A Pilot Trial to Study the Availability and Effect of Post-OP IV Ketorolac on Ovarian, Fallopian Tube or Primary Peritoneal Cancer, Cells Retrieved From the Peritoneal Cavity
Phase: EARLY_PHASE1
Conditions: Fallopian Tube Cancer, Ovarian Cancer, Peritoneal Cavity Cancer
Sponsor: New Mexico Cancer Research Alliance

Protocol Rev #2: v01/29/2013 
 
  
The University of New Mexico Cancer Center 
INST 1115: A PILOT TRIAL TO STUDY THE AVAILIBILITY AND EFFECT OF POST-OP 
IV KETOROLAC ON OVARIAN, FALLOPIAN TUBE OR PRIMARY PERITONEAL 
CANCER CELLS RETRIEVED FROM THE PERITONEAL CAVITY 
 
 
PRINCIPAL INVESTIGATOR: 
Carolyn Y. Muller, MD 
University of New Mexico Cancer Center 
1201 Camino de Salud, NE  
Albuquerque, NM 87131 
505-925-0460 
CMuller@salud.unm.edu 
 
 
CO-INVESTIGATORS: 
 
Teresa Rutledge, MD 
University of New Mexico Cancer Center 
1201 Camino de Salud, NE 
Albuquerque, NM 87131-0001 
505-925-0460 
CMuller@salud.unm.edu 
 
Sarah Adams, MD 
University of New Mexico Cancer Center 
1201 Camino de Salud, NE 
Albuquerque, NM 87131-0001 
505-925-0395 
SAdams@salud.unm.edu 
 
Laurie Hudson, PhD 
Regents Professor 
 University of New Mexico, MSC09 5390, 
Albuquerque, NM 87131 
505-272-6264 
hsc-preaward@salud.unm.edu 
505-272-6267 
 
Angela Wandinger Ness, PhD 
Professor: Pathology 
University of New Mexico, MSC08 4640, 
Albuquerque, NM 87131 
505-272-1459 
AWandinger@salud.unm.edu 
 
Edward Bedrick, Ph.D 
Professor: IM Div of Epidemiology  
University of New Mexico, MSC10 5550, 
Albuquerque, NM 87131 
505-272-2520 
EBedrick@salud.unm.edu 

Protocol Rev #2: v01/29/2013 
 
  
 
 
 
STUDY LOCATION(S): 
 
University of New Mexico Cancer Center 
1201 Camino de Salud, NM 
Albuquerque, NM 87131 
505-272-4946 
 
 
University of New Mexico Hospital 
2211 Lomas Blvd, NE 
Albuquerque, NM 87106 
505-272-2111 
 
 
University of New Mexico College of Pharmacy 
2502 Marble, NE Room 179 NRPH Building 
Albuquerque, NM 87131 
505-272-2482 
 
 
University of New Mexico Microscopy Fluorescence Facility 
University of Mexico School of Medicine 
2325 Camino de Salud, NE CRF225A 
Albuquerque, NM 87131 
505-272-1459 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

Protocol Rev #2: v01/29/2013 
 
  
 
 
Table of Contents 
 
 
1.0 BACKGROUND   
 
 
 
 
 
 
 
 
1 
 
1.1 The Ovarian Cancer Problem 
 
 
 
 
 
 
1 
 
1.2 The Role of GTPase in Ovarian Cancer   
 
 
 
 
2 
 
1.3 Drug Library Screening Predicts R-Entatiomers of Naproxen and Ketorolac as 
 
 
 GTPase Inhibitors  
 
 
 
 
 
 
 
3 
 
1.4 Inhibition of Rac and Cdc2 in Ovarian Cancer – Preclinical Data  
 
5 
 
 
1.4.1 In vitro GLISA Assays   
 
 
 
 
 
5 
 
 
1.4.2 In vivo Studies   
 
 
 
 
 
 
5 
 
1.5 Ketorolac   
 
 
 
 
 
 
 
 
7 
 
 
1.5.1 Ketorolac Description   
 
 
 
 
 
7  
 
 
1.5.2 Ketorolac Pharmacology and Pharmacokinetics  
 
 
7 
1.5.3 Side Effects from Ketorolac 
 
 
  
 
 
8 
1.5.4 Side Effects from Blood Tests or Intravenous Risks 
 
 
8 
1.5.5 Side Effects from Peritoneal Port 
 
  
 
 
8 
1.5.6 Potential Benefits 
 
 
 
  
 
 
9 
2.0 OBJECTIVES  
 
 
 
 
 
 
 
 
 
9 
 
2.1 Primary Objective  
 
 
 
 
 
 
 
9 
 
2.2 Secondary Objective  
 
 
 
 
 
 
 
9 
3.0 INCLUSION OF WOMEN AND MINORITIES   
 
 
 
 
9 
 
4.0 PATIENT ELIGIBILITY AND EXCLUSIONS  
 
 
 
 
 
9 
 
4.1 Inclusion Criteria   
 
 
 
 
 
 
 
9 
 
4.2 Exclusion Criteria  
 
 
 
 
 
 
 
10 
5.0 STUDY DESIGN   
 
 
 
 
 
 
 
 
10 
 
5.1 Study Design  
 
 
 
 
 
 
 
 
10 
 
5.2 Study Population   
 
 
 
 
 
 
 
10 
 
6.0 SUBJECT INFORMATION AND CONSENT 
 
 
 
 
 
11 
 
7.0 STUDY RECRUITMENT PROCESS 
 
 
 
 
 
 
11 
 
8.0 STUDY MODALITY  
 
 
 
 
 
 
 
 
11 
 
8.1 Ketorolac Tromethamine   
 
 
 
 
 
 
12 
 
 
9.0 TREATMENT PLAN  
 
 
 
 
 
 
 
 
13 
 
9.1 Concomitant and Excluded Therapies   
 
 
 
 
13 
 
9.2 Patient Discontinuation   
 
 
 
 
 
 
14 
 
9.3 Study Discontinuation  
 
 
 
 
 
 
 
14 

Protocol Rev #2: v01/29/2013 
 
  
 
10.0 STUDY ASSESSMENTS  
 
 
 
 
 
 
 
14 
 
10.1 Pre-treatment Sample Collection  
 
 
 
 
 
14 
 
10.2 Post-treatment Sample Collection  
 
 
 
 
 
14 
 
10.3 Pharmacokinetic Evaluation  
 
 
 
 
 
 
14 
 
 
10.3.1 Ketorolac Concentration in Peritoneal Fluid - HPLC  
 
15 
 
 
10.3.2 GTPase Inhibitory Activity of Peritoneal Fluid   
 
 
15 
 
 
10.3.3 Efficacy of Peritoneal Fluid containing Ketorolac in Inhibiting Cell  
 
 
Adhesion and Spreading in Vitro  
 
 
 
 
 
15 
 
 
10.3.4 Spreading, Adhesion, and Migration Assays of Patient Derived  
 
 
Peritoneal Cells  
 
 
 
 
 
 
 
16 
 
 
 
10.3.4.1 Spreading  
 
 
 
 
 
 
16 
 
 
 
10.3.4.2 Adhesion  
 
 
 
 
 
 
16 
 
 
 
10.3.4.3 Migration  
 
 
 
 
 
 
16 
 
 
10.3.5 PCR analysis of Patient Derived Peritoneal Cells  
 
 
16 
11.0 STATISTICAL METHODS  
 
 
 
 
 
 
 
17 
 
11.1 Analysis of Demographic and Baseline Characteristics  
 
 
17 
 
11.2 Endpoint Analyses  
 
 
 
 
 
 
 
17 
 
 
11.2.1 Primary Endpoint and Sample Size   
 
 
 
17 
 
 
11.2.2 Secondary Endpoints   
 
 
 
 
 
17 
 
11.3 Safety Analysis   
 
 
 
 
 
 
 
18 
 
12.0 SOURCE DOCUMENTS AND CASE REPORT FORMS 
 
 
 
18 
12.1 Source Documents 
 
 
 
 
 
 
 
18 
12.2 Case Report Forms 
 
 
 
 
 
 
 
18 
 
 
13.0 ASSESSMENT OF SAFETY   
 
 
 
 
 
 
19 
 
13.1 Specification of Safety Variables  
 
 
 
 
 
19 
 
 
13.1.1 Adverse Events  
 
 
 
 
 
 
19 
 
 
13.1.2 Serious Adverse Events  
 
 
 
 
 
19 
 
13.2 Methods and Timing for Assessing and Recording Safety Variables  
20 
 
13.2.1 Assessment of Adverse Events  
 
 
 
 
 
20 
13.2.2 Independent Research Monitoring  
 
 
 
 
 
21 
 
 
14.0 RESPONSIBILITIES OF THE PRINCIPAL INVESTIGATOR  
 
 
21 
 
 
REFERENCES  
 
 
 
 
 
 
 
 
 
23 
 

Protocol Rev #2: v01/29/2013 
 
1 
 
1.0 
BACKGROUND 
 
1.1 
The Ovarian Cancer Problem 
Ovarian cancer is the sixth most common cancer and the seventh most common cause of 
cancer deaths in women across the globe.(1) In the US alone, it is estimated that 21,880 
new cases of ovarian cancer were diagnosed and 13,850 women died of the disease in 
2010. (2) The majority of women, nearly 70%, will present with advanced stage disease that 
heralds a poor prognosis. (3) Despite aggressive treatment that still favors initial debulking 
surgery followed by a platinum and taxane based chemotherapy regimen, most patients 
relapse after achieving a complete clinical response. (3, 4) 
 
The management of advanced stage ovarian cancer continues to evolve. A recent 
randomized trial from the European clinical trials group (EORTC) concludes that 
neoadjuvant chemotherapy followed by surgical debulking is as effective as primary 
cytoreductive surgery followed by chemotherapy with less overall toxicity. (5) However, 
based on some limitations of the trial, most US gynecologic oncologists still favor optimal 
front line cytoreductive surgery followed by adjuvant chemotherapy. Data from several 
randomized trials also support the role of intraperitoneal chemotherapy following tumor 
debulking, demonstrating a significant PFS and overall survival (OS) improvement in 
patients with optimal surgical cytoreduction (all residual tumor < 1 cm). (6) This recent GOG 
trial (GOG172)  led to an NCI alert discussing intraperitoneal chemotherapy as the new 
standard of care and the GOG has adopted IP therapy as the control arm of the subsequent 
front-line trial design (GOG 252).(7,8) 
 
There is a move towards personalized medicine in cancer care, and significant effort is 
underway to evaluate new targeted therapeutics for the treatment of ovarian cancer.(9) 
Novel therapeutics studied by the Gynecologic Oncology Group (GOG) alone in recurrent 
ovarian cancer  include VEGFR inhibitors (bevacizumab, VEGF-TRAP, sorafenib, 
enzastaurin, AMG-706), EGFR inhibitors (gefitinib, imatinib, cetuxumab, pertuzumab, 
tratuzumab, lapatinib), mTOR inhibitors (temsirolimus, everolimus), and  DNA methylation 
inhibitors (vorinostat, dasatinib)[ https://gogmember.gog.org/]. To date, the only agent of any 
particular interest is bevacizumab, as recent studies suggest a progression free survival 
advantage (PFS) in both front line and recurrent ovarian cancer.(10-17) But the cost and 
toxicity of this agent may not warrant generalized use.(18) The number of novel biologic 
agents under development  is staggering, demanding changes in clinical trial design to 
efficiently identify promising targets for ovarian cancer.   
 
