Tag Archives: B-cell

A breakthrough against Chronic Lymphocytic Leukemia…thank the mice!

A challenge that science communicators frequently face when discussing the process whereby a scientific discovery eventually leads to a medical breakthrough is the time that this often takes, indeed by the time that the reports of exciting clinical trial outcomes start to appear in the press the role of the scientists who made the initial discoveries is often relegated to a passing comment…if it is mentioned at all. An example of this comes from the Weizmann Wave blog, produced by the Weizmann Institute of Science.

You may remember reports last month on the very promising results of a small clinical trial where a new immunotherapy technique was used to eradicate cancer cells in patients with Chronic Lymphocytic Leukemia (CLL), a blood cancer for which currently available treatments are often inadequate.  That trial, conducted by scientists at the University of Pennsylvania led by Professor Carl June, involved removing T-cells from the patient, treating the cells with a lentiviral vector that encodes for a Chimeric Antigen Receptor which recognises a protein named CD19 that is found on B-cells, including the cancer cells responsible for CLL, and then infusing the transformed T-cells back into the patients.  As the reported in the Los Angeles Times the results were dramatic, within a few weeks of the infusion the modified T-cells expanded rapidly and targeted the cancer cells in all three paients, so that a year later two of the three patients were still in complete remission.

It’s exciting stuff but as the Weizmann Wave reports the Press Release issued by Penn Medicine noted that this was a “cancer treatment breakthrough 20 years in the making” but “didn’t, however, explain those “20 years in the making.””. The Weizmann Wave goes on to discuss the pioneering basic scientific research undertaken by Professor Zelig Eshhar at theWeizman Institute of Science in the late 1980’s, which you can read about here.

Of course between the basic research undertaken by Prof. Eshhar and his colleagues in the 1980’s and the clinical trial whose outcome was announced last month there was a lot of work to be done. It would be impractical to describe all the different discoveries that made this immunotherapy possible, but one discovery in particular highlights the importance of animal research to this breakthrough.

There have been previous attempts to use Chimeric Antigen Receptors to target T-cells to attack cancer, but these had disappointing results in clinical trials.  A major improvement made by the University of Pennsylvania team was to include an additional motif – named the CD137 co-stimulatory molecule- which greatly enhances the cancer killing ability of the infused T-cells.  In a recent paper published in the Journal of Cancer the University of Pennsylvania team point out that the decision to include CD137 (called 4-1BB in mice) in their Chimeric Antigen Receptor construct was based on promising results in studies undertaken in mice:

 Our group has tested a CAR directed against CD19 linked to the CD137 (4-1BB) co-stimulatory molecule signaling domain to enhance activation and signaling after recognition of CD19. By inclusion of the 4-1BB signaling domain, in vitro tumor cell killing, and in-vivo anti-tumor activity and persistence of CART-19 cells in a murine xenograft model of human ALL (acute lymphocytic leukemia) is greatly enhanced”

Indeed, in a paper published by Professor June and colleagues in the journal Molecular Therapy in 2009 they describe this work in much more detail, highlighting just how groundbreaking the results were:

Previous in vitro studies have characterized the incorporation of CD137 domains into CARs.10,11,29 Our results represent the first in vivo characterization of these CARs and uncover several important advantages of CARs that express CD137 that were not revealed by the previous in vitro studies. We demonstrated that CARs expressing the CD137 signaling domain could survive for at least 6 months in mice bearing tumor xenografts. This may have significant implications for immunosurveillance, as well as for tumor eradication. For example, in a mouse prostate cancer xenograft model, survival of CAR+ T cells for at least a week was required for tumor eradication.30

Long-term survival of the CARs did not require administration of exogenous cytokines, and these results significantly extend the duration of survival of human T cells expressing CARs shown in previous studies.17,31 To our knowledge, this is the first report demonstrating elimination of primary leukemia xenografts in a preclinical model using CAR+ T cells. Furthermore, complete eradication was achieved in some animals in the absence of further in vivo therapy, including prior chemotherapy or subsequent cytokine support.

