KR102041381B1 - Treatment of non-small cell lung carcinoma by active immunotherapy - Google Patents

Abstract

This disclosure is a novel immunotherapeutic regimen for treating patients suffering from cancer such as, for example, non-small cell lung cancer (NSCLC). This regimen involves a combination of GM-CSF and peptides derived from human telomerase for patients who have previously received chemoradiotherapy.

Description

Treatment of non-small cell lung cancer using active immunotherapy {TREATMENT OF NON-SMALL CELL LUNG CARCINOMA BY ACTIVE IMMUNOTHERAPY}

The present invention relates to the field of anticancer treatment and immunology. Specifically, the present invention is directed to non-small cell lung cancer (non-small cell cancer) by active cancer therapy using active immunotherapy, that is, by active immunization with a peptide derived from the catalytic subunit of human telomerase. lung cancer (NSCLC).

Lung cancer is the leading cause of cancer deaths in both men and women worldwide. About 80% of them are non-small cell lung cancer (NSCLC), which is a stage 3 or 4 disease in which most subjects are inoperable. Metastatic disease (stage 4) has a poor prognosis, ie 5 year survival rate of 1%. The success of Phase III treatment depends on whether it can control both local disease and potential metastasis. If the disease can be included in an appropriate radiation volume region, i.e., phases 1-3A, then therapeutic radiation therapy (= 60 Gy) is the preferred method of treatment. However, the 5-year survival rate for stage 3 patients treated with radiation therapy alone is less than 5%. In addition, about one third of these patients recur locally, the other third patients progress to distant metastases, and the other third represent both. Several approaches to complex treatments have been studied. This includes induction chemotherapy and chemo-radiotherapy as well as consolidation chemotherapy. However, progress has been limited to date. Most patients die of disease recurrence and new treatment strategies are required.

Recently, the development of vaccines against lung cancer is receiving more attention. Several large-scale Phase III trials are currently underway for NSCLC patients who are attempting different vaccine strategies.

Enzyme telomerase is expressed in most human cancers, including NSCLC, which is considered an attractive target for general purpose cancer vaccines. Telomeric DNA confers stability to chromosomes, and normal celiac cells can only divide a limited number of cell divisions, because telomeres become shorter with each somatic cell division. Tumor cells evade this biological clock by expressing telomerase, which synthesizes new telomeres subunits.

Peptide GV1001 (SEQ ID NO: 1) consists of 16 amino acids derived from the activation site of hTERT (human telomerase reverse transcriptase). Two GV1001 clinical trials have been reported in this regard. GV1001 peptides are recognized by a variety of human leukocyte antigen class II (HLA class II) molecules encoded by DP, DQ and DR subloci. This unobstructed HLA-binding profile suggests that GV1001 may be an applicable vaccine in the general patient population and may widely elicit T-helper responses between individual individuals. In addition, GV1001 contains a narrow range of HLA class I antigenic epitopes, which allow the adoption of CD8 + cytotoxic T cells.

In a previous NSCLC vaccine clinical trial, 26 pretreatments were performed and vaccination with telomerase peptide I540 and GV1001 was performed in patients with advanced disease (mostly stage 4). No chemo or radiotherapy was involved. Vaccine-specific T cell responses were observed in 13 patients for GV1001 and 2 patients for I540, without any serious side effects associated with treatment.

It is an object of embodiments of the present invention to provide novel therapeutic regimens for patients suffering from NSCLC.

There is increasing interest in combining conventional therapies with cancer vaccines. Existing concerns that chemotherapy inhibits the immune response have been reinforced with considerations of possible synergistic effects.

In many cases, most tumor cells are removed by conventional therapies. But the tumor eventually recurs. Cancer vaccines work through different mechanisms and can be effective against cancer cells that are resistant to chemo- and radiotherapy. In addition, chemo-radiotherapy may increase rather than retard an immune response. Firstly, tissue damage induces a "danger signal", which provides a pro-inflammatory microenvironment. Second, there is evidence suggesting that gene products induced by radiation make tumors more susceptible to T-cell attack. Third, lung tumors harbor regulatory T-cells that are thought to interfere with the host's immune response, and several studies have shown that chemotherapy controls regulatory T-cells and myeloid-derived suppressor cells. Suppresses derived suppressor cells (MSDC).

