Clinical trial NCT03253679
Conditions: Advanced Malignant Solid Neoplasm Refractory Malignant Solid Neoplasm
Sponsor: National Cancer Institute (NCI)
Start date: 2019-01-16; Completion date: 2022-10-31
Enrollment: 31 participants; Allocation: NA; Intervention model: SINGLE_GROUP

Brief summary:
This phase II trial studies how well AZD1775 works in treating patients with solid tumors with CCNE1 amplification that have spread to other places in the body (advanced) and do not respond to treatment (refractory). AZD1775 may stop the growth of tumor cells by blocking some of the enzymes needed for cell growth.

Detailed description:
PRIMARY OBJECTIVE:

I. To evaluate the proportion of patients with the objective response rate (ORR) to adavosertib (AZD1775) in patients with advanced refractory cancers with CCNE1 amplification.

SECONDARY OBJECTIVES:

I. To evaluate the proportion of patients alive and progression free at 6 months of treatment with AZD1775 in patients with advanced refractory cancers with CCNE1 amplification.

II. To evaluate proportion of patients with extended time to progression (time to progression on AZD1775/ time to progression on last line of therapy \>= 1.3).

III. To evaluate time until death or disease progression. IV. To identify potential predictive biomarkers beyond the genomic alteration by which treatment is assigned or resistance mechanisms using additional genomic, ribonucleic acid (RNA), protein and imaging-based assessment platforms.

OUTLINE:

Patients receive adavosertib orally (PO) once daily (QD) on days 1-5 and 8-12. Cycles repeat every 21 days in the absence of disease progression or unacceptable toxicity.

After completion of study treatment, patients are followed up for 4 weeks.

Arms: arm 1: Patients receive adavosertib PO QD on days 1-5 and 8-12. Cycles repeat every 21 days in the absence of disease progression or unacceptable toxicity.
Interventions: intervention 1: Adavosertib
Intervention descriptions: intervention 1: Given PO

Protocol documents:
Prot_SAP_000.pdf: 
NCI Protocol #: 10136 
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NCI Protocol #: 10136 
Local Protocol #: NCI10136 
 
ClinicalTrials.gov Identifier:  TBD 
 
TITLE:  A PHASE 2 STUDY OF AZD1775, A WEE1 INHIBITOR, IN PATIENTS 
WITH CCNE1 AMPLIFICATION  
 
Corresponding Organization: LAO-TX035 / University of Texas MD Anderson Cancer Center 
LAO   
 
Principal Investigator: 
Siqing Fu, MD, PhD 
University of Texas MD Anderson Cancer Center  
1515 Holcombe Blvd. Unit 455 
Houston, TX 77030 
Direct Office: 713-792-4318 
Fax: 713-745-3855 
siqingfu@mdanderson.org 
 
Participating Organizations (Only the participating LAOs should be listed.) 
LAO-11030 / University Health Network Princess Margaret Cancer Center LAO 
LAO-CA043 / City of Hope Comprehensive Cancer Center LAO 
LAO-CT018 / Yale University Cancer Center LAO 
LAO-MA036 / Dana-Farber - Harvard Cancer Center LAO 
LAO-MD017 / JHU Sidney Kimmel Comprehensive Cancer Center LAO 
LAO-MN026 / Mayo Clinic Cancer Center LAO 
LAO-NC010 / Duke University - Duke Cancer Institute LAO 
LAO-NJ066 / Rutgers University - Cancer Institute of New Jersey LAO 
LAO-OH007 / Ohio State University Comprehensive Cancer Center LAO 
LAO-PA015 / University of Pittsburgh Cancer Institute LAO 
LAO-TX035 / University of Texas MD Anderson Cancer Center LAO 
LAO-NCI / National Cancer Institute LAO 
 
 
 
 


NCI Protocol #: 10136 
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Statistician: 
 
  Study Regulatory Coordinator: 
Kenneth Hess, PhD 
  
 
Katherine Torres, MHA 
1515 Holcombe Blvd., Unit 1411 
1515 Holcombe Blvd., Unit 455 
Houston, Texas 77030 
Houston, Texas 77030 
713-794-4168 
832-750-4997 
khess@mdanderson.org 
 Ktorres4@mdanderson.org 
 
Responsible Research Nurse: 
        Responsible Data Manager: 
Valerie Marcott, RN 
Naiyi Shi, MS 
1515 Holcombe Blvd., Unit 455 
1515 Holcombe Blvd., Unit 455 
Houston, Texas 77030 
Houston, Texas 77030 
713-794-1014 
713-794-1897 
VDMarcot@mdanderson.org 
nishi@mdanderson.org 
 
Translational Investigators: 
      
Khandan Keyomarsi, PhD 
Funda Meric-Bernstam, MD 
1515 Holcombe Blvd., Z6.3012 
1515 Holcombe Blvd., Unit 455 
Houston, Texas 77030 
Houston, Texas 77030 
713-792-4845 
713-794-1226 
kkeyomar@mdanderson.org 
fmeric@mdanderson.org 
 
 
 
NCI-Supplied Agent(s):  AZD1775 (Adavosertib) NSC 751084, AstraZeneca Pharmaceuticals 
IND #:  
IND Sponsor: DCTD, NCI 
 
 
Protocol Type / Version # / Version Date:     Original/version date: 06/24/2017 
                                                                             Revision/version date: 09/14/2017 
Revision/version date: 10/12/2017  
Revision/version date: 11/20/2017 
Revision/version date: 02/01/2018 
Revision/version date: 03/02/2018 
Revision/version date: 03/30/2018 
Revision/version date: 04/26/2018 
Revision/version date: 06/12/2018 
Revision/version date: 07/05/2018 
Revision/version date: 07/18/2018 
Revision/version date: 08/17/2018 
Revision/version date: 01/22/2019 
Revision/version date: 06/21/2019 
Revision/version date: 08/21/2019 
Revision/version date: 06/02/2020


NCI Protocol #: 10136 
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SCHEMA 
 
This is an open-label phase II study of AZD1775, a Wee1 kinase inhibitor in patients with 
advanced solid tumors harboring CCNE1 amplification. This study follows a two-stage design to 
assess anti-tumor activity according to RECIST version 1.1, and toxicity profile according to 
NCI-CTCAE 5.0.  
 
The primary endpoint is the objective response rate, and the secondary endpoints are safety, 
tolerability, and survivals. As shown in Figure 1, patients will be treated with AZD1775 at 300 
mg PO daily for 5 days each week for 2 weeks (one cycle of therapy is 21 days). Blood and 
tumor specimens will be collected for translational research.  
 
 
Figure 1. Study Schema 
 
 


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TABLE OF CONTENTS 
 
TITLE:  A Phase 2 Study of AZD1775, A Wee1 Inhibitor, in Patients with CCNE1 
Amplification .................................................................................................................................1 
SCHEMA .........................................................................................................................................3 
TABLE OF CONTENTS .................................................................................................................4 
1. 
OBJECTIVES ......................................................................................................................7 
1.1 
Primary Objectives ...................................................................................................7 
1.2 
Secondary Objectives ...............................................................................................7 
2. 
BACKGROUND .................................................................................................................8 
2.1 
Study Disease(s).......................................................................................................8 
2.2 
AZD1775: an oral, potent Wee1 inhibitor ...............................................................9 
2.3 
Clinical experience with AZD1775 .......................................................................10 
2.4 
AZD1775 Pharmacokinetics ..................................................................................13 
2.5 
AZD1775 Safety ....................................................................................................15 
2.6 
Cyclin E encoded by CCNE1 is a critical G1/S transition regulator .....................16 
2.7 
CCNE1 Amplification Sensitizes Cancer Cells to AZD1775 ................................18 
2.8 
The Feasibility to Conduct a Phase II Basket Study in CCNE1 Amplified 
Solid Tumors ..........................................................................................................21 
2.9 
Correlative Studies – Background and Rationale ..................................................24 
3.   PATIENT SELECTION ..........................................................................................................28 
3.1 
Patient Inclusion Criteria .......................................................................................28 
3.2 
Patient Exclusion Criteria ......................................................................................30 
3.3 
Inclusion of Women and Minorities ......................................................................32 
4.   REGISTRATION PROCEDURES .........................................................................................33 
4.1 
Investigator and Research Associate Registration with CTEP ..............................33 
4.2 
Site Registration .....................................................................................................34 
4.3 
Patient Registration ................................................................................................36 
4.4 
General Guidelines .................................................................................................37 
5. 
TREATMENT PLAN ........................................................................................................38 
5.1 Agent Administration...................................................................................................38 
5.2 Study Procedures .........................................................................................................38 
5.3 General Concomitant Medication and Supportive Care Guidelines ............................41 
5.4 Restrictions during the study .......................................................................................42 
5.5 Duration of Therapy .....................................................................................................44 
5.6 Duration of Follow Up .................................................................................................46 
5.7 Criteria for Removal from Study .................................................................................47 