The long-term control of well-established tumors by immunotherapy has rarely been reported. Most preclinical models in a therapeutic setting have tested tumors that have been implanted for a week or less before initiation of therapy.32 After establishing leukemia 2–3 weeks before T cell transfer, we found that many animals had long-term control of leukemia for at least 6 months. The efficacy of targeted, adoptive immunotherapy in this xenograft model of primary human ALL compares favorably to our prior experience testing the antileukemic efficacy of single cytotoxic (ref. 27 and data not shown) or targeted agents,26 where we have observed extension of survival but not cure of disease. Additionally, we have not previously observed the ability to control xenografted ALL for a period of as long as 6 months.”

These results led directly to the clinical trial reported last month.

So there you have it, behind the headlines are years of graft by hard-working and innovative scientists, who utilised a wide range of experimental approaches – among which animal studies figure prominently – to develop a novel therapy for CLL. As Professor Bruce Levine points out in the video above, the key to success is often keeping one hand in the basic research lab and the other in the clinic.

Paul Browne

Addendum: Scienceblogger Erv has written an excellent commentary on this study

Taming the Wolf: a new treatment for Lupus

Earlier today we posted a commentary on PeTA’s misleading propaganda by Professor Anthony Garro of UMass Dartmouth.   At the time I mentioned that it was a pity that Prof. Garro was not able to write more about the role of animal research in 21st century medicine, but a recent story in Nature News provides an excellent example, showing how research on mice and monkeys was crucial to the development of a new drug for lupus.

The autoimmune disease lupus, or to give it its full name Systemic Lupus Erythematosus (SLE), affects over 100,000 people in the United States, causing damage to a variety of tissues in the body and a wide range of symptoms ranging from fever, headache and  joint pains to anemia and renal failure. While there is no cure for lupus it can be treated successfully, though current treatments do not work well for all patients. Continue reading

From Mouse to Monkey to Humans: The Story of Rituximab

Modern advances in science have meant that our models of diseases have vastly improved. Be that in a dish in the laboratory, a computer simulation or through using a transgenic mouse, there have been developments across the biomedical field that have given us a greater understanding of diseases and how our bodies work.

This increase in knowledge has meant that we are finding may drugs already on the market can treat a variety of diseases – those involving the same pathway or cell type. This is precisely what happened this month with a drug called Rituximab.

Rituximab was licensed in 1997 for use in the treatment of Non-Hodgkin’s lymphoma (NHL) – a cancer where cells of the immune system called B-cells mutate and divide abnormally. The cancer then spreads around the body when the B-cells clone themselves in replication.

Since it’s initial approval for use in NHL, rituximab has been used to successfully treat advanced rheumatoid arthritis and has also been part of anti-rejection treatments for kidney transplants (both involve B cells. Then news came last week that it could even slow the progression of rheumatoid arthritis (RA) in the early stages of the disease.

Rheumatoid Arthritis

Rituximab is an interesting drug, as it is a chimeric antibody. This means that it contains portions of both human and mouse antibodies mixed together. The first papers reporting on rituximab were published in 1994. The first looked at its creation, and the second reported on the phase I clinical trials of the drug.

The human immune system works by using antibodies as their ‘messengers’. The antibodies contain multiple regions that allow them to work effectively. One part of the molecule binds with the foreign molecule; the other part then recruits the immune cells to destroy the molecule and eliminate it from the body.

The B-cells mutated in NHL and involved in RA are part of the human immune system and are responsible for making antibodies against ‘foreign invaders’. Mature forms of B-cells have a protein called CD20 on their surface.

