Applied BioLogics

Investigative Cancer Vaccine

Vaccines are designed to stimulate the immune system to mount an immune response against the target in the vaccine. For instance, the flu vaccine contains pieces of the flu virus, and stimulates the immune system to make cells that fight the flu virus. The flu vaccine needs to be given at least two weeks before exposure to the flu. This is an example of a preventive vaccine. The flu vaccine can stimulate long-lasting immunity to the strain of flu used in the vaccine.

Cancer vaccines are different in that they are not preventive. Rather, cancer vaccines are therapeutic-they are used to treat the disease rather than prevent it. Like the flu vaccine, cancer vaccines are designed to stimulate the immune system to mount a response toward the target-in this case the cancer cells. Unlike flu virus which is foreign to the body, cancer cells are not foreign and generally do not stimulate a strong immune response on their own. So cancer vaccines use other substances or cells to help the immune response along.

Some of the substances in cancer vaccines are called cytokines, which act as "immune hormones." Other substances are called heat shock proteins. Heat shock proteins and cytokines can help alert the immune system to the information about the cancer cells. This alert helps certain immune cells that are sensitive to the cancer cells to divide. This new "army" of cells will kill any cancer cell they come in contact with.

The cells that are most efficient at stimulating an immune response are called dendritic cells. Dendritic cells are a specialized immune cells found throughout the body. To make a vaccine, precursor cells are taken from a blood sample of the patient and grown in the laboratory. Information about the surface of the patient's cancer cells is placed inside dendritic cells that are grown in the laboratory. When the cells are injected they can activate an immune response toward the cancer cells. As with cytokine and heat shock protein activation, the alerted cells divide and kill cancer cells they come in contact with.

Unlike chemotherapy cancer vaccines generally have few side effects. Here at the clinic we use all the types of cancer vaccines described above. The results achieved vary with tumor, type of tumor and stage of disease.

Cancer vaccines represent an innovative potential cancer therapy — a therapy that seeks to harness the body's own defenses to fight the uncontrolled growth and spread of cancer cells.

The immune system has the ability to recognize the difference between "self" and "non self," that which is and is not a naturally occurring molecule in the body. In the case of cancer, the difference between cancer cells and normal healthy cells is sometimes so slight that they go unnoticed by the immune system and no response occurs, or the immune system is overwhelmed. The body is "tolerant" of the cells allowing them to multiply in the body. Cancer vaccines seek to "break" this tolerance.

Cancer vaccines are designed to introduce molecules expressed on cancer cells into the body in a new way that awakens the immune system to respond and destroy the cancer cell. These vaccines attract immune cells such as dendritic cells that engulf the vaccine cells which include "antigens" or proteins on their cell surfaces, and then present (exhibit) fragments of these antigens. These immune cells, known as "antigen presenting cells" (APCs), then signal other immune cells to mature and attack the specific invading antigen. Lymphocytes, including helper T cells, killer T cells, and B cells, are called into action. Helper T cells release cytokines, chemical messages that recruit other immune cells, and killer T cells engulf the antigen (and the cell it is attached to) the APCs presented to it. In addition to awakening the cellular side of the immune system to the tumor cell, some cancer vaccines stimulate the humoral side of the immune system, which includes antibodies, into action as well.

Types of Cancer Vaccines

Research and development efforts are currently under way to develop therapeutic cancer vaccines for the treatment of multiple forms of cancers. Currently, there are two primary approaches being explored in the development of making cancer vaccines—the "antigen-specific" approach, and the "whole cell" approach.

Antigen-Specific Approach

The antigen-specific approach seeks to make a vaccine that stimulates an immune response to a specific antigen or antigens that are believed to be unique to a specific type of tumor. This approach may result in a highly specific antitumor response, however poses the challenge of successfully identifying the specific antigens that are most highly expressed on a given tumor. Failure to identify the appropriate antigens could result in lower or no efficacy.

One approach to developing an antigen-specific vaccine involves the removal and isolation of a patient's dendritic cells, one type of APC. The dendritic cells are exposed to antigens that are believed to be associated with a specific tumor type, and are given time to ingest, process, and "present" the antigens. The cells are then reintroduced into the patient in vaccine form.

Whole Cell Approach

The whole cell approach uses whole cancer cells to make the vaccine, not just a specific antigen. Since whole cells express multiple—sometimes thousands of—antigens, there is potentially a greater chance of stimulating an immune response since this approach does not require choosing specific antigens which may or may not turn out to be appropriate for the patient. Cell Genesys is pursuing a whole cell vaccine approach with its GVAX® cancer vaccines.

