CRYOVAX®

CryoVax® is a therapeutic vaccine made inside the body. The vaccine is designed to direct an immune response against a patient’s own tumors. CryoVax® combines a procedure known as cryoablation with the intratumal injection of a living cell drug called AlloStim®. AlloStim® is derived from the blood of normal blood donors. AlloStim® is a patented, living T-cell with both Th1- and NK-like properties with monoclonal antibodyconjugated microparticles attached. These cells are bioengineered outside the body to elicit a special immune effect known as the ‘Mirror Effect’. This immune effect is designed to elicit the same anti-effect that occurs after stem cell transplant procedures without the associated toxicity. There is no requirement to match the donor with the patient.

Background

Therapeutic cancer vaccines for metastatic cancer have historically not performed well in the clinic. This is believed to be due to the tumors’ ability to modulate the immune response to one that is ineffective, and also due to the ability of the tumors to evade immune attack. In order to design a therapeutic vaccine capable of both modulating the immune response of a cancer patient from an ineffective response to an effective response, and counter-acting the tumor’s ability to escape immune attack, researchers have developed a technology called the “Mirror Effect” and an experimental drug called “AlloStim”. The mechanism of the Mirror Effect is still considered theoretical and AlloStim has not been approved by any regulatory agency any where in the world. The efficacy and toxicity of this novel therapeutic vaccine method is under investigation and can not be stated with certainty.

Mirror Effect

The only mechanism known to elicit an immune response that can effectively eliminate chemotherapy-resistant tumors, is the immune response that occurs after an allogeneic, non-myeloablative stem cell transplant (also known as a “Mini-Transplant”). In these procedures, patients undergo chemotherapy, not for the purpose of eliminating tumor cells, but for the purpose of suppressing the immune system sufficient to allow engraftment of immune cells from a healthy donor (usually a sibling). Under the conditions where the host immune system and the donor immune system are living side-by-side (chimeric immune system), a phenomenon called the graft vs host (GVT) effect occurs. GVT has been described as the most powerful anti-tumor immune response ever discovered, because it is the only mechanism known to be capable of eliminating chemotherapy-resistant metastatic disease. Unfortunately, the GVT effect can not be used routinely in clinical practice to treat metastatic disease due to an intimately and proportionally related toxicity known as graft vs. host disease (GVHD), GVHD is a devastatingly toxic side-effect of Mini-Transplant procedures and is often lethal. The same immune cells responsible for killing tumor cells in the GVT effect are also capable of killing normal tissues which causes GVHD. Despite considerable amount of investigation, researchers have not been able to successfully separate the beneficial GVT effect from the toxic GVHD effect. In fact, the killing of normal tissues that occurs in GVHD is a required step in the immune cascade leading to tumor elimination by GVT.

The Mirror Effect is a technology designed to preserve the anti-tumor effects of GVT while eliminating the risk of GVHD. Instead of the GVT/GVHD immune events flowing from the graft to the tumor and the graft to the host (GVT/GVH), the Mirror Effect reverses the immunological flow so that it occurs from the host to the tumor (HVT) and the host rejection of the graft (HVG). HVT/HVG is the ‘mirror’ of GVT/GVH. In animal models the HVT effect is the same as the GVT effect.

Unlike Mini-Transplant procedures, the Mirror Effect does not require matched tissue donor or chemotherapy conditioning to create a chimeric immune system. The Mirror Effect is believed to still be capable of occurring in heavily pre-treated metastatic cancer patients, whose immune systems have been damaged by chemotherapy and radiation.

AlloStim®

AlloStim® is an experimental living cell drug that has been bioengineered to elicit the Mirror Effect. AlloStim® contains ex-vivo expanded and differentiated Th1 immune cells with tiny monoclonal antibody-coated, 4.5 micron beads attached. AlloStim® is derived from healthy blood donors. The donors are recruited in the Phoenix, Arizona area. The donors and screened and tested for blood-borne diseases. The blood is then processed and shipped to Israel for further manufacturing into a precursor called “T-Stim” over a 9- day culture process. The culture process occurs in clean rooms and is conducted under Good Manufacturing Practice (GMP) standards. T-Stim is aliquoted into individual dose vials and stored frozen until required in the clinic. When required in the clinical, the vials are thawed and the cells activated by incubation with the microbeads in a 4hr process. After 4h, the cells change into AlloStim® and the cells together with the microbeads are packaged into syringes. The syringes are shipped to Phoenix twice a week for use in the clinical trial.

Protocol Design

The Phase IIb clinical trial is evaluating six different dose administration frequencies. All the protocols are designed with a priming stage, a vaccination stage and a booster stage:

Priming

In the priming stage, patients receive intradermal injections of the study drug. This stage is designed to educate your immune system to identify and kill (reject) the foreign cells in study drug. Because the foreign cells are powerful immune stimulants, even patients with weak immune systems should be capable of rejecting the cells. After each intradermal injection, the immune system will reject the AlloStim® cells and the reject response should be stronger and memory of the foreign cells should develop. Blood tests will be drawn before and after the completion of the intradermal dosing. This blood will be processed in the laboratory to isolate the immune cells. The isolated immune cells will be sent to Israel to determine if memory immune cells capable of rejecting the AlloStim® cells are increased in the blood.

Vaccination

The vaccination stage is designed to educate the immune system to identify tumors and be armed to kill a patient’s own tumor cells. Vaccines generally contain a source of tumor antigen mixed with an adjuvant to enhance the immune response to the selected antigen(s). Tumor antigens serve as the ‘bar-code’ of a tumor cell, allowing a killer immune cell to ‘read’ the cells and distinguish between target tumor cells and normal cells. Most previous vaccines have isolated proteins expressed on the surface of tumors to be used as antigens in cancer vaccine formulations. However, immune cells are designed to recognize antigens processed on the inside of cells and presented on the surface only in the context of special molecules called MHC molecules. However, since tumors do not usually express MHC molecules, internal antigens are not routinely used.

In order to use internally processed antigens, a selected tumor lesion is selected for killing using extreme cold (cryoablation). Cryoablation is an FDA approved technology designed for killing small tumor lesions. Cryoablation alone, however, has not been shown to benefit metastatic cancer patients. The freeze-thaw cycles of the cryoablation procedure cause the rupture of tumor cell membranes, spilling their internal contents in the tumor microenvironment. Immediately after the cryoablation procedure, the AlloStim® study drug is injected into the area.

The introduction of the foreign AlloStim® cells into the ablated lesion is then expected to elicit a memory rejection response, causing the killing of the foreign cells and the spilling the internal contents into the microenvironment of the dead tumor cells. The tissue injury caused by the cryoablation should attract specialized immune cells, called dendritic cells, to the area to engulf the cellular debris. The dendritic cells are expected to engulf a mixture of antigens from both the tumor and the foreign cells. The processing of these antigens is expected to trick the immune system to believe that the tumor is the same as the foreign immune cells. Once the immune system is educated to identify the tumor cells as foreign, a systemic immune response to eliminate tumors at any location in the body may be possible.

Blood samples will be taken after the cryoablation in order to determine if tumor-specific memory cells are detectable in the circulation.

Activation and Booster Stages

In the activation and booster stages, the AlloStim® cells are infused intravenously. The infusion of these cells is expected to activate memory cells in circulation, including antitumor memory cells. Activated memory cells are capable of leaving the circulation and trafficking to sites of tumors through-out the body. In addition, the rejection of the AlloStim® cells in circulation is designed to cause the release of inflammatory cytokines which can serve to disable the ability of tumor cells to avoid immune destruction. Blood samples will be taken after intravenous infusions to verify if the desired cytokine