A Broad Based Antiangiogenesis and Antioxidant Protocol
1 Executive Summary:
This Investigational Board Approval (IRB) Submission is centered on antiangiogenesis and antioxidant treatments diminishing tumor growth and metastasis, specifically using tetrathiomolybdate, zinc, ascorbic acid, N-acetylcysteine and vitamin B6. Copper bound to ceruloplasmin increases angiogenic activity and correlates with tumor incidence, burden and malignant progression.2 Copper has been found to behave as a molecular switch for activating cytokines, interleukin-1 (IL-1) and tumor necrosis factor-alpha (TNF-a), and growth factors such as basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF).2 All four of the above signaling factors have been shown to be angiogenic. Copper seems to act as an “obligatory cofactor” allowing for the angiogenic activator to become functional.2 In addition, copper was found to stimulate the directional migration of endothelial cells where other trace metals were not.2 The underlying hypothesis of antiangiogenesis using copper-reduction therapy is that the level of copper required for angiogenesis is higher than that required for essential copper-dependent cellular functions.9 Having established that copper is intimately involved in tumor growth via the angiogenic pathway, it is feasible to propose a method of treatment, which will decrease the body’s concentration of copper. Drs. Brewer and Merajver from the University of Michigan have used TM to reduce copper levels in patients.12,14 TM has been shown to be essentially nontoxic, fast acting, and copper specific.12 The goal of these studies has been to reduce ceruloplasmin levels to 20% of the patient’s normal baseline.9,12 Ceruloplasmin (CP) levels more accurately measure copper depletion than total copper alone.9 As previously mentioned, zinc has been shown to reduce copper absorption, although zinc is not a chelator. Zinc like TM is nontoxic but has the disadvantage of having therapeutic effectiveness much slower than TM.15 Zinc lowers copper levels by inducing hepatic and intestinal metallothionein (MT) synthesis which in turn binds copper, rendering it unavailable for absorption into the bloodstream.16 Enterocytes lining the small intestine are shed periodically into the lumen of the intestine with the intracellular copper-MT complex and subsequently, excreted.15
N-acetylcysteine (NAC) is derived from the sulfur-containing amino acid cysteine. It is found naturally in foods and serves as a powerful antioxidant.19 Furthermore, NAC was shown to prevent angiogenesis of endothelial cells, inhibiting invasion and metastasis of malignant cells.38 The mechanism of this action is believed to be due to the production of angiostatin by NAC.2,44 NAC and ascorbic acid,when used together, have an additive effect in inhibiting lung tumorigenicity.42 Ascorbic acid, like NAC, has been shown to enhance the activity of cytotoxic drugs such as cisplatin, paclitaxel, dacarbazie, 5-FU, and doxorubicin.31 Also, ascorbic acid may act as an antiangiogenic agent by aiding in copper reduction. Ascorbic acid decreases the intestinal absorption of copper and the oxidase activity of serum ceruloplasmin.6,7,11In summary we have combined tetrathiomolybdate, ascorbic acid and N-acetylcysteine to act synergistically in their respective functions to reduce copper and act as antioxidants.
2 Table of Contents
|2||TABLE OF CONTENTS|
|4.1||INTRODUCTION, SCIENTIFIC BACKGROUND AND PROTOCOL|
|4.1.2||Copper’s Role in Angiogenesis and Cancer|
|7||TRIAL DESIGN AND RATIONALE|
|7.1||Protocol for Treatment|
|7.1.3||Antioxidant Treatment- N-acetylcysteine and Ascorbic Acid|
|7.2.1||Protocol for Copper reduction|
|7.3||TM Blood Test Schedule|
|8||SELECTION OF SUBJECTS|
|8.1||SUBJECT INCLUSION CRITERIA|
|8.2||SUBJECT EXCLUSION CRITERIA|
|9.1.3||Storage and Stability|
|9.1.4||Source of Drug|
|9.2||N-acetylcysteine (NAC)-Rx Mucomyst or generic equivalent|
|9.2.3||Storage and Stability|
|9.2.4||Source of Drug|
|9.3||Ascorbic Acid tablets|
|9.3.3||Storage and Stability|
|9.3.4||Source of Drug|
|9.4.3||Storage and Stability|
|9.4.4||Source of Drug|
|10||Clinical Efficacy Assessments: Objective Tumor Response|
|10.1||Methods of Assessments|
|10.2||Definitions of Objective Tumor Response and Progression|
|10.2.3||Evaluation of Overall Objective Tumor Response|
|10.3||Methods and Assessment- Tumor Markers|
|11||Clinical Safety Assessments|
|11.1||Definition of an Adverse Event|
|11.2||Serious Adverse Events|
|11.3||Unexpected Adverse Events|
|11.4||Adverse Events Reporting Period|
|11.5||Eliciting Adverse Event Information|
|11.7||Recording Adverse Events in the CRF’s Procedures|
|11.8||Exposure In Utero|
|12.2||Objective Tumor Response Rate|
|12.3||Time to Objective Tumor Response|
|12.4||Duration of Objective Tumor Response|
|12.5||Time to Objective Tumor Progression|
|12.6||Time to Treatment Failure|
|13||QUALITY CONTROL AND QUALITY ASSURANCE|
|14||Ethical Conduct of the Trial|
|15||Institutional Review Board/ Independent Ethics Committee|
|16||Patient Informed Consent|
|16.2||US- Specific IRB/IEC Requirements|
ARC: AIDS Related Complex
BFGF: basic Fibroblast Growth Factor
CBC: Complete Blood Count
CR: Complete Response
HIV: Human Immunodeficiency Virus
IRB: Institutional Review Board
NE: not evaluable for response
PD: Progressive Disease
PR: Partial Response P & U: Pharmacia and Upjohn
RECIST: Response Evaluation Criteria in Solid Tumors
ROS: Reactive Oxygen Species
SD: Stable Disease
TNF-a: Tumor Necrosis Factor-alpha
VEGF: Vascular Endothelial Growth Factor
4 Background Information
4.1 Introduction, Scientific Background and Protocol
For a tumor to grow and metastasize it must have a blood supply. Angiogenesis is the process by which new blood vessels called capillaries develop. Angiogenesis occurs naturally in reproduction, the healing process of wounds, and in the development of the embryo.1 Angiogenesis occurs also in several unnatural, pathological conditions such as rheumatoid arthritis, diabetic retinopathy, and cancer.1 In the case of cancer, a tumor will not grow larger than a pea size unless it obtains a blood supply through the angiogenic process.
