Telomerase+Peptide+Vaccine—The+Next+Cancer+Therapy?

by Ashley Leslie and Charles Walker

Introduction
We began our investigation into the use of telomerase as a target for cancer therapies with a 2008 review article published in the British Journal of Cancer. Although the article discussed multiple approaches to telomeric therapeutics, we chose to focus on immunotherapy, in particular, the use of peptide vaccines, as shown below. Given the relatively new nature of this type of cancer therapy, we were interested in exploring its efficacy and evaluating whether or not it appeared to be a viable alternative to the current standard of treatment.



=Background =

About Telomerase
Telomerase is an enzyme that maintains telomeres, the sequences of repeated DNA at the ends of chromosomes (Geron). Telomerase consists of an RNA subunit (telomerase RNA, or TERC) and a protein subunit (telomerase reverse transcriptase, or TERT) (Allison). Through the RNA template (AAUCCC) provided by TERC, TERT uses reverse transcriptase to add repeating bases (TTAGGG) to the ends of chromosomes in the 5'-3' direction (Allison, 2012).

Telomerase and Cancer
Telomeres are an essential part of a chromosome, acting to protect and stabilize it, but each time a cell’s DNA is replicated, the chromosome's telomeres are shortened (Shay et al., 2008). With successive divisions, telomeres become so short that the cell either stops dividing (senesces) or dies (apoptosis) (ibid.). Thus, telomerase works to elongate telomeres and stall this cell fate. Telomerase activity normally peaks during embryonic development when cells are dividing extremely rapidly to promote growth (Geron). It is then downregulated in mature cells, with the exception of germ and stem cells (Geron). However, telomerase is also found to be active in 85-90% of tumors from cancers of all types (Harley). It is the upregulation of telomerase that provides cancer cells with the potential for immortality—their telomeres are continually extended, permitting unlimited rounds of cell division (Allison). Given this disparity in telomerase activity between normal cells and cancerous cells, telomerase is an appealing target for cancer therapeutics and is the focus of much current research.



Telomerase Immunotherapy
Immunotherapy involves the treatment of disease by either eliciting or suppressing an immune response. In the case of telomerase immunotherapy, a drug product would be designed that causes an immune response against the hTERT protein (the catalytic subunit of telomerase) (Harley). Thus, any telomerase-positive cells expressing hTERT would be killed. Researchers have already demonstrated that hTERT peptides can be presented as epitopes by major histocompatibility complexes (MHCs), resulting in an immune response (Harley).

Telomerase immunotherapy is promising as hTERT is a widely expressed tumor-associated antigen, but, given manufacturing complexities and complicated biological mechanisms, designing such a therapy has proven difficult. Consequently, there is a need for more innovative clinical trial designs to determine how exactly telomeric immunotherapies could best be used as a treatment against cancer.

Telomerase Peptide Vaccine (GV1001)
One of the most advanced telomeric immunotherapy products is the telomerase peptide vaccine GV1001. GV1001contains the 16 amino acid long peptide hTERT (611-626), which is a "promiscuous HLA (human leukocyte antigen) class II epitope", ("Definition of Telomerase Peptide Vaccine GV1001", Bernhardt et al). This means that it is able to bind various HLA molecules and potentially raise an immune response against telomerase-positive cells ("Definition of Telomerase Peptide Vaccine GV1001"). In 2006, researchers published a paper on a phase I/II clinical trial involving the administration of the telomerase peptide GV1001 as a vaccine in patients with inoperable pancreatic cancer (Bernhardt et al). More recently, phase III trials have been conducted as well.

=The Research =

<span style="font-family: 'Times New Roman',Times,serif;">GV1001 Phase I/II Clinical Trial
<span style="font-family: 'Times New Roman',Times,serif;">The goals of this trial were to determine the safety and tolerability of GV1001, as well as the vaccine's optimal dosage and any potential survival benefits (Bernhardt et al).

<span style="font-family: 'Times New Roman',Times,serif;">Methods
<span style="font-family: 'Times New Roman',Times,serif;">48 patients with inoperable pancreatic cancer were divided into three groups for the trial: low dose (60 nmol, n=11), medium dose (300 nmol, n=17), and high dose (1µmol, n=20). GV1001 vaccinations were therapeutically administered to patients through intradermal injections (injected between the layers of skin). Three vaccinations were given in the first week, followed by only one in weeks 2, 3, 4, 6, and 10. While data from all patients was used for the safety analysis, only those who received at least 6 injections over a four week period were considered for the evaluation of immunogenicity and survival. The number of evaluable patients in the low, medium and high dose groups were eight, 16, and 14, respectively.