One means of identifying potential new drug targets is an in-silico approach, where a 
signaling pathway is chosen for a drug library screen for predicted interaction at a key 
kinase or enzymatic site. Drug repurposing, screening a library of FDA approved agents, will 
identify agents that are clinically available and for which pharmacology and 
pharmacokinetics are known and preclinical data can be generated rapidly without the 
subsequent need for GMP new drug production. Our co-investigator Wandinger-Ness 
funded by R03 and R21 grants  in collaboration with the UNM Center for Molecular 
Discovery (UNMCMD) conducted the first comprehensive high throughput screen for 
inhibitors and activators of small GTPases using purified proteins and libraries of both FDA 
approved and novel small molecules (19,20). This clinical trial is a novel design in order to 
test an FDA approved drug for effect on ovarian cancer cells in the peritoneal cavity.  

Protocol Rev #2: v01/29/2013 
 
2 
 
Fig. 2 . Dysregulation of GTPase targets. (A) Ovarian tumor tissue microarrays were purchased from US Biomax 
and stained for Cdc42 (pAb 10155-1-AP ProteinTech Group, Inc.). Tumor grade and type was confirmed, and 
immunoreactivity scoring was conducted by pathologist Dr. Lesley Lomo. Statistical analyses show p<0.001 for 
low vs. high grade tumors, UNM Cancer Center Biostatistics core. (B) Primary tumor cDNA samples (TissueScan 
Disease Tissue qPCR Arrays, OriGene) were screened for Rac1b expression by qPCR. Normal vs. Gr. 2 p<0.05. 
(C) Rac1 and Cdc42 GTPase activities measured by GLISA in cells from fresh ovarian patient ascites or after 48 
h in culture.  
 
1.2 
The Role of GTPase in Ovarian Cancer 
Small GTPases, including members of the Rab, Ras and Rho families, are attractive 
targets for the development of cancer therapeutics based on their pivotal roles in protein 
trafficking, proliferation/survival and cytoskeletal organization, respectively (Fig. 1). The Ras-
homologous (Rho) family of small GTPases (Rac, Cdc42 and Rho) are key regulators of 
numerous cellular functions including actin reorganization, cell motility, cell-cell and cell-
extracellular matrix (ECM) adhesion, all 
important parameters of ovarian cancer 
metastatic dissemination ) In many 
human cancers (including colon and 
breast), aberrant Rho-family signaling 
due to changes in the GTPase itself or in 
its regulation loops is a critical 
underpinning of tumor growth and 
survival, invasion and metastasis, [24-
26]. Our own studies are the first to 
demonstrate dysregulation of Rac1 and 
Cdc42 GTPase expression and activity 
in ovarian cancer (Fig. 2). Stage and 
grade dependent overexpression of 
Cdc42 protein was detected by 
immunohistochemical staining of human 
tumor samples (Fig. 2A). Cdc42 
overexpression levels were highly 
significant for malignant, high-grade 
tumors (p<0.001). Although there was 
more modest elevation of total Rac1 
protein, expression of a constitutively 
active splice variant Rac1b (first documented in breast tumors, [26]) was detected by qPCR 
(Fig. 2B) and a GLISA activity assay showed both Cdc42 and Rac1 GTPases highly active 
in fresh tumor isolates, but activity declined after 48h in culture (Fig. 2C). Although Rac1 and 
Cdc42 have been recognized as attractive therapeutic targets, specific Rac GTPase 
inhibitors [27,28] are not in clinical use and there are no known  Cdc42 specific inhibitors in 
clinical use.  
 
 
Fig. 1. Rac1 and Cdc42 GTPases integrate signaling 
pathways that are important in cancer growth and 
metastasis. Activation of tyrosine kinase receptors, G-
protein coupled receptors (GPCRs) and integrins causes 
Rac and Cdc42 GTPases to bind GTP and membranes. 
The 
GTP-bound 
proteins 
interact 
with 
specific 
downstream effectors to promote actin reorganization 
that affect changes in cell motility, adhesion, cell growth, 
gene expression and apoptosis. GTPase functions in the 
regulation of proliferation, angiogenesis and metastasis 
are intimately linked to cancer development and 
progression. Adapted from [1]. 

Protocol Rev #2: v01/29/2013 
 
3 
 
1.3 
Drug Library Screening Predicts R-Entatiomers of Naproxen and Ketorolac as 
GTPase Inhibitors 
 
Screening of the Prestwick library of FDA-approved molecules identified the R-
enantiomer of naproxen as active against Rac1 and Cdc42, which we confirmed in 
cellular GLISA assays (Fig. 3A).  R-, and S-ketorolac were identified for structure 
activity testing based on additional 
cheminformatics. Despite the common 
chemotype, S-naproxen and 6-MNA did 
not significantly inhibit Rac1 or Cdc42 
indicating that GTPase inhibition is 
independent of the COX pathway. At 
nontoxic doses that are meaningful with 
respect to human serum levels at 
therapeutic doses (Table 1), the R-
enantiomers of naproxen or ketorolac 
are more effective for inhibition of 
migration in OVCA 429 and SKOV3ip 
(Fig. 4A-B), and primary ovarian tumor 
cells isolated from ascites (not shown). 
The same structure activity relationship 
was observed for multicellular 
aggregate formation (Fig. 4C). 
Inhibition of proliferation was less 
marked (~50%) with R-naproxen>>S-
naproxen. More than 20 other common 
NSAIDS were tested and did not detect 
inhibitory activity in vitro or in cells 
against Rac1 or Cdc42, nor any impact 
on ovarian cell migration or 
proliferation. Thus, the structure activity 
relationships and selectivity suggest 
that the inhibitory actions of R-naproxen 
and R-ketorolac are due to an effect on 
novel Rac1 and Cdc42 GTPase targets 
as first revealed by the molecular library 
screen and predicted by 
cheminformatics.  
Virtual docking predicts that R-
naproxen, but not S-naproxen can bind 
the GDP-bound pocket of Rac1 (Fig. 
3B). In this model R-naproxen is 
stabilized through favorable H-bonds 
with Thr17 and Asp57, as  
 
 
 
Fig. 3. Enantiomer selective inhibition of 
GTPases. (A) GLISA measures cellular effects 
of small molecules on Rac1. Cells were 
untreated or 
incubated with the indicated 
compound (R-Nap=R-naproxen,R-ketorolac, S-
ketorolac 
(enantiomers 
differ 
based 
on 
orientation of circled methyl group on naproxen 
or carboxyl group on ketorolac); or the non-
stereoselective 
6MNA 
= 
6-methoxy-2-
naphthalene) for the indicated times then treated 
+/- 10 ng/ml EGF for 2 min. Equal cell lysate 
protein was assayed for activated Rac1 using a 
commercial, plate based Pak-binding assay 
according to manufacturer’s instructions (GLISA, 
Cytoskeleton, Inc.). Similar results were obtained 
for Cdc42 and S-naproxen like S-ketorolac was 
not inhibitory (not shown) (B) Docking predicts 
R-enantiomer-selective binding of naproxen 
to GDP-bound Rac1. R-naproxen predicted to 
bind GDP-bound Rac1, but not GTP-bound 
conformation. S-naproxen is sterically blocked 
from binding (not shown). Rac1 crystal structure 
used to dock molecules via FRED (OpenEye).  

Protocol Rev #2: v01/29/2013 
 
4 
 
Table 1. Serum concentrations and effective doses of screened targets 
 
S-Nap 
R-Nap 
(S,R) Ket 
S-Ket 
R-Ket 
Target 
COX 1/2 
Rac1/cdc42 
COX 1>2 
COX1>2 
Rac1/Cdc42 
Serum Cmax 
413+/-74µM 
NA 
6.4+/-1.8 µM 
NA 
NA 
Serum Cave 
130-391 µM 
NA 
8.3+/-2.3 µM 
NA 
NA 
IC50COX1 
35.5-48.3 µM 
>100xS-Nap 
0.6 µM 
0.1 µM 
ND 
IC50COX2 
64.6-79.5 µM 
>100xS-Nap 
2.7 µM 
2.5 µM 
ND 
IC50Rac1 
>100 µM 
TBD 
TBD 
TBD 
TBD 
IC50 (est) 
migration 
>300 µM 
100 µM 
34 µM 
34 µM 
7 µM 
Legend Table 1. Serum concentrations (maximum (Cmax) and steady state (Cave)) were based on typical 
oral dosing (S-Naproxen 500 mg; R,S-ketorolac 30 mg) and derived from FDA product literature and 
primary literature (S-Naproxen, [29,30]; ketorolac [31-35]. Note that an IV dose of 30 mg ketorolac 
achieves a Cmax of 13.7 µM [35,36]. IC50  values for COX1/2 in human cells were obtained from the 
literature (R and S-naproxen, [37; 38-40]; R and S-Ketorolac, [31-33; 35; 36; 41].  Migration  IC50 values 
were estimated from limited dose response data (Fig. 4) or calculated by GraphPad Prism5 (ketorolac). 
NA-not applicable, no human dosing; ND=not detected, no COX inhibition; TBD=to be determined. 
well as through interaction with magnesium via the naproxen carboxyl group. 
Interestingly, Thr17 is the mutated residue in dominant negative forms of Rac1. The 
model predicts stabilization of the inactive GDP-bound GTPase and provides a 
testable hypothesis for a novel mechanism of action which is under active 
 
Fig. 
4. 
Enantiomer 
selective 
inhibition of ovarian tumor cell 
migration & aggregation. A) R-
naproxen inhibits ovarian tumor cell 
migration. OVCA 429 cell migration 
was measured under the indicated 
conditions 
using 
modified 
Boyden 
chambers. *P<0.05 Similar results 
were obtained for SKOV3ip and OVCA 
433 cells. (B) R-ketorolac inhibits 
ovarian 
tumor 
cell 
migration. 
SKOV3ip cells were incubated for 24h 
under the indicated conditions and 
migration was measured as in (A). 
Comparable results were obtained for 
OVCA 429 cells. C) Inhibition of 
aggregation. OVCA 429 cells were 
trypsinized and resuspended at 3x106 
cells/ml in medium containing drugs as 
indicated (100 μM). 25 μl drops were 
suspended and imaged after 24h. 
Comparable responses were observed 
in OVCA 433 and SKOV3ip cells.  