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6. 
DOSING DELAYS/DOSE MODIFICATIONS  The principal investigator is responsible 
for ensuring that all staff involved in the study is familiar with the content of this section. .......48 
6.1  Definition of adverse events .......................................................................................48 
6.2 
Definitions of serious adverse event ......................................................................48 
6.3 
Dose Modification .................................................................................................49 
6.4 
Dose modifications due to hematologic toxicity ...................................................50 
6.5 
Dose modifications due to non-hematologic toxicity ............................................51 
6.6 
Recording of adverse events ..................................................................................54 
7. 
ADVERSE EVENTS:  LIST AND REPORTING REQUIREMENTS ............................60 
7.1 
Comprehensive Adverse Events and Potential Risks List (CAEPRs) ...................60 
7.1.1 CAEPR for AZD1775 ............................................................................................60 
7.2 
Adverse Event Characteristics ...............................................................................63 
7.3 
Expedited Adverse Event Reporting ......................................................................65 
7.4 
Routine Adverse Event Reporting .........................................................................67 
7.5 
Secondary Malignancy ...........................................................................................67 
7.6 
Second Malignancy ................................................................................................67 
8. 
PHARMACEUTICAL INFORMATION ..........................................................................68 
8.1 
AZD1775 (NSC #751084) .....................................................................................68 
8.2 
Agent Ordering and Agent Accountability ............................................................70 
8.3 
Investigator Brochure Availability ........................................................................70 
8.4 
Useful Links and Contacts .....................................................................................70 
9. 
Biomarker, correlatives and Special studies ......................................................................72 
9.1 
Collection of Specimens ........................................................................................72 
9.2   Shipping Information .................................................................................................74 
10. 
STUDY CALENDAR .......................................................................................................75 
11. 
MEASUREMENT OF EFFECT........................................................................................76 
11.1 
Definitions..............................................................................................................77 
11.2 
Disease Parameters ................................................................................................77 
11.3 
Methods for Evaluation of Measurable Disease ....................................................79 
11.4 
Response Criteria ...................................................................................................82 
12. 
STUDY OVERSIGHT AND DATA REPORTING / REGULATORY 
REQUIREMENTS .......................................................................................................................85 
12.1 
Study Oversight .....................................................................................................85 
12.2 
Data Reporting .......................................................................................................86 
12.3 
CTEP Multicenter Guidelines ................................................................................87 
12.4 
Collaborative Agreements Language .....................................................................87 
12.5 
Genomic Data Sharing Plan ...................................................................................89 
12.6 
Safety Monitoring ..................................................................................................90 
12.7 
Data Safety Monitoring Plan .................................................................................90 


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13 
STATISTICAL CONSIDERATIONS...............................................................................91 
13.1 
Study Design/Endpoints.........................................................................................91 
13.2 
Sample Size/Accrual Rate ......................................................................................92 
13.3 
Stratification Factors ..............................................................................................92 
13.4 
Analysis of Secondary Endpoints ..........................................................................92 
13.5 
Stopping Rules .......................................................................................................94 
REFERENCES ..............................................................................................................................95 
APPENDIX A: PERFORMANCE STATUS CRITERIA .............................................................98 
APPENDIX B: PATIENT DRUG INFORMATION HANDOUT AND WALLET CARD ........99 
APPENDIX C: MEDICATION DIARY .....................................................................................101 
TABLE OF CONTENTS .............................................................................................................105 
1.0 
PURPOSE ............................................................................................................106 
2.0 
SCOPE .................................................................................................................106 
3.0 
ABBREVIATIONS .............................................................................................106 
4.0 
INTRODUCTION ...............................................................................................106 
5.0 
ROLES AND RESPONSIBILITIES ...................................................................106 
6.0 
MATERIALS AND EQUIPMENT REQUIRED ................................................107 
7.0 
OPERATING PROCEDURES ............................................................................108 
7.4 
Tumor Needle Biopsy Collection and Handling ..................................................108 
8.0 
SHIP TO FNLCR FOR ANALYSIS (OPTIONAL) ...........................................110 
8.6 
Day of Shipment ..................................................................................................110 
APPENDIX 1: BATCH RECORD ..............................................................................................112 
APPENDIX 2: PD SAMPLE SHIPPING MANIFEST ...............................................................114 
 
 
 
 


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1. OBJECTIVES 
 
1.1   Primary Objectives 
•   To evaluate the proportion of patients with the objective response rate (ORR) to AZD1775 in 
patients with advanced refractory cancers with CCNE1 amplification.   
 
 
1.2   Secondary Objectives 
 
•   To evaluate the proportion of patients alive and progression free at 6 months of treatment with 
AZD1775 in patients with advanced refractory cancers with CCNE1 amplification.   
 
•   To evaluate proportion of patients with extended time to progression (time to progression on 
AZD1775/ time to progression on last line of therapy ≥ 1.3). 
 
•   To evaluate time until death or disease progression.  
 
•   To identify potential predictive biomarkers beyond the genomic alteration by which treatment 
is assigned or resistance mechanisms using additional genomic, RNA, protein and imaging-based 
assessment platforms.  
 
 
 


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2. BACKGROUND 
2.1   Study Disease(s) 
 
The study diseases include all histologically advanced solid tumors harboring CCNE1 
amplification. 
 
 
 


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2.2   AZD1775: an oral, potent Wee1 inhibitor 
 
AZD1775 is a potent, highly selective, ATP-competitive, small-molecule inhibitor of Wee1 
kinase, displaying in vitro and in vivo preferential sensitization of tumor cells (relative to normal 
cells) to various DNA-damaging anticancer agents and modalities while sparing normal tissues 
from radiation- and chemotherapy-induced toxicity [1, 2]. Wee1 phosphorylates and inhibits 
cyclin-dependent kinases 1 (CDK1) and 2 (CDK2), and is involved in regulation of the intra-S 
and G2 cell cycle checkpoints [3]. Proper functioning of these checkpoints is essential for the 
DNA metabolism and the DNA damage response. The gene expression profile induced by Wee1 
inhibition revealed dose-dependent inhibition of direct substrate pCDK1 and changes of genes 
regulating S-G2 cell cycle progression or checkpoints, which are consistent with the mode of 
action of Wee1 inhibitors [4]. Inhibition of WEE1 is expected to release a tumor cell from DNA 
damage-induced arrest at the G2/M boundary, so that un-repaired DNA damage may be taken 
into mitosis (M-phase). Since cancer cells exhibit higher levels of endogenous damage than 
normal cells, as well as exhibiting loss of one or more DNA damage response capabilities, this is 
expected to preferentially enhance cancer cell death through mitotic catastrophe compared to 
normal cells. Also, inhibition of WEE1 is expected to cause aberrantly high CDK2 activity in S-
phase cells that will have multiple effects for cancer cells that already have G1/S checkpoint 
aberrations. These include de-regulated replication origin firing before sufficient 
deoxynucleotides are available, resulting in a higher degree of replication stress and ultimately 
replication fork collapse where endonucleases generate DNA damage, which leads to apoptosis. 
 
CDK1 (also called cell division cycle 2, or CDC2) activity drives a cell from the G2 phase of the 
cell cycle into mitosis. In response to DNA damage, WEE1 inhibits CDK1 to prevent the cell 
from dividing until the damaged DNA is repaired (G2 checkpoint arrest). Inhibition of WEE1 is 
expected to release a tumor cell from DNA damage-induced arrest at the G2/M boundary, so that 
un-repaired DNA damage may be taken into mitosis (M-phase). Since cancer cells exhibit higher 
levels of endogenous damage than normal cells, as well as exhibiting loss of one or more DNA 
damage response (DDR) capabilities, this is expected to preferentially enhance cancer cell death 
through mitotic catastrophe compared to normal cells. In vitro experiments demonstrate that 
AZD1775 has synergistic cytotoxic effects when administered in combination with various DNA 
damaging agents that have divergent mechanisms of action. Therefore, the clinical development 
of AZD1775 includes its use as a chemosensitizing drug in combination with a cytotoxic agent 
(or combination of agents) for treatment of advanced solid tumors. 
 
CDK2 activity drives a cell into, and through the DNA synthesis (S-phase) of the cell cycle in 
which the genome is duplicated in preparation for cell division. An important aspect of CDK2 
regulation of replication is the control of replication origin firing. WEE1, under normal 
circumstances, regulates CDK2 to provide sufficient time to generate the optimum environment 
for DNA synthesis, such as the generation of a sufficient pool of deoxynucleotide triphosphates 
(dNTPs), the building blocks of DNA. Inhibition of WEE1 is expected to cause aberrantly high 
CDK2 activity in S-phase cells that will have multiple consequences for cancer cells that already 
have G1/S checkpoint aberrations such as p53 mutations or CDKN2A deletions [5]. These 
include de-regulated replication origin firing before sufficient dNTPs are available, resulting in a 
higher degree of replication stress (already greater in cancer cells). The resulting increase in 
levels of replication stress will result in replication fork stalling, the generation of unstable DNA 


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replication structures and ultimately replication fork collapse where endonucleases generate 
DNA damage. Therefore, it is anticipated that AZD1775 will have independent anti-tumor 
activity in the absence of added chemotherapy, particularly in cancer cells that already have 
significantly higher levels of replication stress.  
 
