Using Nell-1 to harness the osteogenic potential of adult stem cells

Funding Type: 
Disease Team Research I
Grant Number: 
DR1-01477
ICOC Funds Committed: 
$0
Disease Focus: 
Solid Tumor
Cancer
Stem Cell Use: 
Cancer Stem Cell
Public Abstract: 
Traumatic injuries are responsible for over 15% of the global health burden. Many are musculoskeletal injuries that, if untreated, can cause long-term disability. Bone regeneration is a natural process that is critical for fracture repair and for the healing of bone grafts used during skeletal reconstruction. However, aging and metabolic bone diseases like osteoporosis can negatively affect the bone regeneration capacity by reducing stem cell numbers and cell activity. The failure to heal fractures in osteoporotic patients, for instance, has been reported to be as high as 50%. Osteoporosis is by far the most common systemic disease of skeletal fragility. Up to 50% women and 20% men over age 50 will suffer an osteoporosis-related fracture in their remaining lifetime. Furthermore, osteoporosis is frequently associated with other diseases like i] spinal cord injury, in which fractures are the result of limb disuse, ii] Parkinson’s disease and multiple sclerosis, in which there is an increased risk of life threatening fractures from falls, iii] malignant neoplasms invading bone that increase pathologic fracture risk and iv] organ transplant patients on chronic steroid-mediated immunosuppression. Due to changing demographics and increased life expectancy, annual osteoporotic fracture costs are projected to increase from $20 to $240 billion worldwide by 2040. Bone regeneration therapies can be cell- or non-cell based. Cell-based therapies either take the patient’s own cells (which causes significant pain and discomfort) or use cells from deceased donors. The latter requires extensive processing to remove immune components, and do not work as well as the patient’s own bone and stem cells. Non-cell based therapies are either less effective (deceased donor bone with no cells) or have undesirable effects (e.g., bone morphogenetic proteins can cause life threatening neck swelling). Current therapies also do not address the needs of osteoporotic populations where the healing environment is suboptimal and/or bone harvest from the patient’s own hip can further damage already weak bone. To address the unmet medical need, our team has developed an adult stem cell product that exceeds the efficacy and safety of current bone repair therapies. Stem cells hold great promise in their ability to regenerate damaged tissues. However, until now, stem cells have been difficult to identify and isolate in numbers sufficient for medical use without cumbersome cell culture. We have developed a highly effective and reproducible method to isolate and purify high numbers of a person’s own stem cells from fat tissues obtained through liposuction, a relatively safe and common cosmetic surgery procedure. In addition, we have developed a highly potent and bone-specific growth factor to help these stem cells form bone. Combined, the stem cell-growth factor product forms an environment that can significantly regenerate bone in normal as well as osteoporotic patients.
Statement of Benefit to California: 
This well developed proposal aims to enable IND submission for the use a novel growth factor to direct the differentiation of human perivascular stem cells within a osteogenic carrier for bone regeneration. This highly multi-disciplinary project has many near-term and long-term benefits to the State of California. 1. California’s climate is attractive to people all ages, particularly the elderly. Unfortunately, a large fraction of the aged population suffers from osteoporosis. In 1998, the health care burden for osteoporosis exceeded $2.4 billion in California alone. A whopping 64% of the $2.4 billion was caused by hip fracture. By promoting the repair of both normal and healing-impaired bone in a safe and effective manner, our mature technology will reduce the long term health care burden for California’s public health insurance program. 2. Besides direct health cost, bone injuries and diseases can result in hospitalizations and long term disabilities that can lead to sick days and lost productivity. The hard working Californians are responsible for California’s annual gross domestic product of $1.8 trillion, which rank our state among the top eight largest economies in the world. By promoting the repair of both normal and healing-impaired bone in a safe and effective manner, our mature technology will reduce the loss of work productivity at the front end, reduce work disability costs, and reduce the loss of state income tax. 3. California boasts a highly diverse population with numerous ethnic groups. While this diversity offers amazing opportunities for cross cultural discoveries, some ethnic groups (e.g. Latino and some Asian descents) may be disadvantaged in terms of donor matches for traditional stem cell sources. Our stem cell therapy can benefit these minorities and reduce the long term health care burden for California’s public health insurance program. 4. This project directly adds jobs at {REDACTED} and at the California-based companies that are involved in this project. 5. This project will procure supplies and equipment from strategic California-based companies. 6. This project will produce intellectual property that is owned by the {REDACTED}. 7. This mature project is precisely the type of cutting-edge, ethical, multi-disciplinary stem cell project that Californians imagined when they approved proposition 71 in 2004. The establishment of CIRM has transformed the research infrastructure at {REDACTED}, increased our ability to recruit world class stem cell scientists, and attracted the attention of superb scientists from other disciplines to this new field. Working together, our team has compiled an impressive list of accomplishments and we are confident in our abilities to take this project to IND submission in a timely fashion. Funding of this project will fulfill the promise of proposition 71.