<span style="font-family: 'Times New Roman',Times,serif;">As a measure of safety and toxicity, patients were monitored for adverse reactions during and after receiving each of their vaccine injections. Immune response was measured by delayed-type hypersensitivity (DTH) skin test—DTH is an injury of one's tissue caused by the body's mounting of a T cell mediated defense. These tests were performed on each patient at each visit. T cell response was also monitored via proliferation assays of Acid Citrate Dextrose-blood samples.

<span style="font-family: 'Times New Roman',Times,serif;">Results
<span style="font-family: 'Times New Roman',Times,serif;">None of the 48 patients showed any signs of toxicity or a particularly severe reaction to the injection. However, every one of the patients were found to have signs of inflammation at the injection site. Observed side effects included fever in 2% of patients, chills in 10%, pain in 6%, fatigue in 2%, nausea in 12%, and vomiting in 2%. Interestingly, when separated by dose, the low dose group had the highest frequency of such adverse events at 45% of patients, followed by the intermediate group (41%), and then the high dose group with only 10%. Just looking at the number of adverse effects, the intermediate group had approximately four times more than either of the the other groups. This pattern could be partially explained by a sort of culmination effect—the intermediate group received more injections and was followed over a longer period of time.

<span style="font-family: 'Times New Roman',Times,serif;">Patients that tested positive for DTH, or were found to have GV1001-specific T cells in their assayed blood samples, were considered immune responders (Figure 3). In some cases, DTH tests were positive while T-cell responses were negative and vise-versa. Because researchers didn't know what to attribute this discrepancy to—different biological responses or the sensitivity in assays used to test T-cell response—patient cytokine profiles from two different patients with different immune reactions were taken. Researchers were able to conclude that GV1001 vaccinations produced identical cytokine profiles in both patients despite their different immune responses and further testing confirmed that different T-cells could recognize different GV1001 fragments. Overall, immune responses were detected in 63% of the evaluable patients. The intermediate group showed the most promising results with the highest proportion of immune responders—75% of evaluable patients in that group. This group also developed immune responses faster, stronger, and more often than either of the other dose groups.

<span style="font-family: 'Times New Roman',Times,serif;">A treatment-related survival benefit from the GV1001 vaccine was found for the intermediate treatment where the median survival time was 8.6 months, compared to 4.0 months for the low dose and 5.1 months for the high dose, which were not considered statistically significant (Figure 4A). When the data is stratified by immune response, rather than level of dosage, immune responders were found to have a significantly higher median survival time of 7.2 months, compared to 2.9 months for all non-immune responders (Figure 4B).





<span style="font-family: 'Times New Roman',Times,serif;">Discussion
<span style="font-family: 'Times New Roman',Times,serif;">The GV1001 vaccine was found to be a safe therapeutic product without major physiological side effects and the finding of increased survival time points to its efficacy as a possible cancer vaccine. Overall, the result of this study were very encouraging and supported continued research in the form of a phase III trial. Also, given that GV10001 is a promiscuous epitope, it could be used in the creation of a broad spectrum cancer vaccine, but further research is needed.

<span style="font-family: 'Times New Roman',Times,serif;">GV1001 Phase III Clinical Trial
<span style="font-family: 'Times New Roman',Times,serif;">Given the positive results of the combined phase I/II trial, an international, randomized phase III trial was initiated to compare the efficacy of the current standard of treatment, Gemcitabine (a common chemotherapy drug), alone versus GV1001, followed by Gemcitabine, in patients with unresectable, metastatic pancreatic cancer (Pharmexa, Buanes et al). The trial was conducted to compare the primary outcome of overall survival and the secondary outcome of progression free survival.

<span style="font-family: 'Times New Roman',Times,serif;">Methods
<span style="font-family: 'Times New Roman',Times,serif;">520 patients were randomly assigned to one arm of the trial—either Gemcitabine alone or GV1001 plus Gemcitabine. Gemcitabine was only added to the experimental treatment if the patient's condition worsened. Gemcitabine was to be given in accordance with the Burris regime and the SmPC, followed by cycles of one week of rest and three weeks of chemotherapy. 0.56mg of GV1001 would be administered to patients in the experimental group on day one, three, five, eight, 15, 22, week six, and every four weeks after that. CT scans were taken every eight weeks to measure cancer progression.

<span style="font-family: 'Times New Roman',Times,serif;">Results
<span style="font-family: 'Times New Roman',Times,serif;">The study began in June 2006 and ran until May 2008 when a preliminary analysis found that there was no survival benefit for 178 of the patients given GV1001 and the trial was terminated. 365 patients were enrolled in the study at the time of termination. By August 2008, 238 of the patients had died. Median overall survival was 7.3 months for Gemcitabine and 5.9 months for GV1001 and median progression free survival was 3.7 versus 1.9 months.