Protocol Rev #2: v01/29/2013 
 
5 
 
investigation. As predicted by the enantiomer-selective inhibition of Rac1 and Cdc42 
activity, we find that R-naproxen, R-ketorolac and the established Rac1 inhibitor 
NSC23766 inhibit ovarian tumor cell proliferation, migration and MCA formation (Fig. 
4A-C and data not shown) while S-naproxen and 6-MNA display little effect. This 
data clearly shows the enatiomeric specificity of the target GTPases in ovarian 
cancer.   
 
1.4 
Inhibition of Rac and Cdc2 in Ovarian Cancer – Preclinical Data 
 
 
Our group has shown that the R-enantiomers of naproxyn and ketorolac 
inhibit Rac1 and Cdc42 activity which inhibits cell proliferation, migration and multi-
cell aggregate formation. This effect is completely independent of any COX inhibition 
which is a function of the S-enantiomers (Figures 3.4).  R-naproxen was used as the 
prototype drug for the preclinical data, while repeat assays for R-ketorolac are 
underway.  
 
1.4.1 In vitro GLISA Assays 
Our collaborator, Wandinger-Ness, has extensively used GLISA for measuring EGF-
stimulated Rac and Cdc42 activation in Swiss 3T3 cells in the presence and 
absence of test compounds. Preliminary data demonstrate R-naproxen inhibits Rac1 
and Cdc42 GTPase activities in OVCA cell lines (429 and 433).  The GLISAs are 
highly specific, quantitative, can be performed with small sample sizes (<50 µg of 
cell lysate per assay, compared to 1-10 mg required for conventional PAK binding 
assay) and have excellent reproducibility. Preliminary data demonstrate that R-
naproxen, but not 6MNA, inhibits Rac and Tiam membrane association consistent 
with the inhibition of Rac1 activity measured by GLISA (Fig. 5A-C).  In addition, the 
formation of Cdc42-dependent, actin-based invadopodia that are crucial for matrix 
degradation and invasion [42; 43] is also significantly reduced by R-naproxen, but 
not by 6 MNA (Fig. 5D-E). Repeat studies using R-ketorolac are underway 
 
1.4.2 In vivo Studies 
Using R-naproxen as the prototype drug, R-naproxen decreased the number of 
implanted tumors in a xenograft animal model using athymic nude mice injected with 
human ovarian tumor cells adapted to intraperitoneal growth (SKOVip; obtained 
through a formal MTA from MD Anderson). Treatments were incorporated into 
transgenic dough (BioServe, Frenchtown, NJ) and animals were dosed twice per 
day to achieve established therapeutic serum levels [39]. Animals were given 
placebo dough or dough containing R-naproxen, S-naproxen or 6MNA. After 
sacrifice, animals fed R-naproxen exhibited a 4-fold reduction in tumor number and a 
36% decrease in total tumor burden (Fig. 6), while the 6MNA and S-naproxen 
treated animals were similar to the placebo controls. These data demonstrate that 
the enantiomer specific inhibitory properties of R-naproxen are preserved in vivo and 
that this NSAID may reduce tumor growth and spread, a point that will be further 
tested and extended to ketorolac in this proposal. 
 

Protocol Rev #2: v01/29/2013 
 
6 
 
 
Fig. 5. Assays of GTPase and pathway inactivation by drug treatment.  (A) OVCA 433 cells were 
treated with 100 µM R-naproxen for 1 h, fixed and stained for Rac1, actin and Tiam1. (B) 
Quantification of staining shows a notable loss of Rac1 and Tiam membrane association with R-
naproxen, but not S-naproxen or 6MNA treatment.  (C) Total Rac protein levels were unchanged by 
treatment. (D) OVCA 429 cells plated on FITC-fibronectin extend invadopodia into the matrix and 
create holes due to the activity of associated matrix metalloproteinases. (E) R-naproxen, but not 
6MNA decreased invadopodia formation 
 
 
 
 
 
 
 
 
 
 
 
 
 
1.5
Fig. 6. R-Naproxen reduces tumor number in xenograft model. Athymic nude mice were given an 
oral dose (10mg/kg) of the indicated compounds in transgenic dough. Individual dosing was confirmed 
by direct observation. Mice were acclimated with placebo for 3 days and 1 day prior to injections of 
human ovarian GFP-tagged SKOV3ip cells, mice were left on placebo or provided the indicated 
treatments. After 2 weeks of tumor growth the mice were sacrificed, necropsy was performed and 
images were taken of the peritoneal cavity. All tumors were counted. Internal organs were moved to 
visualize the full cavity with quantification given in the chart. 

Protocol Rev #2: v01/29/2013 
 
7 
 
 
1.5 KETOROLAC  
Ketorolac will be used in this study as preliminary data support the enantiomeric specific 
inhibition of cell adhesion and migration. Although R-naproxen was the prototype NSAID R-
enatiomer, clinically available naproxen is the S-enantiomer. Ketorolac is a racemic mixture 
of the R and S-enantiomer, the R-enatiomer is stable during metabolism and ketorolac is 
available for IV administration and is FDA approved for post-operative pain management.  
 
 
1.5.1.  Ketorolac Description  
Ketorolac tromethamine is a member of the pyrrolo-pyrrole group of non-steroidal anti-
inflammatory drugs (NSAIDs). The chemical name for Ketorolac tromethamine is (±)-5-
benzoyl-2,3-dihydro-1H-pyrrolizine-1-carboxylic acid, compound with 2-amino-2-
(hydroxymethyl)-1,3-propanediol. The structural formula is: 
 
C19H24N2O6          M.W. 376.41 
Ketorolac tromethamine is a racemic mixture of [-]S and [+]R Ketorolac tromethamine. 
Ketorolac tromethamine may exist in three crystal forms. All forms are equally soluble in 
water. Ketorolac tromethamine has a pKa of 3.5 and an n-octanol/water partition coefficient 
of 0.26. Ketorolac is available for oral, intramuscular and intravenous use.  
 
1.5.2. Ketorolac Pharmacology and Pharmacokinetics 
Ketorolac tromethamine is a non-steroidal anti-inflammatory drug. The biological activity of 
Ketorolac tromethamine for pain relief is associated with the S-form. The peak analgesic 
effect of Ketorolac tromethamine occurs within 2 to 3 hours and is not statistically 
significantly different over the recommended dosage range of Ketorolac tromethamine. The 
greatest difference between large and small doses of Ketorolac tromethamine is in the 
duration of analgesia. 
Ketorolac tromethamine is a racemic mixture of [-]S- and [+]R-enantiomeric forms, with the 
S-form having analgesic activity and the R-form the proposed cancer inhibition effect. The IV 
dosing of ketorolac is 100% bioavailable. The pharmacokinetic parameters for IV dosing is 
shown in Table 2. In adults, following IV administration of Ketorolac tromethamine in the 
recommended dosage ranges, the clearance of the racemate does not change. This implies 
that the pharmacokinetics of Ketorolac tromethamine in adults, following single or multiple  
IV doses of Ketorolac tromethamine are linear. At the higher recommended doses, there is a 
proportional increase in the concentrations of free and bound racemate. 
The mean apparent volume (Vß) of Ketorolac tromethamine following complete distribution 
was approximately 13 liters. This parameter was determined from single-dose data. The 
Ketorolac tromethamine racemate has been shown to be highly protein bound (99%). 
Nevertheless, plasma concentrations as high as 10 mcg/mL will only occupy approximately 
5% of the albumin binding sites. Thus, the unbound fraction for each enantiomer will be 
constant over the therapeutic range. A decrease in serum albumin, however, will result in 

Protocol Rev #2: v01/29/2013 
 
8 
 
increased free drug concentrations. The half-life of the Ketorolac tromethamine S-
enantiomer was approximately 2.5 hours (SD ± 0.4) compared with 5 hours (SD ± 1.7) for 
the R-enantiomer. In other studies, the half-life for the racemate has been reported to lie 
within the range of 5 to 6 hours. 
 
Tmax (min) 
Cmax (mcg/ml) 
single dose 
Cmax (mcg/ml) 
steady state qid 
Cmin (mcg/ml) 
steady state qid 
Cavg (mcg/ml) 
steady state qid 
15 mg 
1.1 + 0.7 
2.47 + 0.51 
3.09 + 0.1.17 
0.61 + 0.21 
1.09 + 0.30 
30 mg 
2.9 + 1.8 
4.65 + 0.7 
6.85 + 0.7 
1.04 + 0.7 
2.17 + 0.7 
Tmax = time to peak plasma concentration, Cmax = peak plasma concentration, Cmin = trough 
plasma concentration, Cavg = average plasma concentration 
Table 2. Average Pharmacokinetic Parameters for IV Ketorolac 
The intraperitoneal distribution of ketorolac and its racemic mixture is unknown and 
represents the primary endpoint of this trial. Based on the known pharmacokinetics, it is 
favorable that the half-life of the R-enantiomer is longer and that both are protein bound. We 
anticipate that the post-op decrease in serum albumin will favor free drug with a high rate of 
transport into the peritoneal cavity.  
1.5.3 Side Effects from Ketorolac 
There are few side effects that are expected after a single dose.  Side effects that are likely 
in 1% -10% chance include: pain at the IV site, rashes, itching, abdominal pain, gas, 
bleeding, mouth sores, constipation, diarrhea, bloating, heartburn, vomiting, upset stomach, 
ulcers, high liver function tests, low red cell count, swelling, increased bleeding time, purple 
spots, drowsiness, ringing in the ears, dizziness, sweating and abnormal kidney function. 
Side effects that have a greater than 10% chance of happening include: nausea and 
headaches. It is very unlikely that these side effects will occur from a single dose or even 
several doses over 24 hours. Many of these side effects will also be present just from the 
surgery and other medications given after the surgery.  
 