Whole cell vaccines can be either patient-specific (made completely from the individual's own tumor cells), non patient-specific (made from a "cell line"—tumor cells that are grown in a laboratory), or a mixture of the two. Patient-specific vaccines may offer some advantages over non patient-specific vaccines when treating cancers that involve many different cell types with few like characteristics (e.g. non small-cell lung cancer). Using the patient's own tumor cells may increase the likelihood of creating an individualized vaccine that effectively stimulates an immune response against all cell types associated with specific form of cancer being treated.

Activating an Immune Response

While some cancer vaccines are designed to stimulate an immune response based solely on the presence of antigens, others are being developed that utilize antigens as well as cytokines to mount an attack against cancer cells. Cytokines are chemical messages that stimulate other immune cells to attack antigens. Some researchers are exploring the idea of creating vaccines comprised of cells that have been genetically modified to secrete a cytokine such as GM-CSF, interleukins, and interferons. The presence of these cytokines may potentially help "jump start" the immune system to launch a more robust and efficacious immune response.

Possible Benefits of Cancer Vaccines

In addition to providing a new treatment option for patients who have failed other therapies, clinical data suggest that cancer vaccines may offer therapeutic advantages over existing therapies:

1. Favorable Side Effect Profile: Unlike many traditional cancer treatments such as chemotherapy and radiation therapy, cancer vaccines have generally been associated with very few side effects. This favorable side effect profile may potentially enable patients to maintain a higher quality of life during the course of treatment.
2. Combination Therapy: Numerous clinical trials are being conducted evaluating the use of cancer vaccines in combination with other traditional therapies such as chemotherapy, radiation therapy, and stem cell transplantation. Combination therapies offer the potential of improving/enhancing the efficacy of these traditional treatments.

The Current State of Cancer Vaccines

Research and development efforts are currently under way at numerous organizations to thoroughly evaluate the safety and efficacy of different approaches to cancer vaccines. Currently, cancer vaccines are being evaluated in multiple human clinical trials for many types of cancer and are available only in the clinical trial setting.

Cytokines & Cancer Vaccine Treatment

What are Cytokines?

Cytokines are the messengers of the immune system-they are sometimes called immune hormones. They can act either locally or at a distance. Cytokines can either enhance or suppress immunity. In cancer treatment they are generally used to enhance immunity.

Two cytokines, interleukin-2 (IL-2) and interferon-alfa 2b are approved by the FDA for use against certain cancers. IL-2 is used to treat renal cell carcinoma, melanoma, lymphoma, and leukemia. Interferon has been useful against those diseases and Kaposi's sarcoma, chronic myelogenous leukemia, and hairy cell leukemia.

The response rate of individual cytokines is generally low. Cytokines are produced by white blood cells in combinations-in nature they work together. Studies using combined cytokines-in the ratios they are produced naturally have shown that the combinations have synergistic effects. For instance, IL-2 is used to stimulate certain white blood cells to divide. When used alone, a very high dose of IL-2 is required to make the cells divide. High doses of IL-2 can cause serious negative side effects. When a natural combination of cytokines produced by white blood cells is used, the dose of IL-2 can be lowered by a factor of 5000 producing minimal side effects. This is one of the combinations of cytokines used at Stowe BioTherapy.

These combinations of cytokines can also be used to enhance the effects of a vaccine designed to stimulate the immune system to mount a cancer cell specific immune response. Heat shock protein vaccines and dendritic cell vaccines can both be enhanced when given with natural cytokine mixtures.

The immune system is the body's defense system. It works on three different levels. The first level is the anatomic response. It consists of anatomical barriers to foreign particles and includes the skin and acid in the stomach. Anatomic barriers prevent foreign substances from entering the body. If foreign particles pass through the first line of defense the second line of defense called the inflammatory response kicks in. The third line of defense is the immune response. It is the main player in specific immune defense.

The cells of the immune system mount the immune response. These cells are also called white blood cells.

There are several types: The neutrophils are responsible for killing bacteria and yeast and are the first white blood cells at the site of an infection. The eosinophils play a part in delayed reactions to foreigners. The key players that will be discussed here are the monocytes and the lymphocytes. Monocytes are scavangers. They scour the body for anything out of place. They can engulf foreign particles and chew off pieces of tumor cells. Lymphocytes are not able to engulf any foreign particles or eat tumor cells. They take the information given to them by monocytes and monocyte-like cells and do their job. There are several types of lymphocytes. The types essential to this topic are the B lymphocytes, and the T lymphocytes. The B lymphocytes get their characteristics after being nurtured in the bone marrow, hence the B. B lymphocytes are primarily responsible for producing antibodies. Antibodies can inactive bacteria, fungi, and viruses and make them and other foreign particles easier to see by the rest of the immune system. T lymphocytes mature in the thymus gland, which is located under the breast bone, hence the T. For the purposes of this topic they can be divided into three major categories: the T helper cells, the T suppressor cells, and the cytotoxic T cells. The T helper and suppressor cells do exactly what their names imply. The cytotoxic T cells are primarily responsible for killing virally infected, and tumor cells.