In sarcoma cell lines, AZD1775 treatment at clinically relevant concentrations led to unchecked 
entry into mitosis associated with increased phosphorylated CDK1Tyr-15 and histone 3 (pH3Ser-10); 
DNA damage associated with phosphorylated -H2AXSer-139; and initiation of apoptosis 
associated with cleaved caspase-3 [6]. Clonogenic survival and tumor xenograft analyses showed 
that AZD1775 abrogated radiation-induced G2 blockade by accelerating the progression of cells 
harboring radiation-induced DNA lesions into premature mitosis and inducing apoptosis in p53-
defective tumor cells but not in p53-wt cell lines [7]. AZD1775 selectively sensitizes p53-
deficient tumor cells to DNA-damaging agents such as gemcitabine, carboplatin, cisplatin, 
pemetrexed, doxorubicin, camptothecin, mitomycin C, and 5-fluorouracil [8-11]. Further 
investigation showed that AZD1775 displayed single-agent activity and enhanced the response 
of HPV-positive HNSCC to cisplatin both in vitro and in vivo, which was associated with G2 
checkpoint abrogation, persistent DNA damage, and apoptosis induction through decreased 
expression of the anti-apoptotic proteins MCL-1 and XIAP [12]. The therapeutic effects of 
AZD1775 as single agent or combination with other agents are actively being evaluated in more 
than 30 clinical trials across multiple tumor types. A phase I study of 25 patients with a 
refractory solid tumor showed that single-agent AZD1775 can be safely administered as 225 mg 
twice per day orally over 2.5 days per week for 2 weeks every 21 days, and confirmed partial 
responses were observed in two patients carrying a BRCA mutation (one with head and neck 
cancer and one with ovarian cancer), providing preliminary clinical evidence of its preferential 
cytotoxicity to DNA damage repair–deficient cancer cells [13].  
 
In vitro experiments demonstrated that AZD1775 inhibits WEE1 activity and induces DNA 
damage as well as G2/M checkpoint escape. AZD1775 sensitizes tumor cells to cytotoxic effects 
of different DNA damaging agents, including chemotherapies. In mouse and rat xenograft 
models, anti-tumor efficacy of chemotherapy was significantly enhanced by AZD1775.  
Treatment with AZD1775 in nude mouse xenograft models resulted in significant tumor growth 
inhibition at tolerated doses, and also enhanced the anti-tumor growth effect of gemcitabine, 
carboplatin, radiation therapy, and olaparib, a PARP inhibitor that also induces replication 
associated DNA damage. Moreover, in preclinical cancer cell models associated with high levels 
of endogenous replication stress resulting from a combination of G1/S checkpoint deficiencies 
due to p53 mutations or CDKN2A deletions and the over-expression of oncogenic drivers such 
as MYC, mutant KRAS or the amplification of Cyclin E, AZD1775 also demonstrated 
significant single-agent anti-tumor activity. 
 
2.3   Clinical experience with AZD1775 
 
AZD1775 has been administered to patients in 12 AstraZeneca-sponsored or Merck-sponsored 
clinical studies, 6 of which are ongoing. As of 11 November 2016, a total of approximately 551 
patients have been exposed to AZD1775 in AstraZeneca-sponsored or Merck-sponsored clinical 
studies. Of these 551 patients, 103 received AZD1775 monotherapy, 407 patients received 
AZD1775 in combination with cytotoxic chemotherapy agents and the remaining 41 patients 


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received AZD1775 in combination with targeted therapies MEDI4736 or olaparib.   
 
These patients have received single doses per cycle as high as 1300 mg of AZD1775 as 
monotherapy, 325 mg of AZD1775 in a single-dose in combination with chemotherapy, and 325 
mg twice a day (BID) in a multiple-dose regimen in combination with chemotherapy. In 
addition, approximately 350 patients have also received AZD1775 as part of externally-
sponsored scientific research.   
 
The completed or terminated early studies include: 
 
• PN001 (NCT00648648) (except for Part 3): a first-time-in-patients (FTIP), Phase I, dose-
escalation study evaluating AZD1775 both as monotherapy and combination therapy with 
gemcitabine, cisplatin, or carboplatin in adult patients with advanced solid tumors. 
 
• PN004 (NCT01357161): a Phase II study evaluating AZD1775 combined with carboplatin and 
paclitaxel in patients with platinum-sensitive p53-mutant ovarian cancer 
 
• PN005 (NCT01047007): a Phase I, dose-escalation study evaluating AZD1775 as monotherapy 
(Part 1), combination therapy with 5-FU (Part 2), and combination therapy with 5 FU plus 
cisplatin (Part 3) in adult Japanese patients with advanced solid tumors was terminated early due 
to portfolio prioritization in oncology at Merck after 3 patients had been enrolled in Part 1 and 8 
patients had been enrolled in Part 2. Part 3 was not initiated. 
 
• PN008 (NCT01076400): a Phase I/IIa, dose-escalation study evaluating AZD1775 in 
combination with topotecan plus cisplatin in adult patients with cervical cancer was terminated 
early due to portfolio prioritization in oncology at Merck after 7 patients had been enrolled in the 
dose-escalation part of the study. The Phase IIa part was not initiated. 
 
• PN011 (Investigator Sponsored Study): a Phase I study of single-agent AZD1775, in patients 
with refractory solid tumors, sponsored by the National Cancer Institute (NCI) Cancer Therapy 
Evaluation Program in collaboration with AstraZeneca and Merck. This study reported 
AZD1775 monotherapy activity in patients carrying BRCA mutations for the first time. 
 
• D6011C00001 (NCT02087176; SCRI LUN 262): a lead-in Phase II multicenter, randomized, 
double-blind study comparing AZD1775 plus docetaxel with placebo plus docetaxel in 
previously treated patients with non-small-cell lung cancer (NSCLC) 
 
• D6011C00002 (NCT02087241; SCRI LUN 261): a Phase II study of AZD1775 plus 
pemetrexed and carboplatin followed by a randomized comparison of pemetrexed and 
carboplatin with or without AZD1775 in patients with previously untreated stage IV non-
squamous NSCLC 
 
Ongoing: 
• D6010C00004 (NCT02272790; SCRI GYN 49): a multicenter Phase II study of AZD1775 plus 
either paclitaxel, gemcitabine, carboplatin, or pegylated liposomal doxorubicin in patients with 
platinum-resistant epithelial ovarian, fallopian tube, or primary peritoneal cancer 


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• D6010C00005 (NCT02511795; SCRI REFMAL 384): a Phase I study evaluating AZD1775 in 
combination with olaparib in refractory solid tumors. 
 
• D6011C00003 (NCT02341456): a Phase Ib dose-finding study evaluating AZD1775 as 
monotherapy and in combination with carboplatin and paclitaxel in adult Asian patients with 
advanced solid tumors 
 
• D6015C00001 (NCT02482311; SCRI REFMAL 383): a Phase I, dose escalation, safety and 
pharmacokinetic study of AZD1775 monotherapy (Schedule 1) in patients with advanced or 
metastatic solid tumors 
 
• D6015C00002 (NCT02617277; SCRI REFMAL 412): a Phase I study assessing the safety, 
tolerability, and pharmacokinetics of AZD1775 in combination with MEDI4736 in patients with 
advanced solid tumors 
 
• D6015C00003 (NCT02610075; SCRI REFMAL 398): a Phase Ib study to determine the 
maximum-tolerated dose (MTD) of AZD1775 monotherapy (Schedule 2) in patients with locally 
advanced or metastatic solid tumors. 
 
• NCI130032 (NCT01748825): AZD1775 for Advanced Solid Tumors (CTEP Sponsored 
study) 
 
In Study PN001, of 176 evaluable patients who received AZD1775 (either single or multiple 
doses) as monotherapy or in combination with gemcitabine, cisplatin, or carboplatin, a partial 
response (PR) (confirmed and unconfirmed) was observed in 17 (9.7%) patients, and stable 
disease (SD) was observed in 94 (53.4%) patients (AZD1775 Investigator’s Brochure[IB])In 
Study PN001, 9 patients received AZD1775 monotherapy. Single ascending doses of AZD1775 
up to 1300 mg were well tolerated; the maximum tolerated dose (MTD) was not established. 
 
