oncology – Page 2 – Drug Discovery Opinion

Once Bitten…

The concept of bispecific antibodies – monoclonal antibodies able to recognise and engage two different antigens – has been explored for over twenty years. Development of therapies based on the approach has, however, been hampered by difficulties in their construction, poor efficacy and undesirable side-effects.

One particular subset of bispecific antibodies, the so-called bispecific T-cell engager (BiTE®), has nevertheless begun to show promise. Blinatumomab, developed by Micromet, targets the CD19 receptor of B-cells and CD3 on T-cells and is designed to direct cytotoxic T-cells to B-cell tumours. Interim data from a phase I trial in Non-Hodgkin’s Lymphoma patients have shown signs of clinical efficacy and additional clinical trials in acute lymphoblastic leukemia (ALL) and chronic lymphocytic leukemia (CLL) are ongoing.

More recently, the Micromet team have reported on preclinical data using BiTE® antibodies targeting the EGFR receptor and CD3, incorporating the binding domains of either panitumumab or cetuximab. Panitumumab and cetuximab, as well as EGFR kinase inhibitors, are marketed for treatment of colorectal cancer (CRC) and primarily inhibit CRC growth by interfering with EGFR signalling. CRC patients whose tumours have mutated KRAS or BRAF, however, are resistant to treatment. This latest study, published in Proceedings of the National Academy of Sciences, showed that both EGFR-specific BiTE® antibodies mediated potent redirected lysis of KRAS– and BRAF-mutated CRC lines by human T cells at subpicomolar concentrations. The cetuximab-based BiTE® antibody also inhibited growth of tumours from KRAS– and BRAF-mutated human CRC xenografts, whereas cetuximab was not effective. The researchers also report preliminary safety data in non-human primates and conclude that EGFR-specific BiTE® antibodies may have potential to treat CRC that does not respond to conventional antibodies.

NSAIDs and Cancer – Another Piece of the Puzzle

Regular use of NSAIDS has been linked to reduced incidence of certain types of cancer but the underlying protective mechanisms are unclear. Some of the anticancer effects are believed to be mediated through inhibition of COX-2, but a study led by investigators at Sanford-Burnham Medical Research Institute has now identified another mechanism by which the sulindac sulfide (the NSAID metabolite of sulindac) inhibits tumour growth. The team found that sulindac sulfide induces apoptosis by binding to retinoid X receptor-α (RXRα), a member of the nuclear hormone receptor family which had been already been identified as a potential target for cancer therapy. In cancer cells, levels of RXRα are often reduced, at least in part because of proteolytic processing to a truncated form, tRXRα. As with other nuclear receptors, RXRα regulates transcription of target genes by binding to DNA response elements but accumulating evidence suggests that RXRα may also have extranuclear activity. Both RXRα and tRXRα can exist in the cytoplasm and the study showed that cytoplasmic tRXRα can activate the PI3K/AKT survival pathway by interaction with the p85a subunit of PI3K, leading to anchorage-independent cell growth in vitro, and tumour growth in animals. Sulindac sulfide was found to inhibit the tRXRα-mediated PI3K/AKT activation, suggesting that the compound could provide a useful lead for anti-cancer drugs targeting this pathway.

The use of NSAIDs to reduce the incidence of cancer has been limited by the risk of major cardiovascular events and the Sanford-Burnham have identified an analogue of sulindac sulfide, K-80003 which has improved affinity for RXRα but lacks significant COX-2 inhibitory activity. K-80003 inhibited the growth of cancer cells in vitro and in animals and would be expected to have reduced COX-2-associated side effects.

The study is published in the journal Cancer Cell.

Role for PEAK1 Kinase in Tumour Metastasis

The cytoskeleton plays a key role in regulating many cellular functions; it maintains cell shape, protects the cell, enables cellular motion, and has important roles in proliferation and differentiation. Metastasising cancer cells exploit the cytoskeleton to produce protrusions that allow them to invade surrounding tissue and enter the blood system from where they can spread to distant tissues and seed new tumours.