1.5.4 Side Effects from Blood Tests or Intravenous Risks   
Frequent blood samples will be collected during this study. The subject may experience 
pain, bleeding from the puncture site or in tissues surrounding the puncture site, blood clot 
formation, or local infection and inflammation at the puncture site (very rare). An intravenous 
catheter (plastic tube) will be in place for routine postoperative care and will be the site used 
for administration of the ketorolac. It is typical that a second IV is in place. If able, this IV will 
be used to draw the study samples in an attempt to avoid additional needle sticks. In the 
event that a second IV is not in place, the investigator will draw blood using a small needle.  
Bleeding at the site where the catheter is or where needle sticks are made into the skin can 
cause bleeding under the skin causing a bruise. There is a possibility of infection, with 
swelling, redness and pain at the site where the ketorolac goes into the vein. It is extremely 
rare but possible that a serious infection of the blood stream or the heart valves or the 
formation of a blood clot in the lungs. The subjects will be re-admitted to the hospital to get 
treatment for these rare but serious problems if they occur after hospital discharge.  
 
1.5.5 Side Effects from Peritoneal Port   
Frequent fluid samples from the subjects’ abdomen will be collected during this study. The 
subject may experience pain, bleeding from the puncture site at the port or in tissues 

Protocol Rev #2: v01/29/2013 
 
9 
 
surrounding the puncture site, blood clot formation, or local infection and inflammation at the 
puncture site (very rare). A Huber needle is inserted in the port reservoir at the time of 
catheter placement in the operating room and left there during the time of the collection.. 
The subject may feel some irritation in abdomen during the time when the fluid is being 
drawn into a syringe. There is a very small possibility that they subject will develop 
peritonitis. This would need to be treated with antibiotics and would likely become evident 
during the hospitalization.  
 
1.5.6 Potential Benefits 
Ketorolac will improved pain control in the post-operative period. It is also possible that use 
of ketorolac in ovarian cancer patients may have a benefit in cancer outcome as is 
suggested in one breast cancer study.  
 
2.0 
OBJECTIVES 
2.1 
Primary Objective  
The primary objective of this study is to determine the concentration of measurable levels of 
R and S- Ketorolac (and the percent racemic mixture) in the peritoneal cavity after IV 
administration in the post-operative patient after cytoreductive surgery.  
 
2.2 
Secondary Objectives 
The secondary endpoint of this study is to determine the effect of IV ketorolac on cell 
adhesion and migration of ovarian cancer cells retrieved from the peritoneal cavity after 
cytoreductive surgery 
.  
3.0 
INCLUSION OF WOMEN AND MINORITIES 
The University of New Mexico Cancer Center will not exclude potential subjects from 
participating in this or any study solely on the basis of ethnic origin or socioeconomic status. 
Every attempt will be made to enter all eligible patients into this protocol and therefore 
address the study objectives in a patient population representative of the entire ovarian, 
fallopian tube, and primary peritoneal cancer population treated by participating institutions. 
Only women will be recruited to this trial as only women can be diagnosed with ovarian, 
fallopian tube or primary peritoneal cancer.  
 
4.0 
PATIENT ELIGIBILTY AND EXCLUSIONS 
4.1 
Inclusion Criteria  
This protocol will have a 2 stage eligibility as consent will be done in the preoperative 
planning. Patients must meet all of the following criteria to be eligible for study entry. 
 
Primary Eligibility (pre-op) 
 
 
Patients must be suspected of having a diagnosis of ovarian, fallopian tube or 
primary peritoneal cancer with a planned cytoreductive surgery. 
 
Borderline ovarian cancer with ascites. 
 
ECOG/Zubrod/SWOG Performance Status <2 (Karnofsky Performance Status > 
70%) 
 
Female‚ age >18 years 
 
Ability to provide informed consent  
 
Baseline laboratory values (bone marrow, renal, hepatic):   
o Adequate bone marrow function: 

Protocol Rev #2: v01/29/2013 
 
10 
 
 
Absolute neutrophil count >1000/µL 
 
Platelet count >100‚000/µL 
o Renal function: 
 
Serum creatinine < 1.5 x ULN 
o Hepatic function: 
 
Bilirubin <1.5x normal  
 
Serum glutamic-oxaloacetic transaminase (SGOT) (aspartate 
aminotransferase [AST]) or serum glutamic-pyruvic transaminase 
(SGPT) (alanine aminotransferase [ALT]) levels 2 x ULN 
 
No known bleeding disorders 
 
No known sensitivity to NSAIDs 
 
No active peptic ulcer disease 
 
No active bleeding 
 
Secondary Eligibility 
 
 
Histologic diagnosis of epithelial ovarian, fallopian tube or primary peritoneal cancer 
on frozen section diagnosis 
 
Placement of an intraperitoneal port at the time of surgery for anticipated use for 
adjuvant chemotherapy or management of post-operative ascites 
 
If epidural catheter was used – the catheter must be removed prior to treatment 
 
No active bleeding 
 
4.2 
Exclusion Criteria 
Patients meeting any of the following criteria are ineligible for study entry: 
 
Non-epithelial ovarian cancer or metastatic cancer to the ovaries 
 
Borderline ovarian cancer without ascites 
 
Presumed early stage ovarian cancer 
 
No clinical indication for a peritoneal port 
 
Active use of an epidural catheter  
 
Uncontrolled or unstable medical conditions 
 
Off study use of ketorolac or other NSAIDs prior to study administration  
 
Active bleeding or high risk of bleeding 
 
Active therapeutic anticoagulation 
 
Known hypersensitivity to NSAIDs 
 
Chronic or acute renal insufficiency as defined by a preoperative serum creatinine 
greater than 1.5mg/dL or creatinine clearance of < 60 ml/min 
 
Any co-morbid condition that‚ in the view of the attending physician‚ renders the 
patient at high risk from treatment complications 
 
5.0 
STUDY DESIGN 
5.1 
Study Design 
This is a pilot (feasibility) trial to determine the availability, concentration and racemic 
mixture of a clinically indicated pain medication, ketorolac, in the peritoneal cavity and the 
subsequent effect on free floating ovarian cancer cells in patients after cytoreductive surgery 
for ovarian cancer.  
 
5.2 
Study Population 
Adult female subjects at least 18 years of age suspected of having a diagnosis of ovarian, 
fallopian tube or primary peritoneal cancer with a planned cytoreductive surgery will be 
eligible for this study. 

Protocol Rev #2: v01/29/2013 
 
11 
 
 
Subjects will undergo a pre-operative and a post-operative eligibility evaluation. Subjects 
who meet all of the inclusion criteria and do not meet any of the exclusion criteria will be 
eligible for enrollment into the study. 
 
6.0 
SUBJECT INFORMATION AND CONSENT 
The Investigator or her representative will explain the nature of the study to the subject, and 
answer all questions regarding this study.  Prior to any study-related pre-operative and post-
operative procedures being performed on the subject, the informed consent statement will 
be reviewed and signed and dated by the subject and the person who administered the 
informed consent.  A copy of the informed consent form will be given to the subject and the 
original will be placed in the subject’s medical record. 
 
Spanish speaking subjects will be consented using a Spanish Short Form. A Certified 
Spanish Translator from the University of New Mexico will translate the study processes and 
procedures to the subject and family member. 
 
The subjects will be re-consented if revisions are made to the consent form that would alter 
the subjects decision to participate in the trial. 
 
7.0 
STUDY RECRUITMENT PROCESS 
The Principal Investigator, Dr. Carolyn Muller and Co-Investigator, Dr. Teresa Rutledge will 
identify potential study participants through medical record review from patients within their 
practices. The Investigators will discuss this study with subjects during the subjects pre-
operative counseling visit. 
 
Subjects will not be compensated for participation in the research study. 
 
 
8.0 
STUDY MODALITY 
8.1  
Ketorolac tromethamine   (Torodol® Roche Pharmaceuticals) 
 
Formulation: Ketorolac is supplied in sterile single use 1mL vials of 15 mg/mL( NDC 
55390-480-01) and 30 mg/mL, 1 mL single-dose vials of 30mg/mL (NDC 55390-481-
01). Supplies are obtained by the University Hospital (University of New Mexico) and 
stored in the Pyxsis until ordered by the treating physician. 
 
Preparation: Ketorolac tromethamine for IV use is supplied in 1 mL vials of either 
15mg or 30mg individual dose which requires no reconstitution.  
 
Storage: Vials of ketorolac tromethamine are stored at room temperature protected 
from light in the carton until use. 
 
Dosing:  Ketorolac tromethine will be dosed according to FDA approved 
recommendations. For planned single dose (IV) based on clinical scenario: 
• Patients < 65 years of age: One dose of 30 mg. 
• Patients ≥ 65 years of age, renally impaired and/or less than 50 kg (110 lbs)   
of body weight: One dose of 15 mg. 
For planned Multiple-Dose Treatment (IV) for indicated pain management 
• Patients < 65 years of age: The recommended dose is 30 mg ketorolac 
tromethamine injection every 6 hours. The maximum daily dose for these 
populations should not exceed 120 mg. 

Protocol Rev #2: v01/29/2013 
 
12 
 
• For patients ≥ 65 years of age, renal impaired patients, and patients less 
than 50 kg (110 lbs): The recommended dose is 15 mg ketorolac 
tromethamine injection every 6 hours. The maximum daily dose for 
these populations should not exceed 60 mg. 
 
Adverse Effects:  Complications of treatment with ketorolac tromethamine, such as 
G.I. ulceration, bleeding and perforation, postoperative bleeding, acute renal failure, 
anaphylactic and anaphylactoid reactions and liver failure have been described.  
These NSAID-related complications can be serious in certain patients for whom 
ketorolac tromethamine is indicated, especially when the drug is used 
inappropriately. 
 