<span style="font-family: 'Times New Roman',Times,serif;">Discussion
<span style="font-family: 'Times New Roman',Times,serif;">In comparison with the current standard of treatment for pancreatic cancer, GV1001 failed to demonstrate efficacy. It is possible that the GV1001 treatment would prove to be beneficial if given in parallel with chemotherapy rather than as a precursor to it. [Why wasn't the trial conducted in such manner?]

=<span style="font-family: 'Times New Roman',Times,serif;">Conclusions =

<span style="font-family: 'Times New Roman',Times,serif;">Final Thoughts on the GV1001 Peptide Vaccine
<span style="font-family: 'Times New Roman',Times,serif;">The GV1001 peptide vaccine originally appeared to be an effective treatment. In the phase I/II trial, the peptide vaccine was found to be safe and nontoxic—even at the highest dosage (Bernhardt et al). However, the phase III trial debunked those previous suppositions by showing that GV1001 was not any better of a treatment for pancreatic cancer than current chemotherapy drugs (Buanes et al). <span style="font-family: 'Times New Roman',Times,serif; line-height: 1.5;">Even given this failure, the preliminary success and sound conceptual basis for this treatment make it worth pursuing. Cancer vaccines are a hot button research topic and only time will tell if the GV1001 peptide vaccine is viable as an effective cancer therapy.

<span style="font-family: 'Times New Roman',Times,serif;">Going forward, it would be interesting for studies to focus on administering the vaccine to patients with various stages of pancreatic cancer. The vaccine could even be tested prophylactically, rather than therapeutically, for individuals with a genetic predisposition. In addition, it was mentioned earlier that a GV1001 peptide vaccine could potentially be used on a wide variety of cancers—we would be curious to see whether it shows any better efficacy when used as a treatment for a different type of cancer.

<span style="font-family: 'Times New Roman',Times,serif;">Innovative clinical trials must be done to determine the vaccine's efficacy in other cancers, at different stages, and in combination with other cancer therapies. Testing for <span style="font-family: 'Times New Roman',Times,serif; line-height: 1.5;">any synergistic effects when the vaccine is combined with standard cancer treatments, for example chemotherapy or surgical debulking of tumors, could be the most promising research avenue to pursue as it wouldn't negatively interfere with patients' treatment plans. Additionally, teasing out biomarkers corresponding to patients who did respond positively to GV1001, and thereby making the vaccine as specialized as possible, could prove extremely useful.

<span style="font-family: 'Times New Roman',Times,serif;">References

 * 1) <span style="font-family: 'Times New Roman',Times,serif;">Allison, Elizabeth A. Fundamental Molecular Biology, 2e. Hoboken, NJ: Wiley, 2012. Print.
 * 2) <span style="font-family: 'Times New Roman',Times,serif;">Bernhardt, S. L., M. K. Gjertsen, S. Trachsel, M. Moller, J. A. Eriksen, M. Meo, T. Buanes, and G. Gaudernack. "Telomerase Peptide Vaccination of Patients with Non-resectable Pancreatic Cancer: A Dose Escalating Phase I/II Study." British Journal of Cancer 95 (2006): 1474-482. Web. 25 May 2014.
 * 3) <span style="font-family: 'Times New Roman',Times,serif;">Buanes, T., J. Maurel, W. Liauw, M. Hebbar, and J. Nemunaitis. "A Randomized Phase III Study of Gemcitabine (G) versus GV1001 in Sequential Combination with G in Patients with Unresectable and Metastatic Pancreatic Cancer (PC)." Journal of Clinical Oncology 27.15S (2009): n. pag. Web. 25 May 2014.
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 * 6) <span style="font-family: 'Times New Roman',Times,serif;">Harley, Calvin B. "Telomerase and Cancer Therapeutics." Medscape.com. Nature Publishing Group, 2008. Web. 25 May 2014. <[]>.
 * 7) <span style="font-family: 'Times New Roman',Times,serif;">Pharmexa A/S. "GV1001 and Gemcitabine in Sequential Combination to Gemcitabine Monotherapy in Pancreatic Cancer." Clinicaltrials.gov. U.S. National Institutes of Health, May 2008. Web. 01 June 2014. <[]>.
 * 8) <span style="font-family: 'Times New Roman',Times,serif;">Shay, J. W., and W. N. Keith. "Targeting Telomerase for Cancer Therapeutics." British Journal of Cancer 98 (2008): 677-83. Print.
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