Immunomodulators

1. Immunomodulators Phase III Committee III December 2013

2. Immunomodulators • The development of agents that modulate the immune response rather than suppress it has become an important area of pharmacology. • The rationale underlying this approach is that such drugs may increase the immune responsiveness of patients who have either selective or generalized immunodeficiency. • The major potential uses are in immunodeficiency disorders, chronic infectious diseases, and cancer. • The AIDS epidemic has greatly increased interest in developing more effective immunomodulating drugs.

3. Cytokines INF-a INF-b INF-g Interleukins TNF-a TNF-b Colony-stimulating factors Granulocyte colony-stimulating factor G-CSF Erythropoietin (epoetin, EPO) Thrombopoietin (TPO) Immunomodulators

4. Cytokines • The cytokines are a large and heterogeneous group of proteins with diverse functions. • Some are immunoregulatory proteins synthesized within lymphoreticular cells and play numerous interacting roles in the function of the immune system and in the control of hematopoiesis. • In most instances, cytokines mediate their effects through receptors on relevant target cells and appear to act in a manner similar to the mechanism of action of hormones. • In other instances, cytokines may have antiproliferative, antimicrobial, and antitumor effects.

5. Cytokines (2) • The first group of cytokines discovered, were the interferons (IFNs), followed by the colony-stimulating factors. • Colony-stimulating factors. regulate the proliferation and differentiation of bone marrow progenitor cells. • Most of the more recently discovered cytokines have been classified as interleukins (ILs) and numbered in the order of their discovery. Cytokines are produced using gene cloning techniques. • Most cytokines (including TNF-a, IFN-g, IL-2, granulocyte colony-stimulating factor [G-CSF], and granulocyte-macrophage colony-stimulating factor [GM-CSF]) have very short serum half-lives (minutes). • The usual subcutaneous route of administration provides slower release into the circulation and a longer duration of action. • Each cytokine has its own unique toxicity, but some toxicities are shared. • For example, IFN-a, IFN-b, IFN-g, IL-2, and TNF-a all induce fever, flulike symptoms, anorexia, fatigue, and malaise.

6. Interferons (IFNs) • Interferons are proteins that are currently grouped into three families: • IFN-a, IFN-b, and IFN-g. • The IFN-a and IFN-b families comprise type I IFNs, ie, acid-stable proteins that act on the same receptor on target cells. • IFN-g, a type II IFN, is acid-labile and acts on a separate receptor on target cells. • Type I IFNs are usually induced by virus infections, with leukocytes producing IFN-a. • Fibroblasts and epithelial cells produce IFN-b. IFN-g is usually the product of activated T lymphocytes.

7. Interferons (IFNs) 2 • IFNs interact with cell receptors to produce a wide variety of effects that depend on the cell and IFN types. • IFNs, particularly IFN-g, display immune-enhancing properties, which include; increased antigen presentation and macrophage, NK cell, and cytotoxic T-lymphocyte activation. • IFNs also inhibit cell proliferation. • In this respect, IFN-a and IFN-b are more potent than IFN-g. • Another striking IFN action is increased expression of MHC molecules on cell surfaces. • While all three types of IFN induce MHC class I molecules, only IFN-g induces class II expression.

8. Interferons (IFNs) 3 • IFN-a is approved for the treatment of several neoplasms, including; • hairy cell leukemia, • chronic myelogenous leukemia, • malignant melanoma, and • Kaposi's sarcoma, • IFN-a is approved for the treatment of hepatitis B and C infections. • It has also shown activity as an anticancer agent in • renal cell carcinoma, • carcinoid syndrome, and • T cell leukemia.

9. Interferons (IFNs) 4 • IFN-b is approved for use in relapsing-type multiple sclerosis. • IFN-g is approved for the treatment of chronic granulomatous disease. • IL-2 is approved for the treatment of metastatic renal cell carcinoma and malignant melanoma. • Numerous clinical investigations of the other cytokines, including IL-1, -3, -4, -6, -11, and -12, are still in progress.

