RNA interference is a naturally occurring means to block the function of genes in our body. We propose that RNA interference
can be used to block HIV-1 infection and its reproduction within the body. When RNA interference is introduced into a stem cell,
its blocking activity will be present throughout the lifetime of the stem cell, theoretically the lifespan of a human being. Thus, in
theory an effective stem cell RNA interference therapy will require only a single treatment as opposed to the current lifetime
administration of anti-HIV-1 drugs often accompanied by serious side effects.
In nature, some individuals carry a genetic mutation that renders them resistant to HIV-1 infection. This mutation prevents
HIV-1 from attaching to the white blood cells. One RNA interference approach will be to mimic this natural situation by blocking
the activity of this “co-receptor” within infected individuals by creating a new blood system that carries the RNA interference
therapy. Other RNA interference therapies will be directed against genes of HIV-1 itself.
The need for novel approaches to the treatment of HIV infection has never been greater, because new infections continue to
occur at undiminished rates, in California and across the nation, despite decades of prevention efforts. Moreover, the number of
people living with HIV is rising steadily, thanks to improved management of HIV infection. As a result, California, which ranks
second in the nation in diagnosed cases of HIV infection, behind only New York, has identified 63,289 men, women, and children
who carry the virus. (Estimates of the number of Californians who are infected but have yet to be diagnosed range as high as
25,000.) Not all of the state’s HIV-positive residents are currently on therapy, but eventually virtually all of them will be—and
many of them will receive their drugs through government-supported programs. In addition, the longer these infected
individuals live, the more likely they are to avail themselves of a range of support services that the state pays for. The drugs
themselves, which are routinely administered in combination, are initially effective in suppressing viral replication in infected
individuals, but their potency diminishes over time, even as the toxic effects of therapy accumulate.
California, which is supporting the nation’s second-highest case-load of HIV-positive individuals, can expect to see that
number grow at a rate of 4,000-5,000 a year for the foreseeable future. The cost of providing life-sustaining medications and
social services to this burgeoning population will also continue to rise, not arithmetically but exponentially—because as
increasing numbers of infected individuals fail standard drug regimens, it will be necessary to shift them to newer, more
expensive treatments, and as the side effects of therapy become more manifest, it will be necessary to prescribe drugs to
combat those toxicities, adding to the overall drug burden for patients and the overall cost of drug therapy for the state.
In such circumstances, the prospect of stem-cell based therapy that will require “only a single treatment” is especially
compelling. In theory, RNA interference might effectively cure individuals infected with HIV, by blocking the ports through
which the virus enters CD4 cells and destroys the body’s immune system. And even if RNA interference proves only partially
effective in blocking viral entry, it could significantly suppress viremia while sparing patients the toxicities associated with drug
therapy. This would be a boon to infected individuals, but it would also be a benefit to uninfected Californians, because
reductions in viremia in infected individuals translate into a reduction in the community burden of HIV infection—and that, in
turn, reduces the overall rate of new infections statewide.
The PI proposes to develop a strategic plan to use RNA interference (RNAi) to block a co-receptor expression and HIV replication in a stem-cell-based therapeutic modality for HIV disease. Individuals homozygous for a defective gene, are protected from HIV infection and heterozygous individuals have an attenuated disease course. Inhibitory RNAs will be introduced into autologous hematopoietic stem cells mobilized from bone marrow. In later iterations, blood stem cells will be derived from human embryonic stem cells (hESC) or induced pluripotent cells (iPSC). Success would require reconstitution of the immune system by the recipient’s thymus from the protected hematopoietic stem cells. A team of basic scientists, clinical investigators, and business executives will be assembled to develop an approvable regulatory filing for a Phase I or Phase I-II clinical trial during the term of the Disease Team Research Award. Several research institutions will be involved.
Reviewers generally agreed upon the validity of the scientific rationale of the proposal and the use of RNAi, which is relatively mature and ready to apply in this context. A strength of the proposal is that the investigators have demonstrated proof of concept: that down regulation of the targeted co-receptor prevents HIV infection in vitro, and that they can achieve and maintain (for more than two years) down regulation of the co-receptor in a large animal model. An advantage of this approach is that gene transduced stem cells should be able to reconstitute immunity without being susceptible to infection, and could ultimately replace on-going anti-retroviral therapy with a single therapy.