In patients taking ketorolac tromethamine or other NSAIDs in clinical trials, the most 
frequently reported adverse experiences and frequency are shown in the table 
below: 
 
 
< 1% 
1%-10% 
>10% 
cardiovascular 
CHF, palpitation, 
pallor, tachycardia, 
syncope 
hypertension 
 
dermatologic 
Alopecia, urticaria, 
photosensitivity 
Injection site pain, 
rashes, pruritis 
 
gastrointestinal 
Anorexia, dry mouth, 
eructation, 
esophagitis, 
excessive thirst, 
gastritis, glossitis, 
hematemesis, 
hepatitis, increased 
appetite, jaundice, 
melena, rectal 
bleeding 
Abdominal pain, 
flatulence, bleeding, 
perforation, 
stomatitis, 
constipation, 
diarrhea, GI fullness, 
heartburn, vomiting, 
dyspepsia, ulcers, 
elevated liver 
functions 
nausea 
heme and lymph 
Ecchymosis, 
eosinophilia, 
epistaxis, 
leukopenia, 
thrombocytopenia 
Anemia, edema, 
increased bleeding 
time, purpura 
 
metabolic/nutritional 
Weight change 
 
 
nervous system 
abnormal dreams, 
abnormal thinking, 
anxiety, asthenia, 
confusion, 
depression, 
euphoria, 
extrapyramidal 
symptoms, 
hallucinations, 
hyperkinesis, inability 
to concentrate, 
insomnia, 
nervousness, 
Drowsiness, tinnitus, 
dizziness, sweating 
headaches 

Protocol Rev #2: v01/29/2013 
 
13 
 
paresthesia, 
somnolence, stupor, 
tremors, vertigo, 
malaise 
reproductive 
infertility 
 
 
respiratory 
Asthma, cough, 
dyspnea, pulmonary 
edema, rhinitis 
 
 
urogenital 
cystitis, dysuria, 
hematuria, increased 
urinary frequency, 
interstitial nephritis, 
oliguria/polyuria, 
proteinuria, 
renal failure, urinary 
retention 
 
Abnormal renal 
function,  
 
others 
Fevers, infections, 
sepsis, conjunctivitis 
 
 
Table 3. Reported toxicities of racemic ketorolac 
 
Supplier: Manufactured by Bedford Laboratories™ Bedford, OH 44146  Div-KRL-
P05. Vials are supplied directly to University Hospital.  
 
Administration:  Ketorolac 15mg or 30 mg doses will be administered via IV Push 
undiluted over 15 – 30  seconds.  Prior to administration, the IV line will be flushed with 5-10 
ml's NS.  The ketorolac dose will be followed with a 5-10 ml NS push again to ensure the 
ketorolac completely clears the IV line.   
 
9.0 
TREATMENT PLAN 
All patients will receive a single dose of IV ketorolac for pain management for the indication 
of post-operative pain control. The single dose will be considered the study dose for which 
all consenting eligible patients will receive. The dose of IV ketorolac will be determined per 
the recommendations in the package insert. Patients less than 65 years of age and in 
otherwise good health will receive a 30mg IV single dose. Patients 65 years of age or 
greater, or who have mild renal insufficiency or are of low weight (as per section 3.2) will 
receive a single 15 mg dose IV ketorolac. In patients who have a clinical pain response and 
have no contraindications to multi-dose (every 6 hours over 24 hours), additional doses will 
be given per physician discretion based on clinical indication. Patients receiving 24 hour 
dosing will be eligible for sample time points after 24 hour dosing.  
 
9.1 
Concomitant and Excluded Therapies 
Use of other non-steroidal (NSAIDs) medications during the time-points of this trial is not 
permitted while participating in this study. Narcotic regimens with or without acetaminophen 
is allowed. If patients require additional NSAIDs for pain control, the patients will be 
withdrawn from study although data collected prior to withdrawal will be included in the 
analysis. Use of cytotoxic chemotherapy during this trial is not allowed, however if the 
clinical situation warrants immediate chemotherapy, the patient will be withdrawn from the 
trial. Prophylactic anticoagulation is allowed but therapeutic anticoagulation is a direct 
exclusion criteria.  
 

Protocol Rev #2: v01/29/2013 
 
14 
 
Use of concurrent investigational agents is not permitted. 
 
9.2 
Patient Discontinuation  
Patients may discontinue study treatment and sample collection at any time.  The primary 
reason for discontinuation should be recorded.  Reasons for discontinuation of a patient by 
the investigator include, but are not limited to, the following:   
 
Need for additional use of ketorolac or other NSAIDs over the outlined collection period 
 
Clinically significant deterioration of the patient’s condition during the study 
 
Patient noncompliance 
 
Peritoneal port failure 
 
Development if intra-abdominal complication 
 
Development of infectious morbidity 
 
Acute blood loss 
 
Investigator determination that it is not in the patient’s best interest to continue 
participation 
 
9.3 
Study Discontinuation 
It is unlikely that the trial itself will require a premature termination. However, the trial will be 
stopped prematurely (prior to completion of accrual, protocol therapy, and follow-up) for 
safety reasons. 
 
10.0  STUDY ASSESSMENTS 
All samples from the peritoneal cavity will be retrieved by the PI (Dr. Muller) or co-PI ‘s (Dr. 
Rutledge and Dr. Adams) gynecologic oncologists who are skilled in the access and 
management of newly placed intraperitoneal ports. The ports will ideally be accessed at the 
time of surgery and access maintained until completion of the study endpoint samples.   
 
10.1   Pre-treatment Sample Collection 
 
Collection of ascites cells at the time of tumor debulking 
 
Collection of peritoneal fluid and cells by the PI (Dr. Muller) or the co-PI’s (Dr. 
Rutledge and Dr. Adams) and blood prior to ketorolac administration.  
10.2  Post-treatment Sample Collection 
 
Collection of peritoneal fluid after ketorolac administration will be done at select time-
points after ketorolac administration (1 hour, 6 hours, and 24 hours) an additional 48 
hour sample (after the initial dose) will be done in patients having elective multiple 
ketorolac dosing.  
 
Cells and effluent will be separated from fluid at the above select time-points (1 hour, 
6 hours, 24 hours) and at 48 hours after the initial ketorolac dose in patients who 
receive elective multiple ketorolac dosing.  
 
10.3 
Pharmacokinetic Evaluation: 
Blood and 10-20 ccs of peritoneal fluid will be retrieved at the following time-points. Serum 
will be isolated and peritoneal fluid will be isolated from cellular material via centrifugation.  
Timepoint 
Time after ketorolac 
1 
Predose 
2 
1 hour 
3 
6 hours 
4 
24 hours 
5 
48 hours* 
* elective time-point for patients receiving clinically indicated multiple dosing of ketorolac 

Protocol Rev #2: v01/29/2013 
 
15 
 
10.3.1  Ketorolac Concentration in Peritoneal Fluid - HPLC 
Peritoneal concentrations of the R(-) and S(+) enantiomers of ketorolac will be determined 
by using published procedures for measuring plasma concentrations (44). High pressure 
liquid chromatography will be performed using UV detection at 313 nm. Standards will be 
prepared at 7 ketorolac plasma concentrations using 100 mcL steroid-free plasma to attain 
final concentrations of 0.025, 0.05, 0.10, 0.25, 0.5, 1.0, and 2.0 mcg/mL. Peritoneal samples 
(100 mcL) and standards will be added to silanized tubes containing the internal standard, 
loxapine HCL. Before extraction with 4 mL of methyl tert-butyl ether, proteins will be 
precipitated with 30 mcL of 10% weight to volume solution of trichloroacetic acid. Samples 
will be shaken for 10 min, centrifuged for 10 min, and the organic layer transferred to 
silanized culture tubes and evaporated under nitrogen. Mobile phase (50 mcL) will be 
added, the samples then will be transferred to autosampler vials, and 30 mcL will be injected 
onto the high pressure liquid chromatography column. Baseline separation of the isomers 
will be achieved using a chiral column (Astec Chirobiotic R (250 _ 4.6 mm2) obtained from 
Advanced Separation Technologies, Inc. Whippany, NJ) using a binary gradient mobile 
phase of 0.07 g ammonium formate and 24 mcL glacial acetic acid per liter methanol and 
increasing to 100% methanol for 17.5 min at a flow rate of 1.0 mL/min. Samples will be 
performed in duplicate.  
 
10.3.2 GTPase Inhibitory Activity of Peritoneal Fluid 
Rac1 activity will be measured in cells using effector binding assays (GLISA) that are 
commercially available (Cytoskeleton, Inc) as an independent measure that there is 
sufficient bioactive R-ketorolac in the peritoneal fluid to inhibit GTPase activity in cultured 
cells (as illustrated in Fig 3A, ketorolac panel). Because Rac1 inhibition is most robust we 
will focus on this GTPase. 3T3 cells will be cultured without serum in 6-well dishes in 
duplicate and subsequently incubated with 500mcL/well peritoneal fluid from each time-point 
for 1 hour prior to stimulation with 10ng/ml EGF for 2 minutes 3T3 cells will be harvested, 
lysed and assayed for activated Rac1 GTPase per manufacturer’s instructions. Results will 
be reported as ng Rac1-GTP normalized to +EGF drug samples. Controls will be included to 
monitor EGF responsiveness of Rac1 activation and ketorolac inhibition of Rac1 activation. 
Negative controls: peritoneal fluid prior to ketorolac treatment and cells without EGF 
stimulation. Positive controls: 1 ng purified Rac1-GTP; +EGF treated cells with serum in lieu 
of peritoneal fluid; cells treated with 100uM R-ketorolac or S-ketorolac for one hour as 
shown in Fig. 3A.  
 
 
10.3.3 Efficacy of Peritoneal Fluid containing Ketorolac in Inhibiting 
Cell Adhesion and Spreading in Vitro.   
The peritoneal fluid will be tested on immortalized cells in vitro as an alternate measure that 
there is sufficient bioactive R-ketorolac in the fluid to inhibit the adhesion/migration of 
immortalized mouse Swiss 3T3 or human ovarian cancer cells (OvCa429, SKOV or 
OvCa433) in culture.  The assay will utilize CYTOO micropatterned chips that are coated 
with fibronectin in defined patterns (144 micropatterns/sector with 12 sectors/chip) (Fig. 7).  
Immortalized cells will be mixed with 500 mcL/5x103 immortalized cells that have been  
 
 
 
 
 
 
 

Protocol Rev #2: v01/29/2013 
 
16 
 
 
trypsinized and washed with PBS+.  The mixture will be allowed to adhere to a CYTOO chip 
for 15-60 min.  Cells will be fixed directly, imaged and evaluated for cell number and cell 
shape on each of four micropatterns.  It is expected that the presence of R-ketorolac will 
reduce speed of cell adhesion and or cell spreading on the substrate.  Positive controls: 
peritoneal fluid prior to ketorolac treatment; cells in serum in lieu of peritoneal fluid; Negative 
controls: cells plated in the presence of 100 µM R-ketorolac or S-ketorolac for one hour 
prior.   
 
10.3.4. Spreading, Adhesion, and Migration Assays of Patient Derived 
Peritoneal Cells 
The impact of ketorolac treatment on residual ovarian tumor cell behavior will be assessed 
by measuring time for cell adhesion and spreading, as well as migration distance as follows.  
All of the assays require only small numbers of cells.  CYTOO chips (see 5.6.5 and Fig. 7) 
will serve as an alternate measure for assessing cell adhesion and spreading. Controls: 
patient cell adhesion prior to treatment; cell adhesion when incubated with 100 µM R-
ketorolac.   
 
