Indian Journal of Cancer, Vol. 47, No. 4, October-December, 2010, pp. 443-451
Biological response modifiers: Current use and future prospects in cancer therapy
M Bisht1, SS Bist2, DC Dhasmana1
1 Department of Pharmacology, Himalayan Institute of Medical Sciences, HIHT University Jolly grant, Doiwala, Dehradun, Uttarakhand 248 140, India
Code Number: cn10104
AbstractOver the past few decades, considerable success has been achieved in the field of cancer treatment with biological response modifiers (BRM), which are agents that improve the body's ability to fight cancer by immunostimulation. Biological agents, such as interferons and interleukins, provide nonspecific active immunity, whereas the monoclonal antibodies provide passive immunity. Apart from this, other biological agents, such as antiangiogenic agents, matrix metalloprotease inhibitors, tyrosine kinase inhibitors, and tumor vaccines, are also increasingly being used in cancer treatment. Hematopoietic factors, such as granulocyte colony-stimulating factor, are used to increase the general immunity and prevent opportunistic infection. BRM are basically used alone or as adjuvants to cancer chemotherapeutic agents. This review sheds light on the current use and the future development of cancer immunotherapy. Search strategy included Pubmed, using the terms "Biological response modifiers in cancer" citations relevant to the topic were screened.
Keywords: Biological response modifiers, cancer, monoclonal antibodies
Cancer at present is a major global problem. Most cancers are treated by combined treatment modalities, such as chemotherapy, radiotherapy, and surgery. Till now, the main pharmacologic strategy against human cancers included the use of chemotherapeutic agents, which at times is incapable in completely eradicating the cancer cells. Therefore, there is still a need to increase the armamentarium of anticancer agents. Biological response modifiers (BRM) are the naturally occurring agents present in the body to destroy cancer cells by immunostimulation. The National Cancer Institute defines BRM as "agents or approaches that modify the relationship between tumor and host by modifying the host′s biological response to tumor cells with resultant therapeutic effects."  The primary aim of these agents is to provide passive or active immunization against cancer cells, although some of these agents may also have antiproliferative, antiangiogenic, and direct cytotoxic action. Recent advances in tumor immunology have enabled the development of specific agents targeted against cancer cells. Today BRM are an important target for cancer research and treatment. Biological therapy, immunotherapy, and biotherapy are the other synonyms of BRM. Searching the Pubmed with the key words "Biological response modifiers in cancer" revealed many articles, and the relevant articles were screened.
In 1893, William B Coley who is now recognized as the "Father of BRM" observed that many cancer patients recovered after developing postoperative infections. He, thereby, concluded that fever upregulates the immune system in cancer patients, which then recognizes and destroys the invading tumor cells.  The use of BRM in cancer has developed during past 3 decades. Officially, BRM were first studied for the cancer treatment in 1960, when Bacillus Calmette-Guι rin and Corynebacterium parvum were used to stimulate the immune system of cancer patients.  This therapy, termed as immunotherapy, soon lost its momentum, as the results of clinical trials were not encouraging. In late 1970s, the use of several immunomodulating cytokines, including interferons (IFN), interleukins (IL), and tumor necrosis factor (TNF) was introduced for cancer treatment.  The use of colony-stimulating factors for mitigating the side effects of cancer therapy started around 1980s.  In 1990s, various antigens associated with cancer cells were identified and the mechanisms of antigen presentation and recognition was understood deeply at the molecular level. As a result of this, various monoclonal antibodies against cancer cells are being developed. The future of biotherapy involves the use of gene therapy in cancer using cytokine genes and protooncogenes and vaccine therapy using antigenic epitope peptides and antigen presenting cells or cancer antigen genes.
The various BRM being used in cancer can be categorized as follows:
The mechanism by which most BRM act can be broadly divided into 3 major categories:
The agents having direct cytotoxic action on the tumor cells are included in this category. For example, monoclonal antibodies.
These agents restore, augment, or modulate the immune system to facilitate the destruction of tumor cells. For example, IFNs and ILs.