Finally, docetaxel, applied as a therapeutic regimen in the present invention (see below), may potentially increase the vaccine response through other mechanisms.

The present invention is based on phase 2 clinical trial data of a telomerase peptide GV1001 (EARPALLTSRLRFIPK, SEQ ID NO: 1) conducted in Phase 3 NSCLC patients. This study was evaluated for GV1001 vaccination immediately following chemo-radiotherapy.

Specifically, the results of a phase 2 clinical trial that studied the GV1001 vaccination of NSCLC patients were provided. The clinical trial study included 23 patients and did not show any serious adverse effects related to treatment. This study showed an 80% immune response per protocol. This response rate is significant when compared to most cancer vaccine clinical trials, including those that have studied telomerase-based approaches. In addition, this study also demonstrated the generation of durable GV1001-specific T cell memory responses.

Of particular interest are the various approaches, the possibility of long-term data and the association between immune response and clinical outcome.

While there are many theoretical debates that suggest that cancer vaccines may be most effective if they are combined with drug use, clinical trial data on how these aspects interact is sparse. Interestingly, the frequency of immune responders in the clinical trials conducted in the present invention was superior to the three ongoing GV1001 clinical trials of the inventors who studied vaccination with monotherapy. This finding suggests that applying chemo-radiotherapy does not prevent immunization and may have contributed to improving the immune response.

While most vaccines display antigenic determinants that conform to HLA-class I, GV1001, a long peptide that conforms to HLA class II (HLA II), may be particularly suitable for combination protocols, according to the findings. . Long peptides are adopted by CD4 + T-helper cells, which are known to interact extensively with other immune cells. In tumor tissues irradiated with radiation, GV1001-specific T-helper cells recruit antigen presenting cells (APCs) that display antigens coming from apoptotic tumor cells and reduce the distribution of antigenic determinants. Induce.

The inventors of the present invention address these topics for studies conducted in long-term survivors in GV1001 clinical trials and reveal responses that actually counteract hTERT antigenic determinants outside GV1001. In a new half-phase clinical trial in NSCLC patients, we plan to use vaccination with these new hTERT peptides in combination with chemo- or radiotherapy.

There is limited knowledge about the long-term development of cancer vaccine responses and how to design vaccine boosting schedules. In the GV1001 clinical trial presented herein, some subjects were tested negative after the first vaccination, but T cell responses developed detectably several months after the booster vaccination. As such, we found previous studies of GV1001 and other vaccines that seem to enhance T-cell responses by boosting vaccination. These studies suggest that repeated vaccination over an extended period of time results in a higher immune response and longer response. The question of how long to keep on vaccination remains to be answered.

However, the inventors of the present invention demonstrated that the immunized patients maintained the GV1001 response for 6-12 months through samples obtained up to 43 months after the last vaccination. The last study suggested that GV1001-vaccin vaccination may provide long lasting T cell memory. Given this vaccination, it may be necessary to provide booster injection only to patients who do not have a strong response in subsequent samples.

High levels of major Th1 (T-helper 1) reactor cytokines, Interferon-gamma (INFγ) and Tumor necrosis factor alpha (TNFα), and low levels of interleukin -4) and IL-10 (interleukin-10). These cytokine patterns may suggest a desirable match between immunity and T cells. If these reactions were only analyzed for IFNγ, IL-4 and / or IL-10 in the usual vaccine clinical trial, they would have been easily designated as “Th1”. Thus, it can be noted that the inventors of the present invention detected significant levels of the major Th2 (T-helper 2) cytokines IL-5 (interleukin-5) and IL-13 (interleukin-13). Subsequent observations are consistent with those found in other previous studies that the cytokine profiles of vaccine clinical trials often do not follow the Th1 / Th2 delineation. This suggests that Th2 cytokines may occur in response to potent immune activity. Long term survivors have been observed by the inventors of the present invention, and a broad range of Th1 / Th2 cytokines may indicate a polyfunctional response. Several studies, particularly those for infectious diseases, suggest that multifunctional cytokine profiles are associated with protective immunity.