The protrusions, known as pseudopodia, are highly specialised ‘feet’ that the cell uses to pull itself forward across the underlying surface. A team led by researchers at the University of California, San Diego has now identified a previously unknown kinase – termed pseudopodium-enriched atypical kinase one or PEAK1 – that regulates the cytoskeleton and plays a central role in the formation of pseudopodia. Preliminary studies in mice suggest that PEAK1 is important during tumour growth and the team also showed that PEAK1 levels are increased in primary and metastatic samples from human colon cancer patients. Whether PEAK1 is capable of transforming non-tumour cells into cancer cells has not yet been determined but the fact that PEAK1 has kinase activity suggests that it may be possible to design specific inhibitors which could help to elucidate its role in both normal and cancer cells. PEAK1, which is a 190-kDa non-receptor tyrosine kinase, could serve as a clinical biomarker that predicts whether a cancer is likely to metastasise and could also be a target for future cancer treatments.

The study is published in the Early Edition of the Proceedings of the National Academy of Sciences.

CD95: A Question of Life or Death

death becomes you by Esther Kirby — Image: Flickr - Esther Kirby

Programmed cell death (apoptosis) is essential to maintain homeostasis within living organisms and is controlled by a variety of intra- and extra-cellular signals. Activation of the death receptor CD95 (also known as Fas or Apo-1) by its physiological ligand, CD95 ligand (Fas ligand), leads to apoptosis in many tissues and is especially important in the immune system.

Resistance to apoptosis is also key to the survival of malignant cells but the role of CD95 in cancer progression is complex: although the down-regulation of CD95 that is frequently observed in tumour cells could contribute to their survival, complete loss of CD95 is rare in human cancers. On the other hand, many cancer cells express large quantities of CD95 and are highly sensitive to CD95-mediated apoptosis in vitro. Cancer patients often have high levels of CD95 ligand, suggesting that CD95 could perhaps promote the growth of tumours through non-apoptotic activities. Scientists from the University of Chicago and Northwestern University Feinberg School of Medicine have further investigated the role of CD95 in several human cancer cell lines and in mouse models of liver and ovarian cancer. Cancer cells – regardless of their sensitivity to CD95-mediated apotosis – were found to produce CD95 ligand and to depend on constitutive activity of CD95 for optimal growth. In the mouse models, deletion of CD95 reduced both the incidence and size of tumours. In further studies, the tumour promoting activity of CD95 was shown to be mediated by pathways involving JNK and Jun but not caspase-8.

The study, which is published in Nature, suggests that, paradoxically, reducing rather than enhancing activity of the death receptor CD95 may be an effective way to control the growth and proliferation of cancer cells. Further research is needed to understand the switch between signalling ‘die’ and ‘grow’, but eventually soluble CD95 or antibodies against CD95 ligand could find a role in the treatment of cancer.

Loss of microRNA in Malignant Mesothelioma

Scanning electron micrograph showing asbestos fibres; Source – Wikimedia Commons

MicroRNAs (miRNAs) are small (21-23 nucleotides), single-stranded RNA molecules that function as regulators of gene expression. The human genome encodes several hundred miRNAs and abnormal expression of these has been associated with cancer progression. We have previously reported on miRNA involvement in cholesterol regulation, amyotrophic lateral sclerosis and liver cancer. Now a collaboration between Rosetta Genomics, NYU Langone Medical Center and Vanderbilt School of Medicine has identified the potential utility of miR-33 for development of therapies targeting malignant mesothelioma (MM).

MM is a rare cancer that has been associated with exposure to asbestos dust (although a small proportion of patients have never been exposed). Taking anywhere between 20 and 50 years for symptoms to develop, the cancer affects the mesothelium, a thin layer of tissue surrounding the internal organs. The most common form of MM is pleural, involving the lining of the lungs, but it may also affect other tissues such as the peritoneum. Current treatment options, including surgery and chemotherapy, are limited and often confounded by late diagnosis because of the absence of symptoms.