In Study PN004, all patients were treated at the 225 mg AZD1775 BID 2.5-day dose level in 
combination with paclitaxel and carboplatin. Of the 14 evaluable patients by RECIST v1.1 in 
Part 1, there were 11 PRs (6 confirmed and 5 unconfirmed), and 3 SDs; 7 patients were 
evaluable by CA-125 with 3 CRs and 4 PRs. Final data for Part 2 is not available as of the cut-
off date for this protocol. 
 
In Study PN005, patients in Part 1 received single-cycle BID dosing of AZD1775 for 5 days at 1 
of 2 dose levels as monotherapy. A cohort of 3 patients was enrolled at the starting dose level of 
AZD1775 65 mg BID and no serious adverse events (SAEs) were experienced. 
 
No complete responses (CRs) or PRs were observed in either of Studies PN005 or PN008 at the 
time that they were terminated. 
 
In Study PN011, patients received single-agent AZD1775 PO BID over 2.5 days per week for 2 
weeks in 3-week treatment cycles. Twenty-five patients were enrolled to determine the MTD 
using a 3+3 design. The MTD was established at 225 mg by mouth (PO) BID for 5 doses on 


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Weeks 1 and 2 of a 3-week schedule. Six patients with BRCA-mutated solid tumors were 
enrolled at the MTD. Partial responses were confirmed in two of the patients carrying BRCA 
mutations (ovarian cancer patient and head/neck cancer patient). Paired tumor biopsies were 
obtained from 5 patients treated at the MTD at baseline and after the 5th AZD1775 dose to 
determine the levels of pY15-Cdk and γH2AX. The biopsies showed a decrease in pY15-Cdk 
levels (2/5 paired biopsies). The same biopsies were analyzed for increases in γH2AX, an 
indicator of DNA damage. Three of the 5 biopsy pairs showed an increase in γH2AX levels. 
DNA damage response was observed in this study through provided paired tumor biopsies. 
 
In Study D6011C00001, 32 patients with NSCLC were treated with 225 mg AZD1775 BID over 
2.5 days in combination with docetaxel (75 mg/m2 IV) administered on Day 1 followed by 
pegfilgrastim on Day 4 of each 21-day cycle. The 3 patients (9.4%) that achieved PR by 
RECIST v1.1 had TP53 mutations. Twenty-one patients (65.6%) had SD and 10 (47.6%) of these 
patients had TP53 mutations. The planned Interim Analysis of 32 patients in the single cohort 
lead-in (Part A) suggested that toxicities associated with AZD1775 given in combination with 
docetaxel were greater in frequency and severity than with docetaxel alone. Additionally, the 
analysis revealed that it was very unlikely the target response rate would be reached in this study 
and a decision was made to terminate enrolment. 
 
In Study D6011C00002, 14 patients with NSCLC were treated with 225 mg AZD1775 BID over 
2.5 days in combination with pemetrexed 500 mg/m2 IV and carboplatin AUC 6 IV, both 
administered on Day 1 of each 21-day cycle. Enrolment was stopped because of the introduction 
of new therapies for the treatment of first-line NSCLC, such as immunotherapy, which resulted 
in challenges in patient recruitment. In addition, the planned Interim Analysis of Study 
D6011C00001 revealed that it was very unlikely that the target response rate would be reached, 
and increased gastrointestinal and hematologic toxicities associated with AZD1775 were 
observed. 
 
In study NCI130032, the MTD was determined to be 300mg daily on days 1-5 and 8-12 every 21 
days (EORTC poster, Oct 2017). In monotherapy studies carried out by AstraZeneca, a similar 
dosing regimen was also determined to be the MTD (data on file at AstraZeneca), therefore it 
will be the selected starting dose for this study.  
 
 
 
2.4   AZD1775 Pharmacokinetics 
 
The pharmacokinetic (PK) data of AZD1775 following a single oral administration (Study 
PN001) showed a moderate rate of absorption with a time of maximum concentration (Tmax) 
occurring at 3 to 4 hours, 2 and 4 hours on day 1 and 3 respectively (IB v15 p. 119). Post-peak 
plasma concentrations declined essentially in a mono-exponential manner with a terminal 
elimination half-life (t1/2) between 9.02 to 12.3 hours 13.2 hours at the 225 mg PMF dose (IB 
v15 p. 119). Exposure as measured by maximum plasma drug concentration observed (Cmax) 
and area under the curve AUC(0-∞) increased in a dose-proportional manner over the dose range 
of 325 to 1300 mg.  
 


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Following single (100 to 325 mg) and multiple dose administrations of AZD1775 (25 to 325 mg 
BID and 100 to 200 mg once daily [QD]) with carboplatin, cisplatin, and gemcitabine, plasma 
exposure of AZD1775 was consistent with predictions based on the single-dose regimen. 
Preliminary investigation of drug-drug interactions (DDIs) in Study PN001 suggest a ~40% 
increase in the exposure of AZD1775 in the presence of aprepitant (moderate CYP3A4 inhibitor) 
on both Day 1 and Day 3 of dosing, but no effect with the concomitant administration of steroids 
(moderate CYP3A4 inducers). Preliminary studies also suggested that the Pre-marketed Oral 
Formulation of AZD1775 was similar to that of the Fit-For-Purpose formulation.   
 
Based on the preliminary comparison of the results of AZD1775 PK parameters at the 225 mg 
dose, PK estimates in Asian patients were higher than in Western patients. After single dose 
administration on Cycle 0 Day 1 (monotherapy), Cmax and AUC at the 225 mg dose were 45% 
and 35% higher, respectively, in the Asian population as compared to the Western population 
(Study PN011). At steady state (Cycle 3 Day 1), a similar trend of higher exposure in Asian 
patients was observed. Additional analysis/investigation will be conducted based on the 
emerging data to understand the exposure differences between the populations. 
 
 


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2.5   AZD1775 Safety 
 
Overall risks and benefits support the administration of oral AZD1775 according to this clinical 
protocol. The identified risks (expected events) for AZD1775 are described in section 5.4 
(Emerging Safety Profile) of the IB. Section 6.4 (Risk Management) of the IB.  
 
Based on the safety data from the completed AZD1775 clinical studies and preliminary data 
from ongoing studies, adverse drug reactions to AZD1775 monotherapy include: anemia, 
neutropenia, thrombocytopenia, QTc prolongation, gastrointestinal events such as dyspepsia, 
diarrhea, nausea and vomiting (with or without dehydration or serum electrolyte decreases), as 
well as decreased appetite. 
 
Based on information emerging during the clinical development program of AZD1775, potential 
risks with AZD1775 monotherapy include asthenia/fatigue, febrile neutropenia, gastrointestinal 
hemorrhage, lymphopenia/lymphocyte count decreased, leukopenia/WBC count decreased, 
myalgia, stomatitis, sepsis and transaminases elevation. 
 
 
 


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2.6  Cyclin E encoded by CCNE1 is a critical G1/S transition regulator 
 
Human cyclin E1 was initially discovered as one of several substitutes (cyclins A, B1, B2, C, D, 
and E) for G1 cyclin genes in yeast, and accumulated periodically through the cell cycle, 
initiating the G1 to S transition [14]. CCNE1 encodes the full-length 50-KD E1 protein that 
interacts with cyclin-dependent kinases 1 and 2 (CDK1 and CDK2) to activate their kinase 
activity, a requirement for G1-S transition [15]. In normal dividing cells, cyclin E1 is highly cell 
cycle-dependent, peaking in late G1 and reaching its nadir in G2-M and early G1 phases [16]. 
Early in the G1 phase, increases in c-Myc levels promote cyclin D transcription, which activates 
CDK4 and CDK6 to phosphorylate their target protein, such as retinoblastoma (Rb) and its 
complex partners (p130 and p107) [17]. Activated Rb acts as a tumor suppressor to maintain 
cells in the G1/G0 phase through binding the inhibiting E2F in the absence of cyclin E/CDK 
complex. Upon Rb hyperphosphorylation by cyclin D/CDK complexes, E2Fs are released from 
its inhibitory complex and can initiate CCNE1 expression [18]. Once activated, cyclin E1/CDK2 
becomes independent of mitogenic stimuli and can further upregulate its own expression by 
increased phosphorylating and dissociating Rb/E2F complexes, triggering the G1-S transition. 
Cyclin E1/CDK complexes can be silenced by CCNE1 promoter silencing, protein inhibition by 
p21Cip1 and p27Kip1, degradation by the SCFFbw7 tumor suppressor pathway [19], and microRNA-
mediated inhibition (miR-15a, miR-15b, miRNA-16) [20].  
 
The function for cyclin E1 to induce S phase entry depends on its interactions with CDK1 and 
CDK2 [21], mediating cell cycle progression from G1 to S phase; DNA replication and 
transcription; centriole duplication, centrosome separation, and endoreduplication; activating 
BRCA1/2 transcription via E2F transcription factors to trigger important components of the 
homologous recombination pathway for DNA repair and confer relative chemotherapy 
resistance; and many other biological processes such as apoptosis, pre-mRNA splicing, synaptic 
function and epigenetic silencing, as described in Figure 2 [22]. 
 