In GV1001 clinical trials conducted without prior chemotherapy, the frequency of immune responders is similar to that assessed via Delayed-type hypersensitivity (DTH) recordings or T-cell assays. In contrast, most of the subjects of clinical trials vaccinated immediately following chemotherapy, presented in most of the present invention, were DTH negative. This observation suggests possible immunomodulatory effects of chemotherapy on GV1001 responses.

Telomerase is expressed by normal stem cells. It is noteworthy that when the GV1001 vaccination began immediately after chemo-radiotherapy in the example clinical trials mentioned below, no toxicity associated with stem cells was achieved. In view of the long-term safety, we monitored 19 study subjects for which no toxicity was detected for more than two years (see Table 2).

In addition, long-term data from patients in a relevant study (see end of this example below) suggested that boost immunization and sustained immune responses beyond 8 years were well tolerated. No toxicity was found in the continuous monitoring of bone marrow samples and in peripheral blood counts. We also observed retained blood counts of melanoma and colorectal cancer patients vaccinated with GV1001 for many years. In all, our data suggested a weak toxicity profile of GV1001 vaccination.

Most patients in the NSCLC clinical trial experienced an ongoing disease despite their immune response. This observation reflects the aggressive nature of progressive NSCLC. On the other hand, we observed long-lasting tumor responses in some subjects and found that almost all long-term survivors belonged to immune responders.

We concluded that GV1001 vaccination immunized NSCLC patients at high rates. High immune response rates are encouraging and indicate that the vaccine is applicable to the general patient population without prior HLA (Human leukocyte antigen) discrimination. In addition, while bone marrow function is not at stake, it induces long-term T cell memory for telomerase antigens. In phase II clinical trials involving the concept of a combination of vaccine and chemo- or radiotherapy, high immune response rates and low toxicity were observed. This is of interest for clinical development of both GV1001 and other cancer vaccines. Importantly, the study also provides signs of clinical experimental effects.

Progressive disease In patients with NSCLC, a series of clinical phase 1/2 trials (performed in patients who have never received concomitant chemo- or radiotherapy) showed a strong link between immune response and survival. We believe that the results further warrant research on the NSCLC of GV1001. High immune response rates, low toxicity, and clues to clinical activity provide the basis for conducting larger randomized trials. In this context it is interesting that 12/13 patients who survived more than 1000 days to date are immune responders. The observed association suggests that the evaluation of GV1001 for NSCLC in a randomized clinical trial is warranted. In pancreatic cancer, a GV1001 phase 3 clinical trial is currently ongoing.

For this reason, based on the above-referenced results obtained from a phase 2 clinical trial, the present invention discloses a method of treating cancer in a human patient, the method comprising an effective amount of the EARPALLTSRLRFIPK (SEQ ID NO: 1) peptide and immunological adjuvant And administering radiotherapy and optionally chemotherapy, wherein the patients receive radiotherapy for 4 to 28 days prior to the start of treatment.

As clearly set forth herein, this combination therapy has been shown to be promising for NSCLC patients, but SEQ ID No. 1 has shown beneficial anticancer immunity in patients suffering from a large number of different cancer types and being treated with radiotherapy or chemotherapy. It is considered to have the potential to induce. As noted here, it has recently been shown that it is possible to noticeably increase the immune response even in disease progression in patients treated with aggressive cytostatic / cytotoxic drugs and / or radiotherapy.

For this reason, according to the present invention, SEQ ID NO: 1 will be useful for combination therapy involving chemotherapy and / or radiotherapy, in most but not all cancer types, if chemotherapy and / or radiotherapy is used to combat disease. . It is preferred if the patients in question have been or have undergone chemotherapy and had received radiation therapy for 4 to 28 days prior to starting treatment with SEQ ID NO: 1.

According to a preferred embodiment of the invention, the cancer to be treated is non-small cell lung cancer (NSCLC).

