Conditions: Ovarian Cancer
Keywords: epithelial ovarian fallopian tube primary peritoneal carcinoma peritoneal ovary
Allocation: RANDOMIZED | Intervention model: PARALLEL
Enrollment: 3
Healthy volunteers: no

Brief summary:
The main purpose of this Pilot Study is to test the safety, tolerability and quality of life in women who take Fermented Wheat Germ Extract (FWGE), to determine if an active form of FWGE can be detected in the blood, and determine whether short-term therapy with FWGE has any effect on the tumor marker, cancer antigen 125 (CA-125).

Detailed description:

Arms:
arm 1: FWGE administration 2 - 4 weeks prior to planned surgery, + Quality of Life (QoL) Surveys: FACT-O. arm 2: Placebo administration 2 - 4 weeks prior to planned surgery, + Quality of Life (QoL) Surveys: FACT-O.

Interventions: intervention 1: Fermented Wheat Germ Extract (FWGE) intervention 2: Placebo intervention 3: Standard of Care: Planned Surgery intervention 4: Quality of Life (QoL) Surveys: FACT-O
Intervention descriptions: intervention 1: 5.53 grams of FWGE combined with natural orange flavor and stevia Reb-A sweetener to create an instant drink mix, dissolved in 150 mL of water, orally once-daily, for 2 to 4 weeks. intervention 2: Dried and pulverized, white and black rice, 5.53 grams, combined with natural orange flavor and stevia Reb-A sweetener, to create an instant drink mix, dissolved in 150 mL of water, orally once-daily, for 2 to 4 weeks. intervention 3: Participants will undergo their planned surgery during the 2nd to 4th week after entry into the study. However, if there are delays, patients can continue to take drug/placebo for a time period no greater than 2 months. If surgery has not occurred after this 2-month period, the patient will be removed from the study. Participants will take the study drug up until the day prior to surgery. The minimum amount of time on study drug is 2 weeks. A separate consent form will be obtained for the surgical procedure. intervention 4: Quality of life questionnaire (FACT-O) at the time of enrollment, then weekly and the day of surgery.

Protocol document:
Prot_SAP_000.pdf: 
 
 
 
 A PILOT RANDOMIZED, PLACEBO CONTROLLED, TRIAL OF FERMENTED WHEAT 
GERM EXTRACT IN WOMEN WITH OVARIAN CANCER  
NCT02411565 
Version Date: 
June 9, 2017 




 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 2 
Table of Contents 
 
                 
Page 
1.  
PROTOCOL SCHEMA 
3 
2.  
HYPOTHESIS and OBJECTIVES 
4 
3.  
BACKGROUND AND RATIONALE 
5 
4.  
PATIENT ELIGIBILITY AND EXCLUSIONS 
11 
5.  
STUDY MODALITIES 
12 
6.  
TREATMENT PLAN AND ENTRY PROCEDURE   
14 
7.  
TREATMENT MODIFICATIONS 
19 
8.  
STUDY PARAMETERS & SERIAL OBSERVATIONS  
19 
9.  
EVALUATION CRITERIA 
20 
10.  DURATION OF STUDY 
21 
11.  STUDY MONITORING & REPORTING PROCEDURES  21    
12.  STATISTICAL CONSIDERATIONS 
22 
13.  BIBLIOGRAPHY 
25 
 
Appendix I  FACT-O 
27 
Appendix II  Letter of Support 
31 
Appendix III  Compositional Analysis FWGE 
32 
Appendix IV   Compositional Analysis Placebo 
33 
 
 




 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 
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2.  HYPOTHESIS and OBJECTIVES 
 
  
2.1 Hypothesis 
Fermented Wheat Germ Extract (FWGE) has activity against epithelial ovarian cancer 
(OVCA).  FWGE may increase sensitivity to chemotherapy and may decrease side 
effects related to chemotherapy.  FWGE active components (2,6-dimethoxy-p-
benzoquinone) may be detected in serum of women receiving FWGE. 
 
In an effort to generate preliminary data to support a future peer-reviewed grant-funded 
large phase II clinical trial of adjuvant FWGE in women undergoing therapy for ovarian 
cancer and to test the safety and tolerability of FWGE, we propose a pilot randomized, 
placebo-controlled study of FWGE in women with a suspected diagnosis of OVCA. We 
propose to test the safety and tolerability of FWGE, and to determine if we can detect an 
active product of FWGE (2,6-dimethoxy-p-benzoquinone) in the serum, and whether 
short-term therapy with FWGE has an effect on the tumor marker, CA125.  We will 
analyze changes in tissue proliferative assays and gene expression in women who 
receive FWGE (n=10) and those who receive placebo (n=10).   
 
We recognize that the sample size in the current study will prohibit definitive statistical 
conclusions to be drawn on our primary objective (to test the safety and tolerability of 
FWGE), or our secondary objectives (to compare levels of FWGE active product, CA125 
levels, proliferation, and gene expression in OVCAs from subjects receiving FWGE 
versus placebo).  The study is neither designed nor powered to develop statistically 
significant results, rather, we seek to develop preliminary/pilot data that will enable us to 
pursue a future large phase II, peer-reviewed grant-funded trial of adjuvant FWGE in 
women undergoing therapy for ovarian cancer.  Preliminary data developed in the 
current proposal will support such grant applications. 
 