Cyclin E demonstrates high oncogenic potential and their amplification can induce tumor 
formation both in vivo and in vitro [23-26]. CCNE1 is frequently amplified in many human 
tumors, with the frequency of amplification ranging from 2 to 40% [27]. CCNE1 amplification 
results in cyclin E1 overexpression, which play an important role in cancer progression, and 
correlates with worse overall survivals [28-34], as well as disease survival as shown in Figure 3 
(data from Dr. K. Keyomarsi’s laboratory). 
 
 
 
 
 
 
 
 
 
 
 


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Figure 2. Summary of cyclin E1 functions. Cyclin E1 functions in 
cell cycle progression, inhibition of apoptosis, and DNA 
transcription, replication, and repair. Less characterized functions 
may include centriole duplication, pre-mRNA splicing, and 
epigenetic silencing. Kanska J et al: Gyn Onc, 2016 
Figure 3: Overexpression of cyclin E and 
pCDK2 was associated with worse disease-
free survival in patients with triple negative 
breast cancer. (Chen X et al: unpublished 
data). 


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2.7  CCNE1 Amplification Sensitizes Cancer Cells to AZD1775 
 
Overexpression of cyclin E has been shown to promote genomic instability by causing DNA 
replication stress and deregulating the G1 to S transition [35-37]. Wee1 kinase, which prevents 
premature mitotic entry by inhibiting CDKs, may be essential when cyclin E is overexpressed in 
order to prevent massive DNA damage and cell death [38-40]. Recent findings from a genome-
wide knockout screen revealed that absence of CDK2 decreases sensitivity to Wee1 kinase 
inhibition [41]. Furthermore, Dr. Khandan Keyomarsi and her group at MD Anderson have 
shown that overexpression of cyclin E is significantly associated (85%; p<0.001) with active 
CDK2 (phoshpo-CDK2) in tumor samples from 1,678 breast cancer patients [28, 42].   
 
Collectively, these studies suggest that overexpression of cyclin E could predict response to Wee 
1 kinase inhibition. To test this hypothesis, we initially evaluated the sensitivity of 6 TNBC cell 
lines to a clinically available Wee 1 kinase inhibitor, AZD1775 [43], as a function of several cell 
cycle modulators.  Results revealed that the expression of cyclin E was highly predictive (r=0.90) 
of response to AZD1775 in all 6 TNBC cell lines (Chen X et al: unpublished data, Figure 4). 
Consistent with this conclusion, the levels of p-CDK1 (Y15, a known downstream effector of 
Wee1 kinase) and p-CDK2 (T160) were moderately (2/6 cell lines) predictive of AZD1775 
sensitivity among other key cell cycle regulators.  
 
Figure 4: Correlation between cyclin E normalized protein 
level and AZD1775 sensitivity (r=-0.90) from IC50 values. 
(Chen X et al: unpublished data). 


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In two of the TNBC cell lines (HCC1806 and MB231), the knockdown of cyclin E by shRNA, 
desensitized both cell lines to AZD1775, compared to their scrambled controls. Induction of 
cyclin E expression in 76NE6 (a human mammary epithelial cell line) and MDA231 cells 
significantly (p <0.001) increased AZD1775 sensitivity, compared to their uninduced controls. 
Furthermore, induction of the tumor specific cytoplasmic-localized low molecular weight cyclin 
E (LMWE) [44, 45] in 76NE6 cells also increased sensitivity to AZD1775, but not more than full 
length cyclin E. We interrogated the sensitivity of AZD1775 in vivo, by developing several 
patient derived xenografts (PDX) models from TNBC patients: PDX1 has normal copy number 
and low cyclin E expression, while PDX2 and PDX3 exhibit amplification and overexpression of 
cyclin E. CDK2, p-CDK2 and p-CDK1 levels were also co-overexpressed with cyclin E in PDX2 
and PDX3. Each PDX model was treated with either vehicle or 50mg/kg AZD1775 for 4 cycles 
and sacrificed when maximum allowable tumor burden were reached. Consistent with our in 
vitro results, AZD1775 treatment significantly decreased tumor burden and improved survival of 
cyclin E overexpressing PDX2 and PDX3 models, with little to no inhibition of PDX1 model 
(Chen X et al: unpublished data, Figure 5). Collectively, these results suggest cyclin E 
overexpression can predict response to Wee1 kinase inhibition. 
The clinical data to use AZD1775 in patients with advanced solid tumors is emerging from the 
ongoing phase I study (NCT02482311; AstraZeneca Study D6015C00001; REFMAL 383), 
which is a single agent AZD1775 study. In a phase II trial PN004 that investigated the 
combination of AZD1775 and platinum-containing chemotherapy for the treatment of patients 
with TP53 mutant recurrent platinum-sensitive ovarian cancer (NCT01357161), enrollment 
Figure 5: PDX1 has normal copy number and low cyclin E expression, while PDX2 and 
PDX3 exhibit amplification and overexpression of cyclin E. Each PDX model was 
treated with either vehicle or 50mg/kg AZD1775 via oral gavage BID x 2 days every 
week for 4 weeks. AZD1775 treatment significantly decreased tumor burden and 
improved survival of cyclin E overexpressing PDX2 and PDX3 models, with little to no 
inhibition of PDX1 model. (Chen X et al: unpublished data).  


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started with 15 patients for open-labeled safety run-in study (Part A) followed by 121 patients 
(Part B) who were randomized to receive AZD1775 plus paclitaxel and carboplatin or placebo 
plus paclitaxel and carboplatin (Laing N et al: Genetic analysis of tumors from a Phase II trial 
evaluating AZD1775, carboplatin and paclitaxel in patients with TP53-mutant ovarian cancer. 
AACR 2016 Abstract #337). Two patients (#10, and #13) were identified to have CCNE1 
amplification in Part A: both achieved partial responses for more than one year. In Part B, two 
patients were found to have CCNE1 amplification in AZD1775 plus paclitaxel and carboplatin 
group: one had stable disease (#25), and the other achieved complete remission (#29); three 
patients were identified to have CCNE1 amplification in placebo plus paclitaxel and carboplatin 
group: one had stable disease (#38), and the other two achieved partial responses (#35, and #44).  
 
 
 
 
 


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2.8   The Feasibility to Conduct a Phase II Basket Study in CCNE1 Amplified Solid 
Tumors 
The above preclinical studies have provided the mechanistic and preclinical evidence to support 
the use of cyclin E a biomarker of response to AZD1775 in TNBC [43]. We show that the 
replication stress pathway activated upon cyclin E overexpression results in sensitivity to Wee1 
kinase inhibition. Evaluation of cyclin E overexpression resulted from CCNE1 amplification can 
readily be used as an integral biomarker in future clinical trials, to guide identification of patients 
who are likely to respond to AZD1775. As in Figure 6, CCNE1 amplifications have been 
observed in more than 7.5% of many tumor types: uterine, breast, ovarian, pancreatic, bladder, 
gastric, esophageal, lung cancers and sarcoma. Therefore, it might be appropriate to conduct a 
phase II basket study in patients with advanced solid tumors harboring CCNE1 amplification. 
 
In our own institutional experience at MD Anderson, Dr. Funda Meric-Bernstam and her 
colleagues have collected tumor specimens from 3807 patients with advanced solid tumors. 
Cyclin E amplifications were identified on multiple genomic testing platforms, as shown in 
Table 1, ranging from 1.3% to 6.1%. In these 3807 patients, Cyclin E amplifications were 
detected in 108 patients (2.8%), frequently in patients with bladder (n=3), breast (n=10), bile 
duct | cholangiocarcinoma (n=6), colorectal (n=9), endometrial (n=12), gall bladder (n=11), 
gastric (n=4), liver (n=8), lung (n=7), pancreatic (n=4), primary peritoneal (n=6), ovarian cancer 
(n=14), and sarcoma (n=4), as shown in Figure 7. 
 
Figure 6: Frequency of CCNE1 amplification in different pathologic 
diagnoses. (K. Keyomarsi et al: unpublished data). 