Promotion of cell differentiation: The agents that help in replacing the damaged cells in the body are included in this category. For example, CSF.
Interference with neoplastic changes: Agents, such as retinoid facilitate the differentiation of the tumor cell into normal cell types.
Prevention of metastasis: Angiogenic inhibitor prevents tumor metastasis.
IFNs are the small proteins synthesized by the immune cells in response to various stimuli, such as viral infections and cytokines (IL-1, IL-2, TNF-α). They inhibit viral replication and promote cellular (T cell) immune response.  There are 3 major classes of human IFNs: alpha, beta, and gamma. All the 3 types have distinctive antiproliferative and immunomodulatory actions. IFNs act via specific cellular receptors linked with JAK-STAT pathway to stimulate the formation of specific proteins, which mediate their actions.
Their antitumor action is quite complex, which include antiproliferative effects, promotion of differentiation, immunomodulation, alteration in tumor cell surface antigen expression, inhibition of oncogene activation, and angiogenesis. 
A high response rate of approximately 75-90% is seen with IFN-α in previously untreated patients of chronic myelogenous leukemia, hairy cell leukemia, myeloproliferative disorders, and cutaneous T-cell lymphomas.  The response rate is nearly 40-50% in low-grade lymphomas and multiple myeloma (MM). Apart from this, it also has a limited activity in renal carcinoma, melanoma, Kaposi sarcoma, and carcinoid syndrome.
IFN-α is available in recombinant, natural, and pegylated forms. The main advantage of pegylated IFN-α is prolonged half-life. Depending on the condition, a wide range of doses and schedules of IFN can be used, ranging from 3 MU three times weekly to 36 MU daily. It can be administered by subcutaneous (SC) or intravenous (IV) route. Apart from this, they can also be given intranasally and recently, the oral use of IFN-α is also recommended.  Side effects depend on the dose and schedule of IFN therapy and consist of fever, chills, myalgia, headache, nausea, anorexia, and weight loss (flu-like syndrome).  Many of these side effects are commonly experienced by the patient on first exposure to IFN, but they usually decrease in intensity with continued therapy. In a few patients myelosuppression might occur, which is reversible within 1-3 days of discontinuation of IFN therapy. Rarely, interstitial nephritis, confusion, coma, hypotension, and arrhythmias may occur.
The cytokines produced in the body by the lymphocytes are known as ILs. They mediate their actions through the cell surface receptors in relevant target cells. Many ILs have been identified and numbered in the order of their discovery (IL-1 to IL-23). Out of these, some ILs, such as IL-2, stimulate the growth and activity of immune cells, which target cancer cells. IL-2 acts as an antitumor agent by increasing the cytolytic activity of antigen-specific cytotoxic T lymphocytes and natural killer (NK) cells and by increasing the gene expression responsible for encoding the lytic component of cytotoxic granules, that is, perforin and granzymes.  It also increases the expression of adhesion molecules, thereby facilitating binding of activated leukocytes to tumor endothelium and cells. Lymphocytes stimulated by IL-2, known as lymphokine-activated killer (LAK) cells, are also found to be effective in destroying tumors. Lymphocytes are isolated from cancer patient′s blood, stimulated with high doses of IL-2 in the laboratory, and reinjected in the patient as LAK cells with the aim of improving the patient′s anticancer immune response. However, randomized trials have not shown any significant improvement in the response rates with LAK cells as compared with IL-2 therapy. 
IL-2 is approved by US Food and Drug Administration (FDA) for the treatment of advanced metastatic renal cell carcinoma and malignant melanoma.  Besides these conditions it has also been tried in acute myelocytic leukemia (AML) and bone marrow transplant. 