 
2.2 Objectives 
 
2.21 Primary Pilot Objective: To assess the safety, toxicity and quality of life (QoL) of 
women with suspected ovarian cancer scheduled to undergo surgery, administered 
FWGE versus placebo daily for up to 4 weeks. 


 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 
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2.22 Seconday Pilot Objectives:  
2.221 To develop pilot data on the impact of FWGE on: the tumor marker CA125, 
tumor proliferation. 
2.222  To develop pilot data on changes in gene expression in ovarian cancer 
exposed to FWGE versus placebo. 
 
 
3.0  BACKGROUND AND RATIONALE 
 
3.1  Ovarian Cancer  
Ovarian cancer (OVCA) is the 5th leading cause of cancer death among women in the 
United States and Europe and has the highest mortality of all gynecologic cancers.  It is 
estimated that 21,980 women in the U.S. will be diagnosed with epithelial OVCA in 2014 
and 14,270 women will die of the disease [1].  Approximately 75% of cases are 
diagnosed at an advanced-stage (III/IV) with disseminated intraperitoneal metastasis [2]. 
Women with OVCA undergo major surgery followed by chemotherapy in an attempt to 
prolong survival.  Although approximately 70% of women will demonstrate a clinical 
response to this primary therapy, the majority will develop progressive or recurrent 
disease that is platinum-resistant.  Women with platinum-resistant OVCA often 
demonstrate cross-resistance to most other chemotherapeutic agents.  Once platinum-
resistance has developed, few active therapeutic options exist and patient survival is 
generally short-lived.  Throughout treatment for OVCA, prolongation of survival and the 
successful maintenance of quality of life remain important goals.  Improving our ability to 
manage the disease by optimizing the use of existing drugs and/or developing new 
agents is therefore essential in this endeavor.  
 
3.2  Use of natural products  
Up to 90% of patients with cancer use alternative therapies, often herbal or natural 
products [3,4].  The majority of these products have not been subjected to 
comprehensive study for efficacy or potential negative interactions with chemotherapy.  
Natural, non-toxic regimens that enhance standard-of-care therapy and/or prolong 
survival are highly desirable in management of patients with cancer.   
 


 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 
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3.3  Fermented Wheat Germ Extract 
Fermented wheat germ extract (FWGE, Avemar®) was developed in Hungary in the early 
1990’s [5].  It is produced by extraction of wheat germ, fermentation of the extract, 
separation of the fermentation liquid, concentration, and drying.  The chemical 
composition of FWGE is a mixture of molecules including 2-methoxy-p-benzoquinone 
and 2,6-dimethoxy-p-benzoquinone, which are felt to contribute to its biological 
properties [6].  FWGE has been available as a dietary supplement in Hungary since 
1998 and was approved in 2002 as a “medical nutriment for cancer patients.”  It is 
classified by the European Union as a “dietary food for special medical purposes”.  
FWGE has been shown to be safe when taken orally with no toxicity, mutagenicity, or 
genotoxicity [6]. 
 
3.31  
In Vitro Studies 
FWGE has been evaluated in-vitro and shown to induce apoptosis in some cancer 
cell types, including: leukemia, breast, colon, testicular, head and neck, cervical, 
ovarian, gastric, thyroid, melanoma, brain and hepatocellular carcinomas [7-10].  The 
addition of FWGE to 5-fluorouracil, oxaliplatin or irinotecan chemotherapy has been 
shown to have no negative impact on effectiveness or toxicity profiles, and in colon 
cancer cell lines has additive to synergistic effects [7].  
 
With this data suggesting that FWGE has activity against many cancer cell types, we 
studied the broader spectrum of activity against OVCA.   
 
3.32  
Our Ovarian Cancer In Vitro Studies [11] 
Our in-vitro data demonstrates that FWGE induces OVCA cell line growth arrest and 
apoptosis and that it sensitizes OVCA cells to cisplatin-induced cell death. We also 
have data to suggest that the sensitivity of OVCA cells to FWGE may be determined 
by a discrete number of molecular signaling pathways.   
 
FWGE induces growth arrest and cell death in ovarian cancer cell lines  
We evaluated the effects of FWGE on cell growth arrest in a panel of 12 OVCA cell 
lines using MTS cell viability assays (Fig 1). Several of these cell lines were paired, 
mother/daughter, sensitive/resistant lines. Notably, cisplatin-resistance, a surrogate 
marker of greater chemotherapy-resistance, did not predict FWGE sensitivity. This 




 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 
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ZNF254  ZNF708  COPG2  SHF  
tcag7.1228 SLC25A13 
ZCCHC17 
DYNC2LI1 
GPATCH2 FAM133B 
ARL5B  FAM117B NRF1  PHF21A POLR2J  SOX12  DBF4  FMC1  KLHL7  DUXAP10 
LSM8  PHF21A  SLC7A7 SMAD2 SMARCD1 SUZ12P  ZBTB44  ZFP106  TRIM24  RPE 
ZNF322A ZSCAN21 ACVR2B APOC1 C10orf111 C20orf74 MIDN  PLXND1 AGFG1  CCDC5 
Table 1. Genes correlated with FWGE sensitivity. 
 