Endothelial cells line blood vessels. The tumor produces angiogenic substances to initiate endothelial cell growth with subsequent capillary growth. These signals may be growth factors, proteases, trace elements, oncogenes, signal transduction enzymes, cytokines, and endogenous modulators.2 Some antiangiogenic agents have been found over the past few years inhibiting some of these signals. This Investigational Board Approval (IRB) Submission is centered on antiangiogenesis and antioxidant treatments diminishing the impact of tumor growth and metastasis, specifically using tetrathiomolybdate, zinc, ascorbic acid, N-acetylcysteine and vitamin B6.
4.1.1 Copper Metabolism
Copper is found in whole grains, shellfish, legumes, liver, and nuts while poor sources of copper seem to be dairy products.4 Copper is rapidly absorbed from the stomach and small intestine and mainly stored in the liver.4 Most of the copper in the body is bound to proteins (Table 1) leaving a small amount unbound.4
Proteins Bound to Copper4
|Transport proteins||Storage proteins||Cu-containing Enzymes||Enzyme function|
|Metallothioneins||Cytochrome C oxidase
Clotting factor V
Blood clotting Antioxidant
However, antagonists of copper absorption may be present and therefore impede copper absorption. Zinc and vitamin C have been shown to be strong antagonists of copper absorption.4 Zinc supplements have been shown to decrease copper status in adult males.5 Furthermore, ceruloplasmin activity, which is one of the main copper binding proteins and is directly proportional to bound copper concentration, has been shown to decrease when supplements of vitamin C were taken for 64 days.6 Also, a study done on laboratory rats reflected that ascorbic acid induced a decrease in the intestinal absorption of copper.7
Drs. Brewer and Prasad have worked extensively on Wilson’s disease, a genetic disease in which there is excessive accumulation of copper in the liver. Dr. Brewer and his team treated patients with 50mg of zinc three times a day while on a low copper diet.8 This protocol has been used successfully for the past 15 years.
In addition to zinc and vitamin C, sulfur and molybdate have been shown to be antagonists of the absorption of the trace element. A complex of sulfur and molybdate known as tetrathiomolybdate (TM) has proven to be extremely safe for copper reduction9 and has been shown to occur naturally in ruminant animals for copper removal.3 TM’s activity lies in that it is a potent chelator for copper. If taken at mealtime, it prevents the body form absorbing copper in food and copper found in gastric and salivary secretions.10 Taken between meals, TM in the blood would bind copper to a blood protein forming a TM-copper-protein complex.10 Therefore, ascorbic acid, zinc, and TM may act indirectly as inhibitors of angiogenesis by inhibiting copper absorption.
4.1.2 Copper’s Role in Angiogenesis and Cancer
Copper levels are usually elevated in cancer.2,4,9 Plasma levels of copper have, however, proven to be somewhat misleading. Ruminants form thiomolybdates which complex with copper and inhibit absorption of the trace mineral. There is an initial increase of copper plasma levels due to copper release from the liver.3 Likewise, patients on TM therapy actually have increased blood copper levels.9 Ascorbic acid does not affect copper absorption, retention, total serum copper or the serum level of ceruloplasmin.11 However, the oxidase activity of ceruloplasmin was decreased significantly.11 The most reliable biomarker for monitoring copper reduction is ceruloplasmin.9
Ceruloplasmin synthesis by the liver is controlled by the amount of copper available to the liver; therefore, as copper is reduced, serum ceruloplasmin is reduced.12 This copper-binding molecule is non-angiogenic when not bound to copper but upon binding, becomes angiongenic.2 Ceruloplasmin has been investigated as a diagnostic marker of cancer showing significant elevations in advanced stages of solid malignant tumors.13 It has also been shown to be a good marker for lymphoma.9 This principal transport protein of copper has been shown to increase four to eight-fold during malignant progression.2 Thus, increased copper bound to ceruloplasmin increases angiogenic activity correlating with tumor incidence and burden and malignant progression.2
Copper has been found to behave as a molecular switch for activating cytokines, interleukin-1 (IL-1) and tumor necrosis factor-alpha (TNF-a), and growth factors such as basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF).2 All four of the above signaling factors have been shown to be angiogenic. Copper seems to act as an “obligatory cofactor” allowing for the angiogenic activator to become functional.2 In addition, copper was found to stimulate the directional migration of endothelial cells where other trace metals were not.2 The level of copper required for angiogenesis is higher than required for essential cellular functions which are copper-dependent.9
In conclusion, all four of the above angiogenic pathways could be inhibited by simply reducing the copper levels significantly while maintaining a baseline of copper for essential enzymatic pathways. Ongoing clinical trials are now testing a complex of sulfur and molybdate known as tetrathiomolybdate (TM), and the lack of toxicity in TM indicates that it may be extremely safe for copper reduction.9
5 Trial Objectives
5.1 Primary Objective
Reduce tumor load by antiangiogenic treatments
5.2 Secondary Objective
Utilize tumor markers in the assessmemt of residual tumor
6 Trial Endpoints
6.1 Primary Endpoints
Image reduction of tumor mass
6.2 Secondary Endpoints
The reversal of tumor marker levels to normal values
7 Trial Design and Rationale
7.1 Protocol for Treatment
7.1.1 Tetrathiomolybdate (TM)