According to a preferred embodiment of the invention, SEQ ID NO: 1 is administered intradermal but other convenient routes of administration may be applied. For example, SEQ ID 1 may be administered subcutaneously or through other commonly used parenteral routes of administration, such as intraperitoneal or intramuscular routes.

The preferred dosing position is the lower abdomen, but the dosing position can be conveniently and suitably modified. This means that the site of administration may be limbs or dorsal.

Usually the immune adjuvant is administered at the same site as SEQ ID NO: 1. "Identical site" herein means both the same site and the same kind of administration in the human body. Thus, if SEQ ID NO: 1 is administered intradermally in the lower abdomen, the adjuvant is also administered intradermally in the lower abdomen.

According to an embodiment of the invention, the immunological adjuvant is generally just prior to the administration of SEQ ID NO: 1, eg 1-30, 2-29, 3-28, 4-27, 5-26, 6-25, 7- Administered between 26, 8-25, 9-24 minutes prior to SEQ ID NO: 1 administration. Preferably the adjuvant is administered 10-15 minutes before the SEQ ID NO: 1 administration.

The dosage of SEQ ID NO: 1 must be effective, which should be at least 10 nmol, for example at least 50 nmol, or 200, or 250 nmol. On the other hand, the amount administered is usually at most 2000 nmol, for example at most 1500 nmol, at most 1000 nmol, or at most 800 nmol, for example at most 500 nmol. According to a particularly preferred embodiment the amount is about 300 nmol.

The adjuvant may be any convenient immunological adjuvant, but particularly immunostimulatory adjuvant. Examples of suitable immunological adjuvants are mentioned in WO 98/20027. A particularly immunologically interesting adjuvant is Granulocyte macrophage colony-stimulating factor (GM-CSF), which was administered in an amount of 75 μg per dose in one embodiment of the present invention. Dosages may be high or low as determined to be tested according to the efficacy of GM-CSF when injected through human sites and certain routes.

As mentioned above, patients treated by the present invention have previously been treated by chemotherapy and radiotherapy. Any chemotherapeutic regimen for the treatment of NSCLCL may be used, but in embodiments of the present invention, when immunized with SEQ ID NO: 1 after treatment with docetaxel, in detail, 20 mg / m ² (m ² label is treated Beneficial effect was observed after docetaxel administration.

In addition, the radiotherapy regimen may be any convenient regimen in NSCLC treatment, but in the examples of the present invention, the results after radiotherapy consisting of 2Gy × 30 3D radiotherapy have been reported as follows.

The method of the present invention involves repeated immunization following immunization 1 to induce an effective immune response. This immunization scheme allows patients to receive the same dose of SEQ ID NO: 1 and an immunological adjuvant for the first immunization via the same route, followed by the following scheme: twice a week after the first dose, 2 Follow the examples below, followed by dosing once per week 3, 4, 6, 8, 10, 14, 18, 22, once every 6 months and once every 9 months. However, such an immune scheme can be optimized, and as shown in the examples, there is the possibility that the number of subsequent or supporting immunizations can be reduced-a skilled immunologist can make a decision from a more optimized immunization scheme. . In general, the immunological status of an individual patient is monitored and based on the results of this monitoring, the immunization protocol can be adjusted individually for each patient.

The present invention also refers to GV1001 (SEQ ID NO: 1) used for the methods disclosed herein and in the claims. The present invention furthermore refers to the use of SEQ ID NO: 1 for the preparation of pharmaceutical compositions for the treatment of NSCLC according to the methods disclosed in the specification and claims of the invention.

[Definition of Terms]

"GV1001" refers to peptide EARPALLTSRLRFIPK (SEQ ID NO: 1), which is derived from the amino acid sequence of human telomerase protein (hTERT).

"Radiotherapy" generally refers to treating a tumor with the use of ionized radiation, which can be applied locally or systemically. The exact type of effective radiotherapy is cancer-specific and generally well known to those skilled in the art.