context, an analysis of biologic pathway relationships was performed using 
commercially available software (GeneGo Meta-Core™, (genego.com). GeneGo 
identified 16 pathways represented by genes associated with FWGE sensitivity 
p<0.05. As described above, one of the active components of FWGE is thought to be 
2,6-dimethoxy-p-benzoquinone. Analysis of publically available Affymetrix expression 
data (http://discover.nci.nih.gov/cellminer/loadDownload.do) and NCI60 cell line GI50  
 
 
 
 
 
 
 
values for 2,6-dimethoxy-p-benzoquinone 
(http://dtp.nci.nih.gov/dtpstandard/cancerscreening data/index.jsp), identified 3839 
genes representing 267 pathways associated with sensitivity to 2,6-dimethoxy-p-
benzoquinone (data not shown). Comparison of signaling pathways correlated with 
sensitivity to FWGE in OVCA cells and 2,6-dimethoxy-p-benzoquinone in NCI60 cell 
lines identified 9 common biological pathways 
 
3.33   
In Vivo Studies 
FWGE has been shown in vivo to decrease metastatic spread of cancer, decrease 
carcinogenesis and to be non-toxic and non-inhibitory when administered with 
chemotherapy.  
 
Synchronous oral FWGE and 5-fluorouracil intraperitoneal delivery significantly 
reduced the metastatic spread of c38 colorectal cancer in mice. [12]  
 
In a study on F-344 rats, the administration of azoxymethane induced colon 
carcinogenesis. When FWGE was fed to the rats prior to the azoxymethane 
administration, significantly fewer rats developed colon tumors (83% vs 45% p< 
0.001) [13] 
 
In vivo, FWGE was administered in mice simultaneously with cyclophosphamide, 


 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 
9 
vinorelbine, or doxorubicin. There was no increased toxicity or decreased 
antiproliferative activity [14]. 
 
3.34  
Human Clinical Studies 
When taken orally in addition to chemotherapy, FWGE has been shown to decrease 
adverse events, inhibit cancer cell proliferation and metastatic spread, and increase 
overall survival in patients with colorectal cancer and melanoma. 
 
One hundred and seventy-six colorectal cancer patients with histologically confirmed 
(Dukes A–D) colorectal cancer were enrolled. No restrictions were made for prior 
radiotherapy and/or chemotherapy, but all the patients had to undergo curative 
surgery at the time of diagnosis. There was no randomization; cohorts were formed 
based on patient preference. FWGE was taken orally once daily. Patients received 
adjuvant therapy by physician preference.  No grade 3 or 4 adverse events were 
observed. Patients receiving FWGE had a significant prolongation of progression 
free survival (p = 0.0184) and overall survival (p = 0.0278) [15] 
A trial performed in pediatric patients with solid malignancies undergoing 
chemotherapy was conducted to assess whether the combined administration of 
FWGE with chemotherapy would reduce the incidence of treatment-related febrile 
neutropenia.  Twenty-two patients were enrolled; they were not randomized, but 
allowed to choose their treatment arm.  The patients were matched by diagnosis, 
stage of disease, age, and gender.  During the treatment period, there was no 
progression of the malignant disease.  The frequency of febrile neutropenia was 
significantly decreased in patients receiving FWGE: 30 febrile neutropenic episodes 
(24.8%) in the FWGE group versus 46 (43.4%) in the control group (p < 0.05) [16] 
 
An open-label, randomized, phase II trial to assess the supportive value of FWGE in 
patients with stage III melanoma scheduled to receive adjuvant therapy was 
performed. Post-operatively, patients were randomized to FWGE simultaneous with 
adjuvant dicarbazine (DTIC) and FWGE continued for 12 months or DTIC alone. 
Fifty-eight patients were enrolled. Three from each treatment arm withdrew from 
study, for 52 patients analyzed. Adverse events included: grade1 and 2 
nausea/vomiting, and grade 1 diarrhea. There were no grade 3 or 4 adverse events.   
They noted fewer toxic side effects in patients receiving the FWGE than in those of 


 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 
10 
the control group. There was a statistically significant improvement in progression-
free survival 55.8 months (FWGE) versus 29.9 months (control) p = 0.0137. There 
was a statistically significant improvement in overall survival of 66.2 months (FWGE) 
versus 44.7 months (control) p = 0.0298 [17]. 
 
3.35 Summary and Rationale:  
Fermented wheat germ extract has been demonstrated in vitro to induce ovarian 
cancer cell growth arrest and apoptosis.  When FWGE is combined with cisplatin 
chemotherapy it enhances cell killing.  In animal studies, FWGE has shown no 
toxicity and anticancer effects.  Observations from human clinical trials demonstrate 
FWGE to be safe and without serious adverse events.  The data also suggest 
beneficial effects of FWGE on disease progression and survival in patients with 
melanoma and colorectal cancer.   
 