10.3.4.1. Spreading: Cells (100-5,000/sample) will be rinsed in Dulbecco's 
PBS (DPBS) and plated on coverslips without or with extracellular matrix coating and 
images will be obtained at regular intervals. Spreading is defined as flattened cells 
without the refractile, fibroblastic morphology determined and expressed as a 
percentage of the total cells ± SD.  
 
10.3.4.2. Adhesion: Cells (100-5,000/well) will be plated onto 96 well plates 
without or with extracellular matrix coating and incubated for 30 min. Cells will be 
fixed and non-adherent cells will be removed by gentle washing. Colorimetric 
quantification of adherent cells will be performed using standard protocols.  
 
10.3.4.3 Migration: Cells (100-5,000/sample) will be placed on collagen-
coated coverslips in culture medium and migration will be measured by time lapse 
photography on a heated microscope stage (UNM Microscopy Facility). Distance 
traversed for individual cells will be quantified by image analysis software.   
 
 
10.3.5.  
PCR analysis of Patient Derived Peritoneal Cells  
Genes downstream of Rac1 and Cdc42 activation, which have been validated in cell culture 
and through in vivo studies (e.g. PAK, IQGAP, DOCK-180, CRK, WAVE3) will be tested by 
quantitative real time PCR using human genome predesigned and validated primer sets and 
assay kits from Origene (Rockville, MD).  Each assay will be performed in duplicate and 
ketorolac response will be established by comparison of pretreatment versus treatment 
samples.  A >2-fold change between 0 treatment and subsequent treatment time-points 
Fig. 7.  Cytoo micropatterened chips for analysis of cell adhesion and spreading.  Chips have 
arrays of adhesive patterns as illustrated in red that enable cells to adhere and spread into characteristic 
shapes.  The chips have 144 micropatterns/sector, there are four different shapes arrayed in three sizes 
(small, medium and large).  Thus, a single chip allows the analysis of >1700 cells.  We have tested cell 
shape changes in response to EGF stimulation +/- a Cdc42 inhibitor. Inhibition of Cdc42 caused cells to 
look like unstimulated control cells, and fail to adopt a migratory phenotype + EGF that was readily 
quantified using the chips (not shown).  

Protocol Rev #2: v01/29/2013 
 
17 
 
would be considered significant. This assay will be performed only when sufficient cell 
numbers are present in the peritoneal fluid (>10,000 cells) to allow purification of RNA.      
 
  
 
11.0 
STATISTICAL METHODS 
11.1 
Analysis of Demographic and Baseline Characteristics 
The analysis of demographic characteristics (age, gender, tobacco abuse history and 
ethnicity) and baseline characteristics, including weight change, performance status, and 
histologic subtype, will be primarily descriptive.  
 
11.2 
Endpoint Analyses 
11.2.1 Primary Endpoint and Sample Size 
Ketorolac availability, concentration and racemic mixture of Toradol® ([R,S]-ketorolac) in the 
peritoneal cavity. Intraperitoneal concentration and racemic mixture of ketorolac is currently 
unknown although serum pharmacokinetic data suggests a slower clearance of the R-
racemate. The primary objective of this study is to determine the concentration of 
measurable levels of R and S- Ketorolac (and the percent racemic mixture) in the peritoneal 
cavity after IV administration in the post-operative patient after cytoreductive surgery. Since 
this is a pilot trial for feasibility of obtaining the desired measures, a true sample size 
calculation is difficult to assess. We anticipate that we will need to recruit 60 patients pre-
operatively who meet the first phase of eligibility. We anticipate that 60% will have a 
diagnosis other than advanced stage invasive epithelial ovarian cancer, fail optimal 
debulking, have a clinical decision to not proceed with a port placement or not enough 
ascites or negative cytology of the ascites that will warrant no further participation in the 
study. This data is based on our initial experience with the first 12 patients enrolled on the 
trial. We then anticipate that 25 patients will have intraoperative port placement and be 
eligible for secondary eligibility. We anticipate that and additional 15-18 patients will either 
fail secondary eligibility, decline sample collection, decline ketorolac or have indications to 
withhold ketorolac, decline blood sampling, have inadequate re-accumulated fluid for 
sampling, or failed extraction of ascites samples from the port. The sample target remains 7-
10 complete sets of blood and ascites samples for primary and secondary endpoint 
analyses.  
 
Data will be analyzed as a repeated measure ANOVA followed by Tukey's post hoc analysis 
in order to evaluate whether there are significant differences between groups (pre-and post-
ketorolac treatment). We plan to group both dose groups together as we anticipate similar 
drug measures based on known prior serum pharmacokinetic studies. If we find a large 
discrepancy in peritoneal ketorolac concentrations between the 15mg and 30mg treated 
group, we may amend the protocol to increase sample size to evaluate the groups 
independently.  The analysis of demographic characteristics (age and ethnicity) and clinical 
characteristics, including histologic tumor type, and stage of disease will be descriptive. For 
the peritoneal distribution study, seven patients should be sufficient for statistically 
meaningful results based on a study of similar design to measure concentration of 
doxorubicin in ovarian ascites following IV administration (45). 
 
11.2.2 Secondary Endpoints  
We will determine the effect of IV ketorolac on cell adhesion and migration on ovarian 
cancer cells retrieved from the peritoneal cavity after cytoreductive surgery using several 
different biologic assays. Isolated cells will be tested for evidence of target inhibition and 
functional changes in adhesion and migration. Target activity assays: Rac1 and Cdc42 
activity will be measured in cells using effector binding assays (GLISA) that are 

Protocol Rev #2: v01/29/2013 
 
18 
 
commercially available (Cytoskeleton, Inc) and are used to determine activity of primary 
cells ex vivo as we have demonstrated in (Fig. 1C).  In addition, we will conduct qRT-PCR 
analysis of cells isolated pre and post ketorolac administration to test for expression of 
genes downstream of Rac1 and Cdc42 activation that were validated in cell culture and in 
vivo studies. Adhesive properties of cells will be measured by assays for spreading and 
adhesion. For spreading analysis, cells will be rinsed in Dulbecco's PBS (DPBS) and plated 
on coverslips without or with extracellular matrix coating and images will be obtained at 
regular intervals. Spreading is defined as flattened cells without the refractile, fibroblastic 
morphology determined and expressed as a percentage of the total cells ± SD as we’ve 
described previously (46). For adhesion, cells will be plated onto 96 well plates without or 
with extracellular matrix coating and incubated for 30 min. Cells will be fixed and non-
adherent cells will be removed by gentle washing. Colorimetric quantification of adherent 
cells will be performed using standard protocols. Migration will be measured after plating 
cells on collagen-coated coverslips in assessing migration by time lapse photography on a 
heated microscope stage (UNM Microscopy Facility). Distance traversed for individual cells 
will be quantified by image analysis software. Data will be analyzed as a repeated measure 
ANOVA followed by Tukey's post hoc analysis in order to evaluate whether there are 
significant differences between groups (pre-and post-ketorolac treatment). Trend analyses 
of response will also be performed based on peritoneal R-ketorolac concentrations.  
 
 
11.3 
Safety Analysis 
Patients will be examined at each time-point for subjective/objective evidence of developing 
drug toxicity according to NCI-CTC toxicity criteria or complications from the blood draws or 
port access 
 
12.0 
SOURCE DOCUMENTS AND CASE REPORT FORM COMPLETION 
 
12.1 
Source Documents 
Source documents are defined as original documents, data and records. This may include 
hospital records, clinical and office charts, laboratory data/information or evaluation 
checklists, pharmacy dispensing and other records recorded from automated instruments. 
Data collected during this study must be recorded on the appropriate source documents. 
 
 
12.2 
Case Report Forms  
Case report forms (CRF) must be completed for each subject enrolled in this study. The 
case report forms will contain the subjects unique indentifiers, demographic characteristics 
(age, gender, and ethnicity) and baseline characteristics, including vital signs, performance 
status, and histologic subtype, will be primarily descriptive The information on the case 
report forms will be collected and used for data analysis.  
 
Information obtained from subjects will be used by study staff and it will be shared with the 
sponsor of the study (representatives of the Department of Defense). The University of New 
Mexico Health Sciences Center Human Research Review Committee (HRRC) which 
oversees human subject research, the New Mexico Cancer Care Alliance, the UNM CC 
Clinical Trials Office, and the Food and Drug Administration and/or other entities that may 
have access will be permitted to access to records. 
 
Study records and specimens will be assigned a code number by the study team and the 
subjects will not be identified by name in the study records.  
 
Study records will be retained for 2 years following the date of study completion. 
 

Protocol Rev #2: v01/29/2013 
 
19 
 
13.0 
ASSESSMENT OF SAFETY 
The safety of this therapy will be assessed through collection and analyses of adverse 
events (AEs) and laboratory tests. 
  
13.1 
Specification of Safety Variables  
Safety assessments will consist of monitoring and recording protocol-defined adverse 
events (AEs) and serious adverse events (SAEs); measurement of clinically-specified 
hematology, clinical chemistry, and urinalysis variables; measurement of clinically-specified 
vital signs; and other clinically-specified tests that are deemed critical to the post-operative 
patient care plan. 
 
           13.1.1 Adverse Events 
An AE is any unfavorable and unintended sign, symptom, or disease temporally associated 
with the use of any medication or other protocol-imposed intervention, regardless of 
attribution. 
This includes the following: 
 
AEs not previously observed in the patient that emerge during the protocol-specified 
AE reporting period, including signs or symptoms associated with the malignancy 
that were not present prior to the AE reporting period  
 
Complications that occur as a result of protocol-mandated interventions  
 
If applicable, AEs that occur prior to assignment of study treatment associated with 
medication washout, no treatment run-in, or other protocol-mandated intervention 
 
Preexisting medical conditions (other than the condition being studied) judged by the 
investigator to have worsened in severity or frequency or changed in character 
during the protocol-specified AE reporting period 
 
Diagnoses and/or symptoms associated with malignancy should be reported as AEs 
if they worsen or change in character.  Clinical progression of malignancy should not 
be reported as an AE. 
 
           13.1.2 Serious Adverse Events 
An AE should be classified as an SAE if: 
 
It results in death (i.e., the AE actually causes or leads to death).  
 
It is life threatening (i.e., the AE, in the view of the investigator, places the patient at 
immediate risk of death.  It does not include an AE that, had it occurred in a more 
severe form, might have caused death.). 
 
It requires or prolongs inpatient hospitalization. 
 
It results in persistent or significant disability/incapacity (i.e., the AE results in 
substantial disruption of the patient’s ability to conduct normal life functions). 
 