This latest study found that MM cell lines derived from patients with aggressive disease failed to express miR-31, a microRNA that has also been linked to suppression of breast cancer metastasis. Functional studies, where miR-31 was reintroduced to the cells, showed that the microRNA could inhibit proliferation, migration, invasion, and clonogenicity of mesothelioma cells. miR-31 suppressed expression of a number of factors associated with maintenance of DNA replication and cell cycle progression, including the pro-survival phosphatase PPP6C. The mRNA for PPP6C, which contains three miR-31-binding sites in its 3′-UTR, was down-regulated when miR-31 was present and up-regulated in clinical MM specimens compared to matched normal tissues.

Whilst the study, published in the Journal of Biological Chemistry, reveals a key role for miR-31 in MM, considerable challenges remain to exploit this finding for therapy of the disease.

It May Not Be the Clock that’s Wrong

To ensure normal growth and avoid tumour formation, cell division must be tightly regulated. Remarkably, many cells from species as diverse as single celled organisms and humans only divide at certain times of the day, suggesting that division is under the control of a circadian clock. The evolutionary explanation put forward for this is a selective advantage for organisms with cells that divide at night when the mutational effects of ultraviolet light are lowest.

It has been proposed that the uncontrolled growth shown by tumour cells is caused by fault in the biological clock but a study by researchers at Vanderbilt University has now shown that although immortalised rat fibroblasts have functioning clocks, the clocks don’t control the rate at which the cells divide and grow. Similar results were seen in preliminary experiments with lung cancer cells. If follow-up studies confirm that control of cell cycle by circadian rhythm has been lost in immortalised cells, the research may suggest new targets for cancer therapy.

The study is published in PNAS.

Stopping Cancer Spread

Fascin crystal structure — Human fascin complexed with macroketone, PDB ID=3LNA

The deadliest feature of cancer is its ability to spread, or metastasise. Migrastatin, a compound isolated from Streptomyces, was found to weakly inhibit tumour cell migration and, in 2005, researchers from Weill Medical College of Cornell University and the Sloan-Kettering Institute for Cancer Research described simplified analogues of migrastatin, including a compound they called macroketone, that inhibit mammary tumour metastasis in mice. Although the compounds were effective in preventing the spread of cancer cells, it wasn’t known how they achieved this. In a new study, published in the journal Nature, the team have revealed that macroketone exerts its anti-metastatic effect by targeting the actin-bundling protein, fascin. Cancer cells use invasive finger-like protrusions called invadopodia to spread into and degrade extracellular matrix and recent studies have shown that fascin is important for their assembly and stability.

Mice implanted with cancer cells and treated with macroketone lived out a full lifespan without any spread of the cancer whilst untreated animals all died from metastases. When treatment was delayed for one week after introduction of the cancer cells, metastasis was still blocked by more than 80%. Macroketone did not prevent implanted cancer cells from forming tumours or growing, suggesting that such compounds would need to be used in combination with chemotherapy drugs acting on the primary tumour. Because fascin is overexpressed in metastatic tumour cells but is only expressed at very low levels in normal epithelial cells, treatments that target fascin should have comparatively little effect on normal cells and may have fewer side effects than other treatments.

X-ray studies showed that macroketone binds to one of the actin-binding sites on fascin which prevents the actin fibres from bundling together and could form the basis for further drug design.

Lipid Genes Linked to Cancer

Although clinical and epidemiological studies have linked cancer with other chronic conditions such as inflammatory and metabolic diseases, the pathways linking different diseases are poorly understood. Inflammation is commonly associated with the development and progression of cancer and increased cancer risk is also linked to metabolic syndrome which encompasses obesity, type II diabetes, high cholesterol, and atherosclerosis. To overcome limitations in previous approaches to identifying common genes and signalling pathways, researchers at Harvard Medical School have carried out transcriptional profiling in two experimental isogenic models of cellular transformation. Using two isogenic models in which the transformed and non-transformed tissues are genetically identical makes it is easier to identify genes involved in transformation.

The team used a computational approach to organise the genes identified by transcriptional profiling into networks with central nodes. Comparison with a gene set describing metabolic syndrome revealed a high overlap between the central nodes of cancer and metabolic syndrome. Inflammatory factors such as IFN-γ, IL-1β, IL-6, and NF-κB as well as insulin and low-density lipoprotein (LDL) appeared as central nodes in cancer gene networks, suggesting the importance of inflammatory processes in both cancer and metabolic diseases and also a link between protein and lipid metabolism and cellular transformation. Lipid-related genes that had not previously been linked to cancer included OLR1, SNAP23, VAMP4, SCD1, SREBP1 and GALNT2.