Having established that copper is intimately involved in tumor growth via the angiogenic pathway, it is feasible to propose a method of treatment, which will decrease the body’s concentration of copper. Drs. Brewer and Merajver from the University of Michigan have used TM to reduce copper levels in patients.12,14 TM has been shown to be essentially nontoxic, fast acting, and copper specific.12 The goal of these studies has been to reduce ceruloplasmin levels to 20% of the patient’s normal baseline.9,12 Ceruloplasmin levels more accurately measure copper depletion than total copper alone.9 By reducing to such a level, the body’s normal important copper-based reactions could carry on normally. It has been reported that mammary cancer in transgenic mice has been prevented using tetrathiomolybdate (TM).2 TM binds with copper making a stable compound, which is cleared by the body.12
By giving patients 20mg of TM three times daily with meals and dose level of 20mg three times between meals, a reduction of ceruloplasmin to 20% normal was achieved in 60-90 days.12 This 20% normal is referred to by Brewer and Merajver as a “window” in which the copper levels are low enough to initiate antiangiogenesis activity but high enough for normal metabolic functions of copper.10 Iron levels should be monitored and an iron supplement may be warranted because one of the first signs of copper deficiency is that of anemia.9 This is due to copper’s importance in the synthesis of heme, a protein imperative for red blood cell formation. The hematocrit should be monitored in copper-reduction therapy. Brewer and Merajver set ceruloplasmin reduction to 20% of baseline while only reducing the hematocrit to 80% of baseline.12 Thus, TM has been shown to be an effective chelator of copper with few side effects. Therefore, for antiangiogenic copper-reduction therapy, TM may prove to be the safest and most potent cancer therapy.15
As previously mentioned, zinc has been shown to reduce copper absorption, although zinc is not a chelator. Zinc like TM is nontoxic but has the disadvantage of having therapeutic effectiveness much slower than TM.15 Zinc lowers copper levels by inducing hepatic and intestinal metallothionein (MT) synthesis which in turn binds copper, rendering it unavailable for absorption into the bloodstream.16 Enterocytes lining the small intestine are shed periodically into the lumen of the intestine with the intracellular copper-MT complex and subsequently, excreted.15
Drs. Brewer and Prasad used 150mg of elemental zinc in 50mg doses given three times daily one hour before or after a meal in the treatment of Wilson’s disease, a genetic disease in which there is excessive accumulation of copper in the liver.9,17This dose of zinc is approximately ten times the RDA of 15mg/day, however, zinc toxicity occurs very rarely in humans. Doses of greater than 2 grams daily (13 times Dr. Brewer’s recommended dose) will lead to gastrointestinal irritations such as nausea, stomach upset and vomiting.18 With zinc having such a low potential for toxicity, Brewer and Prasad successfully treated Wilson’s disease with zinc and found it to be as effective as treatment with TM, although significantly slower.9 With TM, ceruloplasmin levels reach the target range within one and a half to two months whereas, zinc may take up to six months or more to reach the therapeutic levels of copper reduction.9
In order to lower the copper to a therapeutic level in a timely manner, a combination of TM and zinc may be used. This can be accomplished by a patient taking 20mg of TM at mealtime three times per day and a combination of 20mg of TM and 50mg zinc 3 times daily at one hour or more before or after a meal. The supplementation with zinc would only be used if the patient did not reduced their copper levels to the therapeutic 20% of baseline within 60 days.
7.1.3 Antioxidant Treatment – N-acetylcysteine and Ascorbic Acid
Antioxidants neutralize free radicals, which are produced by normal metabolic activity. Free radicals such as superoxide radicals, hydroxyl radicals, peroxyl radicals, and alkoxyl radicals when left unchecked wreak havoc on cells causing damage to membranes and DNA. Since these free radicals have the potential for such devastating damage, they have been considered by scientists to be a major factor in the cancer and aging processes.
N-acetylcysteine (NAC) is derived from the sulfur-containing amino acid cysteine. It is found naturally in foods and serves as a powerful antioxidant.19 NAC is a precursor of intracellular glutathione which behaves as an antioxidant as well, functioning to remove toxic peroxides.20 NAC is readily absorbed, quickly converted to L-cysteine and then intracellular glutathione, thus, replenishing and maintaining healthy levels of glutathione.20 In a study done by De Flora et al., NAC induced an increase in oxidized glutathione reductase activity in rats.21 NAC and glutathione are very important antioxidant, detoxifying agents of the body.
Furthermore, NAC was shown to prevent angiogenesis of endothelial cells, inhibiting invasion and metastasis of malignant cells.38 The mechanism of this action is believed to be due to the production of angiostatin by NAC. 2,44
Both NAC and glutathione are sulfur-containing compounds. Sulfhydryl groups react with heavy metals such as mercury, lead, boron, cadmium, and chromium.20 In addition, NAC and glutathione have been found to conjugate with metabolites of valproic acid in rats and humans, ridding the body of these compounds which may contribute to hepatotoxicity.22 Furthermore, orally administered NAC seems to have a protective effect against gastric damage induced from ethanol.23 In addition, NAC is used to prevent damage to the liver caused by acetaminophen overdose.20,24-26
NAC affects the immune system by increasing intracellular glutathione. Reduced intracellular glutathione levels are present in the Human Immunodeficiency Virus (HIV), which is reversed when NAC is administered and thereby blocking the AIDS virus production in vitro.19 Furthermore, NAC enhances T cell colony formation in vitro of AIDS and ARC (AIDS related complex) patients.27 Not only does NAC work to enhance the immune system, it protects the immune cells from free radical damage via its antioxidant activity.