3.4 Proposal Overview: In this pilot clinical trial, we will evaluate the safety, toxicity and 
quality of life (QoL) of ten women with ovarian cancer scheduled to undergo surgery, 
administered FWGE versus ten women receiving placebo daily for up to 4 weeks.  We 
hypothesize that FWGE will be safe and non-toxic and that administration may decrease 
some of the symptomatology associated with ovarian cancer.  As secondary objectives, 
we will seek to determine whether we can detect differences in levels of 2,6-dimethoxy-
p-benzoquinone (one of FWGE’s active products), in the serum of women receiving 
FWGE versus placebo.  We will also evaluate the impact of FWGE on the tumor serum 
marker CA125, as well as histologic tumor proliferation and gene expression in ovarian 
cancer.  As noted above, we recognize that the sample size in the current study will 
prohibit definitive statistical conclusions to be drawn on our primary objective (to test the 
safety and tolerability of FWGE), or our secondary objectives (to compare levels of 
FWGE active product, CA125 levels, proliferation, and gene expression in OVCAs from 
subjects receiving FWGE versus placebo).  The study is neither designed nor powered 
to develop statistically significant results, rather, we seek to develop preliminary/pilot 
data that will enable us to pursue a future large phase II, peer-reviewed grant-funded 
trial of adjuvant FWGE in women undergoing therapy for ovarian cancer.  Preliminary 
data developed in the current proposal will support such grant applications. 
 
3.5 Rationale for the Dosage Used in this Study 


 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 
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The dose of FWGE that has been chosen for the study, is the dose used in each of the 
three previous clinical trials.  This consists of 5.5 grams of FWGE formulated as a water-
soluble granulate with added 0.03 grams of stevia sweetener and orange flavorings 
(taking it to approximately 5.53 grams), dissolved in 150 mL of water, orally once-daily. 
 
This dose has been tolerated well in previous clinical trials with no grade 3 or 4 toxicity, 
even when combined with chemotherapy. 
 
With regard to length of the intervention, most patients are scheduled for surgery within  
4 week interval.  We do not want to delay a patient’s surgery for the intervention.   
 
 
4.  
PATIENT ELIGIBILITY 
 
  
4.1 Inclusion Criteria   
 
4.11  Women with suspected epithelial ovarian, fallopian tube or primary peritoneal 
carcinoma scheduled to undergo surgical exploration with no prior treatment for the 
cancer.  Signs of ovarian cancer include, but are not limited to: an elevated CA125, a 
complex pelvic mass, ascites, and carcinomatosis.   These signs are not necessary 
for suspicion or enrollment in this protocol.   
 
4.12 Age > 18 years and competent to give informed consent 
 
4.13 Patient must have a ECOG Performance sttus of 0, 1, or 2 and a life-
expectancy of at least 60 days. 
 
4.14. Patients must have adequate:  
       4.141 Bone marrow function: Hemoglobin greater than or equal to 9gm/dl. 
Absolute neutrophil count (ANC) greater than or equal  to 1,500/ul, 
equivalent to Common Toxicity Criteria (CTCAE v4.0) grade 1.  Platelets 
greater than or equal to 100,000/ul. 
 
      4.142 Renal function:  creatinine less than or equal to 1.5 x institutional upper 
limit normal (ULN), CTCAE v4.0 grade 1. 


 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 
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      4.143 Hepatic function: Bilirubin less than or equal to 1.5 x ULN (CTCAE v4.0 
grade 1).  SGOT and alkaline phosphatase less than or equal to 2.5 x ULN 
(CTCAE v4.0 grade 1). 
 
4.15 Study subjects must sign an approved informed consent and authorization 
permitting release of personal health information. 
 
4.16 Women of childbearing potential  must have  a negative serum pregnancy test 
prior to the study entry and be practicing an effective method of birth control 
during the course of the study, in a manner such that risk of failure is 
minimized.  Prior to study enrollment, women of childbearing potential must be 
advised of the importance of avoiding pregnancy during trial participation and 
the potential risk factors for an intentional pregnancy. 
 
 
  
4.2  Exclusion Criteria 
 
4.21  Current use of FWGE  
 
4.22  Known allergy to wheat, rice (contained in the placebo), orange or the 
sweetener, Stevis 
 
4.23 Patient who received neoadjuvant chemotherapy for ovarian cancer 
 
4.24 An upper gastrointestinal or other condition that would impair swallowing or 
absorption of oral medication 
 
4.25 Any serious illness or medical condition that would not permit the patient to be 
managed according to the protocol, including, but not to limited, any the 
following:  
•  History of significant neurologic or psychiatric disorder (e.g., 
uncontrolled psychiatric disorders) that would impair the ability to 
obtain consent or limit compliance with study requirement 


 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 
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•  Active uncontrolled or serious infection 
•  Active peptic ulcer disease 
                  
                   4.26 Uncontrolled hypertension defined as systolic greater than 180 and diastolic 
greater than 100 
 
 
 
5.  
STUDY MODALITIES 
 
  
5.1   FWGE 
 
5.11  Formulation: American BioSciences Inc, who is the exclusive 
manufacturer of FWGE in the Unites States, produces fermented wheat 
germ extract.  See Letter of Support (Appendix II).  It is formulated as a 
granulate. One package contains 5.53 grams of FWGE combined with 
natural orange flavor and stevia Reb-A sweetener to create an instant 
drink mix.  See compositional analysis (Appendix III). 
.  
5.12  Administration: FWGE is dissolved in 150ml water and taken orally daily.  
 
