Signals that affect cell behavior come from adjacent cells, the stroma in which the cells are located, hormonal signals that originate remotely, and from the cells themselves (autocrine signaling). These signals generally exert their influence on the receiving cell through activation of signal transduction pathways that have as their end result the induction of activated transcription factors that mediate a change in cell behavior or function or the acquisition of effector machinery to accomplish a new task. Although signal transduction pathways can lead to a wide variety of outcomes, many such pathways rely on cascades of signals that sequentially activate different proteins or glycoproteins and lipids or glycolipids, and the activation steps often involve the addition or removal of one or more phosphate groups on a downstream target. Other chemical changes can result from signal transduction pathways, but phosphorylation and dephosphorylation play a major role. The proteins that add phosphate groups to proteins are called kinases. There are two major distinct classes of kinases; one class acts on tyrosine residues, and the other acts on serine/threonine residues. The tyrosine kinases often play critical roles in signal transduction pathways; they may be receptor tyrosine kinases, or they may be linked to other cell-surface receptors through associated docking proteins (Fig. 26-2).
Therapeutic targeting of signal transduction pathways in cancer cells. Three major signal transduction pathways are activated by receptor tyrosine kinases (RTK). 1. The protooncogene Ras is activated by the Grb2/mSOS guanine nucleotide exchange factor, which induces an association with Raf and activation of downstream kinases (MEK and ERK1/2). 2. Activated PI3K phosphorylates the membrane lipid PIP2 to generate PIP3, which acts as a membrane-docking site for a number of cellular proteins including the serine/threonine kinases PDK1 and Akt. PDK1 has numerous cellular targets, including Akt and mTOR. Akt phosphorylates target proteins that promote resistance to apoptosis and enhance cell cycle progression, whereas mTOR and its target p70S6K upregulate protein synthesis to potentiate cell growth. 3. Activation of PLC-γ leads to the formation of diacylglycerol (DAG) and increased intracellular calcium, with activation of multiple isoforms of PKC and other enzymes regulated by the calcium/calmodulin system. Other important signaling pathways involve non-RTKs that are activated by cytokine or integrin receptors. Janus kinases (JAK) phosphorylate STAT (signal transducer and activator of transcription) transcription factors, which translocate to the nucleus and activate target genes. Integrin receptors mediate cellular interactions with the extracellular matrix (ECM), inducing activation of FAK (focal adhesion kinase) and c-Src, which activate multiple downstream pathways, including modulation of the cell cytoskeleton. Many activated kinases and transcription factors migrate into the nucleus, where they regulate gene transcription, thus completing the path from extracellular signals, such as growth factors, to a change in cell phenotype, such as induction of differentiation or cell proliferation. The nuclear targets of these processes include transcription factors (e.g., Myc, AP-1, and serum response factor) and the cell cycle machinery (CDKs and cyclins). Inhibitors of many of these pathways have been developed for the treatment of human cancers. Examples of inhibitors that are currently being evaluated in clinical trials are shown in purple type.
Normally, tyrosine kinase activity is short-lived and reversed by protein tyrosine phosphatases (PTPs). However, in many human cancers, tyrosine kinases or components of their downstream pathways are activated by mutation, gene amplification, or chromosomal translocations. Because these pathways regulate proliferation, survival, migration, and angiogenesis, they have been identified as important targets for cancer therapeutics.
Inhibition of kinase activity is effective in the treatment of a number of neoplasms. Lung cancers with mutations in the epidermal growth factor receptor are highly responsive to erlotinib and gefitinib (Table 26-2). Lung cancers with activation of anaplastic lymphoma kinase (ALK) or ROS1 by translocations respond to crizotinib, an ALK and ROS1 inhibitor. A BRAF inhibitor is highly effective in melanomas and thyroid cancers in which BRAF is mutated. Targeting a protein (MEK) downstream of BRAF also has activity against BRAF mutant melanomas. Janus kinase inhibitors are active in myeloproliferative syndromes in which JAK2 activation is a pathogenetic event. Imatinib (which targets a number of tyrosine kinases) is an effective agent in tumors that have translocations of the c-Abl and BCR gene (such as chronic myeloid leukemia), mutant c-Kit (gastrointestinal stromal cell tumors), or mutant platelet-derived growth factor receptor (PDGFR; chronic myelomonocytic leukemia); second-generation inhibitors of BCR-Abl, dasatinib, and nilotinib are even more effective. The third-generation agent bosutinib has activity in some patients who have progressed on other inhibitors, whereas the third-generation agent ponatinib has activity against the T315I mutation, which is resistant to the other agents. Sorafenib and sunitinib, agents that inhibit a large number of kinases, have shown antitumor activity in a number of malignancies, including renal cell cancer (RCC) (both), hepatocellular carcinoma (sorafenib), thyroid cancer (sorafenib), gastrointestinal stromal tumor (GIST) (sunitinib), and pancreatic neuroendocrine tumors (sunitinib). Inhibitors of the mammalian target of rapamycin (mTOR) are active in RCC, pancreatic neuroendocrine tumors, and breast cancer. The list of active agents and treatment indications is growing rapidly. These new agents have ushered in a new era of personalized therapy. It is becoming more routine for resected tumors to be assessed for specific molecular changes that predict response and to have clinical decision-making guided by those results.