In addition to the antioxidant and detoxifying roles of NAC, this compound has been found to be effective as a mucolytic agent breaking up mucus in patients with pulmonary disorders such as chronic fibrosis, asthma, chronic bronchitis, and pneumonia.19 NAC functions to reduce the viscosity of the mucus allowing for easier expectoration.20
Cancer research has shown NAC to be an important protective agent due to its antioxidant and detoxifying abilities. NAC has been shown to protect against carcinogens found in air which lead to respiratory complications due to their DNA-damaging effects.28 NAC was shown to protect rat liver and lung mitochondrial DNA from damage caused by toxic carcinogens such as cigarette smoke.29 NAC could protect from interferon (IFN)-induced reduction of cytochrome P450 which is vital to drug metabolism in the liver.30 This is significant because cancer patients who may be undergoing IFN treatment should take NAC because of its antioxidant potential. IFN increases a liver enzyme, xanthine oxidase, which is known to produce reactive oxygen species which attack harm the cytochrome P450 system and other cellular components.30
In addition to interferon, the cytotoxic effects of antineoplastic agents are affected by reactive oxygen species (ROS), implying that for the most effective response to chemotherapy, antioxidants should be used.31 Antineoplastic drugs depend on rapid proliferation of cells for optimal activity.31 Oxidative stress, ROS production leads to lipid peroxidation which adversely affects cell proliferation.31 If cells aren’t proliferating rapidly, antineoplastic drugs are not as effective. Cancer cells have mechanisms, which have allowed them to prevent lipid peroxidation, allowing for rapid proliferation. If excessive oxidative stress occurs in cancer cells, the proliferation will diminish, and therefore, the cancer is less responsive to chemotherapy.31 However, antioxidant supplements during chemotherapy may optimize the activity of the antineoplastic agent. Doxorubicin (DOX) is a cytotoxic drug shown to have a synergistic effect against metastasis in mice when combined with N-acetylcysteine.32 NAC was shown to also prevent DOX-induced myelogenotxicity and alopecia while inhibiting tumor cell metastasis and invasion.33 The activity of cyclophosphamide and ifosfamide was not impaired when taken with NAC.31 Furthermore, supplementation with NAC during chemotherapy treatment may also reduce chemotherapy-induced side effects. NAC reduces cisplatin-induced nephrotoxicity and also protects against bleomycin-induced genotoxicity.31
Free radicals induce expression of cyclooxygenase (COX-2) which has been shown to play a role in colorectal cancer.34,35 COX-2 acts as an angiogenic agent.36,37 NAC was shown to decrease COX-2 expression in a human colorectal cancer cell line and therefore, may act as an antiangiongenic factor.35 Furthermore, NAC was shown to prevent angiogenesis of endothelial cells, inhibiting invasion and metastasis of malignant cells.38 Also, NAC was found to suppress development of tumors or preneoplastic lesions in rodents39 and to induce apoptosis in transformed cell lines by increasing p53 expression, a tumor suppressor gene.40
Ascorbic acid (vitamin C) has been known for years to be a powerful antioxidant but also may act as a pro-oxidant producing radicals when combined with transition metals as in metal-overload states.41 With this in mind, D’Agostini et al. (2000), investigated the combined effects of NAC and ascorbic acid and found that when used together were shown to have an additive effect in inhibiting lung tumorigenicity of urethane in mice while also inhibiting mutagenicity of chromium(VI).42 Ascorbic acid, like NAC, has been shown to enhance the activity of cytotoxic drugs such as cisplatin, paclitaxel, dacarbazie, 5-FU, and doxorubicin.31 Also, ascorbic acid may act as an antiangiogenic agent by aiding in copper reduction. Ascorbic acid decreases the intestinal absorption of copper and the oxidase activity of serum ceruloplasmin.6,7,11
7.2.1 Protocol for Copper Reduction
(one capsule per meal and one between meals for a total of six per day)
Listed below is the protocol for treating cancer patients by a copper-reduction regiment based on the information above.
· 20 mg of TM should be taken with meals six times per day.
Serum ceruloplasmin levels must be monitored weekly from the beginning to establish baseline so that 20% baseline is determined.12 In addition, it is important to monitor iron levels and hemocrit.
This regiment should be taken for 60-90 days – EVERY day until copper ceruloplasmin levels are within 20% of baseline.9 To maintain the 20% level, a maintenance level dosage is utilized by taking 20 mg TM 3-4 times per day. About two weeks into treatment, antioxidant capsules consisting of 300mg N-acetylcysteine (NAC) and 300mg of ascorbic acid should be taken six times per day. Because copper has been shown to react with NAC and ascorbic acid to cause DNA damage,43 the antioxidant treatment should start after copper reduction begins.
7.3 TM Blood Test Schedule
You must find a lab that will give you a turn-around time of no more than 48 hours on ceruloplasmin.
Before starting the Copper Reduction/Antioxidant Protocol, establish baselines:
End of 4th week:
Ceruloplasmin and CBC
End of 6th week:
Ceruloplasmin and CBC
End of 8th week:
Once Ceruloplasmin is in the teens:
Ceruloplasmin and CBC every week
Copper and Zinc once per month
The zinc to copper ratio should be 3:1
Target ceruloplasmin is 20% of baseline reading
Selection of Subjects
Subject Inclusion Criteria
|Patients must meet all of the following inclusion criteria:|
|1.||Patient must have histologically cancer.|
|2.||Patient must be greater than or 18-years-old.|
|3.||3. Patients must have an Eastern Cooperative Oncology Group (ECOG) performance status of 0, 1, or 2.|
|4.||Measurable disease is required.|
|5.||All acute toxic effects of any prior radiotherapy or 5-FU given as a radiation sensitizer must have resolved to National Cancer Institute Common Toxicity Criteria (Version 2.0) grade < or equal to 1.|
|6.||Required initial laboratory data for patients include:CBC
Absolute neutrophil count
Serum glutamate-oxaloacetate transaminase
Serum pregnancy test for females of childbearing potential
Tumor markers depending on the tumor type
|7.||Patients must sign an informed consent that they are aware of the neoplastic nature of their disease and must willingly provide written consent after being informed of the procedures to be followed, the experimental nature of the therapy, alternatives, potential benefits, side-effects, risks, and discomforts. Patients must be willing and able to comply with scheduled visits, treatment plan, and laboratory tests.|
|The presence of any of the following will exclude a patient:
20 mg of TM9.1.2 DRUG ADMINISTRATION
Oral gel capsule9.1.3 STORAGE AND STABILITY
Se manufacturers recommendation. All TM, whether powder or capsule form, is stored under argon. Orders are prepared fresh from argon storage with a three month shelf life.9.1.4 SOURCE OF DRUG – Physician prescription
9.1.5 TOXICITY – Anemia
N-acetylcysteine (NAC)-Rx Mucomyst or generic equivalent
Mucomyst or generic equivalent (20% solution)9.2.2 DRUG ADMINISTRATION
Oral solution made by diluting 10 mL with 50 mL of cola9.2.3 STORAGE AND STABILITY
Store diluted solution in the refrigerator.