   
  
5.13  Adverse effects  
The clinical safety experience of FWGE is derived from trials in which 
FWGE has been used as a single agent or in combination with 
chemotherapeutic agents. FWGE has not been associated with any grade 
3 or 4 toxicities  
 
5.14  Storage: FWGE is to be maintained at <80 degrees F 
 
5.15  Quality assurance of the manufacturing of FWGE in the U.S. 
 
5.151  The purchased wheat germ and yeast, which are the raw materials 
of FWGE production, are certified for human consumption. The origin of 
the wheat germ and yeast, used in the fermentation, are all U.S. certified. 
 


 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 
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5.152   After each batch of FWGE has been freeze-dried, it is tested for 
hazardous contaminates, such as toxic elements, including heavy metals, 
and for toxic microorganisms, such as E.Coli, Listeria, etc. Once, the 
lyophilizate has been found free from toxic contaminants, and has been 
qualified as microbiologically safe (such as complete lack of toxic 
micriorganisims, etc.), it Is standardized to ensure batch-to-batch 
chemical compositional homogeneity and biological efficacy-related 
reproducibility. 
 
5.153   For this purpose, a characteristic high performance liquid 
chromatography (HPLC) fingerprint UV chromatogram of FWGE is 
compared with the HPLC fingerprint UV chromatogram of an established 
standard marker “core” of FWGE (reference pattern). As a result of a 2-
years research collaboration between the manufacturer  and an NCI-
designated Cancer Center in the U.S., the marker “core” of the FWGE 
has been developed and validated to possess the characteristic biological 
effects of FWGE, including testing by in vitro antiproliferative assays 
using human cancer cell lines; kinase panel tests; robust gene expression 
assays by quantitative proteomic analysis, using isobaric tag for relative 
and absolute quantitation labeling technique with electrospray mass 
spectrometry; and by laboratory animal cancer experiments. The said 
reference pattern of the standard marker “core” of FWGE has been 
established to comprise peaks characterized by retention times and 
relative retention quotients, respectively. 
 
The said fingerprint of the representative sample of the FWGE batch must 
be identical with the reference pattern. (Chemical 
standardization/Qualitative chemical assay.) 
 
5.154  The quality of the marker “core” in each FWGE batches must also 
meet a fixed reference value. (Chemical standardization/Quantitiative 
chemical assay.) 
 
5.155  FWGE batches are also characterized by quantitative biological 
assays according to in vitro anticancer protocols. Each and all batches 
must show the required quantitative biological efficacy. (biological 
standardization.) 
 
  
5.2 Placebo 
5.21 Formulations: American BioSciences Inc, who is the exclusive manufacturer 
of FWGE in the Unites States, produces  the placebo.  See Letter of Support 
(Appendix II).  It is formulated as a granulate. One package contains cooked, 


 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 
15 
dried and pulverized, white and black rice, 5.53 grams, combined with natural 
orange flavor and stevia Reb-A sweetener, to create an instant drink mix.  See 
compositional analysis placebo (Appendix IV).  
 
5.22 Administration: FWGE Placebo is a dissolved in 150ml water and taken 
orally daily.  
 
   
  
5.23 Supplier:  American BioSciences Inc 
 
   
  
5.24 Storage:  to be maintained at <80 degrees F 
 
5.3 Drug Accountability: The investigator, or a responsible party designated by the 
investigator, will maintain a careful record of the receipt, disposition, and return of all 
drugs received from American Biosciences Inc.  
 
5.4 Drug Dispensing: FWGE and placebo will be in unidentified packages.  A random 
number table will be used to assign patients to the FWGE versus placebo group.  The 
key will be held by the investigational pharmacy and is not revealed to the researchers 
or patients until the study is complete. 
   
 
5.4 Unblinding: This is a double-blind study. The treating physician and/or patient may 
be informed as to which regimen the patient was assigned if either of two situations 
arises: serious adverse reaction occurs or study completion. In the case of an adverse 
reaction, the treating physician must contact the Study Chair to obtain permission to 
break the blind. The Study Chair will notify the Pharmacy which in turn will notify the 
treating physician. The patient will be considered off study treatment at that time. 
 
6.  
TREATMENT PLAN AND ENTRY PROCEDURE 
Prior to enrollment, approval will be obtained from the scientific review committee and the 
institutional review board.   
 
  
6.1   Patient Entry and Registration 
 
6.11  An informed consent and personal health information release will be 
signed by each patient enrolled. 


 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 
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   6.12  All eligibility requirements indicated in Section 4.0 must be  
   
 
  satisfied. 
 
  
6.2  Treatment plan 
6.21  All subjects will undergo: 
 
6.211 history and physical examination within 28 days of study entry. 
 
6.212 laboratory evaluation: complete blood count, differential, 
comprehensive metabolic panel and CA125 test prior to starting 
protocol therapy and preoperatively. 
 
6.213 serum pregnancy test (within 7 days of starting study drug), for 
women of child-bearing potential. 
 