The Immune System and Cancer

In order for cancer to occur, the immune system must have failed. The normal sequence of events when the immune system comes across tumor cells follows.

An immune cell called the macrophage (also called a monocyte) comes into contact with a cancerous or precancerous cell. This cell has some strange surface features. The strange features signal the macrophage that the cell is not healthy and that the macrophage should take a bite out of it.

The macrophage then begins to digest the bite of the tumor cell. Several little packets of enzymes act like a cellular stomach and break down the piece into smaller and smaller pieces.

There are two possible scenarios that can happen next. The macrophage can either hand off these little pieces of tumor cell to another type of immune cell, or it can transform itself into another, specialized immune cell called a dendritic cell. There is more and more evidence that the latter happens most often.

Dendritic cells are found in all tissues of the body, and many of them began as macrophages. The first dendritic cell discovered is found throughout the skin and is called a Langerhan's cell.

Now that the macrophage has digested pieces of the tumor cell, it transforms into the dendritic cell. The dendritic cell is a much more effective messenger. When it is fully mature, it gives the information about the tumor contained in the small digested packets to the rest of the immune system. A key point here is that the dendritic cell must be mature to effectively present the tumor information. The cell needs to have additional markers on its surface that the other immune cells can recognize. These markers are called co-stimulatory molecules and are shown as white crosses on the picture of the mature dendritic cell below.

When the dendritic cell begins to mature, it also starts moving, or migrating toward a lymph node. The lymph nodes contain large numbers of lymphocytes, another type of immune cell. Everyone probably remembers having enlarged lymph nodes in their neck when they had a sore throat. The lymph nodes are where the action is when it comes to the immune system. There are areas in the body that contain large numbers of lymph nodes. The neck, armpits, and groin areas all have clusters of nodes that lie close to the skin.

So the mature dendritic cell has migrated to the lymph node. There it comes in contact with different kinds of lymphocytes. If it has matured properly, the co-stimulatory molecules on its surface will help pass the tumor information along to the cytotoxic T lymphocytes, or CTLs. The CTLs are the body's main defense against tumor cells. When the right CTL comes in contact with the dendritic cell, it will become activated and begin to divide, effectively making an army of cloned soldiers ready to kill any cancerous or pre-cancerous cell having the same altered membrane discovered by the macrophage.

When the CTL soldiers come in contact with cells that have the same surface as the original cancerous cell, they bind to it. They then release a chemical that pokes tiny holes in the membrane of the tumor cell, and the tumor cell spills its guts and dies.

Let's summarize what happens normally in the body after a normal cell turns cancerous. First, a macrophage comes in contact with the tumor cell, which has a different type of membrane that signals the macrophage to eat part of it. The macrophage then digests the eaten tumor cell fragment and starts to turn into a dendritic cell. It then begins to mature, and travels to a nearby lymph node and hands off the tumor cell information to CTLs. The CTLs then divide, circulate throughout the body, and kill any tumor cells they come in contact with.

Above we covered what happens normally in the body when a cell becomes cancerous. This process occurs countless times as cells get genetic mutations and become cancerous. But, if you have cancer, then something must have gone wrong. Did the macrophage fail to recognize the funny cell surface? Did macrophages not become dendritic cells? Or did the T cells not do their job? It is impossible to tell for sure but there are some clues that the problem is with the dendritic cells.

Lately several research groups have been looking at the dendritic cells in and around tumors. What they're finding is that there are dendritic cells there, but they are immature. They don't have the co-stimulatory molecules necessary for the successful hand off of the tumor cell membrane information to the T cells. Moreover, because they are immature, they are much less likely to migrate to the lymph nodes to make the hand off.

To make a football analogy, the dendritic cell is the quarterback and needs to hand off the football to the running back (the T cell). In order to do that, he needs to move toward the running back and hand him the ball without fumbling. When the dendritic cell is immature, it just stands in one place and drops the ball. If that continues to happen, your team never scores and ultimately loses the game.

If you cut up a piece of tumor from kidney cancer or renal cell carcinoma and look at it under the microscope, you'll find millions of dendritic cells many more than in any other type of tumor. Expectedly, the majority of these dendritic cells are immature they don't have co-stimulatory molecules on them. What makes this more interesting is the fact that kidney cancer is the most likely type of cancer to disappear without a trace without any treatment, or spontaneously remiss. What I believe happens when someone has a spontaneous remission is the conditions in and around the tumor change enough to allow at least some of the dendritic cells to mature. This is more likely to induce a remission in renal cell carcinoma simply because of the larger numbers of dendritic cells.

So, what can you do to get dendritic cells to hand off information about your tumor cells to your CTLs? Both animal and human trials of using dendritic cells in the treatment of cancer have shown promising results and give us a direction in which to go.