Discard diluted solution after 96 hrs.9.2.4 SOURCE OF DRUG
Ascorbic Acid Tablets – OTC
Vitamin C 500mg per tablet9.3.2 DRUG ADMINISTRATION
Oral tablet9.3.3 STORAGE AND STABILITY
See manufacturer’s recommendations9.3.4 SOURCE OF DRUG
Over the counter
Zinc Sulfate (or Zinc Acetate)
220 mg of sulfate or 50 mg as zinc9.4.2 DRUG ADMINISTRATION
Oral capsule9.4.3 STORAGE AND STABILITY
See manufacturer’s recommendations9.4.4 SOURCE OF DRUG
Over the counter
Clinical Efficacy Assessments: Objective Tumor Response
Methods of Assessment
|The principle evaluation of antitumor activity will be based on serial objective determination of changes in measurable lesion size. The newly developed World Heath Organization Response Evaluation Criteria in Solid Tumors (RECIST) will be employed.Radiographic evaluation is preferred to evaluation by clinical examination. Tumor assessments should preferably include abdominal CT scans with contrast. Abdominal MRI scans are acceptable. Conventional CT and MRI should be performed with cuts of 1.0cm or less in slice thickness contiguously. Spiral CT should be performed using a 0.5cm contiguous reconstruction algorithm. Lesions on chest x-ray are acceptable as measurable lesions when they are clearly defined and surrounded by aerated lung. However, CT is preferable for assessment of chest lesions. Chest CT scans or chest X-rays should be followed in patients with known pulmonary metastases at baseline or in whom pulmonary symptoms develop during treatment.Documentation by color photography including a ruler is necessary to estimate the size of skin lesions. Ultrasound is a possible alternative to clinical measurements for superficial palpable nodes, subcutaneous lesions, and thyroid nodules. Ultrasound may also be used to confirm the complete disappearance of superficial lesions usually assessed by clinical examination. Ultrasound should not be used to measure tumor lesions that are clinically not easily accessible. Endoscopy, laparoscopy, and radionuclide scan should not be used for response assessment.The same method of assessment and the same technique should be used to characterize each identified and reported lesion at baseline and during follow-up. To ensure compatibility, the baseline x-rays/scans to assess response must be performed using identical techniques. Copies of the scans must be available for review.
Malignant tumor masses (target lesions) should be selected on the basis of their size and their suitability for accurate repetitive measurements. All patients must have at least one target lesion that can be accurately measured in at least one dimension (longest diameter to be recorded) greater than or equal to 2.0cm with conventional radiographic techniques or MRI, or greater than or equal to 1.0cm with spiral CT scan. To assess objective response, it is necessary to estimate the overall tumor burden at baseline and use this as a comparator for subsequent measurements. Whenever possible, several measurable lesions up to a maximum of 10 target lesions representative of involved organs should be identified, measured and recorded at baseline. To allow later retrieval, lesions should be clearly marked on the films or otherwise identified. If the lesion is detected by more than one method at baseline, the investigator should select at baseline the method to be used at subsequent evaluations. All measurements should be performed using a caliper or ruler and should be recorded in metric notation in centimeters. A sum of the longest diameter for all target lesions will be calculated and reported as the baseline sum longest diameter. The baseline sum longest diameters will be used as reference to further characterize the objective tumor response of the measurable dimension of the disease by repeated assessment of the sum of the longest diameters during treatment.
All other lesions (or sites of disease) should be identified as non-target lesions and should also be recorded at baseline. Measurements are not required and these lesions should be followed as “present” or “absent”.
All baseline evaluations should be performed as closely as possible to the treatment start and never more than 14 days before the beginning of the treatment. After randomization, oncologic assessment will be repeated every 6 weeks, for patients in both treatment arms.
10.2 Definitions of Objective Tumor Response and Progression-Imaging
Measurable lesions are defined as malignant tumor masses that can be accurately measured in at least one dimension (longest diameter to be recorded) as greater than or at 2.0cm with conventional techniques or as greater than or equal to 1.0cm with spiral CT scan. Clinical lesions will only be considered measurable when they are superficial (e.g. skin nodules, palpable lymph nodes).
Nonmeasurable lesions are defined as malignant tumor masses that cannot be accurately quantified for tumor response, and would include bone lesions, leptomeningeal disease, ascites, pleural or pericardial effusions, lymphangitis of the skin or lung, abdominal masses that are not confirmed and followed by imaging techniques, cystic lesions, irradiated lesions, and disease documented by indirect evidence only (e.g. by laboratory tests such as alkaline phosphatase.)
10.2.1 Target Lesions
|Complete response (CR) is defined as the disappearance of all target lesions.Partial response (PR) is defined as a greater than or equal to 30% decrease in the sum of the longest dimensions of the target lesions taking as reference the baseline sum longest dimensions.Progressive disease (PD) id defined as a greater than or equal to 20% increase in the sum of the longest dimensions of the target lesions as reference the smallest sum of the longest dimensions recorded since the treatment started, or the appearance of one or more new lesions.Stable Disease (SD) is defined as neither sufficient shrinkage to qualify for PR nor sufficient increase to qualify for PD taking as references the smallest sum of the longest dimensions since the treatment started.|
10.2.2 Non-Target Lesions
|Complete response (CR) is defined as the disappearance of all nontarget lesions.Non-Complete Response ((Non-CR)/Non-Progressive disease (Non-PD) is defined as a persistence of one or more nontarget lesions.Progressive disease (PD) is defined as unequivocal progression of existing nontarget lesions, or the appearance of one or more new lesions.Patients will be defined as Not Evaluable for Response (NE) if there is no post-randomization oncologic assessment.