It is considered a significant medical event by the investigator based on medical 
judgment (e.g., may jeopardize the patient or may require medical/surgical 
intervention to prevent one of the outcomes listed above). 
 
All AEs that do not meet any of the criteria for serious should be regarded as nonserious 
AEs. 
 
The terms “severe” and “serious” are not synonymous.  Severity (or intensity) refers to the 
grade of a specific AE, e.g., mild (Grade 1), moderate (Grade 2), or severe (Grade 3) 
myocardial infarction.  “Serious” is a regulatory definition (see previous definition) and is 
based on patient or event outcome or action criteria usually associated with events that 
pose a threat to a patient’s life or functioning.  Seriousness (not severity) serves as the 

Protocol Rev #2: v01/29/2013 
 
20 
 
guide for defining regulatory reporting obligations from the Sponsor to applicable regulatory 
authorities.  
 
Severity and seriousness should be independently assessed when recording AEs and 
SAEs. 
 
13.2 
Methods and Timing for Assessing and Recording Safety Variables 
The investigator is responsible for ensuring that all AEs and SAEs that are observed or 
reported during the study are recorded and reported to the sponsors (UNM CRTC) in 
accordance with protocol instructions. Since this protocol does not include an investigational 
agent, the SAE period will include the time of sample collection and 24 hours after the last 
collection at which time the port is de-accessed.  
 
           13.2.1 Assessment of Adverse Events 
Investigators will assess the occurrence of AEs and SAEs at all patient evaluation time 
points during the study.  All AEs and SAEs whether volunteered by the patient, discovered 
by study personnel during questioning, or detected through physical examination, laboratory 
test, or other means will be recorded in the patient’s medical record. 
 
Each recorded AE or SAE will be described by its duration (i.e., start and end dates), 
severity, regulatory seriousness criteria if applicable, suspected relationship to the 
investigational product (see following guidance), and actions taken. 
 
The AE grading (severity) scale found in the National Cancer Institute Common Terminology 
Criteria for Adverse Events (NCI-CTCAE), Version 3.0, will be used for AE reporting. 
 
Table 3 
Adverse Event Grading (Severity) Scale 
Grade 
Severity 
Alternate Description  
1 
Mild (apply event-specific 
NCI-CTCAE grading criteria) 
Transient or mild discomfort 
( 48 hours); no interference 
with the patient’s daily activities; 
no medical intervention/therapy 
required 
2 
Moderate (apply event-specific  
NCI-CTCAE grading criteria) 
Mild to moderate interference 
with the patient’s daily activities; 
no or minimal medical 
intervention/therapy required 
3 
Severe (apply event-specific  
NCI-CTCAE grading criteria) 
Considerable interference with 
the patient’s daily activities; 
medical intervention/therapy 
required; hospitalization 
possible 
4 
Very severe, life threatening,  
or disabling (apply event-specific 
NCI-CTCAE grading criteria) 
Extreme limitation in activity; 
significant medical 
intervention/therapy required, 
hospitalization probable 
5 
Death related to AE 
 
Note:  Regardless of severity, some events may also meet regulatory serious 
criteria.  Refer to definitions of an SAE. 
Use these alternative definitions for Grade 1, 2, 3, and 4 events when the 

Protocol Rev #2: v01/29/2013 
 
21 
 
observed or reported AE is not in the NCI-CTCAE listing. 
 
          13.2.2 Independent Research Monitoring 
The University of New Mexico Cancer Center (UNM CC) places a high priority on ensuring 
the safety of patients participating in clinical trials.  All institutional trials require monitoring 
commensurate with the degree of risk, as identified by the Protocol Review & Monitoring 
Committee.  The Independent Research Monitoring is the responsibility of the Data Safety 
Monitoring Committee, and for this protocol will be under the authority of the Vice-Chair of 
the DSMC is Elizabeth McGuire, MD, who is the independent research monitor.  The UNM 
CC Independent Research monitoring is a process for routine real time data monitoring and 
subject safety review of Investigator Initiated trials in accordance with the Essential 
Elements of the National Cancer Institute Guidelines, the FDA monitoring regulations, Good 
Clinical Practice Guidelines and functions under the requirements of the UNM CC Data 
Safety Monitoring Plan (DSMP), as approved by the National Cancer Institute on September 
26, 2011.  
 
The duties and responsibilities of the Independent Research Monitoring Committee and Vice 
Chair include: 
 
 
Monitoring the progress of the trial and the safety of participants 
 
Monitoring of subject eligibility 
 
Monitoring of consent and data quality 
 
Monitoring of study conduct/compliance (accruals, treatment, toxicity and study 
outcome) 
 
Plans for assuring compliance with requirements regarding the reporting of adverse 
events (AEs) 
 
Plans for assuring data accuracy and protocol compliance, including minimization of 
risks. 
 
Determines whether the trial should be suspended or terminated based on ongoing 
reviews of safety data 
 
Has the authority to remove individual human subjects from the research protocol 
and take whatever steps is necessary to protect the safety and well-being of human 
subjects until the IRB assesses the Independent Research Monitor’s report 
 
The study will be monitored continuously for the first three patients of the trial (and reported 
to the PI no later than the time the third consecutive patient completes the study). After the 
third patient completes the study, the Independent Research Monitor will submit a study 
report to the Data Safety Monitoring Committee and the local Human Research Protections 
Office (HRPO). After the first oversight review, the Data Safety Monitoring Committee, Vice-
Chair, Elizabeth McGuire, MD will designate the continued frequency with which the study 
will be monitored with no less than a quarterly review. 
 
14.0 
RESPONSIBILITIES OF THE PRINCIPAL INVESTIGATOR 
The Principal Investigator is agreeing to comply with GCP and local regulations and 
guidelines as summarized below. 
 
1. To conduct the study in accordance with the relevant, current protocol(s) and 
only make changes in a protocol after notifying the sponsor of the study (representatives of 
the Department of Defense), except when necessary to protect the safety, rights, or welfare 
of subjects. 
 

Protocol Rev #2: v01/29/2013 
 
22 
 
2. To personally conduct or supervise the described investigation(s). 
 
4. To report to adverse experiences that occur in the course of the investigation(s) in 
accordance with GCP and local law. 
 
5. To read and understand the information in the study drug package insert, including 
the potential risks and side effects of the drug. 
 
6. To ensure that all associates, colleagues, and employees assisting in the conduct of 
the study are informed about their obligations in meeting the above commitments 

Protocol Rev #2: v01/29/2013 
 
23 
 
References 
1. 
Parkin, D.M., Bray, F., Ferlay, J., Pisani, P. (2005) Global Cancer Statistics, 2002, CA 
Cancer J Clin 55:74-108. 
2. 
Jemal, A., Siegel, R., Xu, J., Ward, E. (2010) Cancer Statistics, 2010, CA Cancer J 
Clin 5: 277-300. 
3. 
Piccart, M. J., Bertelsen, K., Stuart, G., Cassidy, J., Mangioni, C., Simonsen, E., 
James, K., Kaye, S., Vergote, I., Blom, R., Grimshaw, R, Atkinson, R., Swenerton, K., 
Trope, C., Nardi, M., Kaern, J., Tumolo, S., Timmers, P., Roy, J. A., Lhoas, F, Lidvall, 
B., Bacon, M., Birt, A., Andersen, J., Zee, B., Paul, J., Pecorelli, S., Baron, B., 
McGuire, W. (2003) Long-term follow-up confirms a survival advantage of the 
paclitaxel-cisplatin regimen over the cyclophosphamide-cisplatin combination in 
advanced ovarian cancer, Int J Gynecol Cancer 13 Suppl 2:144-8. 
4. 
Ozols, R.F., Rubin, S.C., Thomas, G., et al.  Epithelial ovarian cancer, in Hoskins, 
W.Y., Perez, C.A., Young, R.C. (eds). (2005) Principles and Practice of Gynecologic 
Oncology, (ed 4) Philadelphia, PA, Lippincott Williams & Wilkins, 919-922. 
5. 
Vergote, I., Trope, C. G., Amant, F., Kristensen, G. B., Ehlen, T., Johnson, N., 
Verheijen, R. H., van der Burg, M. E., Lacave, A. J., Panici, P. B., Kenter, G. G., 
Casado, A., Mendiola, C., Coens, C., Verleye, L., Stuart, G. C., Pecorelli, S., Reed, N. 
S. (2010) Neoadjuvant chemotherapy or primary surgery in stage IIIC or IV ovarian 
cancer, N Engl J Med 10:943-53. 
6. 
Runowicz, C. D. (2008) Intraperitoneal chemotherapy in ovarian cancer: an update, 
Cancer J 1:7-9. 
7. 
Trimble, E. L., Christian, M. C. (2008) National Cancer Institute-United States strategy 
regarding intraperitoneal chemotherapy for ovarian cancer, Int J Gynecol Cancer 18 
Suppl 1: 26-8. 
8. 
Fujiwara, K., Aotani, E., Hamano, T., Nagao, S., Yoshikawa, H., Sugiyama, T., 
Kigawa, J., Aoki, D., Katsumata, N., Takeuchi, M., Suzuki, M. (2011) A randomized 
Phase II/III trial of 3 weekly intraperitoneal versus intravenous carboplatin in 
combination with intravenous weekly dose-dense paclitaxel for newly diagnosed 
ovarian, fallopian tube and primary peritoneal cancer, Jpn J Clin Oncol 2:278-82 
9. 
Barrena Medel, N. I., Wright, J. D., Herzog, T. J. (2010) Targeted therapies in 
epithelial ovarian cancer, J Oncol 2010:314326. 
10. 
Burger, R. A., Sill, M. W., Monk, B. J., Greer, B. E., Sorosky, J. I. (2007) Phase II trial 
of bevacizumab in persistent or recurrent epithelial ovarian cancer or primary 
peritoneal cancer: a Gynecologic Oncology Group Study, J Clin Oncol 33: 5165-71. 
11. 
Cannistra, S. A., Matulonis, U. A., Penson, R. T., Hambleton, J, Dupont, J., Mackey, 
H., Douglas, J., Burger, R. A., Armstrong, D., Wenham, R., McGuire, W. (2007) Phase 
II study of bevacizumab in patients with platinum-resistant ovarian cancer or peritoneal 
serous cancer, J Clin Oncol 33:5180-6. 
12. 
Garcia, A. A., Hirte, H., Fleming, G., Yang, D., Tsao-Wei, D. D., Roman, L., Groshen, 
S., Swenson, S., Markland, F., Gandara, D., Scudder, S., Morgan, R., Chen, H., Lenz, 
H. J., Oza, A. M. (2008) Phase II clinical trial of bevacizumab and low-dose 
metronomic oral cyclophosphamide in recurrent ovarian cancer: a trial of the 
California, Chicago, and Princess Margaret Hospital phase II consortia, J Clin Oncol 
26:76-82. 