The similarities between the pathways in cancer and metabolic diseases led the team to test whether drugs used to treat inflammation or aspects of metabolic disease might also affect cellular transformation and tumorigenicity. Four of the compounds that performed best in the cell experiments, metformin, sulindac, simvastatin, and cerulenin were tested for their ability to suppress tumour growth in nude mice: tumour growth was completely suppressed by metformin and sulindac and significantly delayed by cerulenin and simvastatin, suggesting that drugs designed to combat metabolic diseases may also be useful in treating some types of cancer.

The study is published in Cancer Cell.

Enhancing Drug Penetration of Tumours

perforation — Image: Flickr - elektroholunder

One problem associated with the treatment of solid tumours is that chemotherapeutic agents have difficulty in penetrating more than a few cell diameters from the vasculature. Higher doses of drug are required and, because some tumour cells are not reached, the risk of recurrence is high. To address this issue, researchers at Sanford-Burnham Medical Research Institute have now described a peptide able to enhance the ability of drugs to access tumour tissue.

Some years ago it was shown that peptides containing the RGD (Arg-Gly-Asp) sequence recognise a family of cell-surface receptors, integrins, which mediate the interaction of cells with the extracellular matrix (ECM) components fibronectin and type I collagen and are important for the migration and invasion of tumour cells. RGD peptides have been used for homing to malignant tissue and the Sanford-Burnham team have taken this a step further. The new agent is a cyclic nona-peptide (CRGDK/RGPD/EC), referred to as iRGD. The contained RGD sequence targets the agent to tumour tissue where it is cleaved to reveal a ‘CendR’ sequence that binds to neuropilin-1, mediating an active transport system.

In a paper published in Cancer Cell late last year, the research team showed that coupling iRGD to anti-cancer drugs allowed them to penetrate deep into tumours, effectively increasing the activity of the drugs. In their latest study, published in Science, the team have shown that the chemotherapeutic agents do not need to be conjugated to the peptide. Co-administration of iRGD with a variety of drugs, including a small molecule (doxorubicin), nanoparticles (nab-paclitaxel and doxorubicin liposomes), and a monoclonal antibody (trastuzumab), improved their therapeutic index.

The team hope that iRGD may be a valuable adjunct to enhance efficacy of anti-cancer agents, whilst reducing side-effects.

The Mysteries of Arsenic

arsenolite — Arsenolite

Image: Wikimedia Commons

For many people, the word ‘arsenic’ conjures up thoughts of murder mysteries and, in fact, arsenic has been a popular murder weapon since the middle ages. In the Victorian era, arsenic trioxide found favour as a cosmetic and it has also been used in both Chinese and Western medicine. Most recently, arsenic trioxide has been used to treat acute promyelocytic leukaemia (APL) that is unresponsive to first line therapies.

Arsenic trioxide is able to induce complete remission in patients with relapsed or refractory APL and is generally well-tolerated with minimal chemotherapy-related side effects. How arsenic trioxide kills cancer cells is not clear but scientists in China and France believe they have made a key step towards solving the mystery. APL cells are characterised by the occurrence of chromosomal translocations involving the retinoic acid receptor α gene (RARα)and the promyelocytic leukaemia gene (PML). These translocations lead to production of a fusion protein, PML-RORα that has altered functions and protects the cells from apoptosis. Arsenic trioxide triggers small ubiquitin-like modifier (SUMO) proteins to tag PML-RORα as part of a degradation mechanism. The new study has shown that the arsenic binds directly to cysteine residues in zinc fingers located within RBCC (N-terminal RING finger/B-box/coiled coil) domains of PML causing cross-linking and oligomerisation. The aggregated protein then undergoes SUMO modification and degradation. The identification of PML as a direct target of arsenic trioxide provides new insights into how the drug is able to treat APL and may lead to new treatment options.

The study is published in the journal Science.