It is available as recombinant IL-2 (aldesleukin, Proleukin). Due to its short half-life (T1/2α = 13 min; T1/2β = 85 min), it is administered as continuous IV infusion or as multiple intermittent dosing. Pegylated and liposomal forms of IL-2 are under development to extend the half-life and to enhance its delivery to immune cells in tumors. Investigators have developed 2 strategies for administering IL-2. The first strategy is the high-dose bolus IL-2 regimen.  IL-2 is administered at a dose of 600,000-720,000 IU/kg IV every 8 h on Days 1-5 and 15-19 of treatment. Treatment is repeated at 8- to 12-week intervals in patients who demonstrate good response. High-dose IL-2 regimens are associated with more adverse effects. The second strategy is low-dose IL-2 therapy, which can be administered either as IV bolus or continuous IV/SC injection.  With continuous infusion, the IL-2 dose is 18 MIU/m 2 /day on Days 1-5 and 15-19. The response rate as well as toxicity profile with a low dose IL-2 therapy is similar to that seen with a high-dose IL-2 therapy. Recently, a very promising approach of combining IL-2 immunotherapy with cytotoxic immunotherapy, known as biochemotherapy was investigated. IL-2 has also been combined with cytotoxic chemotherapies for the treatment of cancer. The efficacy of this therapeutic approach is yet to be confirmed by large-scale multicentric trials. The most common toxicity due to IL-2 results from its ability to increase the capillary permeability. The symptoms include hypotension, ascites, anasarca, and pulmonary edema. Early side effects include infusion reaction, flu-like symptom, and gastrointestinal effects. Supportive therapy with acetaminophen, H 2 -blockers, antiemetics, and antidiarrheals should be given to mitigate these side effects. Hypotension can be managed by administering IV fluids. Besides IL-2, other ILs used in cancer patients are IL-6 and IL-11. In vitro as well as in vivo observations have demonstrated that IL-6 plays a key role in the pathogenesis of MM. Endogenous IL-6 production is 2-30 times higher in MM patients than in healthy individuals. The anti-IL-6 chimeric monoclonal antibody strongly suppresses this endogenous IL-6 production.  IL-11 induces megakaryocytic proliferation. Oprelvekin is the recombinant form of IL-11 approved for malignancy-induced thrombocytopenia. The other ILs currently under evaluation are IL-3, IL-4, IL-6, IL-12, IL-5, and IL-18. 
Monoclonal antibodies are the clones of similar antibodies that are directed against specific target antigens.  Cancer cells express a wide variety of antigens that are attractive targets for monoclonal antibody-based therapy. These antibodies can attack the cancer cells in different ways [Figure - 1]. They can activate the immune functions and facilitate the destruction of malignant cells by complement-dependent cytotoxicity and antibody-dependent cell-mediated cytotoxicity. Antibodies can also be attached to cytotoxic molecule, which kills the cancer cells. Another way is to tag the bispecific antibody with a killer T cell. Antibodies can also inhibit the action of certain growth factors, such as vascular endothelial growth factor (VEGF). Lastly, antibodies can be conjugated with radioactive substance to facilitate its targeted delivery to the cancer cells.  [Table - 1] summarizes a list of monoclonal antibodies being used in cancer. 
Colony Stimulating Factors
CSFs are growth factors that mediate the proliferation, maturation, regulation, and activation of hematopoietic cells.  Generally, CSFs are named after the major cell lineages, which are affected by them; myeloid growth factors include GM-CSF, which affects both granulocyte and macrophage lineage and G-CSF, which targets only granulocytes; EPO targets erythrocyte production; and TPO is associated with platelet generation. Recombinant human G-CSF (filgrastim) and GM-CSF (sargramostim) are approved by the FDA for the management of neutropenic fever secondary to cytotoxic chemotherapy. They are administered as IV bolus, continuous infusion, or SC by daily injection. They are to be administered 24-72 h after chemotherapy until high neutrophil count (1000/μL) has persisted for 3 consecutive days. G-CSF is generally well tolerated. The main side effect reported with it is mild to moderate bone pain. The pattern of toxicity of GM-CSF is similar to other cytokines, demonstrating an acute reaction at first dose characterized by fever, chills, hypotension, and dyspnea. EPO was FDA-approved in 1989 for the treatment of renal anemia. Nowadays, it is also used for the treatment of malignancy or chemotherapy-induced anemia in the dose of 150-300 U/kg SC 3-5 times a week. It stimulates erythropoiesis in 50-70% of patients and reduces the transfusion requirement, along with improving their quality of life. However, there are some safety concerns associated with the use of EPO. There are reports of increased thromboembolic and cardiovascular events with the use of EPO in patients with hemoglobin concentration of >12 g/dL.  rhuTPO is being investigated to increase the platelet count, but there are reports of patients developing neutralizing antibodies against it.  The efforts are under way to develop small molecular mimics of rhuTPO.