The protein CD20 was the target for a team in San Diego (1) in 1994. Because NHL and RA are characterised by excessive levels of, or mutated B-cells, they looked at ways to reduce their numbers. The researchers determined that CD20 was the perfect target on the human B-cells as it was located on the surface of the cell and it didn’t mutate, move inside the cell or fall off in the life cycle of the B-cell. The team then produced an antibody that would attack CD20 itself, so it would bind to the outside of B-cells, flagging them to the immune system to be eliminated. They identified a mouse antibody that had high anti-CD20 activity.

They then constructed a “chimeric” antibody containing the variable domain of the mouse antibody, the portion that specifically binds CD20, along with the constant domain of human antibody, the portion that recruits other components of the immune system to the target.

The construction of a chimeric antibody (later named rituximab) was crucial, as the mouse antibody was unsuitable for direct use in humans. While the mouse antibody was able to bind to human CD20, it would not be able to then recruit the human complement system and immune cells that are needed destroy the “targeted” B cells. It would also quickly be recognised as foreign in the human body, and destroyed by the immune system, therefore by using a chimeric antibody with enough human characteristics, the antibody would not only recognise the human CD20 and target the immune system to it but would remain in the body long enough to destroy the B cells.

To test whether rituximab would work as hoped, they performed studies in cynomolgus monkeys. They choose this species because the constant domains of their antibodies are very similar to those in humans, unlike those of the mouse, allowing the chimeric antibody to function as it would in humans. Following administration of rituximab the number of B cells in the monkey’s bloodstream fell dramatically. The numbers were also reduced in the bone marrow (where B cells are produced) and the lymph nodes (where they are activated to target foreign molecules). Rituximab administration was non-toxic and in the weeks after treatment finished the number of B-cells slowly recovered. This is important as it demonstrates that the treatment didn’t harm the monkey’s bone marrow stem cells, an important consideration since these cells are required for a healthy immune system.

Rituximab was an ideal candidate to treat NHL and the promising results in monkeys prompted the scientists to conduct phase I clinical trials inhuman patients which confirmed that rituximab was safe and indicated that it could shrink tumors.

Evaluation of the effectiveness of rituximab involved many studies of patients with Non-Hodgkin’s lymphoma. While the initial clinical trial results varied, likely due to the differing sizes of tumors between the patients, they showed it was effective at reducing B-cell numbers and tumor size. Since it’s approval numerous clinical trials have confirmed that rituximab is an effective treatment for Non-Hodgkin’s lymphoma (3).

This month’s exciting study by Professor Paul-Peter Tak from the University of Amsterdam showed that rituximab in combination with the drug methotrexate could slow the progression of early stage rheumatoid arthritis (RA).

The study involved 755 patients diagnosed with RA within the last year. Methotrexate is already considered to be the best treatment for these patients and 12.5% of the patients taking only methotrexate in this study experienced significant reduction of their symptoms. However, compare this to the 30.5% of patients taking a combination of methotrexate and rituximab, and it is clear that rituximab is effective. Issues of cost have been raised in relation to rituximab, but if it turns out to be as effective in treating early RA as this study suggests, then it may ultimately save the health services and insurance companies money as slowing or stopping the progression of the disease will result in fewer patients needing the more expensive treatment and care required in advanced RA.

Emma Stokes

1) Reff M.E. et al. “Depletion of B cells in vivo by a chimeric mouse human monoclonal antibody to CD20.” Blood Volume 83(2), Pages 435-445 (1994) PubMed: 7506951

2) Maloney D.G. et al. “Phase I clinical trial using escalating single-dose infusion of chimeric anti-CD20 monoclonal antibody (IDEC-C2B8) in patients with recurrent B-cell lymphoma.” Blood Volume 84(8), Pages 2457-2466 (1994) Pubmed: 7522629

3) Schultz H. et al. “Chemotherapy plus Rituximab versus chemotherapy alone for B-cell non-Hodgkin’s lymphoma.” Cochrane Database of Systematic Reviews 2007, Issue 4. Art. No.: CD003805. DOI:10.1002/14651858.CD003805.pub2.