"Chemical therapy" has generally been used to treat cancer patients with various cytotoxic or cytostatic drugs. That is, in the context of the present invention, chemotherapy refers to an anticancer therapy using, for example, the following groups of drugs:

Anti-metabolites (L01B) mimic purines (eg, azathioprine and mercaptopurine) or pyrimidines, which are the building of DNA Block. Anti-metabolites prevent these substances from being included in DNA during the "S" phase of the cell cycle and prevent normal development and division. They also affect RNA synthesis.

Vegetable alkaloids and terpenoids (L01C). These alkaloids interfere with the function of the microtubules. The main examples are vinca alkaloids and taxenes. The vinca alkaloids (L01CA) are from Madagascar periwinkle, Catharanthus roseus (formally known as Vinca rosea). Derived, and

Vincristine, vinblastine, vinorelbine, vindesine.

Podophyllotoxin (L01CB) is a plant-derived compound known to aid digestion and is used to produce two different cell proliferative inhibitors, etoposide and teniposide. They prevent cells from entering the G1 phase (DNA replication initiation phase) and DNA replication (S phase).

Taxane (L01CD) is derived from the natural product paclitaxel, originally known as Taxol, and first from the bark of the Pacific Yew tree. Docetaxel is a semi-synthetic analog of paclitaxel. Taxanes enhance the stability of microtubules to prevent chromosomes from separating during the anaphase of the cell cycle.

Topoisomerase inhibitors (L01CB and L01XX). These include type I topoisomerase inhibitors such as camptothecin, irinotecan, topotecan, and type II inhibitors such as amsacrine, etoposide, eto Epoposide phosphate, teniposide.

Cytotoxic antibiotics (L01D) are actinomycin (L01DA01), anthracyclines such as doxorubicin (L01DB01), daunorubicin (L01DB02), Valrubicin, idarubicin, epirubicin (L01DB03), and other cytotoxic antibiotics such as bleomycin (L01DC01), plicacamine (L01DC02) , Mitomycin (L01DC03).

"Immunoadjuvant" has its usual meaning: when a substance or composition, when administered to an individual, helps induce / induce a specific immune response to an antigen. Preferred adjuvant in the present invention is GM-CSF.

1 is a graph showing GV1001-specific T cell responses.
Peripheral blood mononuclear cells (PBMCs) were obtained before starting therapy, at week 6, week 10 and immediately after each vaccine was administered. PBMC was stimulated once in vitro and was used in proliferation experiments against radioactively exposed PBMC ± peptide GV1001. The columns represent the average SI (stimulatory index) (divided into responses with and without GV1001). If the recorded SI value exceeds the upper limit of each chart, the correct SI value is displayed at the top of the columns. The diagram shows the T cell responses before, after and after vaccination of all evaluable patients. In each individual patient, the time point with the highest SI value is indicated. Reactions with SI values> 2 were considered GV1001 specific.
2 is a graph showing GV1001-specific T cell responses.
PBMCs were obtained before the start of therapy, week 6, week 10 and immediately after each vaccine was administered. PBMCs were stimulated once in vitro and were used for proliferation experiments against radiation exposed PBMCs ± GV1001. Columns represent mean cpm (counter per minute) or SI values (divided into reactions with and without GV1001). If the recorded cpm value or SI value exceeds the upper limit of each chart, the exact cpm value or SI value is displayed at the top of the columns. B and C represent long-term T cell memory. The diagram shows the development of the T cell response recorded from subsequent samples. DF shows that most of the samples were stored negatively overnight before PBMC isolation (marked), which was the same even in newly isolated and tested positive samples. G depicts parallel experiments of PBMC samples separated or stored overnight upon arrival.
3 is a graph showing the survival of patients.
Progression-free survival (PFS) values were analyzed by Kaplan-Meier analysis. Immune responders were compared to non-immune responders. PFS values were defined as clinical endpoints in the protocols disclosed herein since standard treatment after progression appears to affect OS (Stage 3 patients). Plots show PFS values in studies with or without patients with GV1001 T-cell responses. For immune responders, the observed PFS value is prolonged.

