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Revamping Drug Discovery Strategies

dvances in finding novel drug targets in such areas as pain and cancer as well as inflammatory and CNS degenerative diseases were key topics discussed at IBC's annual "Drug Discovery & Development of Innovative Therapeutics" conference in Boston.

Regarding peripheral pain, for example, novel targets based on the role of acidosis, or relatively lower pH in tissues around peripheral pain receptors, have emerged. These targets, proton activated receptors composed of six protein subunits, have been the focus of a small molecule drug discovery program at PainCeptor Pharma (www.painceptor.com). The firm develops therapies for acute and chronic pain focused on parasympathetic nervous system (PNS) targets, acid-sensing ion channels (ASICs), and nerve growth factors.

PainCeptor Pharma currently has small molecule programs targeting NGF/p75 TrkA and ASICs that are in late preclinical development. "By focusing on PNS receptors and ion channels we expect to avoid many of the side effects associated with centrally acting pain therapeutics," explained Kazimierz Babinski, PainCeptor's vp of corporate affairs. "The idea is to focus on targets at the beginning of the pain cascade."

PainCeptor's closest-to-the-clinic compound, PPC-5650, targets ASIC receptors, a class of ion channels that encode noxious stimuli into electrical impulses through ion flow across the cell membrane. The company identified its small molecule drug candidate from compound libraries with high-throughput screening against cell lines expressing a recombinant version of the ASIC receptor subtype ASIC1a.

These recombinant ASICs mimic native proton-gated currents recorded from sensory neurons. "By measuring proton-gated currents with the patch-clamp technique, we validate compounds based on their ability to block ion flow through ASIC channels into neuronal cells," Babinski said. "Tissue acidosis is a feature of many painful pathologies and disease states including inflammation, osteoarthritis, cardiac ischemia, bone cancer, and skin incisions. By potently and selectively blocking these acid-sensing ion channels with our lead compound, we have been able to block pain in several animal models."

It is important to note, he added, that PPC-5650 does not affect basal nociception, which is the ability to respond to a noxious stimulus such as heat (a defense mechanism), but blocks the exacerbated pain sensation (hyperalgesia) associated with chronic conditions such as inflammation or injury.

In addition to the ASIC program, PainCeptor has also developed several series of potent small molecules that interact with nerve growth factor (NGF) and prevent NGF from binding to its p75/TrkA receptors. PainCeptor is developing these small molecule NGF antagonists for oral administration. Of these compounds, PPC-1807 is closest to the clinic.

The company anticipates commencement of the exploratory-IND studies and the initiation of the first human proof-of-concept studies on the ASIC program later in 2007. NGF antagonists are currently completing IND-enabling preclinical studies.
Cancer Stem Cells

Conference speakers in the cancer drug development track offered some frustrated commentary about progress in cancer drug development, emphasizing the need to consider completely new drug targets. Several speakers pointed out that most oncology drugs that fail do so late in the drug development cycle, typically in Phase III trials, and that few cancers are cured or have been transformed by drug therapy, despite decades of research. One reason put forth for this is that current chemotherapeutic drugs target the wrong cancer cells.

OncoMed Pharmaceuticals' (www.oncomed.com) vp of cancer biology, Timothy Hoey, Ph.D., described his company's isolation, characterization, and validation of cancer stem cells (CSCs) as potential therapeutic targets. Oncomed's scientific founders, Michael Clarke, M.D., and Max Wicha, M.D., originally demonstrated the presence of these cells in solid tumors of epithelial origin in a 2003 study of breast cancer. Since then, CSCs have been identified in other solid tumors including prostate, brain, colon, and pancreatic cancers.

While normal human embryonic stem cells are healthy cells found in fetal embryos that give rise to normal body tissues and organs, CSCs comprise a small fraction of tumor cells and, among tumor cells, uniquely have the capacity for self-renewal.

"A limited subpopulation of tumor cells can drive tumor growth and generate tumor heterogeneity and have the ability to regenerate a tumor when transferred to mice," Dr. Hoey noted. "We now have evidence that these CSCs preferentially survive chemotherapy."

According to Dr. Hoey, OncoMed has isolated and characterized CSCs from a variety of tumors and is developing novel therapeutics targeting these cells. In collaboration with Dr. Clarke's lab, they also developed an invasiveness gene signature from CSCs that provides prognostic insight into patient outcome.

In his presentation Dr. Hoey described techniques for manipulation and quantitation of the tumorigenic potential of these cells. In order to validate CSC targets, he and his colleagues had to overcome several technical hurdles including the ability to culture them while retaining their tumorigenicity; manipulating them, for example by introducing exogenous genes and shRNAs; and the ability to study single CSCs to monitor their fate and potential.

OncoMed isolates CSC populations in sufficient numbers for study by FACS (fluorescence activated cell sorting) and then infects isolated CSCs in vitro with lentiviral vectors encoding a reporter gene that would allow identification of CSC-dependent tumor growth in vivo.

Tumors produced in mice by the labeled CSCs allowed scientists to distinguish among tumors that were responsive or unresponsive to various shRNAs, overexpressed proteins, or drug candidates.

"We have tested about 150 antibodies in xenograft models derived from CSC-driven human tumors and identified a number of antibodies with reproducible antitumor activity," Dr. Hoey reported. "In our most advanced project, we have discovered an antibody that binds with high affinity and selectivity to its target and functions to completely block ligand-receptor binding and subsequent signaling in CSCs. This antibody has shown activity in a variety of tumor models both as a single agent and in combination with approved cancer treatments."
Gene Splice Variants

Matt Pando, Ph.D., vp of R&D at ExonHit Therapeutics (www.exonhit.com), described the use of his company's SpliceArray™ technology to explore and discover unique therapeutic targets for Parkinson's disease. ExonHit uses its gene-profiling technology to identify the deregulation of gene splice variants that can result in altered protein function and contribute to or possibly trigger disease development.

"A major limitation with standard expression profiling microarray designs is that they produce a composite expression score from a specific gene locus query and can't monitor individual alternative isoforms produced from a single locus. Therefore, specific transcripts that result from alternative splicing events can occur under different conditions, for example during oxidative stress, and can't be detected. Our patented SpliceArray design allows us to monitor individual transcripts being transcribed from a given gene."

According to Dr. Pando, the company has taken the genomic sequence and aligned all of the publicly available human expressed sequences that met certain quality parameters. "We then looked at the resulting alignment data with a proprietary algorithm that identifies differences in sequence content between overlapping transcripts. These structural differences correspond to the alternative splice events.

"The algorithm then produces full annotations describing the identified splice events, produces splice site scores, and extracts appropriate target seque