10. Interferons (IFNs) 5 • Toxicities of IFNs include; • fever, • chills, • malaise, • myalgias, • myelosuppression, • headache, and • depression • These side effects can severely restrict their clinical use.

11. The hematopoietic growth factors • The hematopoietic growth factors are glycoprotein hormones that regulate the proliferation and differentiation of hematopoietic progenitor cells in the bone marrow. • The first growth factors to be identified were called colony-stimulating factors because they could stimulate the growth of colonies of various bone marrow progenitor cells in vitro. • Many of these growth factors have been purified and cloned, and their effects on hematopoiesis have been extensively studied. • These growth factors are produced by recombinant DNA technology.Of the known hematopoietic growth factors, • erythropoietin (epoetin alfa), • granulocyte colony-stimulating factor (G-CSF), • granulocyte-macrophage colony-stimulating factor (GM-CSF), and • interleukin-11 (IL-11) are currently in clinical use. • Thrombopoietin and other potentially useful hematopoietic factors are still in development.

12. Erythropoietin • Erythropoietin, a 34-39 kDa glycoprotein, was the first human hematopoietic growth factor to be isolated. • It was originally purified from the urine of patients with severe anemia. • Recombinant human erythropoietin (rHuEPO, epoetin alfa) is produced in a mammalian cell expression system. • After intravenous administration, erythropoietin has a serum half-life of 4-13 hours in patients with chronic renal failure. • It is not cleared by dialysis. • It is measured in international units (IU). • Darbepoetin alfa is a glycosylated form of erythropoietin and differs from it functionally only in having a twofold to threefold longer half-life.

13. Erythropoietin • Erythropoietin stimulates erythroid proliferation and differentiation by interacting with specific erythropoietin receptors on red cell progenitors. • The erythropoietin receptor is a member of the JAK/STAT superfamily of cytokine receptors that use protein phosphorylation and transcription factor activation to regulate cellular function. • Erythropoietin also induces release of reticulocytes from the bone marrow. • Endogenous erythropoietin is primarily produced in the kidney. • In response to tissue hypoxia, more erythropoietin is produced through an increased rate of transcription of the erythropoietin gene. • Normally, an inverse relationship exists between the hematocrit or hemoglobin level and the serum erythropoietin level.

14. Erythropoietin • Nonanemic individuals have serum erythropoietin levels of less than 20 IU/L. • As the hematocrit and hemoglobin levels fall and anemia becomes more severe, the serum erythropoietin level rises exponentially. • Patients with moderately severe anemias usually have erythropoietin levels in the 100-500 IU/L range, and patients with severe anemias may have levels of thousands of IU/L. • The most important exception to this inverse relationship is in the anemia of chronic renal failure. • In patients with renal disease, erythropoietin levels are usually low because the kidneys cannot produce the growth factor. • These are the patients most likely to respond to treatment with exogenous erythropoietin.

15. Erythropoietin • Anemia can result from any of a large number of underlying conditions that either interrupt the normal process of erythropoiesis or result in the premature loss or destruction of mature erythrocytes. • One common indication for erythropoietin therapy is chronic kidney disease, in which the loss of functional kidney tissue results in loss of the cells responsible for erythropoietin production. • Another potential indication for erythropoietin is cancer, which can induce a state of relative resistance to endogenous erythropoietin by mechanisms that may involve proinflammatory cytokines, oxidative stress, and antierythropoietin antibodies. • Cancer can also cause anemia through bleeding, poor nutrition, and infiltration of the bone marrow by tumor cells; these causes can often be diagnosed and treated directly. • Often, cancer-related anemia results from the bone marrow toxicity of the chemotherapeutic agents used to treat the cancer.