Progress Report: 
  • The objective of our collaborative project is the development of therapeutic candidates that will form the basis of IND submissions designed to test a novel class of drugs for the treatment of tumor initiating cells (TICs) in three solid human malignancies where TICs have been implicated in the pathogenesis of disease. The target profile is the TIC population in colon cancer, ovarian cancer and glioblastoma. The therapeutic compounds that have been developed in the course of the collaboration target a pair of serine-threonine kinases that act at the nexus of mitosis, hypoxia, and DNA repair. These enzymes are over-expressed in many forms of cancer and alterations in their expression patterns correlate with dysregulation of a number of genes that are significantly linked to poor patient outcome.
  • Compounds against the first target have been developed to the point at which a developmental candidate can be selected. The compounds show single digit nanomolar potency in vitro, adequate specificity, appropriate pharmacokinetics to support oral delivery, and the ability to trigger growth inhibition and cell death in a wide panel of tumor cell lines and TICs from the three targeted histologies. Recently completed dose and schedule studies have been used to design and implement tumor model studies. The compound that demonstrates the widest therapeutic index will be selected for IND enabling studies. These IND enabling studies will include synthetic scale-up, toxicity evaluations, combination studies, mechanism of action studies, and a biomarker identification program that will be used to identify a targeted population for optimal clinical trial design.
  • The medicinal chemistry program against the second target was started approximately 15 months after the initiation of the effort against the first target. Sufficient potency, specificity, and activity against tumor cell lines and TICs have been demonstrated with novel molecules. Current efforts are focused on improving the pharmacokinetic properties of the drug candidates.
  • A phospho-flow platform to measure mRNA levels, protein levels, and enzymatic activity using a mass spectrometric readout has also been tested. This system enables the simultaneous measurement of up to 35 different biomolecules. A data management system has been developed to facilitate the associated complex data analysis. Proof or principle experiments have demonstrated that this experimental paradigm can be used to reconstruct the developmental lineages of all progeny downstream of hematopoietic stem cells from human and mouse bone marrow. This approach has recently been applied to the analysis of ovarian cancer cells taken directly from patients. The results of these studies suggest that cancer cells are clearly heterogenous, but perhaps most importantly can be organized into developmental lineages that are formally similar to those seen in bone marrow development. Furthermore, this platform can assess the response of individual subcomponents of the oncological lineage to both approved and experimental drugs. We will be using this platform to gain insight into how tumors respond to individual drugs, including our drug candidates, and combination studies. It is reasonable to expect that it will be possible to not only assess the response of the cancer stem cells, but all subtypes of the tumor lineage.
  • Slamon Mak Cancer Stem Cell Abstract
  • Drug discovery programs against two different mitotic kinases are being pursued. Both programs follow the same general process flow in which lead optimization experiments culminate in the selection of a single small molecule candidate for advancement to preclinical development. The development candidate then proceeds through a standard series of evaluations to establish its suitability for an IND submission and use in subsequent clinical trials.
  • CFI-003 was selected as a clinical development candidate and is progressing through investigational new drug application (IND)-enabling studies. Chemistry activity in the past year has included the selection of the fumarate salt as the final salt form, and production of two kilogram-scale clinical batches, the first of which is scheduled to be released at the end of April. The compound is stable when stored under typical storage conditions, and has an impurity profile that is safe for clinical dosing. In cancer models, CFI-003 was shown to be particularly effective against tumors deficient for the tumor suppressor gene PTEN; this is important given that deficiencies in this gene are generally considered to be an indicator of poor prognosis in the clinic. Experiments are ongoing to determine biomarkers of response to CFI-003 for application in the clinic. Other work includes selection and management of contract research organizations (CROs) for critical IND-enabling studies. For example, Pharmatek has been engaged to assist in the development of a drug formulation that enhances the stability of CFI-003, and maximizes bioavailability of the compound when dosed orally. Other CRO work that is ongoing involves in vitro pharmacology experiments geared toward understanding how CFI-003 might interact with co-administered drugs, and performing key toxicology experiments for determination of a safe and effective clinical dose of the compound. An important milestone was reached in the previous reporting period in that the patent application covering CFI-003 was allowed by the US patent office. The CFI-003 IND development team will continue to move the project forward planning for a successful IND submission toward the end of Q1 2013.