All patients who have a reduction in tumor lesions sufficient to meet criteria for CR or PR on at least one occasion will be considered to have objective evidence of response. To be considered a confirmed objective response, these same criteria for response must be confirmed with a follow-up scan obtained greater than or equal to 4 weeks from the scan documenting the response.
The reasons for discontinuation of treatment without objective evidence of disease progression must be reported on the Study Termination CRF. If a patient is removed from therapy for any reason other than radiographic evidence of tumor progression, every —– should be made to radiographically document disease progression even after discontinuation of treatment.
Evaluation of Overall Objective Tumor Response
|The overall objective response at each oncologic assessment is based on the criteria in Table 11. The best overall response is the best response noted while the patient is in the study.|
Methods of Assessment- Tumor Markers
|A secondary evaluation of antitumor activity will be the monitoring of tumor markers that are positive before treatment and are judged to be reliable indicators of tumor load. While the selection of tumor markers are extensive, every effort will be made to use reasonable judgment in determining potentially positive tumor markers.|
Clinical Safety Assessments
11.1 Definition of an Adverse Event
|An adverse event is any untoward medical occurrence in a patient or trial subject administered a drug or biologic (medicinal product) or using a medical device; the event does not necessarily have a causal relationship with that treatment or usage.Adverse events include the following:
11.2 Serious Adverse Events
|A Serious Adverse Event is an untoward medical occurrence, regardless of whether or not it is considered related to the study medication, which results in:
11.3 Unexpected Adverse Events
|Unexpected adverse events are defined as those events that were not previously reported with study drug as referenced in the protocol, investigator’s brochure, consent form, or package insert or which are symptomatically and pathophysiologically related to a known toxicity but differ because of greater severity or specificity.|
11.4 Adverse Event Reporting Period
|The adverse event reporting period for this trial begins upon receiving the first dose of study medication and ends 30 days after termination of study medication.All adverse events that occur in trial subjects during the adverse event reporting period specified in the protocol must be reported to P&U, whether or not the event is considered medication related.In addition, any known untoward event that occurs subsequent to the adverse event reporting period that the investigator assesses as possibly related to the study medications should also be reported as an adverse event.|
11.5 Eliciting Adverse Event Information
|The investigator is to report all directly observed adverse events and all adverse events spontaneously reported by the trial subject. In addition, each trial subject will be questioned about adverse events at each clinic visit following initiation of treatment. The question asked should be, “Since your last clinic visit have you had any health problems?”, or a similar type of open-ended query.|
11.6 Reporting Requirements
Reporting Requirements for Adverse Events
11.7 Recording Adverse Events in the CRF’s Procedures
|Data on all adverse events (serious and non-serious, expected and unexpected, related or not related) will be collected in the CRF’s. Minimum requirements of adverse event data to be recorded are type of event, start and stop dates, highest severity grade, seriousness, action taken, outcome and relatedness to either study medication or tumor.· Symptoms or Signs Associated with Chemotherapy, Radiotherapy or Concomitant MedicationsSymptoms or signs associated with administration of chemotherapy, radiotherapy, or concomitant should be reported as adverse events.· Symptoms of Neoplastic disease
Symptoms of pancreatic cancer are not to be considered adverse events in this trial, provided that they are present at baseline (before treatment with study medication is initiated) and that they do not worsen during the treatment. In cases where an increase in severity occurs, the event should be reported in the appropriate section of the CRF.
· Abnormalities in Physiological Testing or Physical Examination
Grading of Adverse Events
Note the distinction between the gravity and the intensity of an adverse event. Severe is a measure of intensity; thus, a severe reaction is not necessarily a serious reaction. For example, a headache may be severe in intensity, but would not be classified as serious unless it met one of the criteria for serious events listed above.
11.8 Exposure In Utero
|If any trial subject becomes or is found to be pregnant while receiving a study medication or within 3 months of discontinuing study medication, the investigator is to submit an adverse CRF that includes the anticipated date of birth or pregnancy termination. The subject is then to be followed by the investigator until completion of the pregnancy. If the pregnancy ends for any reason before the anticipated date provided, the investigator should notify the P&U monitor.If the outcome of the pregnancy meets the criteria for immediate classification as a serious medical event (i.e., spontaneous abortion, stillbirth, neonatal death, or congenital anomaly [including that in an aborted fetus]), the investigator should follow the procedures for reporting serious medical events; i.e., report the event to the principal monitor by telephone and follow up by submission of appropriate adverse event CRF’s.Note that “spontaneous abortion” includes miscarriage and missed abortion.All neonatal deaths that occur within one month of birth should be reported, without regard to causality, as serious adverse events. In addition, any infant death after one month that the investigator assesses as possibly related to the in utero exposure to the study medication should also be reported.
In the case of a live birth, the “normality” of the newborn can be assessed at birth (i.e., there is no required minimum follow-up of a presumably normal infant before the Exposure in Utero form can be completed).
The “normality” of an aborted fetus can be assessed by gross visual inspection unless there are pre-abortion laboratory findings suggestive of a congenital anomaly.