Protocol Rev #2: v01/29/2013 
 
24 
 
13. 
Tillmanns, T.D., Lowe, M.P., Schwartzberg, L.S., Walker, M.S., Stepanski, E.J.  
(2010) A phase II study of bevacizumab with nab-paclitaxel in patients with recurrent, 
platinumresistant primary epithelial ovarian or primary peritoneal carcinoma, J Clin 
Oncol 28, 15 suppl, Abstract 5009. 
14. 
Penson, R. T., Dizon, D. S., Cannistra, S. A., Roche, M. R., Krasner, C. N., Berlin, S. 
T., Horowitz, N. S., Disilvestro, P. A., Matulonis, U. A., Lee, H., King, M. A., Campos, 
S. M. (2010) Phase II study of carboplatin, paclitaxel, and bevacizumab with 
maintenance bevacizumab as first-line chemotherapy for advanced mullerian tumors, J 
Clin Oncol 1: 154-9. 
15. 
Burger, R.A., Brady, M.F., Bookman, J.L., Walker, H.D., Homesley, J., Fowler, B.J., 
Monk, B.E., Greer, M., Liang, S.X. (2010) Phase III trial of bevacizumab (BEV) in the 
primary treatment of advanced epithelial ovarian cancer (EOC), primary peritoneal 
cancer (PPC), or fallopian tube cancer (FTC): A Gynecologic Oncology Group study, J 
Clin Oncol 28, 18 suppl, abstr LBA1. 
16. 
Kristensen, T., Perrem, T., Qian, W., Pfisterer, J., Ledermann, A., Joly, F., Carey, 
M.S., Beale, P.J., Cervantes, A., Oza, A.M. (2011) Results of interim analysis of 
overall survival in the GCIG ICON7 phase III randomized trial of bevacizumab in 
women with newly diagnosed ovarian cancer, J Clin Oncol 29: 18 suppl, abstr 
LBA5006. 
17. 
Aghajanian, C., Finkler, N.J.,  Rutherford, T., Smith, D.A., Yi, J., Parmar, H., Nycum, 
L.R., Sovak, M.A. (2011) OCEANS: A randomized, double-blinded, placebo controlled 
phase III trial of chemotherapy with or without bevacizumab (BEV) in patients with 
platinum-sensitive recurrent epithelial ovarian (EOC), primary peritoneal (PPC), or 
fallopian tube cancer (FTC), J Clin Oncology 29, 18 suppl, abstr LBA5007. 
18. 
Cohn, D. E., Kim, K. H., Resnick, K. E., O'Malley, D. M., Straughn, J. M., Jr. (2011) At 
what cost does a potential survival advantage of bevacizumab make sense for the 
primary treatment of ovarian cancer? A cost-effectiveness analysis, J Clin Oncol 29: 
1247-51. 
19. 
Surviladze, Z., Waller, A., Wu, Y., Romero, E., Edwards, B. S., Wandinger-Ness, A., 
and Sklar, L. A. (2010) Identification of a small GTPase inhibitor using a high-
throughput flow cytometry bead-based multiplex assay, J Biomol Screen 15:10-20. 
20. 
Agola, J. O., Surviladze, Z., Buranda, T., Ursu, O., Hong, L., Waller, A., Strouse, J. J., 
Simpson, D. S., Schroeder, C. E., Golden, J. E., Oprea, T. I., Sklar, L. A., and 
Wandinger-Ness, A. (2011) A Competitive Nucleotide Binding Inhibitor: In vitro 
Characterization of Rab7 GTPase Inhibition, ACS Chemical Biology (Submitted)  
21. 
Jaffe, A. B., and Hall, A. (2005) Rho GTPases: biochemistry and biology, Annu Rev 
Cell Dev Biol 21:247-269. 
22. 
Karlsson, R., Pedersen, E. D., Wang, Z., and Brakebusch, C. (2009) Rho GTPase 
function in tumorigenesis, Biochim Biophys Acta 1796: 91-98. 
23. 
Ridley, A. J. (2006) Rho GTPases and actin dynamics in membrane protrusions and 
vesicle trafficking, Trends Cell Biol 16: 522-529. 
24. 
Vega, F. M., and Ridley, A. J. (2008) Rho GTPases in cancer cell biology, FEBS Lett 
582:2093-2101. 

Protocol Rev #2: v01/29/2013 
 
25 
 
25. 
Wennerberg, K., Rossman, K. L., and Der, C. J. (2005) The Ras superfamily at a 
glance, J Cell Sci 118: 843-846. 
26. 
Fritz, G., and Kaina, B. (2006) Rho GTPases: promising cellular targets for novel 
anticancer drugs, Curr Cancer Drug Targets 6:1-14. 
27. 
Ferri, N., Corsini, A., Bottino, P., Clerici, F., and Contini, A. (2009) Virtual screening 
approach for the identification of new Rac1 inhibitors, J Med Chem 52:4087-4090. 
28. 
Nassar, N., Cancelas, J., Zheng, J., Williams, D. A., and Zheng, Y. (2006) Structure-
function based design of small molecule inhibitors targeting Rho family GTPases, Curr 
Top Med Chem 6: 1109-1116. 
29. 
Cohen, A., and Basch, C. (1988) Steady state pharmacokinetics of naproxen in young 
and elderly healthy volunteers, Semin Arthritis Rheum 17: 7-11. 
30. 
Furst, D. E., Sarkissian, E., Blocka, K., Cassell, S., Dromgoole, S., Harris, E. R., 
Hirschberg, J. M., Josephson, N., and Paulus, H. E. (1987) Serum concentrations of 
salicylate and naproxen during concurrent therapy in patients with rheumatoid arthritis, 
Arthritis Rheum 30:1157-1161. 
31. 
Handley, D. A., Cervoni, P., McCray, J. E., and McCullough, J. R. (1998) Preclinical 
enantioselective pharmacology of (R)- and (S)- ketorolac, J Clin Pharmacol 38:25S-
35S. 
32. 
Jett, M. F., Ramesha, C. S., Brown, C. D., Chiu, S., Emmett, C., Voronin, T., Sun, T., 
O'Yang, C., Hunter, J. C., Eglen, R. M., and Johnson, R. M. (1999) Characterization of 
the analgesic and anti-inflammatory activities of ketorolac and its enantiomers in the 
rat, J Pharmacol Exp Ther 288:1288-1297. 
33. 
Buckley, M. M., and Brogden, R. N. (1990) Ketorolac. A review of its 
pharmacodynamic and pharmacokinetic properties, and therapeutic potential, Drugs 
39:86-109. 
34. 
Mroszczak, E. J., Jung, D., Yee, J., Bynum, L., Sevelius, H., and Massey, I. (1990) 
Ketorolac tromethamine pharmacokinetics and metabolism after intravenous, 
intramuscular, and oral administration in humans and animals, Pharmacotherapy 10: 
33S-39S. 
35. 
Pallapies, D., Peskar, B. A., Brune, K., and Geisslinger, G. (1994) Effects on platelet 
functions and pharmacokinetics of azapropazone and ketorolac tromethamine given as 
single parenteral doses, Br J Clin Pharmacol 37:335-339. 
36. 
Pallapies, D., Salinger, A., Meyer zum Gottesberge, A., Atkins, D. J., Rohleder, G., 
Nagyivanyi, P., and Peskar, B. A. (1995) Effects of lysine clonixinate and ketorolac 
tromethamine on prostanoid release from various rat organs incubated ex vivo, Life Sci 
57: 83-89. 
37. 
Kean, W. F., Lock, C. J., Rischke, J., Butt, R., Buchanan, W. W., and Howard-Lock, H. 
(1989) Effect of R and S enantiomers of naproxen on aggregation and thromboxane 
production in human platelets, J Pharm Sci 78:324-327. 
38. 
Hinz, B., Cheremina, O., Besz, D., Zlotnick, S., and Brune, K. (2008) Impact of 
naproxen sodium at over-the-counter doses on cyclooxygenase isoforms in human 
volunteers, Int J Clin Pharmacol Ther 46:180-186. 
39. 
Huntjens, D. R., Spalding, D. J., Danhof, M., and Della Pasqua, O. E. (2006) 
Correlation between in vitro and in vivo concentration-effect relationships of naproxen 
in rats and healthy volunteers, Br J Pharmacol 148:396-404. 

Protocol Rev #2: v01/29/2013 
 
26 
 
40. 
Kean, W. F., Lock, C. J., and Howard-Lock, H. E. (1991) Chirality in antirheumatic 
drugs, Lancet 338:1565-1568. 
41. 
Mroszczak, E., Combs, D., Chaplin, M., Tsina, I., Tarnowski, T., Rocha, C., Tam, Y., 
Boyd, A., Young, J., and Depass, L. (1996) Chiral kinetics and dynamics of ketorolac, 
J Clin Pharmacol 36:521-539. 
42. 
Fisher, K. E., Sacharidou, A., Stratman, A. N., Mayo, A. M., Fisher, S. B., Mahan, R. 
D., Davis, M. J., and Davis, G. E. (2009) MT1-MMP- and Cdc42-dependent signaling 
co-regulate cell invasion and tunnel formation in 3D collagen matrices, J Cell Sci 122: 
4558-4569. 
43.  Machesky, L.M. (2008) Lamellipodia and filopodia in metastasis and invasion, FEBS 
Lett 582:2102-2111. 
44. 
Lynn Am, Bradford H, Kantor ED, Kok-Yong S, Salinger DH, Chen J, Ellenbogen RG, 
Vicini P, Anderson GD. (2007)  Postoperative ketorolac tromethaminine use in infants 
aged 6-18 months: the effect of morphine usage, safety assessment and stero-specific 
pharmacokinentics. Anesthesia and Analgesia 104:1040-1051. 
45. 
Gotlieb WH, Bruchim I, Ben-Baruch G, Davidson B, Zeltser A, Andersen A, Olsen H. 
(2007) Doxorubicin levels in serum and ascites of patients with ovarian cancer. EJSO 
33:213-215.   
46. 
Ning Y, Zeineldin R, Liu Y, Rosenberg M, Stack MS, Hudson LG. (2005) Down-
regulation of integrin alpha2 surface expression by mutant epidermal growth factor 
receptor (EGFRviii) induces aberrant cell spreading and focal adhesion formation. 
Cancer Rse 65:9280-9286.