TABLE 26-2Some FDA-Approved Molecularly Targeted Agents for the Treatment of Cancer ||Download (.pdf) TABLE 26-2 Some FDA-Approved Molecularly Targeted Agents for the Treatment of Cancer
|Drug ||Molecular Target ||Disease ||Mechanism of Action |
|All-trans retinoic acid ||PML-RARα oncogene ||Acute promyelocytic leukemia M3 AML; t(15;17) ||Inhibits transcriptional repression by PML-RARα |
|Imatinib ||Bcr-Abl, c-Abl, c-Kit, PDGFR-α/β ||Chronic myeloid leukemia; GIST ||Blocks ATP binding to tyrosine kinase active site |
|Dasatinib, nilotinib, ponatinib, bosutinib ||Bcr-Abl (primarily) ||Chronic myeloid leukemia ||Blocks ATP binding to tyrosine kinase active site |
|Sunitinib ||c-Kit, VEGFR-2, PDGFR-β, Flt-3 ||GIST; renal cell cancer ||Inhibits activated c-Kit and PDGFR in GIST; inhibits VEGFR in RCC |
|Sorafenib ||RAF, VEGFR-2, PDGFR-α/β, Flt-3, c-Kit ||RCC; hepatocellular carcinoma; TC ||Targets VEGFR pathways in RCC. Possible activity against BRAF in thyroid cancer |
|Regorafenib ||VEGFR-1 to -3, TIE-2, FGFR1, KIT, RET, PDGFR ||Colorectal cancer; GIST ||Competitive inhibitor of ATP binding site of tyrosine kinase domain multiple kinases |
|Axitinib ||VEGFR-1 to -3 ||RCC ||Competitive inhibitor of ATP binding site of tyrosine kinase domain VEGF receptors |
|Erlotinib ||EGFR ||Non-small-cell lung cancer; pancreatic cancer ||Competitive inhibitor of the ATP-binding site of the EGFR |
|Afatinib ||EGFR (and other HER family) ||Non-small-cell lung cancer ||Irreversible inhibitor of ATP-binding site of HER family members |
|Lapatinib ||HER2/neu ||Breast cancer ||Competitive inhibitor of the ATP binding site of HER2 |
|Crizotinib (Xalkori) ||ALK ||Non-small-cell lung cancer ||Inhibitor of ALK tyrosine kinase |
|Bortezomib, carfilzomib ||Proteasome ||Multiple myeloma ||Inhibits proteolytic degradation of multiple cellular proteins |
|Vemurafenib, dabrafenib ||BRAF ||Melanoma ||Inhibitor of serine-threonine kinase domain of V600E mutant of BRAF |
|Trametinib ||MEK ||Melanoma ||Inhibitor of serine-threonine kinase domain of V600E mutant of MEK |
|Cabozantinib ||RET, MET, VEGFR ||MTC ||Competitive inhibitor of ATP binding site of tyrosine kinase domain multiple kinases |
|Vandetanib ||RET, VEGFR, EGFR ||MTC ||Competitive inhibitor of ATP binding site of tyrosine kinase domain multiple kinases |
|Temsirolimus ||mTOR ||RCC ||Competitive inhibitor of mTOR serine-threonine kinase |
|Everolimus ||mTOR ||RCC; breast cancer ||Binds to immuophilin FK binding protein-12, which forms complex that inhibits mTOR kinase |
|Vorinostat, romidepsin ||HDAC ||CTCL ||HDAC inhibitor |
|Ruxolitinib ||JAK-1, -2 ||Myelofibrosis ||Competitive inhibitor of tyrosine kinase |
|Vismodegib ||Hedgehog pathway ||Basel cell cancer (skin) ||Inhibits smoothened in hedgehog pathway |
|Monoclonal Antibodies Alone |
|Trastuzumab ||HER2/neu (ERBB2) ||Breast cancer ||Binds HER2 on tumor cell surface and induces receptor internalization |
|Pertuzumab ||HER2/neu (ERBB2) ||Breast cancer ||Binds HER2 on tumor cell surface at distinct site from trastuzumab and prevents binding to other receptors |
|Cetuximab ||EGFR ||Colon cancer, squamous cell carcinoma of the head and neck ||Binds extracellular domain of EGFR and blocks binding of EGF and TGF-α; induces receptor internalization; potentiates the efficacy of chemotherapy and radiotherapy |
|Panitumumab ||EGFR ||Colon cancer ||Similar to cetuximab but fully humanized rather than chimeric |
|Rituximab ||CD20 ||B cell lymphomas and leukemias that express CD20 ||Multiple potential mechanisms, including direct induction of tumor cell apoptosis and immune mechanisms |
|Alemtuzumab ||CD52 ||Chronic lymphocytic leukemia and CD52-expressing lymphoid tumors ||Immune mechanisms |
|Bevacizumab ||VEGF ||Colorectal, lung cancers, RCC, glioblastoma, cervical cancer ||Inhibits angiogenesis by high-affinity binding to VEGF |
|Ziv-aflibercept ||VEGF-A, VEGF-B, PLGF ||Colorectal cancers ||Inhibits angiogenesis by high-affinity binding to VEGF-A, VEGF-B, and PLGF |
|Ipilimumab ||CTLA-4 ||Melanoma ||Blocks CTLA-4, preventing interaction with CD80/86 and T cell inhibition |