|Survival is defined as the time from date of randomization to date of death. In the absence of confirmation of death, survival time will be censored at the last date of follow-up.|
Objective Tumor Response Rate
|Objective response rate is defined as the proportion of patients who have any evidence of objective CR or PR.Confirmed objective response rate is defined as the proportion of patients who have evidence of objective CR or PR that is confirmed by follow-up tumor assessment greater than or equal to 4 weeks from the tumor assessment documenting the initial response.|
Time to Objective Tumor Response
|Time to response is defined as the time from date of randomization to first objective documentation of response.|
Duration of Objective Tumor Response
|Duration of response is defined as the date of first objective documentation of response until the first objective documentation of tumor progression or death due to tumor progression in the absence of previous documentation of tumor progression. For patients with responding tumors who do not have objective evidence of tumor progression and who are removed from study, who die of causes not related to pancreatic cancer, or who are given antitumor treatment other than the study treatment, duration of response will be censored. Patients who die of unknown causes will be considered to have experienced tumor progression.|
Time to Objective Tumor Progression
|Time to objective tumor progression is defined as the time from date of randomization to first objective documentation of tumor progression or death due to tumor progression in the absence of previous documentation of tumor progression. For patients who do not have objective evidence of tumor progression and who are removed from study, who die of causes not related to pancreatic cancer, or who are given antitumor treatment other than the study treatment, time to tumor progression will be censored. Patients who die of unknown causes will be considered to have experienced tumor progression. A CA19-9 increase meeting criteria for tumor marker progression does not constitute adequate objective evidence of tumor progression. However, such a CA 19-9 increase should prompt a repeat radiographic evaluation to document whether or not objective tumor progression has occurred.|
Time to Treatment Failure
|Time to treatment failure is defined as the time from date of randomization to first objective documentation of tumor progression, or off-treatment date, or death, whichever comes first. Patients who are still on treatment at the time of the analysis and patients who are removed from therapy by their physicians during an objective response and who, at the off-treatment date, have no evidence of objective tumor progression will not be considered to have experienced treatment failure, unless the withdrawal is due to the occurrence of a medical event. For these patients, time to treatment failure will be censored at the off-study date. Censoring for time to treatment failure will also be performed in those patients who are given antitumor treatment other than the study treatment before the first objective tumor progression, off-study date, or death. A CA19-9 increase meeting criteria for tumor marker progression does not constitute adequate objective evidence of treatment failure. However, such a CA19-9 increase should prompt a repeat radiographic evaluation to document whether or not objective tumor progression (and thus treatment failure) has occurred.|
|Time to first definitive performance status worsening is the time from randomization until the last time the performance status was no worse than at baseline.Time to first definitive weight loss is defined as the time from randomization until the last time the percent weight decrease from the baseline was less than 5%.Time to first albumin decline is defined in terms of the time from randomization until decline in albumin by 10%, by 20%, and by 40% below baseline (based on central laboratory data).|
|All treatments are given orally at 20 mg gel capsule dosage for TM and 300m mg each ascorbic acid/N-acetylcysteine for antioxidant treatment. Initial dosages are six TM capsules and three antioxidant capsules per day, respectively. These are given regardless of weight or age. The cerauloplasm levels will determine if the TM dosage is changed over time as judged by the physician.|
|The as-treated population consists of all patients who actually receive study drug (at least one injection), with treatment assignments designated according to actual study treatment received.The primary endpoint of survival will be assessed in the as-randomized and the as-treated patient populations.No placebo groups will be in this study.|
QUALITY CONTROL AND QUALITY ASSURANCE
|An investigators meeting will be held prior to the initiation of the study at which the investigators and site coordinators will be informed regarding details of the protocol, medication supplies, and monitoring responsibilities. The CRF’s will be explained in detail.Each clinical site conducting this study has agreed to conduct this study according to GCP standards and in conformity with local and national requirements (e.g., FDA or ICH standards). Monitoring visits to the trial site will be made periodically during the trial to ensure that all aspects of the protocol are followed. Source documents will be reviewed for verification of agreement with data on the CRF’s. Original patient records (e.g., hospital charts, clinic records, laboratory printouts) should be available at each study site for source document review by a P&U monitor. Source document review is the cross-checking of information recorded on study CRF’s with that recorded in the original patient records. It is recognized that for observations and evaluations conducted solely for the purposes of the study, the CRF may be the original patient record. It is not the purpose of source document review to ensure that all information on CRF’s is also recorded elsewhere in the patient’s records; the purpose is to help ensure that the CRF’s accurately reflect information generated during the study.The investigator/institution must guarantee access to source documents by appropriate regulatory agencies. The trial site may also be subject to quality assurance audits by P&U as well as inspection by appropriate regulatory agencies. It is important that the investigators and their relevant personnel are available during the monitoring visits and possible audits and that sufficient time is devoted to the process.|
Ethical Conduct of the Trial
|The trial will be performed in accordance with the recommendations guiding physicians in biomedical research involving human subjects adopted by the 18th World Medical Assembly, Helsinki, Finland, 1964 and later revisions.|
Institutional Review Board/Independent Ethics Committee
|It is the responsibility of the investigator to obtain approval of the trial protocol/amendments from the IRB/IEC. All correspondence with the IRB/IEC should be filed by the investigator.The study will not be started until approval of the protocol, the patient information and the informed consent form has been obtained from the appropriate IRB/IEC. It is the responsibility of the investigator to forward a copy of the written approval and a list of the members, their titles or occupation, and their institutional affiliations, to P&U. The approval should include study identification and the date of review.Protocol amendments are to be submitted to the IRB/IEC (and other local authorities, according to local regulations) prior to implementation.The investigator will make required progress reports to the IRB/IEC, as well as report any serious adverse events, life-threatening problems or deaths. The investigator will also inform the IRB/IEC of reports of serious adverse events (provided to him/her by the sponsor) occurring in other clinical studies conducted with the study drug. The IRB/IEC must be informed by the investigator of the termination of the study.
The investigator should file all correspondence with the IRB/IEC.
The study will not be started until written approval receipt acknowledgment or elapse of the statutory waiting period from the approved IRB committee.
Patient Informed Consent
|It is the responsibility of the investigator to give each patient (or the patient’s acceptable representative) prior to inclusion in the trial, full and adequate verbal and written information regarding the objective and procedures of the trial and the possible risks involved. The patients must be informed about their right to withdraw from the trial at any time. Refusal to participate will involve no penalty or loss of benefits to which the subject is otherwise entitled, and will not prejudice further medical treatment. Written patent information as contained in the Sample Informed Consent (Appendix D) must be given to each patient before enrollment. Furthermore, it is the responsibility of the investigator to obtain signed informed consent from all patients prior to inclusion in the trial.The responsible physician will inform the patient about the background and present knowledge of the drug under study and will give the patient pertinent information as to the intended purpose, possible benefits, and possible adverse experiences. The procedures and possible hazards to which the patient will be exposed will be explained.An explanation of whom to contact for answers to pertinent questions about the research and research subject’s rights, and who to contact in case of research-related injury will be given.Written patient information, when indicated, must be given to each patient before enrollment.