Several limitations to the approach were noted. Reviewers expressed concern with this approach in advanced stage HIV patients, in which blockade of the co-receptor is unlikely to have much impact on disease progression. It seems most likely that initial patients in a trial of inhibitory RNA therapy would be likely to have advanced disease. The investigator argues that it would be advantageous in the future to use allogeneic hematopoietic cells derived from embryonic stem cells and/or induced pluripotent cells instead of autologous hematopoietic stem cells; however, this approach is not practical given the high level of polymorphism at HLA and minor histocompatibility loci. As pointed out in the proposal, if somatic cell nuclear transfer becomes successful in the future, this approach might then be applied using autologous nuclei. Currently, this approach is most appropriate for autologous transplantation. The concept does seem likely to be clinically applicable within five years, but the subset of patients for whom it might be beneficial is perhaps more limited than indicated by investigators.
The principal investigator (PI), a thought leader in the field of retrovirology, has extensive experience in leading multi-displinary teams. For many years the PI has directed a center for AIDS Research at the applicant institution. Thus, he/she has extensive leadership experience and is well qualified to bring together a team for this planning process. The PI has published over 200 papers in this field, and has made many notable scientific contributions in this field. The PI has already assembled a strong team of investigators, as well as a strong clinical advisory board.
The planning process for the first phase 1 clinical trial is underway, and is expected to be completed by 2011. This study will be funded through venture capital. The planning process under the CIRM Award will be directed toward subsequent modification of this protocol and pursuit of preclinical studies to develop more potent therapies. The outcome will be a detailed plan to develop second generation vectors in a novel delivery system and determination of how to best test them in clinical trials. Overall reviewers were impressed by the scientific rationale, maturity, planning approach and significance of this proposal.
Dr. Chen proposes to develop a strategic plan to use RNA interference to block CCR5 expression and HIV replication in a stem-cell-based therapeutic modality for HIV disease. Individuals homozygous for a defective CCR5 gene, CCR5 32, are protected from HIV infection and heterozygous individuals have an attenuated disease course. CCR5 shRNA will be introduced into autologous hematopoietic stem cells mobilized from bone marrow. In later iterations, blood stem cells will be derived from HES or iPS. Success would require reconstitution of the immune system by the recipient’s thymus from the protected hematopoietic stem cells. A team of basic scientists, clinical investigators, and business executives will be assembled to develop an approvable regulatory filing for a Phase I or Phase I-II clinical trial during the term of the Disease Team Research Award. Several research institutions will be involved.
Reviewer One Comments
The CCR5 co-receptor is required for HIV infection of macrophages and some T cells. Individuals with a homozygous defective CCR5 gene, CCR5 32, are protected from HIV infection and heterozygous individuals have a substantially attenuated disease course. RNAi is sequence-specific and is a promising approach to modifying gene expression. The investigators have developed shRNAs that downregulate CCR5 in primary T cells and protect them from HIV infection. They have successfully knocked down CCR5 in primary T cells in vitro, protecting them from HIV-1 infection. They have also used shRNA to knockdown CCR5 in autologous hematopoietic stem cells that were transplanted back into a rhesus macaque, which maintained CCR5 downregulation for more than two years. Thus, the concept is mature and should soon be ready for clinical evaluation. However, one limitation to the approach is that CXCR4 is the co-receptor used in the setting of advanced HIV infection and it seems most likely that initial patients on the trial of siRNA therapy would be likely to have advanced disease. In such patients, the absence of CCR5 is unlikely to have much impact on disease progression. An advantage of the approach is that gene transduced stem cells should be able to reconstitute immunity without being susceptible to infection, assuming the virus has not yet mutated to the X4-tropic stage. The investigators argue that the relatively low frequency (1-10%) of transduction anticipated will be sufficient to allow immune reconstitution due to the fact that HIV resistant cells will be selected for survival. However, they do not consider the thymic failure that characterizes advanced HIV disease. The investigator argues that it would be advantageous in the future to use embryonic stem cells and/or induced pluripotent stem cells instead of autologous hematopoietic stem cells because the recipient will require less manipulation and cells can be banked and grown to large numbers. They propose that genetic matching to the patient may be achieved by having a large pool of ES or iPS from different sources. This, in fact, is not practicable given the high level of polymorphism at HLA and minor histocompatibility loci. The approach is most appropriate for autologous transplantation. As pointed out, if somatic cell nuclear transfer becomes successful in the future, this approach might then be applied using autologous nuclei. The proposed concept addresses the problem of requirements for ongoing anti-retroviral therapy in order to suppress HIV disease and proposes to replace it with a single treatment. The concept does seem likely to be clinically applicable within five years, but the subset of patients for whom it might be beneficial is perhaps more limited than indicated by investigators.