16. Erythropoietin • Patients with endogenous erythropoietin levels of less than 100 IU/L have the best chance of response, although patients with erythropoietin levels between 100 and 500 IU/L respond occasionally. • These patients generally require higher erythropoietin doses (150-300 IU/kg three times a week) to achieve a response, and responses are often incomplete. • Erythropoietin has been used successfully to offset the anemia produced by zidovudine treatment in patients with HIV infection and in the treatment of the anemia of prematurity. • Erythropoietin is one of the drugs banned by the International Olympic Committee. • The use of erythropoietin by athletes is based on their hope that increased red blood cell concentration will increase oxygen delivery and improve performance. • The most common adverse effects of erythropoietin are associated with a rapid increase in hematocrit and hemoglobin and include hypertension and thrombotic complications. • These difficulties can be minimized by raising the hematocrit and hemoglobin slowly and by adequately monitoring and treating hypertension. • Allergic reactions have been infrequent and mild.

17. Myeloid Growth Factors • G-CSF and GM-CSF, the two myeloid growth factors currently available for clinical use, were originally purified from cultured human cell lines. • Recombinant human G-CSF (rHuG-CSF; filgrastim) is produced in a bacterial expression system. • It is a nonglycosylated peptide of 175 amino acids, with a molecular weight of 18 kDa. • Recombinant human GM-CSF (rHuGM-CSF; sargramostim) is produced in a yeast expression system. • It is a partially glycosylated peptide of 127 amino acids, with three molecular species with molecular weights of 15,500; 15,800; and 19,500. • These preparations have serum half-lives of 2-7 hours after intravenous or subcutaneous administration. • Pegfilgrastim, a covalent conjugation product of filgrastim and a form of polyethylene glycol, has a much longer serum half-life than recombinant G-CSF, and so it can be injected once per myelosuppressive chemotherapy cycle instead of daily for several days.

18. Myeloid Growth Factors • The myeloid growth factors stimulate proliferation and differentiation by interacting with specific receptors found on various myeloid progenitor cells. • Like the erythropoietin receptor, these receptors are members of the JAK/STAT superfamily. • G-CSF stimulates proliferation and differentiation of progenitors already committed to the neutrophil lineage. • It also activates the phagocytic activity of mature neutrophils and prolongs their survival in the circulation. • G-CSF also has a remarkable ability to mobilize hematopoietic stem cells, to increase their concentration in peripheral blood. • GM-CSF has broader biologic actions than G-CSF. • It is a multipotential hematopoietic growth factor that stimulates proliferation and differentiation of early and late granulocytic progenitor cells as well as erythroid and megakaryocyte progenitors. • Like G-CSF, GM-CSF also stimulates the function of mature neutrophils. • GM-CSF acts together with interleukin-2 to stimulate T-cell proliferation and appears to be a locally active factor at the site of inflammation. • GM-CSF mobilizes peripheral blood stem cells, but it is significantly less efficacious than G-CSF.

19. Myeloid Growth Factors CANCER CHEMOTHERAPY-INDUCED NEUTROPENIA • Neutropenia is a common adverse effect of the cytotoxic drugs used to treat cancer and increases the risk of serious infection in patients receiving chemotherapy. • Unlike the treatment of anemia and thrombocytopenia, transfusion of neutropenic patients with granulocytes collected from donors is performed rarely and with limited success. • The introduction of G-CSF in 1991 represented a milestone in the treatment of chemotherapy-induced neutropenia. • This growth factor dramatically accelerates the rate of neutrophil recovery after dose-intensive myelosuppressive chemotherapy. • It reduces the duration of neutropenia and usually raises the low neutrophil count seen following a cycle of chemotherapy. • The ability of G-CSF to increase neutrophil counts after myelosuppressive chemotherapy is nearly universal, but its impact on clinical outcomes is more variable. • Some clinical trials have shown that G-CSF reduces episodes of febrile neutropenia, requirements for broad-spectrum antibiotics, and days of hospitalization; however, other trials failed to find these favorable outcomes. • To date, no clinical trial has shown improved survival in cancer patients treated with G-CSF.

