tools that enable them to make decisions about cancer therapy that are informed and evidence-based. Only a learning system which gathers and organizes information in ways that are accessible, reliable, and rapid will be useful to the practicing oncologist. ⁶⁰ This is a challenge for the future.

The implementation of genomics-based personalized cancer care raises a number of ethical issues: confidentiality of information; clarity on the level of certainty about clinical inferences that can be made from the results of tests for biomarkers; and the level of patient counseling appropriate for proper understanding of genomic data. Interpretation of genomic data is rendered difficult by intrinsic errors in the methodology, the high number of apparent aberrations observed in DNA from most cancer cells, the challenge of determining which genetic abnormality is a relevant driver of a patient’s cancer, and the heterogeneity within each human cancer that can invalidate predictions based on data from only a single sampling of the tumor. Genomic data are not perfectly predictive, and both physicians and patients must understand and accept this in order to use the data appropriately to improve decisions about cancer therapy.

Many advances have been made, but many challenges remain to be addressed before personalized, precision treatment based on a more complete knowledge of a patient’s cancer at the molecular level becomes the standard of practice. Table 45-3 lists some of these challenges.

Table 45-3

Challenges for Personalized Cancer Therapy

CLIA, Clinical Laboratory Improvement Amendment; CMS, Centers for Medicare and Medicaid Services; FDA, U.S. Food and Drug Administration.

Although much can be learned by sampling a patient’s cancer for performance of genomic analysis, the procedure of obtaining a biopsy presents risks that must be taken into consideration, and biopsies cannot be performed repeatedly. Noninvasive ways of obtaining actionable information about a patient’s cancer are desirable, and two that are being explored are promising. The most advanced noninvasive test for assessing molecular and metabolic properties of cancers involves medical imaging, especially positron emission tomographic (PET) scanning and magnetic resonance imaging (MRI). ⁶⁹ The measurement of fluorodeoxyglucose (FDG) uptake with PET scanning to detect changes in glucose metabolism and measurement of deoxyfluorothymidine (FLT) uptake to estimate the rate of proliferation can help in the differential diagnosis of an abnormal mass and can provide a rapid demonstration of the effect of a therapy on cancer in a patient. Some very promising progress has been demonstrated in the imaging of gene products and molecular abnormalities in animal cancer models, and this work is in early stages of evaluation in the clinical setting.

The second noninvasive test that holds great promise is the evaluation of circulating cancer cells and circulating fragments of cancer cell DNA that are released into the blood.^70,71The presence of circulating cancer cells and changes in their number have been shown to be predictive of prognosis and response to therapy in selected situations. DNA fragments are released into the circulation as a natural result of cell death in the body, and cancer cell DNA can be identified by the presence of genomic aberrations known to exist on the basis of prior assays of the patient’s tumor. This may enable tracking of a patient’s tumor burden by periodic sampling of DNA in the blood, as well as screening for a cancer recurrence or documenting a response to therapy. It is hoped that these technologies will develop to the point where they can be advanced into standard practice during the next 5 to 10 years.

Conclusion

A striking lesson relevant to clinical oncology that comes from the expansion of research on human cancer during the past decade is the value of genetic and molecular biomarkers for identifying the most useful targets for new drug development, and for screening patients to identify those most likely to bear cancers that will respond to particular targeted therapies.

In parallel, advancements in understanding signaling and regulatory pathways in cells and their interactions in complex systems have enabled identification of targets for therapy that are most likely to be driving the behavior of cancer cells.

What is especially exciting is that knowledge and technology are enabling sophisticated laboratory research and diagnostic molecular assays to be performed on specimens of primary human cancers. The knowledge gained from those studies is more likely to be relevant to development and selection of effective therapies than the data from cell lines and xenografts, which we were forced to depend on for so many years.

Information that guides selection of therapy will continue to expand as the status of critical RNA and protein molecules is assayed on primary tumor specimens and metastases, and further understanding leads to measures of the patient’s immune/inflammatory system as well as the influence of the microenvironment around the tumor.

The confluence of these approaches will lead to increasingly effective personalized care and precision medicine, moving us forward toward the ultimate goal of understanding how to give the right therapy to the right patient at the right time.

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