Mayordomo et al.¹ inoculated mice with different types of cancer and allowed the tumors to develop for one to two weeks. Dendritic cells were isolated from the bone marrow of these mice, cultured with some growth factors, and exposed to tumor peptides (information about the tumor cell membranes). These Δprimed' dendritic cells were then injected back into the tumor-bearing mice every four to seven days. Recovery, measured as halting of tumor growth and subsequent regression, was seen 7-10 days after the first injection of dendritic cells. Using this treatment, cure rates of 80% for mice with Lewis lung carcinoma and 90% for mice with sarcoma were achieved.

In a similar study, Nair, et al.² induced malignant melanoma lung metastases (new tumors that spread from the first tumor) in mice, and then surgically removed the primary tumor. The mice were then treated with dendritic cells which had been Δprimed' in a manner similar to that described above. Of the seven treated animals, four had no visible lung tumors, two had fewer than five remaining tumor nodules, and one mouse had 15 nodules. The number of nodules in control mice, those that did not receive dendritic cell therapy, were too many to count, but comprised approximately three-quarters of the lung by weight.

Hsu et al.³ at Stanford University pioneered the use of dendritic cell therapy of cancer in humans. They purified dendritic cells from the circulating blood of four patients with B cell lymphoma previously treated with chemotherapy. The dendritic cells were cultured and treated with antigen (tumor cell membrane information) derived from the patients' tumors. The dendritic cells were given using vein injections on 4 occasions; subcutaneous (under the skin) injections of the tumor antigen and a protein that helps stimulate an immune response were injected two weeks after each dendritic cell injection. All of the patients developed measurable T cell immune responses after one or two vaccinations. Meaningful clinical responses were seen. There was one partial response, one minor response, and disease stabilization in three patients with progressive measurable disease, and a complete response in a patient with minimal detectable disease. All of the patients have remained progression-free for two years.

Gerald Murphy, M.D.⁴, and his team at Northwest Hospital's Pacific Northwest Cancer Foundation have been testing the use of dendritic cells in patients with advanced prostate cancer. They cultured monocytes (macrophages) from circulating blood with growth factors and small pieces of protein found on the surface of prostate tumor cells. These dendritic cells were then reinfused into the patients through an intravenous drip. They performed two studies. More than 27% of study patients who participated in both clinical trials showed some improvement and the disease was stable in another 33%. All of the patients in the study had advanced prostate cancer and were unresponsive to conventional therapies, including hormone treatment.

In addition to lymphoma and prostate cancer, the deadly skin cancer malignant melanoma has been treated successfully using dendritic cell therapy. In a recent human study by Nestle et al⁵, dendritic cells were used to treat sixteen patients with advanced metastatic (the cancer has spread) melanoma. Objective responses were seen in 5 of the 16 patients. There were two complete responses and three partial responses with regression of metastases in several organs, including skin, lung, and pancreas. The participants were followed for 15 months and no cases of autoimmunity a potential side effect of the therapy were found in any of the patients. The authors concluded, vaccination with autologous [derived from the person's own body] dendritic cells generated from peripheral blood is a safe and promising approach in the treatment of metastatic melanoma.

In the studies quoted above, there were little to no side effects. Murphy reports transient hypotension (temporary low blood pressure) as the only side effect seen in his study.

Given all of this compelling evidence that dendritic cells may hold a key position in effective, non-toxic treatments for cancer, we began studying them.

Our research to date has focused mainly on methods of:

1. Producing large numbers of dendritic cells from the circulating blood of cancer patients;
2. Finding the source of tumor material (antigen) for each type of tumor that will best stimulate the T cells to proliferate and kill tumor cells; and,
3. Stimulating the dendritic cells already in and around the tumor to mature, and become better T cell stimulators.

What we have found so far is that we can produce large numbers of dendritic cells from the circulating blood, give them tumor antigen, and mature them. These dendritic cells in culture are able to stimulate large numbers of T cells to become active against tumors. We are now setting out to determine if this is possible in humans.

This study begins when monocytes are harvested from the peripheral blood. The monocytes are cultured with cytokines and tumor cell antigens are added. The cells are then allowed to mature. Instead of using the dendritic cells, the liquid in which the cells are growing is collected. The exosomes are then removed from the liquid. They are sterile filtered and injected into the skin just above the lymph glands in the groin or other areas such as the abdomen. The theory is that there they interact with the dendritic cells in the skin, which move into the lymph glands and present tumor antigen to the lymphocytes. Which then divide, circulate in the body, and kill the tumor cells they come in contact with.

We rely heavily on cancer vaccines to stimulate a patient's own immune system to recognize and destroy tumor cells and adjunctive therapy that can fully activate the immune response.