  • The drug discovery efforts in the second program have been focused on improving the pharmacokinetic properties of the lead series molecules while maintaining excellent in vitro activity. Approximately 400 new chemical entities have been synthesized during the last reporting period. Progress to date has been measured by an increase in potency in the biochemical assay and improved anti-proliferative potency in cancer cell growth assays. Activity toward Aurora B has simultaneously been attenuated, and current compounds demonstrate improved selectivity against a diverse panel of kinases. Progress was aided by the acquisition of multiple co-complex x-ray structures which allowed for further refinement of binding models to the target’s active site. Compounds to be qualified for further study must continue to induce an aneuploidy phenotype at least an order of magnitude above the HCT116 (colon adenocarcinoma cell line) GI50, and importantly must also demonstrate adequate plasma levels upon oral dosing. A lead series compound has been shown to have oral efficacy in a cancer model. To follow up this result, additional compounds have been scaled up for testing. Experiments to determine the tolerability have been completed for the latest candidates and further efficacy studies have been initiated. Results from these efficacy studies will aid in the identification of a development candidate for subsequent IND enabling studies.
  • Drug discovery programs against two different mitotic kinases are being pursued. Both programs follow the same general process flow in which lead optimization experiments culminate in the selection of a single small molecule candidate for advancement to preclinical development. The development candidate then proceeds through a standard series of evaluations to establish its suitability for an IND submission and use in subsequent clinical trials.
  • CFI-400945 was selected as a clinical development candidate. The IND-enabling studies included the selection of the fumarate salt as the final salt form, and the production of two kilogram-scale clinical batches, which have been released during the past year. The compound is stable when stored under typical storage conditions, and has an impurity profile that is safe for clinical dosing. In cancer models in mice, CFI-400945 was shown to be particularly effective against specific subsets of tumor cell lines in both tumor cells grown in soft agar and in xenograft models. Experiments are ongoing to determine biomarkers of response to CFI-400945 for application in the clinic. Pharmatek was engaged to assist in the development of a drug formulation that enhanced the stability of CFI-400945, and maximized the bioavailability of the compound when dosed orally. Other CRO work that was completed included in vitro pharmacology experiments geared toward understanding how CFI-400945 might interact with co-administered drugs, and performing key toxicology experiments in animals for determination of a safe and effective clinical dose of the compound. This work culminated in an IND submission in the second quarter of 2013.
  • The drug discovery efforts in the second program has focused on improving the pharmacokinetic properties of the lead series molecules while maintaining excellent in vitro activity. Approximately 400 new chemical entities were synthesized and tested using a battery of biochemical and cell-based assays. Off target activity towards Aurora B has simultaneously been attenuated, and current compounds demonstrate improved selectivity against a diverse panel of kinases. Progress was aided by the acquisition of multiple co-complex x-ray structures which allowed for further refinement of binding models to the target’s active site. Compounds were qualified for in vivo study based on the induction of an aneuploid phenotype at an order of magnitude above the HCT116 (colon adenocarcinoma cell line) GI50, and importantly the demonstration high mouse plasma levels upon oral dosing. Mouse xenograft studies based on a number of tumor cell lines were used to select a short list of compounds. The aggregate data was then used to select a developmental candidate CFI-1870. IND enabling studies have been launched. In parallel, detailed dose and schedule studies are underway along with approaches to identify susceptible tumor subpopulations and associated biomarkers that will eventually support a targeted clinical trial.
  • The Slamon/Mak cancer stem cell drug discovery program funded by CIRM/CSCC has achieved two important milestones in the past year. Our first therapeutic candidate was approved by the FDA and first-in-human dosing of CFI-400945 has taken place as part of the Phase I clinical trial. In our second program we have selected a development candidate that is now in the midst of IND ennabling studies
  • The clinical trial is being carried out at Princess Margaret Cancer Centre (Principal Investigator, Dr Philippe Bedard) and UCLA (Principal Investigator, Dr Zev Wainberg). This clinical trial was initiated after a number of milestones were successfully met following the submission of the IND and CTA in 2013. These have included making improvements to the formulation of the CFI-400945 tablets resulting in the successful reduction of the appearance of a degradant that was slowly accumulating in the initial formulation. This enabled the manufacturing of the cGMP tablets for use in the clinic in September 2013. These formulation changes and the Certificates of Analysis of these tablets were submitted to the FDA and permission was granted to begin clinical evaluation. In December 2013, we were awarded the CIRM Disease Team III funding to continue the CFI-400945 program which enabled the planning and initiation of this Phase I clinical evaluation and additional non-clinical studies.
  • In our second program, Pyrazolo-pyrimidines have emerged as the most promising class of 3rd series TTK inhibitors. TTKis with potent in vitro activity, excellent oral exposure in rats and in vivo efficacy were identified. A short list of 5 pyrazolo-pyrimidines was identified as potential third series development candidates. After further characterization it was determined that 4 or 5 compounds met the preponderance of the selection criteria, 2 of which had outstanding PK properties. The TTK inhibitor CFI-402257 had the best balance of efficacy, PK and off-target activities and was selected as the development candidate. IND enabling studies with 402257 have been initiated, and will continue during the no cost extension period of the grant.

© 2013 California Institute for Regenerative Medicine