If during the study the patient refuses to continue taking part in the study, the investigator will respect this and the patient will receive optimal and appropriate care.
The signed informed consent forms should be filed by the investigator. Where confidentiality laws preclude this, they should not be made available, but filed for possible future audits. The investigator will confirm the receipt of informed consent from each patient by completing and signing the appropriate CRF.
The patient information and the informed consent form to be used must be approved by the same IRB/IEC approving the conduct of this study.
The patient should also be informed that the data will be stored and analyzed by computer in a national database at cancerprotocol.com, that country-specific regulations for the handling of computerized data will be followed and described in the patient information and that identification of individual patient data will only be possible for the investigator. Furthermore, the patients should be informed about the possibility of inspection of relevant parts of the hospital records by health authorities.
US-Specific IRB/IEC Requirements
|This protocol and the consent form that will be used must be approved by an IRB/IEC before the study is initiated. The IRB/IEC must comply with current FDA regulations. Other investigator responsibilities relative to the IRB/IEC include the following:
The language in the informed consent must be understandable and in no way coercive.
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PATIENT INFORMATION AND INFORMED CONSENT FORM
|I, _________________________, do hereby consent to participate in a research study for cancer. This study has been explained to me and I fully understand the following:
To determine if anti angiogenesis treatment with copper reduction therapy and antioxidant therapy will reduce tumor load.
Tetrathiomolybdate (TM) will bind copper and remove it from the body. Copper reduction is a broad- band method to reverse angiogenesis. Ascorbic acid- N-acelylcysteine are potent antioxidates that work synergistically. These have an effective anti-tumor activity.
Start 20mg TM at six times/day, once with each meal and three times between meals. Two weeks into the TM treatment, the antioxidant treatments begins with one capsule every six hours.
D. DURATION OF THERAPY
Therapy will continue until cerauloplasin is 20% of baseline value or 8.0 mg/dL. TM dosage may be capsules to maintain the 20% baseline level for 90 more days to allow the blood vessels feeding the tumor to die. If my disease is stable or shrinking and there is no other reason to stop the drug therapy. Treatment will continue and be evaluated every 60-90 days by the physician.
I understand the antioxidant treatment will continue until my physician judges it should stop.
When my participation in the study is to be terminated, I will complete one last questionnaire, have blood drawn, and undergo assessment of my tumor response to therapy.
If conditions occur which would make my participation detrimental to my health, my doctor may discontinue this treatment even though I might wish to proceed with further therapy. In addition, the entire study may be terminated, at the discretion of the sponsor, if unacceptable risks or side effects develop. In the event of either of these occurrences, my doctor will review my individual treatment options.
My doctor(s) would like to be able to track my cancer after the treatment phase of the study and would also like to keep me informed of any new information regarding the treatment I received while on this clinical study. Therefore, during the first year I will be contacted monthly and during the second year every 3 months by the study nurse and asked about my cancer.
E. POSSIBLE DISCOMFORTS AND/OR RISKS:
Cancer treatments often have side effects. The treatments used in this study may cause all, some or none of the side effects listed in this document. In addition, there is always the risk of very uncommon or previously unknown side effects occurring. Although very unusual, it is even possible that side effects from chemotherapy may result in death. My doctor will be checking me closely to see if any of these side effects are occurring. Routine tests will be done to monitor the effects of treatment. Side effects usually disappear after the treatment is stopped. In the meantime, my doctor may prescribe medication to keep these side effects under control. I understand, I may need to be hospitalized in order for my doctor to treat the side effects.
The effects of chemotherapy on the human fetus are known to be harmful from animal studies. If I am pregnant or nursing, I will not be allowed to participate in this study. If I agree to participate in this study, and am of childbearing potential, I must not become pregnant. If applicable, the initial blood tests may include a pregnancy test. I can avoid pregnancy by using an appropriate method of contraception (e.g., an IUD, partner having had a vasectomy, oral contraceptives, abstinence, or being surgically sterile). If I should become pregnant while participating in this study, I must inform my doctor immediately and my treatment options will be discussed with me. If I am male, I must be either sterile or use an approved contraceptive method (e.g. condom).
· Possible Side Effects: Anemia
Under no circumstances should the cerauloplasmin drop below 8.0 mcg/dL. Ceruloplasmin increases. Lower dosages of TM will be given as judged by the physician
· Prevention/Treatment of Side Effects:
F. POSSIBLE BENEFITS
Suspension of TM treatment until minimal values of ceruloplasmin in are determined.
It is not possible to predict whether or not any personal benefit will result from the use of the treatment program. Possible benefits are reduction in my disease and prolongation of my life.
G. ALTERNATIVE METHODS
Alternative treatments that could be considered in my case include chemotherapy, radiation therapy, or other experimental therapy. I may also choose to have no further therapy and receive supportive care only. This may result in continued growth of my disease. I understand that my doctor will want to discuss these alternatives with me.
H. NEW INFORMATION FOR THE PATIENT
I understand that any new information that is developed during the course of this study that may influence my willingness to continue participation in this study will be made available to me.
I. PATIENT CONTACTS
Dr. ___________________; phone _________________ has agreed to answer any inquires that I may have concerning the procedures and has informed me that I may also discuss with him/her this concerning this research study and my right as a research subject.
J. PATIENT CONFIDENTIALITY
I understand that my clinical trial information will be recorded on forms and sent to the clinical trial sponsor. The data on the forms will be entered into an electronic database for storage and statistical analysis. By signing the informed consent form I authorize this access to my records. All records in which my name appears will be kept confidential. My identity will not be revealed on any forms, or in any report or publication.