Dr. Chen has an extensive history in leading multi-disciplinary teams and has made significant contributions to human retrovirus research. For 16 years he has directed the Center for AIDS Research at UCLA and he is the founding and current director at the UCLA AIDS Institute. Thus, he has extensive leadership experience and is well qualified to bring together a team for this planning process. Dr. Chen has published over 200 papers and is considered a thought leader in this field. He was the first to clone HTLV-II and invented a diagnostic test used worldwide by blood banks to screen for HTLV-1 and II. He has characterized brain infections in patients with AIDS dementia and described the labile latent stage of HIV 1 in quiescent T cells. He has published collaborative studies with Dr. Baltimore’s group on shRNA directed to CCR5. Dr. Chen has already assembled a strong team of investigators, including Louis Breton, Chief Executive of Calimmune, David Baltimore, Inder Verma, Jerome Zack and John Rossi as well as a strong clinical advisory board.
The planning process for the trial is already well underway. A collaboration between UCLA and Calimmune, a for-profit startup biotech company, has already been established with the goal of bringing shRNA directed to CCR5 as a stem cell therapy to treat HIV-1 disease. The first generation therapy will be a single treatment introduced through hematopoietic stem cell transplantation of lentiviral vector-transduced cells. A Phase I clinical trial is planned and is expected to be completed by 2011. This study will be funded through venture capital. The planning process under the CIRM Award will be directed toward subsequent modification of this protocol and pursuit of preclinical studies to develop more potent therapies. The outcome will be a detailed plan to develop second generation vectors in a novel delivery system and determination of how to best test them in clinical trials. The strategic planning steps will include development of a management structure to coordinate the work and strategic planning and development of an advisory group for input and advice. The best candidate therapeutic vectors for transduction of hematopoietic stem cells to inhibit HIV disease will be identified with the intention of combining CCR5 shRNA with other shRNAs directed to HIV sequences. An outline of a clinical protocol will be developed and major regulatory hurdles will be addressed. Intellectual property issues will be addressed as will timelines and milestones for the preclinical and clinical phase of the project. A budget will be developed. This seems a well-planned and a well-thought out planning process. Once it is complete, the disease team should be well-positioned to compete for a Disease Team Research Award.
Reviewer Two Comments
PI: Irvin Chen
Institution: UC Los Angeles
Disease Target: AIDS
Concept: Use RNA interference to block HIV-1 replication in a stem cell based therapeutic transplant strategy. Target gene is CCR5, naturally occurring mutations protect against infection and pace of disease progession. Initial approach with genetically modified autologous hematopoietic stem cells, but will consider hESC or iPS source if necessary. Replacement of existing stem cell pool with resistant population could ultimately lead to complete restoration of immunity and cure.
1. Very impressive list of consultants
2. Preliminary data on shRNAi looks good in primate model.
3. Possibility that single treatment is sufficient.
1. Proposal is clearly fundable by NIH
2. Recognition that transduced cells may be only 1-10%, but suggest
selective advantage for corrected cells. May not be true for
lymphocyte/macrophage/dendritic progenitor cell populations.
3. Hypothesis that protected T-cells would somehow feedback to prevent
parental stem cell senescence is hard to understand?
4. Hypothesis that hESC or iPS may be advantageous as sources is weak.
Adult hematopoietic stem cells are the correct target, starting with a more
primitive precursor raises more safety concerns. Ultimately, the hESC or
iPS will need to be differentiated to create the adult HSC.
a. Perhaps some advantage if can create a more universal cell banking
system without the need for complete tissue matching.
b. Allow for storage and more complete characterization of cells.
5. Commercial collaboration with Calimmune, who is already planning on
clinical studies with shRNAi for CCR5. Why do they need CIRM, and
what impact will this commercial arrangement have on sharing of reagents
with other investigators?
PI: Dr. Chen has been the Director of the UCLA AIDS institute for over 15 years. Very well published (230 papers) with multiple NIH grants on HIV/AIDS research, also strong in the area of molecular characterization and gene therapy. He holds patents on HIV diagnostics, cloned HTLV-II, and is collaborating with David Baltimore on CCR5.
Planning Approach: Recruit a team of basic scientists, clinical investigators, and business executives with goal of developing regulatory filing to support IND application and clinical trial. Involve several research institutions and a biotech company. Plans call for 4 full-day meetings