The perfect drug is a ‘magic bullet’: totally specific, has good activity and pharmacokinetics, and no side effects. In the real world, however, many of the adverse events caused by medicines result from a lack of selectivity: while they have the desired killing, blocking or activating activity, they frequently damage healthy tissue, too. One way of targeting drugs more precisely is to use a monoclonal antibody, either for its own activity or to deliver another agent to the active site. A number of such antibodies are already on the market, and a huge number are in development for a variety of indications.

The market for monoclonal antibodies in Europe has been estimated at about US$1.5 billion for 2004, and Frost & Sullivan expects the total European antibody market to grow at a compound annual growth rate of 34.1% of $11.4 billion by 2011. TAs of January 2005, there were 10 monoclonal antibody therapeutics on the market in Europe. While chimeric MAbs currently dominate, humanised and fully human products are likely to increase in importance.

It is likely that over that period, oncology and autoimmune and inflammatory disorders (AIID) will continue to drive the market. Frost & Sullivan projects that oncology will retain the lead, with sales of $6.5 billion in Europe in 2011, followed by AIID indications at $4.5 billion. It adds that other indications – including cardiovascular disorders, organ transplantation and infectious diseases are likely to show less encouraging growth.

Similar growth is expected in the US. Frost & Sullivan reports that the market generated revenues of $8.7 billion in the past year; it projects that US sales will exceed $16 billion in 2012.

The immune system produces antibodies as a part of its defence strategy. These highly selective proteins bind specifically to other molecules – antigens – on the surface of cells, and then signal to other components within the immune system to destroy the target it is bound to. However, all too often the immune system cannot cope with the infection, and the antibodies it produces are insufficient to remove the infected or injured cells, and disease results.

Therapeutic antibodies, like natural ones, interact with antigens and trigger a biological reaction. However, unlike the body’s own polyclonal antibodies which bind to a variety of antigens, a therapeutic antibody is precisely targeted at a single antigen – either in an infectious agent or on a cell within the body in order to interfere with a cellular process.

Antibodies have a huge number of potential applications, from directly fighting infection, through tagging tumour cells for attack or directing them to self-destruct, to blocking ligands from activating receptors on diseased cells.

Some antigens – tumour associated antigens – are made by both normal and tumour cells, but there are also tumour specific antigens that are only ever produced by tumours. The latter are usually the result of some form of tumour specific mutation, and so present an ideal target for drug therapy. Unsurprisingly, a huge proportion of drug discovery effort involving antibodies is directed at fighting cancer, as tumour antigen specific drugs have great potential as anticancer agents that have minimal effects on normal cells. They are often MHC class I proteins, which are found on the cell surface and interact with killer T cell lymphocytes. They can also come in the form of mutated receptors, which can be recognised by B cells. Either way, they provide drug targets.

If some form of external antigen is introduced into the body, then B cells within the immune system will produce antibodies that bind to the antigen. Although each B cell only makes one type of antibody, other B cells make different antibodies that bind to different parts of the antigen. The resulting mixture of antibodies is termed polyclonal antibodies.

Making specific monoclonal antibodies is a little more complex. First, an animal – usually a mouse – is given an antigen challenge, and B cells are removed from its spleen. These B cells are fused with myeloma tumour cells that are unable to make their own antibodies, which are able to grow indefinitely in a culture, and are termed hybridomas. They multiply rapidly, and make large amounts of antibodies. By diluting them and growing them, a number of different colonies that each make just one antibody can be created. The hybridomas can also be grown in mice by injecting them into the peritoneal cavity, where they produce tumours that contain a fluid rich in antibodies.

The big drawback is that these antibodies are mouse specific, and these are invariably rejected by the human immune system. A number of ways to avoid these problems have been developed. The DNA that encodes the mouse antibody’s binding protein can be merged with DNA that produces human antibodies. This merged DNA can then be expressed in mammalian cell cultures, giving part-mouse, part-human antibodies. An alternative is to genetically engineer the mice so that they produce antibodies that are more like human ones.

The first monoclonal antibody to be given FDA approval was Ortho Biotech’s muronomab-CD3 (Orthoclone OKT3), which was licensed in 1986. It is used to treat rejection of transplanted organs. It is a murine monoclonal antibody to the CD3 antigen of human T cells, and is a biochemically purified immunoglobulin G2a. It is thought to reverse transplant rejection by blocking the function of all T cells that play a major role in acute rejection. It blocks the CD3 molecule in the membrane of human T cells that is associated with the antigen recognition structure of T cells, and is essential for signal transduction. When it binds to T lymphocytes, the result is early activation of the T cells, leading to cytokine release. The drug later blocks all known T cell functions, including those that cause rejection.

The earliest MAbs were so specific that the target population was, by definition, limited. However, there are now a variety of antibodies in clinical development that are being looked at to treat more than one condition, or different forms of the same disease.

An example of this is infliximab (Remicade) from Centocor, which blocks tumour necrosis factor [alpha]. It was first licensed in 1998 to treat Crohn’s disease, but has since been approved for rheumatoid arthritis, psoriatic arthritis and ankylosing spondylitis, all of which are inflammatory conditions mediated by TNF-[alpha].

Antibodies in cancer treatment

There are several ways in which antibodies can be used to treat cancer. The most obvious is to create a monoclonal antibody that binds to the tumour specific antigen, and induces an immunological response within the tumour cell. Another strategy is to use the MAb to deliver some form of cancer destroying agent specifically to the tumour cell. This could be a cytotoxic agent, a cytokine or a radioisotope, for example.