6.214 Evaluation for National Cancer Institute’s Common Toxicity 
Criteria, all grades at the time of enrollment, then weekly and the 
day of surgery. 
 
6.215 Quality of life questionnaire (FACT-O) at the time of enrollment, 
then weekly and the day of surgery (Appendix I). 
 
6.22  Patients will be randomized to treatment with either FWGE or placebo 
and assigned a blinded treatment number corresponding to one of the two 
treatment groups. The pharmacy will label the appropriate blinded drug 
specifically for the patient.  A daily supply of drug will be given to the patient from 
the study pharmacy. The amount will be determined by the time until the surgical 
procedure is scheduled. Neither patients nor treating physicians will be informed 
of patients “treated/control” status. 
 
Participants will undergo their planned surgery within 4th week after entry into the 
study.  However, if there are delays, patients can continue to take drug/placebo 


 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 
17 
for a time period no greater than 2 months. If surgery has not occurred after this 
2-month period, the patient will be removed from the study.  
Participants will take the study drug up until the day prior to surgery. A separate 
consent form will be obtained for the surgical procedure. 
 
6.23  The primary tumor tissue obtained during the subject
's surgery will be 
transferred to the pathology department for processing.  
 
6.231 Formalin Fixation 
Tissue will undergo formalin fixation then embedment in paraffin; tumor 
tissue will be stained using the H&E process and will be examined by light 
microscopy.  The tumor will also be stained by immunohistochemical 
(IHC) stain for proliferation marker Ki-67 (Anti-Ki67 primary antibody, 
catalog number 790-2910, Ventana, Tucson, AZ, USA).  The proliferation 
index will be calculated by percent of nuclear staining of the tumor for 
tumor tissue treated with FWGE versus placebo and the result will be 
compared.  The reading pathologist will be blinded to the treatment group. 
 
6.232  Fresh frozen   
An additional specimen, approximately 30mg, will be flash frozen. 
 
6.233  RNA extraction and Affymetrix gene expression analysis  
A  total  of  20  fresh  frozen  primary  OVCA  tissues  (10  FWGE- and  10 
placebo-treated) will be used for RNA extraction. Tissues (~30mg) will be 
submerged in lysis buffer. Tissues are then pulverized in BioPulverizer H 
tubes  (Bio101)  using  a  Mini-Beadbeater  (Biospec  Products).  Tubes  are 
spun briefly to pellet the garnet mixture and reduce foam.  The lysates are 
then transferred  to  a  new  1.5  ml  tube  using  a  syringe  and  21 gauge 
needle,  followed  by  passage  through  the  needle  10  times  to  shear 
genomic DNA. Total RNA will be collected using the Qiagen RNeasy Mini 
kit  according  to the  manufacturer’s  instructions.  RNA  quality  will  be 
checked  on  an  Agilent  Bioanalyzer  to  assess  quality  of  RNA  via  the 
28S:18S ribosomal RNAs.  
 


 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 
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Microarray  analysis: 10  µg  of  total  RNA  will  be  used  to  develop  the 
targets  for  Affymetrix  microarray  analysis  and  probes  will  be  prepared 
according to the manufacturer’s instructions. Briefly, biotin-labeled cRNA 
is  produced  by in  vitro  transcription,  fragmented,  and  hybridized  to 
customized  Human  Affymetrix  U133-Plus  2.0  gene  chip  arrays  at  45o C 
for 16 hours and then washed and stained using the GeneChip Fluidics.  
 
Method of statistical analysis: The expression value will be calculated and 
normalized  using  iterative  rank-order  normalization (IRON) [18].  IRON 
has been shown to perform better than both MAS5 and RMA and  
is not 
model  based  
like RMA.  Batch  effects  will  be  checked  by  using  PCA; if 
present, they will be removed using COMBAT [19]. The 54,675 probesets 
will  be  screened  to  remove  control  probesets,  probesets  with  a  small 
variance, and probesets expressed at low levels.  
 
6.24  Blood will be drawn at the time of surgery and sent for CA125 and a red 
top tube will be drawn and sent to the laboratory for 2,6, DMBQ testing 
 
6.25  2,6-DMBQ determination 
2,6-DMBQ will be determined in human plasma by high-performance liquid 
chromatography tandem mass spectrometry.  The method will be validated per 
ICH/FDA guidelines and chromatography will be conducted similarly as 
previously published [20].  Plasma samples will be prepared for chromatographic 
injections by solid-supported liquid extraction (SLE).  Calibration and quality 
control (QC) samples will be made by adding known amounts of 2,6-DMBQ to 
blank plasma.  All calibration, QC, and unknown patient samples will be prepared 
in 96 well-plate format.  Aliquots of human plasma and an internal standard will 
be added to a Biotage Isolute ® SLE plate.  A non-polar solvent, similar to the 
previously published method, will be used to extract and elute the 2,6-DMBQ to a 
collection plate. The eluate will be evaporated under a gentle stream of air and 
wells will be reconstituted with mobile phase for injection.  Samples will be 
injected into a Thermo Accela/TSQ Quantum LC/MS/MS system (Thermo 
Scientific, San Jose, CA).  Analytes will be separated using a C18 column.  
Column eluate will enter the mass spectrometer via electrospray ionization and 


 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 
19 
multiple reactions monitoring (MRM) will be employed for identification and 
quantitation. Peaks will be detected and integrated utilizing Thermo LC Quan 
software. Calibration curves will be generated for each run and patient sample 
concentrations back-calculated from the corresponding regression line.   
 