[Preparation and Method]

Patients and Research Protocols

The primary purpose of phase 2 clinical trials is an immunological response. Toxicity and run time are secondary purposes. Twenty-three subjects with inoperable NSCLC tertiary A / B were enrolled at three different centers in Norway between November 2006 and July 2008. Twenty patients were enrolled at the Norwegian Radium Hospital, four St. Olav® Hospitals, and seven Southern Hospitals in seven Kristansands. Clinical trials were approved by the Norwegian Medicines Agency, the Regional committee for Medical Research Ethics and the Hospital Review Board. This was in compliance with the World Medical Association Declaration of Helsinki. All patients received signature consent.

The study population has been treated with 20 mg / m ² docetaxel and 2Gy x 30 three-dimensional (3D) radiation therapy for the last four weeks. Subjects with metastatic disease were excluded from previous studies based on thoracic and upper abdomen CT scans and MRI scans of the brain. Eligibility criteria were Eastern Oncology Group (ECOG) performance status 0-2, age = 18 years, leukocyte count = 1.5 × 10 ⁹ / L; Platelet count = 100 × 10 ⁹ / L, hemoglobin level = 9 g / dL (= 5.6 mmol / L); Creatinine levels = 140 μmol / l (1.6 mg / dl), bilirubin = 20% above normal upper limit, aspartate amino transferase (ASAT) and alanine amino transferase (ALAT) = 1.5, normal albumin Upper limit = 2.5 g / L. Criteria excluded include active infection or significant heart disease requiring antibiotic treatment, except for baseline or skin cell carcinoma or uterine cancer stage IB that has been curable if previously had a malignant tumor history or Other medical conditions, such as severe hemorrhagic heart failure, variable angina, severe arrhythmia, cases of severe negative reactions to vaccines such as anaphylaxis were included. Patients positive for autoimmune disease or hepatitis B, C or HIV infection were also excluded from the study.

Experimental Design

The strategy behind this study is to establish a phase 3 clinical trial in Phase 3 patients with NSCLC and to evaluate the vaccine within various treatment strategies. The dose of GV1001 is based on data from our previous experiments with dose-increasing clinical trials in NSCLC and pancreatic cancer. Chemoradiotherapy was the 2006 institutional standard of care for inoperable NSCLC stage 3. Our decision to include 20 evaluable patients was based on the main research objectives: to show whether treatment with chemotherapy and GV1001 is feasible and to obtain immunization, to provide safety data, and to the PFS and immune response rates. Based on obtaining an estimate of For a given sample size, the number of subjects with immune responses and the number of serious adverse events (SAEs) will follow the binomial distribution. Statistical calculations were based on n = 20 and the binomial distribution shown. Assuming an actual response rate of 54% observed in the Phase 1/2 clinical study, the probability of detecting 5 or more immune responders is 99.8%. Assuming a 10% actual SAE frequency, the probability of detecting one or more SAEs is 87,8%.

cure

In clinical trials, GV1001 vaccination began within 4 days and 4 days after the last radiotherapy treatment. Immunization was given once each week 1 (Monday, Wednesday and Friday) and week 2, 3, 4, 6, 8 and 10, respectively. Dependent immunizations were given at weeks 14, 18, 22, 6 months and 9 months, respectively. GV1001 (300 nmol peptide in 0.20 ml saline) was injected in the lower abdomen by intradermally (id) administration. GM-CSF (75 μg Leukine; Bayer, Oslo Norway) was injected at the same site 10-15 minutes prior to GV1001.

Peptide

Vaccine peptide GV1001 corresponds to 16 amino acid residues 611-626 of hTERT (EARPALLTSRLRFIPK; SEQ ID NO: 1). GV1001 was provided by Pharmexa (HØsholm, Denmark). The manufacturing process was GMP compliant. RAS-peptide 508 (KRAS 52-70, Q61H; Norsk Hydro, Norway) served as a negative control of the T-cell assay.

T-Cell Culture and Assays

Peripheral blood monocytes (PBMCs) were obtained before the start of therapy, at weeks 6 and 10 and immediately after every vaccination. PBMCs were separated and frozen as previously described. Thawed PBMCs were stimulated once in vitro with vaccine peptides prior to T-cell assay, as mentioned previously. After this initial stimulation, PBMCs were incubated with GV1001 (25 μmol / L) for 7-10 days and with added IL-2 (10U / ml) from day 3.