|Denosumab ||RANK ligand ||Breast, prostate cancer ||Inhibits RANK ligand, the primary signal for bone removal |
|Pembrolizumab ||PD-1 ||Melanoma ||Blocks PD-1 preventing interaction with PD-L1 T cell inhibition |
|Antibody-Chemotherapy Conjugates |
|Brentuximab vedotin ||CD30 ||Hodgkin’s disease, anaplastic lymphoma ||Delivery of chemotherapeutic agent (MMAE) to CD30-expressing tumor cells |
|Ado-trastuzumab emtansine ||HER2 ||Breast cancer ||Delivery of chemotherapeutic agent emtansine to HER2-expressing breast cancer cells |
However, none of these therapies has yet been curative by themselves for any malignancy, although prolonged periods of disease control lasting many years frequently occur in chronic myeloid leukemia. The reasons for the failure to cure are not completely defined, although resistance to the treatment ultimately develops in most patients. In some tumors, resistance to kinase inhibitors is related to an acquired mutation in the target kinase that inhibits drug binding. Many of these kinase inhibitors act as competitive inhibitors of the ATP-binding pocket. ATP is the phosphate donor in these phosphorylation reactions. Mutation in the BCR-ABL kinase in the ATP-binding pocket (such as the threonine to isoleucine change at codon 315 [T315I]) can prevent imatinib binding. Other resistance mechanisms include altering other signal transduction pathways to bypass the inhibited pathway. As resistance mechanisms become better defined, rational strategies to overcome resistance will emerge. In addition, many kinase inhibitors are less specific for an oncogenic target than was hoped, and toxicities related to off-target inhibition of kinases limit the use of the agent at a dose that would optimally inhibit the cancer-relevant kinase.
Targeted agents can also be used to deliver highly toxic compounds. An important component of the technology for developing effective conjugates is the design of the linker between the two, which needs to be stable. Currently approved antibody drug conjugates include brentuximab vedotin, which links the microtubule toxin monomethyl auristatin E (MMAE) to an antibody targeting the cell surface antigen CD30, which is expressed on a number of malignant cells but especially in Hodgkin’s disease and anaplastic lymphoma. The linker in this case is cleavable, which allows diffusion of the drug out of the cell after delivery. The second approved conjugate is ado-trastuzumab emtansine, which links the microtubule formation inhibitor mertansine and the monoclonal antibody trastuzumab targeted against human epidermal growth factor receptor 2 (HER2) on breast cancer cells. In this case, the linker is noncleavable, thus trapping the chemotherapeutic agent within the cells. There are theoretical pluses and minuses to having either cleavable or noncleavable linkers, and it is likely that both will be used in future developments of antibody-drug conjugates.
Another strategy to enhance the antitumor effects of targeted agents is to use them in rational combinations with each other and in empiric combinations with chemotherapy agents that kill cells in ways distinct from targeted agents. Combinations of trastuzumab (a monoclonal antibody that targets the HER2 receptor [member of the epidermal growth factor receptor (EGFR) family]) with chemotherapy have significant activity against breast and stomach cancers that have high levels of expression of the HER2 protein. The activity of trastuzumab and chemotherapy can be enhanced further by combinations with another targeted monoclonal antibody (pertuzumab), which prevents dimerization of the HER2 receptor with other HER family members including HER3.
Although targeted therapies have not yet resulted in cures when used alone, their use in the adjuvant setting and when combined with other effective treatments has substantially increased the fraction of patients cured. For example, the addition of rituximab, an anti-CD20 antibody, to combination chemotherapy in patients with diffuse large B cell lymphoma improves cure rates by 15–20%. The addition of trastuzumab, antibody to HER2, to combination chemotherapy in the adjuvant treatment of HER2-positive breast cancer reduces relapse rates by 50%.