6.26  Quality of Life (QoL) 
Patients receiving FWGE may experience a positive or negative affect on QoL 
relative to those who receive placebo.  
 
The measures used in this study to assess QoL are the self-administered 
Functional Assessment of Cancer Therapy questionnaire for ovarian cancer 
(FACT-O) a 39-question assessment. Patients will be asked to complete these 
questionnaires at the following time points during their participation in the study:  
i. 
At the time of enrollment 
ii. 
Weekly 
iii. 
Prior to surgery  
 
Construct and content  
The Functional Assessment of Cancer Therapy scale developed for ovarian 
cancer (FACT-O) is a tool that provides a general QoL score. It consists of 3 
subscales: physical well-being (7 items), social/family well-being (7 items), 
emotional well-being (6 items), functional well-being (7 items), and the Ovarian 
Cancer subscale (12 items).  This questionnaire has been found to be valid and 
reliable [21]. 
 
Descriptive statistics from the baseline QoL data will be calculated. These will 
include descriptions of the distribution of QoL scores (mean, standard deviation, 
median, etc.). For all patients the baseline scores will be calculated using the 
questionnaire completed at the time of enrollment.   
 
Differences in FACT-O scores between patients receiving FWGE and placebo:  
A paired t-test will be used to test the null hypothesis that there is no difference 
between baseline PF scores and the PF scores weekly and prior to surgery for 
those patients randomized to FWGE versus placebo.  


 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 
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7.  
TREATMENT MODIFICATIONS 
 
7.1  It is expected that the dose of FWGE used in this study will be well tolerated; 
therefore there will be no dose modification. Specimens will be collected from all patients 
who undergo surgery even if they do not complete the full course of medication. 
 
7.2   Criteria for removal from study: 
7.31  
Patient refusal to continue treatment. 
7.32  
Patient non-adherence to study plan. 
7.33  
Pregnancy 
7.34  
Patient does not have surgery within 2 months of starting FWGE or 
placebo.  
7.35  
Grade 3 or 4 allergic reaction to FWGE or placebo 
 
 
8.  
STUDY PARAMETERS & SERIAL OBSERVATIONS 
The patient will be evaluated at the time of enrollment.  Quality of life and toxicity will be 
monitored weekly.  The final assessment will occur on the day of surgery.  No further 
surveillance will be performed for this trial. 
 
 
 
 
 
 
 
 
 
 
 
 
8.1 Observations and Tests 


 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 
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The following observations and tests are to be performed and recorded: 
 
 
Baseline  
Weekly 
On the day of 
surgery 
History & Physical Exam  
x 
 
x 
Performance Status  
x 
 
x 
WBC, Diff, Hgb, Hct, Platelets  
x 
 
x 
Chemistries: Na, K, Cl, CO2, 
BUN, creatinine, glucose, 
bilirubin(T/D), SGOT (AST), 
SGPT (ALT), Alk Phos, 
x 
 
x 
CA-125 
x 
 
x 
2,6 DMBQ 
 
 
1 
Toxicity Assessment 
Grade 1-4 AEs 
2 
2 
2 
QoL Survey 
3 
3 
3 
Cancer tissue for paraffin 
embedded and frozen 
 
 
x 
1. drawn in a red top tube 
2. National Cancer Institute’s Common Toxicity Criteria – CTC version 4 
3. Quality of Life Survey (QoL) – FACT-O 
 
 
9.  
EVALUATION CRITERIA 
The evaluation criteria for this study are primarily safety, toxicity, and QoL.  Secondary 
assessments are disease response based on CA125, apoptosis and tumor proliferation, 2,6 
DMBQ serum levels, and molecular changes as indicated below.  
 
9.1 Safety, Toxicity and Quality of Life: will be assessed as reviewed in section 6. 
 
9.2 Response Criteria 
 
9.21 Evaluation of Biomarkers 


 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 
22 
Biomarker-based response involves assessing the patient’s longitudinal CA125 
values. The definition of CA125 response is based on the Gynecologic Cancer 
Intergroup (GCIG 2005) criteria.  CA125 response will be determined and compared 
between FWGE and placebo treated groups 
 
9.22  
2,6-DMBQ determination 
2,6 DMBQ serum presence will be determined in FWGE versus placebo-treated 
groups.  
 
9.23   
Proliferation 
Immunohistochemistry detection of Ki-67 will be carried out in OVCA obtained from 
women following treatment with FWGE or placebo. The percentage of positively 
stained cells relative to total cells in each section will be determined and compared 
between treatment groups.  
 
 
10.  DURATION OF STUDY 
Duration of the intervention, including treatment and surgery will be completed within 4 weeks 
after beginning of study. If a patient’s surgery is delayed, the maximum time a patient will be on 
this study is 2 months.   
 