The loss of differentiation is one of the hallmarks of malignancy. Numerous chemical entities, including vitamin D and its analogs, retinoids, histone deacetylase inhibitors, and inhibitors of DNA methylation have shown to induce differentiation in tumor cell lines in vitro.  The use of tretinoin for inducing remission in acute promyelocytic leukemia is the best example to substantiate the role of differentiating agent in cancer. Other oral vitamin A analogs used in cancer include acitretin, isotretinoin, and bexarotene. Clinical trials have indicated that retinoids have significant activity in the reversal of oral, skin, and cervical premalignancies and in the prevention of head and neck, lung, and skin primary tumor. , Oral retinoids are clinically used for the treatment of acute promyelocytic leukemia, cutaneous T-cell lymphoma and leukoplakia. Retinoids bind with retinoic acid receptor-alpha, which dimerizes with retinoid X receptor, which in turn displaces the repressor of differentiation and promotes the maturation of cancer cells along the expected pattern. The major toxicity associated with retinoids include dry skin, cheilitis, reversible hepatic enzyme abnormalities, bone tenderness, hyperlipidemia, and retinoic acid syndrome.
Tyrosine Kinase Inhibitors
Tyrosine kinases are critical components of signal transduction pathways that transmit information from extracellular ligands or cytoplasm to the nucleus, thereby influencing gene transcription and DNA synthesis. They are subdivided into proteins that have extracellular ligand-binding domain (receptor tyrosine kinase) and enzymes that are confined to the cytoplasm or nuclear compartment (nonreceptor tyrosine kinase).  Epidermal growth factor receptor (EGFR) belongs to a subfamily of 4 closely related tyrosine kinase receptors: EGFR, HER2, HER3, and HER4. Abnormal activation of EGFR is demonstrated in many human neoplasms, including squamous cell cancer of the head and neck, ovarian cancer, colorectal cancer, breast cancer, non-small cell lung cancer (NSCLC), bladder cancer, and therefore is an attractive target for anticancer therapy.  Various TKI available for clinical use and in various phases of clinical trials are listed in [Table - 2]. Currently 6 orally active small molecular weight TKIs, namely imatinib, gefitinib, erlotinib, dasatinib, nilotinib, and lapatinib are FDA approved. They block the phosphorylation and activation of downstream signaling of EGFR and other kinases. Imatinib is approved as a first-line therapy in accelerated phase, chronic phase, and blast crisis of chronic myeloid leukemia (CML). Geftinib is approved for the treatment of patients with advanced NSCLC whose disease has progressed in spite of previous treatments with other agents.  Preclinical studies have demonstrated the efficacy of geftinib in the treatment of esophageal squamous cell carcinomas as well. In a recent phase III trial (IPASS trial), gefitinib was found to be superior to carboplatin-paclitaxel therapy as an initial treatment for pulmonary adenocarcinoma.  Erlotinib is indicated for locally advanced or metastatic NSCLC.  A recent study suggests that erlotinib may be used for the treatment of JAK2V617F-positive polycythemia vera and other myeloproliferative disorders.  Dasatinib and nilotinib are indicated for imatinib-resistant CML. , Lapatinib is approved as a front-line therapy in triple-positive breast cancer and as an adjuvant therapy when patients have progressed on Herceptin.  Sunitinib and sorafanib prevent angiogenesis by inhibiting the downstream signaling associated with VEGF receptor and PDGFR. Sunitinib is approved for advanced renal cell cancer (RCC) and gastrointestinal stromal tumor (GIST).  Sorafenib is approved for imatinib-resistant advanced RCC and hepatocellular carcinoma. ,
Tumor Necrosis Factor-alpha
TNF-α is secreted by macrophages activated by endotoxins. TNF binds to the receptor on cell membranes, initiates cellular activity, and is cytotoxic to that cell. It affects tumor cells by various mechanisms. It can either cause direct destruction of the tumor cell and its vasculature or stimulate NK- and LAK-induced tumor killing.  Although TNF has shown promising antitumor activity in laboratory studies, the dose needed for clinical efficacy is extremely toxic. Phase I/II studies revealed that IV infusion produces severe hypotension and hepatotoxicity. TNF therapy is most effective and least toxic when directed at a specific tumor site; therefore, isolated limb perfusion was tried in the treatment of melanoma and sarcoma.  Another concern is that the long-term use of TNF can be associated with regrowth of resistant cancer and may have tumor promoting action.