20. Myeloid Growth Factors • Clinical guidelines for the use of G-CSF after cytotoxic chemotherapy recommend reserving G-CSF for patients with a prior episode of febrile neutropenia after cytotoxic chemotherapy, • Pegfilgrastim is an alternative to G-CSF for prevention of chemotherapy-induced febrile neutropenia. • Although the two growth factors have similar effects on neutrophil counts, G-CSF is used more frequently because it is better tolerated. • G-CSF can cause bone pain, which clears when the drug is discontinued. • GM-CSF can cause more severe side effects, particularly at higher doses. These include fever, malaise, arthralgias, myalgias, and a capillary leak syndrome characterized by peripheral edema and pleural or pericardial effusions. • Allergic reactions may occur but are infrequent. • Splenic rupture is a rare but serious complication of the use of G-CSF.

21. Megacaryocyte Growth Factors • A low platelet count, or thrombocytopenia, is an important adverse effect of many cancer chemotherapeutic agents, occasionally limiting the doses that can be delivered with acceptable safety and tolerability. • The complications of thrombocytopenia include increased bleeding risk and platelet transfusion requirement; in turn, platelet transfusion is associated with an increased risk of infection, febrile reaction, and, rarely, graft-versus-host disease. • Research into the pharmacologic management of chemotherapy-induced thrombocytopenia has focused on the thrombopoietin analogues recombinant human thrombopoietin (rhTPO) and pegylated recombinant human megakaryocyte growth and development factor (PEG-rHuMGDF).

22. Megacaryocyte Growth Factors • A small-molecule oral drug that directly stimulates the TPO receptor is also in clinical trials. • To date, only recombinant human IL-11 (rhIL-11 or oprelvekin) has been approved by the FDA for clinical use. • These agents all have the potential to increase megakaryocytopoiesis (platelet production) in a dose-dependent fashion; although these drugs stimulate some multipotent as well as committed precursor cells, they do not significantly increase the hematocrit or white blood cell count. • Importantly, these agents must all be administered prophylactically because their activity is delayed in onset, with a 1- to 3-week before the platelet count reaches its peak value.

23. Megacaryocyte Growth Factors • Interleukin-11 (IL-11) is a 65-85 kDa protein produced by fibroblasts and stromal cells in the bone marrow. • Oprelvekin, the recombinant form of interleukin-11 approved for clinical use, is produced by expression in E coli. • The half-life of IL-11 is 7-8 hours when the drug is injected subcutaneously. • Thrombopoietin, a 65-85 kDa glycosylated protein, is constitutively expressed by a variety of organs and cell types. • Hepatocytes appear to be the major source of human thrombopoietin, and patients with cirrhosis and thrombocytopenia have low serum thrombopoietin levels. • Recombinant thrombopoietin is produced by expression in human cells.

24. Megacaryocyte Growth Factors • Interleukin-11 acts through a specific cell surface cytokine receptor to stimulate the growth of multiple lymphoid and myeloid cells. • It acts synergistically with other growth factors to stimulate the growth of primitive megakaryocytic progenitors and, most importantly, increases the number of peripheral platelets and neutrophils. • Acting through its own cytokine receptor, thrombopoietin also independently stimulates the growth of primitive megakaryocytic progenitors. • In addition, it stimulates mature megakaryocytes and even activates mature platelets to respond to aggregation-inducing stimuli. • The critical in vivo role of thrombopoietin has been demonstrated in genetically engineered knockout mice who lack either thrombopoietin or its receptor. • These mice have marked thrombocytopenia but do not display anemia or leukopenia.

25. Megacaryocyte Growth Factors • The most common adverse effects of interleukin-11 are fatigue, headache, dizziness, and cardiovascular effects. • The cardiovascular effects include anemia (due to hemodilution), dyspnea (due to fluid accumulation in the lungs), and transient atrial arrhythmias. • Hypokalemia has also been seen in some patients. • All of these adverse effects appear to be reversible. • Recombinant thrombopoietin and some other potentially useful hematopoietic factors are still in development. • In the limited clinical trial data available so far, recombinant thrombopoietin appears to be well tolerated.