T-cell proliferation assay ( ³ H Thymidine) was done essentially as mentioned previously. Pre- and post-inoculation samples were analyzed in parallel for response to peptides. Autologous PBMCs irradiated with radiation were used as antigen-labeled cells (APCs). Stimulation with Staphylococcal enterotoxin C (SEC) was used as a positive control and immunocompetence. All patients responded to SEC. T-cell cultures were tested three times. SEM was usually less than 10%. Proliferation measurements after irrelevant peptide (K-RAS 508) stimulation were generally not significantly different from controls without peptide. T-cell responses were considered antigen-specific when the stimulatory index (SI; antigen-free and split antigen responses) was higher than 2.

Bioplex cytokine assays were performed in supernatants collected 48 hours after T-cell stimulation and followed manufacturer's protocol (Bio-Rad Laboratories). Supernatants were analyzed twice / three times and each similarity remained separated through T-cell stimulation and bioplex assays.

Delayed type hypersensitivity

Delayed-type hypersensitivity (DTH) skin tests were performed at baseline, weeks 2, 3, 4, 6, 10 and post vaccination. During DTH testing, 60 nmol GV1001 in 0.10 ml saline was injected intradermally without GM-CSF at the site separated from the vaccinated site. Patients registered a DTH skin response 48 hours after dosing. Positive DTH test was defined as erythema / curing with mean diameter = 5 mm.

Clinical evaluation

Adverse drug reactions and ECOG performance were assessed at each visit. Blood screening and general medical examinations were performed at the start of vaccination (week 1), at 2, 3, 4, 6, 8, 10 and each subsequent vaccination. CT scans were performed before vaccination, week 14, and every three months thereafter.

Progression-free survival was defined as the main clinical end point in the CTN-2006 protocol because overall survival tended to be affected by standard treatment after progression. Radioactive fibrosis is generally difficult to distinguish clearly from tumors. At the beginning of vaccination, patients after chemo-radiotherapy with or without viable tumor tissue have residual CT lesions. Thus, the timing of completion, partial response and stable disease was not appropriate. Progressive disease is defined as a new or ongoing lesion identified by CT scan, bronchoscopy and / or histology.

statistics

Progression free survival (PFS) was calculated from the start of vaccination. In view of PFS, Kaplan-Meyer / log-rank analysis was applied from comparing immune responders to non-immune responders. To assess whether the immune response represented an independent diagnostic factor, Cox regression with an entry assay was performed. Subjects were found to be 3A or 3B.

[result]

Patient characteristics and adherence to treatment

Patient characteristics and treatment details are shown in Table 1. In late 2007, Bayer withdrew liquid GM-CSF (Leukine) from the market. This caused a sudden shortage until the lyophilized product was delivered. Therefore, in Radiumhospitalet, one of two patients received GV1001 injection without immunological adjuvant GM-CSF. In St. Olaf Hospital, two other patients did not receive the combined vaccination twice (weeks 8 and 10), respectively. The panel-monitored study replaced patients who did not receive GM-CSF at vaccination 2 (# 203 and 204) and excluded from protocol analysis. Patient 110 was also not evaluable for the protocol because she was withdrawn at 8 weeks due to tumor and disease progression of the right lung.

safety

The safe population includes all patients who have been vaccinated at least once (n = 23). A total of 323 vaccine doses were administered (8-21 doses per patient). Seven serious adverse events (SAEs) were reported in six patients. However, bronchoscopy performed with biopsy showed that the fistula was due to tumor recurrence.

Immune response

GV1001-specific T cell responses were seen in 16 patients after vaccination, compared to no patients in the pre-vaccination sample (see FIG. 1). Positive DTH responses were observed in only one patient. Three subjects were not evaluable according to the protocol (see above). The immunological response rate was 70% by Intention to Treat (ITT) and 80% by protocol.

In order to achieve sustainable clinical effects, development of T cell memory appears to be required. We therefore provided a booster vaccine and monitored the long-term development of the immune response. Subsequent samples were obtained from 15/16 immune responders. The results showed long-lasting GV1001-specific T cell responses in 13/15 subjects, with observation periods up to 91 weeks (see Table 2 and FIGS. 2A-B).