Thalidomide and its Congeners
Thalidomide, a well known teratogen, and its analog (lenalidomide) have antitumor action owing to their immunomodulatory, antiangiogenic, antiproliferative, and proapoptotic properties. It suppresses TNF-α production, reduce the expression of proangiogenic factors, such as VEGF and IL-6, induce NK cells and cause cell growth arrest at the G1 phase. ,, A combination of thalidomide with dexamethasone is the first-line therapy in MM patients.  Significant activity of thalidomide is observed in patients with myelodysplastic syndrome (MDS). Moreover, it is also being investigated for the treatment of metastatic RCC, advanced pancreatic cancer, androgen-independent prostate cancer, glioblastoma multiforme and cancers of the breast and ovary.  Lenalidomide, a thalidomide analog is approved for use in combination with dexamethasone in patients with refractory MM and MDS. 
The growth of new blood vessels, a process termed angiogenesis is necessary for the growth and metastasis of tumors.  Angiogenesis consists of multiple coordinated, sequential, and interdependent steps regulated by a finely balanced equilibrium between proangiogenic and antiangiogenic factors. A wide range of angiogenic factors, including growth factors, chemokines, angiogenic enzymes, endothelial-specific receptors, and adhesion molecules facilitate angiogenesis. Various angiogenic inhibitors that inhibit the angiogenic process have also been described. Pharmacologic manipulation of these factors may lead to antiangiogenic effect. Angiogenesis can be inhibited at different steps of the process. In general, 5 strategies are being used as antiangiogenic therapy, including the inhibition of activated endothelial cells (EC), EC intracellular signaling, extracellular matrix remodeling, adhesion molecules, and angiogenic mediators or their receptors. Most successful strategy till date is the inhibition of angiogenic factors and their receptors. Monoclonal antibodies, such as bevacizumab, cetuximab, panitumumab, trastuzumab and TKIs, such as erlotinib, sorafenib, and sunitinib inhibit the action of angiogenic mediators, such as vascular endothelial growth factors (VEGF) and fibroblast growth factor. Bevacizumab is approved for metastatic colorectal cancer (mCRC), metastatic RCC, non-small cell lung cancer (NSCLC) and advanced breast cancer. , Cetuximab is approved for mCRC and squamous cell cancer of head and neck and panitumumab is approved for mCRC.  Antiangiogenic TKIs have been described in earlier section. Apart from these, other approved antiangiogenic drugs for anticancer effects are highlighted in [Table - 3]. Thalidomide and lenalidomide have also been described in the previous section. Bortezomib, which is a proteasome inhibitor, having indirect antiangiogenic action is approved for MM and mantle cell lymphoma. Temsirolimus and everolimus are small molecule inhibitors of mTOR (mammalian target of rapamycin), which is a part of the PI3 kinase/AKT pathway involved in tumor cell proliferation and angiogenesis. They are approved for use in advanced renal cell carcinoma., Other agents, such as alitretinoin, imiquimod, and IFN-α are used in dermatologic malignancies, such as AIDS-related Kaposi′s sarcoma, malignant skin cancers, and giant cell tumor, respectively. 