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 Table 1: Rates of Cyclin E Amplification- according to NGS platforms 
Platform 
# Amplified/# of Tests 
Rate 
Oncomine 
2/38 
5.3% 
STGAv1 
24/1803 
1.3% 
FoundationOne 
68/1792 
3.8% 
Guardant360 
16/263 (overall) 
6.1% 
6/263  (++) 
2.3% 
0/263 (+++) 
0% 
 
 
 
 
 
 
Bile Duct | Cholangiocarcinoma, 6
Bladder, 3
Breast, 10
Colorectal, 9
Endometrial/Uterine, 
12
Gall Bladder, 11
Gastric/Stomach, 4
Liver, 8
Lung, 7
Ovarian, 14
Pancreatic, 4
Primary Peritoneal, 6
Sarcoma, 4
Other , 10
Figure 7: Disease Types Observed with CCNE1 
Amplification


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Overexpression of cyclin E has been shown to promote genomic instability by causing DNA 
replication stress and deregulating the G1 to S transition. Wee1 kinase, which prevents 
premature mitotic entry by inhibiting CDKs, may be essential when cyclin E is overexpressed in 
order to prevent massive DNA damage and cell death. We hypothesize that cancer cells 
harboring CCNE1 amplification that leads to cyclin E overexpression, will be highly sensitive to 
the treatment with AZD1775, a Wee1 inhibitor, and that the objective response rate (ORR) 
defined as complete responses (CRs), and partial responses (PRs) will be observed in greater 
than 25% of patients with CCNE1 amplified advanced solid tumors who receive the treatment 
with AZD1775. 
 
 
 
 


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2.9   Correlative Studies – Background and Rationale 
 
Introduction 
Overexpression of cyclin E has been shown to promote genomic instability by causing DNA 
replication stress and deregulating the G1 to S transition. Wee1 kinase, which prevents 
premature mitotic entry by inhibiting CDKs, may be essential when cyclin E is overexpressed in 
order to prevent massive DNA damage and cell death. We hypothesize that CCNE1 
amplification in cancer cells leads to an overexpression of cyclin E1 protein, which will sensitize 
cancer cells to the treatment with AZD1775, a Wee1 inhibitor. The consequential disruption of 
intra-S phase and G2/M cell cycle checkpoints by the use of AZD1775 will result in premature 
mitotic entry, inducing significantly greater mitotic catastrophe and apoptosis in cyclin E1 
overexpressed cells. We will test this hypothesis in this clinical study with the help of the 
following correlative studies.  
 
In order to further define the molecular environment of CCNE1 amplification along with Wee1 
inhibition, we will validate several key biological players using next generation sequencing and 
cytogenetic platforms at various treatment time points listed in Table 2. 
 
 
Table 2.  Laboratory Correlates – Biomarker Plan 
Biomarker 
Name 
AND Lead PI 
and Site 
Assay 
(CLIA: Y/N) 
Use (Integral, 
Integrated, or 
Exploratory) AND 
Purpose  
Tissue/Body 
Fluid Tested and 
Timing of Assay 
 
M/O  
Funding 
Source(s)  
CCNE1 
amplification  
(S. Fu /MD 
Anderson) 
Cytogenetics (Y), or a 
next generation 
sequencing platform 
(Y) 
CLIA: Y 
Integral 
To identify patients for 
subgroup enrollment. 
To correlate responses. 
Tumor and blood 
Archival or 
baseline biopsy 
M 
Standard 
of patient 
care 
pCDK, pHH3, 
gH2AX 
(Dr. Robert 
Kinders/PADIS 
Laboratory, 
FNCLR) 
 
Quantitative 
immunofluorescence 
 
CLIA: N 
Integrated 
 
To investigate 
pharmacodynamic 
responses 
Tumor biopsy 
(frozen tissue) 
Pre-study, C1D10 
at 4h post dose, 
and at progression 
M – Pre-
study and 
C1D10 at 
4h post 
dose 
O-at 
progression 
NCI 
Pre-identified 
mutations 
(F. Meric-
Bernstam /MD 
Anderson) 
A next generation 
sequencing platform 
CLIA: N 
Exploratory 
To identify biomarkers 
of responses and 
acquired resistance 
Plasma 
Pre-study, C1D8, 
C2D1, C2D8, and 
at end of study 
M – Pre-
study 
O – All 
other times 
TBA 
Cytokines and 
chemokines 
(F. Meric-
Bernstam /MD 
Anderson) 
Meso Scale Discovery 
platform 
CLIA: N 
Exploratory 
To identify biomarkers 
of responses and 
acquired resistance 
Plasma 
Pre-study, C1D8, 
C2D1, C2D8 and 
at end of study 
M – Pre-
study 
O – All 
other times 
TBA 


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Biomarker 
Name 
AND Lead PI 
and Site 
Assay 
(CLIA: Y/N) 
Use (Integral, 
Integrated, or 
Exploratory) AND 
Purpose  
Tissue/Body 
Fluid Tested and 
Timing of Assay 
 
M/O  
Funding 
Source(s)  
Cyclin E1, 
pCDK2 (K. 
Keyomarsi/ MD 
Anderson  
Immunohistochemistry  
CLIA: N 
Exploratory 
To identify biomarkers 
of response and 
acquired resistance 
Tumor 
Pre-study, C1D10 
at 4h post dose, 
and at progression   
 
O  
TBA  
Molecular 
signatures 
(F. Meric-
Bernstam /MD 
Anderson) 
Targeted exome by a 
next generation 
sequencing platform 
and RNA-seq analysis 
CLIA: N  
Exploratory 
To identify biomarkers 
of responses and 
acquired resistance 
Tumor and blood 
Pre-study, C1D10 
at 4h post dose, 
and at progression  
O 
MD 
Anderson 
IPCT 
 
 
Assay for CCNE1 amplification 
 
Oncomine – a next generation sequencing platform for CCNE1 amplification 
 
Solid Tumor Genomics Assay v1 is one of the Oncomine Cancer Panels conducted at MD 
Anderson. This next generation sequencing-based analysis (PCR-based sequencing) for the 
detection of somatic mutations in the coding sequencing of 128 genes and selected copy number 
variations (amplification) in 49 genes (overlap: 134 genes total) is performed on the DNA 
extracted from the tumor sample in a CLIA-certified molecular diagnostics laboratory at MD 
Anderson. For clinical purposes, the effective lower limit of detection of this assay (analytical 
sensitivity) for single nucleotide variations is in the range of 5% - 10% (one mutant allele in the 
background of nineteen - nine wild type alleles). Sensitivity for amplifications depends on both 
input tumor percentage and the amplitude of the gene amplification. The analytic pipeline in this 
assay attempts to normalize for inter-amplicon performance differences and total sample loading, 
but does not attempt to correct for tumor percentage. A nominal threshold of 7 for the estimated 
copy number is used to restrict reporting to high-confidence amplification calls. 
 
FISH – fluorescence in situ hybridization for CCNE1 amplification 
 
The FISH probe for CCEN1 19p12 (RP11-345 J21) will be acquired from BACPAC Resources 
(Oakland, CA). The probe will be labeled with SpectrumOrange or SpectrumGreen using a nick 
translation kit (Abbott Laboratories, Abbott Park, IL). Tissue sections will be processed for 
removal of protein, denaturation, hybridization, and posthybridization washing, and then will be 
counterstained with DAPI II (Abbott Laboratories) and examined using a fluorescence 
microscope (Olympus, Tokyo, Japan) equipped with a Triple Bandpass Filter set (Abbott 
Laboratories) for DAPI II, SpectrumOrange, and SpectrumGreen. Scoring and evaluation of 
FISH slides will be performed manually by counting the target gene signals and control signals 
in 20 tumor cell nuclei per sample. Gene amplification was determined according to the 
American Society of Clinical Oncology/College of American Pathologists criteria for ERBB2 
amplification. 
 
Assay for pCDK, pHH3, gH2AX 


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Quantitative immunofluorescence assays will be performed as previously described (53). 
 
Assay for pre-identified mutations 
 
Whole blood will be collected in EDTA–containing tubes, centrifuged and spun twice within 2 
hours to yield plasma. The QIAamp Circulating Nucleic Acid kit (Qiagen, Valencia, CA) will be 
used to isolate cfDNA, and 16 -100 ng of unamplified cfDNA will be tested with a multiplex 
ddPCR Screening Kit (Bio-Rad, Pleasanton, CA) using the QX200 Droplet Digital PCR platform 
(Bio-Rad). The lower limit of detection is approximately 0.2% MAF (mutation allelic frequency) 
for the multiplexed screening assay and <0.1% MAF per single well for the mutation-specific 
assays. 
 
Assay for circulating cytokines and chemokines 
 
Circulating cytokines and chemokine will be measured using the MesoScaleDiscovery (MSD) 
platform. The MSD technology uses pairs of antibodies recognizing different epitopes of the 
same cytokine/chemokine. The primary antibodies are spotted onto the well of a 96-well plate 
(several spots recognizing different cytokines/chemokines per well). The secondary antibody (as 
an ELISA sandwich) is labeled with a sulfo-tag that allows using an electrical current to measure 
electrochemiluminescence as the signal. This technology is sensitive and has a broad dynamic 
range (0.2-10000 pg/ml). The samples from each time point of each patient will be analyzed in 
duplicate. Each assay requires 25 µl of plasma/well. Standards for each cytokine (0-10,000 
pg/ml) will be run on each multiplex assay plate. The data will be acquired on the V-plex® 
Sector Imager 2400 plate reader and analyzed using the Discovery Workbench 4.0 software 
(MSD, Rockville, Maryland). 
 