A major effort is under way to develop targeted therapies for mutations in the ras family of genes, which are the most common mutations in oncogenes in cancers (especially kras) but have proved to be very difficult targets for a number of reasons related to how RAS proteins are activated and inactivated. Targeted therapies against proteins downstream of RAS (including mitogen-activated protein [MAP] kinase and ERK) are currently being studied, both individually and in combination. A large number of inhibitors of phospholipid signaling pathways such as the phosphatidylinositol-3-kinase (PI3K) and phospholipase C-gamma pathways, which are involved in a large number of cellular processes that are important in cancer development and progression, are being evaluated. The targeting of a variety of other pathways that are activated in malignant cells, such as the MET pathway, hedgehog pathway, and various angiogenesis pathways, is also being explored.
One of the strategies for new drug development is to take advantage of so-called oncogene addiction. This situation (Fig. 26-3) is created when a tumor cell develops an activating mutation in an oncogene that becomes a dominant pathway for survival and growth with reduced contributions from other pathways, even when there may be abnormalities in those pathways. This dependency on a single pathway creates a cell that is vulnerable to inhibitors of that oncogene pathway. For example, cells harboring mutations in BRAF are very sensitive to MEK inhibitors that inhibit downstream signaling in the BRAF pathway.
Synthetic lethality. Genes are said to have a synthetic lethal relationship when mutation of either gene alone is tolerated by the cell but mutation of both genes leads to lethality, as originally noted by Bridges and later named by Dobzhansky. Thus, mutant gene a and gene b have a synthetic lethal relationship, implying that the loss of one gene makes the cell dependent on the function of the other gene. In cancer cells, loss of function of a DNA repair gene like BRCA1, which repairs double-strand breaks, makes the cell dependent on base excision repair mediated in part by PARP. If the PARP gene product is inhibited, the cell attempts to repair the break using the error-prone nonhomologous end-joining method, which results in tumor cell death. High-throughput screens can now be performed using isogenic cell line pairs in which one cell line has a defined defect in a DNA repair pathway. Compounds can be identified that selectively kill the mutant cell line; targets of these compounds have a synthetic lethal relationship to the repair pathway and are potentially important targets for future therapeutics.
Targeting proteins critical for transcription of proteins vital for malignant cell survival or proliferation provides another potential target for treating cancers. The transcription factor nuclear factor-κB (NF-κB) is a heterodimer composed of p65 and p50 subunits that associate with an inhibitor, IκB, in the cell cytoplasm. In response to growth factor or cytokine signaling, a multi-subunit kinase called IKK (IκB kinase) phosphorylates IκB and directs its degradation by the ubiquitin/proteasome system. NF-κB, free of its inhibitor, translocates to the nucleus and activates target genes, many of which promote the survival of tumor cells. Novel drugs called proteasome inhibitors block the proteolysis of IκB, thereby preventing NF-κB activation. For unexplained reasons, this is selectively toxic to tumor cells. The antitumor effects of proteasome inhibitors are more complicated and involve the inhibition of the degradation of multiple cellular proteins. Proteasome inhibitors (e.g., bortezomib [Velcade]) have activity in patients with multiple myeloma, including partial and complete remissions. Inhibitors of IKK are also in development, with the hope of more selectively blocking the degradation of IκB, thus “locking” NF-κB in an inhibitory complex and rendering the cancer cell more susceptible to apoptosis-inducing agents. Many other transcription factors are activated by phosphorylation, which can be prevented by tyrosine kinase inhibitors or serine/threonine kinase inhibitors, a number of which are currently in clinical trials.
Estrogen receptors (ERs) and androgen receptors (ARs), members of the steroid hormone family of nuclear receptors, are targets of inhibition by drugs used to treat breast and prostate cancers, respectively. Tamoxifen, a partial agonist and antagonist of ER function, can mediate tumor regression in metastatic breast cancer and can prevent disease recurrence in the adjuvant setting. Tamoxifen binds to the ER and modulates its transcriptional activity, inhibiting activity in the breast but promoting activity in bone and uterine epithelium. Selective ER modulators (SERMs) have been developed with the hope of a more beneficial modulation of ER activity, i.e., antiestrogenic activity in the breast, uterus, and ovary, but estrogenic for bone, brain, and cardiovascular tissues. Aromatase inhibitors, which block the conversion of androgens to estrogens in breast and subcutaneous fat tissues, have demonstrated improved clinical efficacy compared with tamoxifen and are often used as first-line therapy in patients with ER-positive disease. A number of approaches have been developed for blocking androgen stimulation of prostate cancer, including decreasing production (e.g., orchiectomy, luteinizing hormone–releasing hormone agonists or antagonists, estrogens, ketoconazole, and inhibitors of enzymes such as CYP17 involved in androgen production) and AR blockers (Chap. 38).