Cancer vaccine is a new type of cancer treatment and is still in its infancy. The identification of various tumor-associated or tumor-specific antigens has facilitated the induction of antitumor immunologic responses in vivo. Cancer vaccines are intended either as preventive or curative agents. Cancer preventive vaccines are designed to target infectious agents that can cause cancer. Recently, human papillomavirus (HPV) vaccines has been developed, which prevent infection against HPV types 16 and 18 that cause nearly 70% of all cervical cancers.  HPV vaccination is now unequivocally recommended for females over the age of 10 years for the prevention of cervical cancer. The impressive range of protection of this vaccine ranges from 86% to 100%. Cancer treatment vaccines are designed to activate cell-mediated immune responses against tumor-associated antigens (TAA). They may prevent the growth of existing tumor, recurrence, or eliminate cancer cells resistant to previous treatments. The antitumor vaccines include peptide vaccination, vaccination with genetically modified organisms (GMO) and application of genetically alterable autologous tumor cells. The peptide vaccination strategy can be used for cancers, such as melanoma where TAA are well defined on the tumor cell surface. The TAA included in vaccines are DNA, RNA, or proteins that are produced by gene technology and administered SC. Vaccination strategy with GMOs includes transfecting viruses with the genetic information to code for TAA. In the third strategy, vaccination with irradiated autologous tumor cells/cell lysates and tumor antigens is tried.  Few therapeutic cancer vaccines have shown promising results in early-stage clinical trials involving different types of cancers, including non-Hodgkin′s lymphoma, NSCLC, and melanoma. Various mechanisms by which the tumor cells may develop acquired resistance to the immune response include the absence of antigen in tumor variants, loss of major histocompatibility complex expression, downregulation of antigen-processing machinery, and expression of local inhibitory molecules. The CD8+ cytotoxic T lymphocytes have a major effect in killing tumor cells. The major focus in the development of cancer vaccines is the generation of tumor-specific T-cell responses. In an attempt to design more effective vaccinations for cancer patients, cytokine gene therapy and dendritic cell vaccination have been explored. Dendritic cells have a key role in the mediation of vaccine action, as they activate the cytolytic T-cell responses by capturing, processing, and presenting the TAA to T lymphocytes.  Cytokines, such as GM-CSF, TNF-α activate dendritic cells, thereby enhancing their ability to activate T cells. The unique part of each antibody is called an idiotype. Anti-idiotype vaccines can be used in tumors, such as B-cell lymphomas where tumor cells express surface antibodies of the same idiotype. The major problem with antigen vaccine is that, they may become less effective over time because the immune system recognizes them as foreign and quickly destroys them. To overcome this, scientists have developed DNA vaccines to provide a steady supply of antigens to keep the immune response going. Cells can be injected with DNA that code for protein antigens and instruct them to keep making more antigens.
Clinical Status of BRM
The therapeutic value of biologic agents in cancer lies in the prolongation of remission of the tumor after chemotherapy, radiotherapy, or surgery. Most of the BRM agents are used as adjuvants in advanced non-treatment responsive tumors.  The success of these agents is established in some malignancies, such as melanoma, breast cancer, certain leukemias, Kaposi sarcomas, and renal cell carcinoma.  However, the results of BRMs in other tumors remain elusive. Some other agents, such as CSFs have a role in palliating the side effects associated with anticancer agents. But the cost remains a major prohibitive factor in the widespread use of these agents.
The influence of BRM on malignant processes has revolutionized cancer therapy. The knowledge regarding biological agents is rapidly expanding, and effective targets are being identified. These agents act by modulation of immune responses, stimulation/inhibition of hematopoiesis, direct regulation of cell growth/differentiation, and modulation of angiogenesis. However, modulation of these complex networks can lead to some unexpected and unwanted side effects, which must be monitored carefully. The past decade has witnessed the development of several BRM, such as mABs, IFNs, ILs, CSFs, TNF, and anticancer vaccines in the treatment of cancers but still there are many more avenues to be explored.
Copyright 2010 - Indian Journal of Cancer
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