Assay for cyclin E1 and pCDK2 
 
Immunohistochemistry for cyclin E and p-CDK2 will be performed using two rabbit polyclonal 
antibodies; sc198 (Santa Cruz Biotechnology, Dallas, TX) at a dilution 1:1000 and Thr160 (Cell 
Signaling Technology, Danvers, MA) at a dilution of 1:100 and developed with the 
VECTASTAIN Elite ABC kit (Dako, Glostrup, Denmark). Nuclear and cytoplasmic staining 
scores will be independently assigned according to their staining intensity and combined to 
generate the low and high values. For p-CDK2 the intensity and the extent of the staining 
generated the final immunoreactivity score will be determined by multiplying the intensity score 
by the staining extent score, with a minimum score of 0 and maximum score of 18; scores of ≥6 
indicated positivity.  
 
Assay for molecular signatures 
 
DNA will be extracted from tumor specimens will be analyzed by next generation sequencing 
(i.e., Solid Tumor Genomics Assay v1) to detect targeted exome aberrations.  
 
RNA will be isolated from tumor specimens using the the Qiagen RNeasy AllPrep method 
(Qiagen Sciences, Valencia, CA) and stored at -80oC until further use. RNA integrity will be 


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monitored using a Bioanalyzer and the Agilent RNA 6000 Pico assay (Agilent Technologies, 
Santa Clara, CA). An RNA integrity number greater than 8 is recommended for all samples. 
Total RNA samples will be fractionated and fragmented to an average length of 150 nt prior to 
the RNA end repair, and the adapter ligation, to generate PCR amplification of RNA-seq library 
post reverse transcription. The library will be quantified for Illumina HiSeq2000 50 base single 
read (SR50) and sequenced on the HiSeq2000 Sequencer, to identify novel genes and splice 
variants, and to detect single nucleotide polymorphisms.  
 
 
 


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3.   PATIENT SELECTION 
3.1   Patient Inclusion Criteria  
 
To be eligible for this trial, patients must meet all of the following eligibility criteria. 
 
1. Patients must have one of the histologically advanced solid tumors harboring CCNE1 
amplification: Their diseases are refractory to, or do not have, standard-of-care therapy; or 
they declined standard-of-care therapy. CCNE1 amplification is defined in a Clinical 
Laboratory Improvement Amendments (CLIA)-certified laboratory: CCNE1 Amplification > 
7 based on targeted custom Ampliseq panel on the Ion Torrent PGM; or CCNE1 
amplification on alternate CLIA platforms such as Foundation One, UW-OncoPlex – Cancer 
Gene Panel, MSK-IMPACT, Solid Tumor Genomic Assay (Life Technologies), etc. will also 
be eligible to be treated after PI approval.  Patients with known CCNE1 amplification on 
local or commercial platforms can start treatment after planned biopsy or submission of 
recent archival sample. Central NGS CCNE1 and FISH testing will be performed to confirm 
the result. 
 
2. Measurable disease per Response Evaluation Criteria in Solid Tumors (RECIST) version 1.1. 
 
3. Male or female sex and age ≥18 years 
 
4. Has read and understands the informed consent form (ICF) and has given written ICF prior to 
any study procedures. Patients with Impaired Decision Making Capacity (IDMC) must have 
a close caregiver or Legally Authorized Representative (LAR). 
 
5. Any prior radiation must have been completed at least 7 days prior to the start of study drugs, 
and patients must have recovered from any acute adverse effects prior to the start of study 
treatment. 
 
6. Eastern Cooperative Oncology Group (ECOG) Performance Status score of 0-1. 
 
7. Baseline laboratory values within 14 days of study drug(s) initiation: 
• Absolute neutrophil count (ANC) ≥1500/ µL 
• Hemoglobin (Hgb) ≥ 9g/dL for mono-therapy 
• Platelets ≥ 100,000/µL 
• Alanine aminotransferase (ALT) and aspartate aminotransferase (AST) ≤ 3 x upper limit 
of normal (ULN) or ≤ 5 x ULN if known hepatic metastases. 
• Serum bilirubin < ULN or < 1.5 x ULN in patients with liver metastases; or total bilirubin 
< 3.0 x ULN with direct bilirubin WNL in patients with well documented Gilbert’s 
Syndrome. Serum creatinine ≤ 1.5 x ULN, or calculated creatinine clearance (CrCl) ≥ 45 
mL/min as calculated by the Cockcroft-Gault method or 24-hour measured urine CrCl ≥ 
45 mL/min. 
CrCl (glomerular filtration rate [GFR]):   a where F= 0.85 for females and F=1 for male 
= (140-age) x (weight/kg) x Fa / (72 x serum creatinine mg/dL) 


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8. Female patients who are not of child-bearing potential and fertile females of childbearing 
potential who agree to use adequate contraceptive measures from 2 weeks prior to the study 
and until 1 month after study treatment discontinuation, who are not breastfeeding, and who 
have a negative serum or urine pregnancy test within 3 days prior to the start of study 
treatment (see Appendix D).  Male patients willing to abstain or use barrier contraception 
(i.e. condoms) for the duration of the study and for 3 months after treatment stops (see 
Appendix D). 
 
9. Willingness and ability to comply with study and follow-up procedures. 
 
10. Ability to take oral medications without medical history of malabsorption or other chronic 
gastrointestinal disease, or other conditions that may hamper compliance and/or absorption of 
the study agent. 
 
11. No prior treatment with Wee1 kinase inhibition.  
 
12. Provision of an archival tissue block, or 10 formalin-fixed paraffin-embedded (FFPE) slides, 
if available, and having biopsiable disease and agreeing to pre-treatment and on-study 
biopsies. 
 
 
 
 
 
 


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3.2   Patient Exclusion Criteria  
 
Patients who meet any of the following criteria will not be eligible for the study.  
 
1. Use of anti-cancer treatment drug ≤ 21 days or 5 half-lives (whichever is shorter) prior to the 
first dose of AZD1775. For drugs for which 5 half-lives is ≤ 21 days, a minimum of 10 days 
between termination of the prior treatment and administration of AZD1775 treatment is 
required.  
 
2. Previous radiation therapy completed ≤ 7 days prior to the start of study drugs. 
 
3. Major surgical procedures ≤ 28 days of beginning AZD1775, or minor surgical procedures ≤ 
7 days. No waiting period required following port-a-cath or other central venous access 
placement. 
 
4. Unresolved Grade 2 toxicity from prior therapy (except alopecia or anorexia). 
 
5. Patient has an inability to swallow oral medications. Note: Patient may not have a 
percutaneous endoscopic gastrostomy (PEG) tube or be receiving total parenteral nutrition 
(TPN). 
 
6. No other anticancer-therapy (chemotherapy, immunotherapy, hormonal anti-cancer therapy, 
radiotherapy [except for palliative local radiotherapy]), biological therapy or other novel 
agent is to be permitted while the patient is receiving study medication. Patients on LHRH 
analogue treatment for more than 6 months are allowed entry into the study and may 
continue at the discretion of the Investigator. 
 
7. Known malignant central nervous system (CNS) disease other than neurologically stable, 
treated brain metastases – defined as metastasis having no evidence of progression or 
hemorrhage for at least 2 weeks after treatment. Must be off any systemic corticosteroids for 
the treatment of brain metastases for at least 14 days prior to enrolment. 
 
8. Patient has had prescription or non-prescription drugs or other products known to be 
sensitive to CYP3A4 substrates or CYP3A4 substrates with a narrow therapeutic index, or to 
be moderate to strong inhibitors/inducers of CYP3A4 which cannot be discontinued 2 weeks 
prior to Day 1 of dosing and withheld throughout the study until 2 weeks after the last dose 
of study drug. Co-administration of aprepitant or fosaprepitant during this study is prohibited. 
The use of sensitive substrates of CYP3A4, such as atorvastatin, simvastatin and lovastatin, 
is also prohibited in this study  
 
9. Herbal preparations are not allowed throughout the study. These herbal medications include 
but are not limited to: St. John's wort, kava, ephedra (ma hung), gingko biloba, 
dehydroepiandrosterone (DHEA), yohimbe, saw palmetto and ginseng. Patients should stop 
using these herbal medications 7 days prior to first dose of study treatment.  
 
10. Any known hypersensitivity or contraindication to the components of the study drug 


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AZD1775. 
 
11. Any of the following cardiac diseases currently or within the last 6 months as defined by 
New York Heart Association (NYHA) ≥ Class 2. 
• Unstable angina pectoris 
• Congestive heart failure 
• Acute myocardial infarction 
• Conduction abnormality not controlled with pacemaker or medication 
• Significant ventricular or supraventricular arrhythmias (patients with chronic rate-
controlled atrial fibrillation in the absence of other cardiac abnormalities are eligible) 
 
12. Mean resting corrected QTc interval using the Fridericia formula (QTcF) >450 msec/male 
and >470 msec/female (as calculated per institutional standards) obtained from 3 
electrocardiograms (ECGs) 2-5 minutes apart at study entry, or congenital long QT 
syndrome. 
 