A huge number of MAbs are under development for treating cancer – and many of the antibodies already on the market are designed to combat one or more forms of the disease, almost all of them for some type of non-solid tumour. Rituxan (rituximab) is used to treat B cell lymphomas, binding to the CD20 molecule. This antigen is found on most B cells – both cancerous and healthy – but after treatment the healthy cells beging to grow again from precursors without the CD20 molecule and thus evaded destruction.

Campath (alemtuzumab) is designed to treat chronic lymphocytic leukaemia, as it binds to the CD52 antigen on white blood cells. It is currently also undergoing trials as a potential treatment for multiple sclerosis.

The treatment of some forms of aggressive breast cancer has been revolutionised by the introduction of Herceptin (trastuzumab). The antibody binds to HER2, which is a growth factor receptor found on a number of tumour cells, notably some breast cancers and lymphomas. It is unusual in that it is active against solid tumours.

Avastin (bevacizumab) binds to and blocks vascular endothelial growth factor (VEGF), which plays an essential role in the growth of the new blood vessels the tumour needs to survive and grow. Licensed for treating metastatic colorectal cancer, in combination with standard chemotherapy drugs, bevacizumab was the first drug that prevents this angiogenesis process to be approved.

One of the most talked about drugs in recent years – Erbitux (cetuximab) – is used to treat colorectal cancers. It targets the epidermal growth factor receptor, which is involved in the regulation of cell growth. It is thought to prevent cancer cells from growing by preventing epidermal growth factor from binding to the cells, thus stopping them from stimulating the cells to grow. It is ideally dosed along with the drug irinotecan.

Conjugated antibodies

Another way in which MAbs can be used to treat cancer is by delivering radioisotopes in a specific way to the tumour. An example, already on the market, is Biogen Idec’s Zevalin (ibritumomab tiuxetan), designed to treat non-Hodgkin lymphoma. Zevalin combines a MAb directed at the CD20 antigen with the radioisotope yttrium-90. CD20 antigens are found on malignant B lymphocytes in patients with NHL, as well as normal mature B lymphocytes. It delivers a therapeutic dose of the radioisotope to the B cells, and the [beta] emission from the radioimmunotherapy induces cellular damage through the formation of free radicals in the target and neighbouring cell. A diagnostic is also available, where the antibody is attached to the gamma emitter indium-111 prior to imaging studies.

In Zevalin’s drug form, the yttrium-90 is linked to the monoclonal antibody, and a powerful crossfire effect delivers radiation to nearby cells, along a mean path length of approximately 5mm, or a range of about 100 to 200 cells. The yttrium-90 has a half life of 64.1 hours.

However, yttrium-90 is less useful for solid tumours because of the time it takes for the radioisotope to penetrate into the tumour – up to 48 hours – means it is likely to have decayed by the time it reaches its target.

A second radioisotope – MAb combination, Bexxar (tositumomab) – is also available for the treatment of non-Hodgkin lymphoma. Here, the antibody is attached to iodine-131, which emits both beta and gamma radiation, with the beta radiation attacking the cancerous cells, and the gamma rays enabling imaging to be carried out, which allows the distribution and clearance of radiation from the body to be evaluated.

Monoclonal antibodies can also be used to target chemotherapy drugs. One of the biggest drawbacks of chemotherapy is the collateral damage that they cause to other tissues. A MAb could be used as a ‘magic bullet’ to direct the cytotoxic drug to the tumour cells instead of healthy ones. As with radioconjugated antibodies, this involves attaching an antibody to a cytotoxic drug or immunotoxon that will kill the cell once it manages to gain entry.

One example is Mylotarg (gemtuzumab ozogamacin). This targets the CD33 molecule, an antigen which is present on the surface of cancerous cells in acute myelogenous leukaemia, but not on the normal stem cells that are involved in the formation of healthy bone marrow. It delivers the complex oligosaccharide cytotoxic agent calicheamicin, which makes double stranded breaks in DNA. It is extremely toxic, so directing it to the cells in this way reduces the damage it causes to healthy cells.

Withdrawal problems

However, as with all drugs, there is always the risk that widespread use in patients after licensing will throw up side effects that cast doubt on a product’s future. This is just what has happened to Tysabri (natalizumab), a monoclonal antibody developed by Elan and Biogen Idec to treat multiple sclerosis. Brain lesions in MS patients are thought to result from autoimmune responses involving activated lymphocytes and monocytes. The glycoprotein [alpha]-4 integrin is expressed on the surface of these cells, and it is critical in the adhesion of the cells to the vascular endothelium, and migration into the parenchyma. Natalizumab Biogen is a humanised monoclonal antibody natalizumab that acts as an [alpha]4 integrin antagonist, and was shown to reduce the development in brain lesions in experimental models.

It was voluntarily withdrawn in February following reports of two serious adverse events in patients who had been given Tysabri in combination with Avonex (interferon [beta]-1a). Two patients developed progressive multifocal leukoencephalopathy (PML), a rare (and frequently fatal) demyelinating disease. Ongoing trials in MS, Crohn’s disease and rheumatoid arthritis have also been suspended, and since then a patient on the Crohn’s trial who had originally been diagnosed as having malignant astrocytoma has been reassessed as suffering from PML.

‘The MAb action mechanism is also dependent on biological milieus and is degraded by body systems and not metabolised in a manner akin to smaller chemical drugs,’ says Frost & Sullivan team leader Patrick Rajan. ‘This subsequently makes their long term safety and efficacy evaluation difficult and risky.’