 
11. STUDY MONITORING & REPORTING PROCEDURES 
11.1  Definition of Adverse Events (AE) 
An adverse event is the development of an untoward medical occurrence, undesirable 
medical condition, or recurrence or deterioration of a pre-existing medical condition 
subsequent to exposure to FWGE.  The adverse reaction must be reported to Moffitt 
Cancer Center IRB. This study will utilize the Common Terminology Criteria for Adverse 
Events version 4.0 (CTCAE v4.0) for toxicity and adverse event reporting. A copy of the 
CTCAE v 4.0 can be downloaded from the CTEP home 
page http://ctep.cancer.gov/reporting/ctc.html  
 
11.2 Definition of Serious Adverse Events (SAE)   
A serious adverse event is any untoward medical occurrence that: 


 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 
23 
•  
Results in death, 
•   
Is life-threatening (defined as an event in which the patient was at risk of death at 
the time of the events; it does not refer to an event which hypothetically might have 
caused death if it were more severe),  
•  
Requires inpatient hospitalization or causes prolongation of existing 
hospitalization. 
•  
Results in persistent or significant disability/incapacity, 
•  
Overdose 
 
All grade 4 and 5 adverse events are by definition serious, and must be reported within 
24 hours of being aware of the event, with follow-up reporting of outcome.  Grade 2 or 3 
adverse events that require or prolong hospitalization, that result in persistent or 
significant disability, or that are events that jeopardize the patient and require 
intervention to avoid their becoming fatal, life-threatening, or disabling are also serious 
and must be reported as soon as the investigator becomes aware of the event.  
 
Collection of complete information concerning grade 3-4 SAEs is extremely important. 
Full description of each event will be followed. Thus, follow-up information which 
becomes available as the SAE evolves, as well as supporting documentation (e.g., 
hospital discharge summaries and autopsy reports), should be collected subsequently, if 
not available at the time of the initial reports, and within 24 hours sent using the same 
procedures as the initial SAE reports. 
 
12.0   STATISTICAL CONSIDERATIONS 
 
12.1 Study Design: This study is a randomized, double-blind, placebo-controlled, trial of 
the effect of FWGE on women with epithelial ovarian cancer scheduled to undergo 
surgery. As noted above, we recognize that the sample size in the current study will 
prohibit definitive statistical conclusions to be drawn on our primary objective (to test the 
safety and tolerability of FWGE), or our secondary objectives (to compare levels of 
FWGE active product, CA125 levels, proliferation, and gene expression in OVCAs from 
subjects receiving FWGE versus placebo).  The study is neither designed nor powered 
to develop statistically significant results, rather, we seek to develop preliminary/pilot 
data that will enable us to pursue a future large phase II, peer-reviewed grant-funded 


 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 
24 
trial of adjuvant FWGE in women undergoing therapy for ovarian cancer.  Preliminary 
data developed in the current proposal will support such grant applications. 
 
12.2  Primary outcome variable: The principal parameters to be collected, analyzed 
and reported will be: 
   
- patient and disease characteristics: age, race, co-morbidities, cancer histologic 
type and stage. Optimal versus suboptimal surgical cytoreduction. 
   
- differences in toxicity 
   
- differences in quality of life 
 
For each measure, Student’s t-test will be used to identify differences between FWGE- 
versus placebo-treated OVCAs. 
 
12.3  Secondary outcome variables for placebo versus FWGE:  
-the presence of 2,6-dimethoxy-p-benzoquinone in the serum  
-the change in the tumor marker CA125,  
-tumor proliferation as measured by Ki-67.  
 
For each measure, Student’s t-test will be used to identify differences between FWGE- 
versus placebo-treated OVCAs. 
 
12.4  Secondary outcome variables for genomic changes in ovarian cancer tissue for 
placebo versus FWGE.  
 
Expression  values  will  be calculated  using  the  robust  multi-array  average  (RMA) 
algorithm  
implemented  
in  
the  Bioconductor  
(http://www.bioconductor.org)  extensions  
to 
the R statistical programming environment.   
Student’s  t-test  will  be  used  to  
identify  differentially  expressed  genes  
in  comparisons 
between  FWGE- versus  placebo-treated genomic  data.  
 Differentially  expressed genes 
will analyzed  using  GeneGO  MetaCore™ software  to identify represented biologic 
pathways.  
12.5  Accrual, Sample Size, and Study Duration: The study plan is a one-stage, 
randomized, double-blind, clinical trial. Patients will be randomized with equal 


 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 
25 
probability to receive either FWGE or placebo.  The study will accrue 20 patients.  It is 
anticipated that the accrual of this many patients will be achieved in 18 months.  Less 
than 10% of the patients registered are expected to be lost due to follow-up or any other 
reason, so 20 accruals should allow for 18 evaluable cases.  
 
 


 
 
 
   
                                                                                                  
MCC 17883 June 9, 2017 
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Montbriand MJ. Alternative therapies as control behaviours used by cancer patients. J 
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Hidvegi M. Current results of FWGE research. Nogyogyaszati Onkol. 1998;3:241-243.  
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MCC 17883 
Protocol Revision 06/09/2017 – Summary of Changes 
•  Changed minimum timeline needed for FWGE and surgery