Compared to other centers, we compared 9/12 patients (9/11 per protocol) of Radiumhospitalet Oslo, 1/4 patients of Trondheim St. Olaf Hospital (1 / per protocol). 2) and GV1001-specific T cell responses of 6/7 patients of Kristiansand. Samples from Trondheim and Kristansand were sent to Oslo for T cell analysis. Some of the first samples were stored overnight prior to PBMC separation. Samples stored overnight gave only negative results. We therefore decided to separate subsequent samples from the same patients upon arrival. Interestingly, most previous negative patients were tested positive. In addition, later samples stored from the same patients were negatively tested upon overnight storage (see FIGS. 2C-E). We also stored the tested samples the same day or overnight at the same time and observed that the response of freshly stored samples was clearly strong (see FIG. 2F). These observations have shown difficulties in managing the multicenter clinical of T cell assays and suggested that T cell data may yield false results if the assay is optimized and performed with untreated samples.

Clinical response

Table 2 shows progression free survival (PFS), overall survival and recurrence points. PFS is a clinical end point following the protocol and was evaluated by CT scans at three month intervals. For date, tumor progression has been recorded in 17/23 patients in the ITT population (median PFS value of 357 days). Five out of six patients with no signs of relapse are immune responders. Considering all patients involved, immune responders recorded increased PFS, with an average value of 371 days versus 182 days compared to non-immune responders (see FIG. 3).

                         SEQUENCE LISTING <110> GEMVAX AS   <120> TREATMENT OF NON-SMALL CELL LUNG CARCINOMA BY ACTIVE IMMUNOTHERAPY <130> PCTEP2012054272 <160> 1 <170> PatentIn version 3.2 <210> 1 <211> 16 <212> PRT <213> Homo sapiens <400> 1 Glu Ala Arg Pro Ala Leu Leu Thr Ser Arg Leu Arg Phe Ile Pro Lys 1 5 10 15

Claims (14)

A composition for treating cancer patients comprising a peptide consisting of SEQ ID NO: 1 as an active ingredient,
The patient has previously received both radiotherapy and chemotherapy to treat cancer,
The composition for treating cancer patients comprising the peptide of SEQ ID NO: 1 as an active ingredient is to start administration within 4 days to 4 weeks after the last radiotherapy treatment,
The peptide is administered three times a week and once at 2, 3, 4, 6, 8 and 10 weeks, and then at 14, 18, 22 weeks, 6 months and 9 months, respectively.
The chemotherapy is a composition for treating cancer patients, characterized in that the administration of Taxane (Taxane). The composition for treating cancer patients according to claim 1, wherein the patient has received radiotherapy for 4 to 28 days before the start of the treatment according to SEQ ID NO: 1. The composition of claim 1, wherein the cancer is non-small cell lung cancer (NSCLCL). The composition of claim 1, wherein the composition is administered intradermally. The composition for treating cancer patients according to claim 4, wherein the composition is administered to the lower abdomen. The composition for treating cancer patients according to claim 1, wherein the composition is administered in combination with an immunological adjuvant. The composition of claim 6, wherein the immunological adjuvant is administered 10 to 15 minutes before administration of the composition. The composition of claim 6, wherein the immunological adjuvant is administered in an amount of 75 μg. 7. The composition of claim 6, wherein the immunological adjuvant is granulocyte macrophage colony stimulating factor (GM-CSF). The composition for treating cancer patients according to claim 1, wherein the peptide consisting of SEQ ID NO: 1 is administered in an amount of 10 nmol to 2000 nmol. The composition of claim 1, wherein the chemotherapy is docetaxel. The composition of claim 11, wherein the docetaxel is administered at 20 mg / m ² per week. A kit for treating cancer patients comprising a composition according to any one of claims 1 to 12 and an immunological adjuvant. The kit for treating cancer patients according to claim 13, wherein the kit further comprises instructions on the dosage and method of administration of the composition and immunological adjuvant.

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