13. Pregnant or breastfeeding women. 
 
14. Serious active infection at the time of study entry, or another serious underlying medical 
condition that would impair the ability of the patient to receive study treatment. 
 
15. Symptomatic and uncontrolled metastasis in the central nervous system or leptomeningeal or 
lymphangitic carcinomatosis. 
 
16. Presence of other active invasive cancers that do not harbor CCNE1 amplification. 
 
17. Grade 2 or higher peripheral neuropathy. 
 
18, Human immunodeficiency virus requiring HAART treatment due to unknown drug-drug 
interactions or has known active hepatitis B (e.g., HBsAg reactive or C virus (e.g., HCV RNA 
(quantitative) is detected) infection 
 


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3.3   Inclusion of Women and Minorities 
 
NIH policy requires that women and members of minority groups and their subpopulations be 
included in all NIH-supported biomedical and behavioral research projects involving NIH-
defined clinical research unless a clear and compelling rationale and justification establishes to 
the satisfaction of the funding Institute & Center (IC) Director that inclusion is inappropriate 
with respect to the health of the subjects or the purpose of the research.  Exclusion under other 
circumstances must be designated by the Director, NIH, upon the recommendation of an IC 
Director based on a compelling rationale and justification.  Cost is not an acceptable reason for 
exclusion except when the study would duplicate data from other sources.  Women of 
childbearing potential should not be routinely excluded from participation in clinical research.  
Please see http://grants.nih.gov/grants/funding/phs398/phs398.pdf. 
 
 
 
 


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4.   REGISTRATION PROCEDURES 
4.1   Investigator and Research Associate Registration with CTEP 
 
Food and Drug Administration (FDA) regulations and National Cancer Institute (NCI) policy 
require all individuals contributing to NCI-sponsored trials to register and to renew their 
registration annually.  To register, all individuals must obtain a Cancer Therapy Evaluation 
Program (CTEP) Identity and Access Management (IAM) account 
(https://ctepcore.nci.nih.gov/iam).  In addition, persons with a registration type of Investigator 
(IVR), Non-Physician Investigator (NPIVR), or Associate Plus (AP) (i.e., clinical site staff 
requiring write access to OPEN or RAVE or acting as a primary site contact) must complete their 
annual registration using CTEP’s web-based Registration and Credential Repository (RCR) 
(https://ctepcore.nci.nih.gov/rcr).  Documentation requirements per registration type are outlined 
in the table below. 
 
Documentation Required 
IVR 
NPIVR 
AP 
A 
FDA Form 1572 
 
 
Financial Disclosure Form 
 
 
 
NCI Biosketch (education, training, employment, license, and 
certification) 
 
 
 
HSP/GCP training 
 
 
 
Agent Shipment Form (if applicable) 
 
CV (optional) 
 
 
 
 
An active CTEP-IAM user account and appropriate RCR registration is required to access all 
CTEP and CTSU (Cancer Trials Support Unit) websites and applications.  In addition, IVRs and 
NPIVRs must list all clinical practice sites and IRBs covering their practice sites on the FDA 
Form 1572 in RCR to allow the following: 
• Added to a site roster 
• Assigned the treating, credit, consenting, or drug shipment (IVR only) tasks in OPEN 
• Act as the site-protocol PI on the IRB approval 
• Assigned the Clinical Investigator (CI) role on the Delegation of Tasks Log (DTL).     
 
Additional information can be found on the CTEP website at 
<https://ctep.cancer.gov/investigatorResources/default.htm>.  For questions, please contact the RCR 
Help Desk by email at < RCRHelpDesk@nih.gov >. 


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4.2   Site Registration 
 
This study is supported by the NCI Cancer Trials Support Unit (CTSU). 
Each investigator or group of investigators at a clinical site must obtain IRB approval for this 
protocol and submit IRB approval and supporting documentation to the CTSU Regulatory Office 
before they can be approved to enroll patients.  Assignment of site registration status in the 
CTSU Regulatory Support System (RSS) uses extensive data to make a determination of whether 
a site has fulfilled all regulatory criteria including but not limited to the following: 
• An active Federal Wide Assurance (FWA) number 
• An active roster affiliation with the Lead Network or a participating organization 
• A valid IRB approval 
• Compliance with all protocol specific requirements 
 
In addition, the site-protocol Principal Investigator (PI) must meet the following criteria: 
• 
Active registration status 
• 
The IRB number of the site IRB of record listed on their Form FDA 1572 
• 
An active status on a participating roster at the registering site 
Sites participating on the NCI CIRB initiative that are approved by the CIRB for this study are 
not required to submit IRB approval documentation to the CTSU Regulatory Office. For sites 
using the CIRB, IRB approval information is received from the CIRB and applied to the RSS in 
an automated process.  Signatory Institutions must submit a Study Specific Worksheet for Local 
Context (SSW) to the CIRB via IRBManager to indicate their intent to open the study locally.  
The CIRB’s approval of the SSW is then communicated to the CTSU Regulatory Office.  In 
order for the SSW approval to be processed, the Signatory Institution must inform the CTSU 
which CIRB-approved institutions aligned with the Signatory Institution are participating in the 
study. 
 
Downloading Regulatory Documents 
 
Site registration forms may be downloaded from the 10136 protocol page located on the CTSU 
Web site.  Permission to view and download this protocol is restricted and is based on person and 
site roster data housed in the CTSU RSS.  To participate, Investigators and Associates must be 
associated with the Corresponding or Participating protocol organization in the RSS.  
 
• Go to https://www.ctsu.org and log in using your CTEP-IAM username and password. 
• Click on the Protocols tab in the upper left of your screen.  
• Either enter the protocol # in the search field at the top of the protocol tree, or 
• Click on the By Lead Organization folder to expand, then select LAO-TX035, and protocol 
10136. 


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• Click on LPO Documents, select the Site Registration documents link, and download and 
complete the forms provided.  (Note: For sites under the CIRB initiative, IRB data will load to 
RSS as described above.) 
 
 
Requirements For NCI 10136 Site Registration: 
• IRB approval (For sites not participating via the NCI CIRB; local IRB documentation, an 
IRB-signed CTSU IRB Certification Form, Protocol of Human Subjects Assurance 
Identification/IRB Certification/Declaration of Exemption Form, or combination is 
accepted ) 
• Local informed consent document 
 
Submitting Regulatory Documents 
Submit required forms and documents to the CTSU Regulatory Office, where they will be 
entered and tracked in the CTSU RSS.   
Regulatory Submission Portal: www.ctsu.org  (members’ area) → Regulatory Tab →Regulatory 
Submission  
When applicable, original documents should be mailed to: 
CTSU Regulatory Office 
1818 Market Street, Suite 3000 
Philadelphia, PA 19103 
 
Institutions with patients waiting that are unable to use the Portal should alert the CTSU 
Regulatory Office immediately at 1-866-651-2878 in order to receive further instruction and 
support. 
 
Checking Site Registration Status 
 
You can verify your site registration status on the members’ section of the CTSU website.   
• Go to https://www.ctsu.org and log in to the members’ area using your CTEP-IAM 
username and password 
• Click on the Regulatory tab at the top of your screen 
• Click on the Site Registration tab 
• Enter your 5-character CTEP Institution Code and click on Go 
 
Note: The status given only reflects compliance with IRB documentation and institutional 
compliance with protocol-specific requirements as outlined by the Lead Network. It does not 
reflect compliance with protocol requirements for individuals participating on the protocol or 


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the enrolling investigator’s status with the NCI or their affiliated networks. 
4.3   Patient Registration 
 
OPEN / IWRS 
 
Patient enrollment will be facilitated using the Oncology Patient Enrollment Network (OPEN).  
OPEN is a web-based registration system available to users on a 24/7 basis.  It is integrated with 
the CTSU Enterprise System for regulatory and roster data interchange and with the Theradex 
Interactive Web Response System (IWRS) for retrieval of patient registration/randomization 
assignment.  Patient enrollment data entered by Registrars in OPEN / IWRS will automatically 
transfer to the NCI’s clinical data management system, Medidata Rave. 
 
For trials with slot reservation requirements, OPEN will connect to IWRS at enrollment initiation 
to check slot availability.  Registration staff should ensure that a slot is available and secured for 
the patient before completing an enrollment. 
 
The OPEN system will provide the site with a printable confirmation of registration and 
treatment information.  Please print this confirmation for your records.  
 
 
OPEN/IWRS User Requirements 
 
OPEN/IWRS users must meet the following requirements: 
• Have a valid CTEP-IAM account (i.e., CTEP username and password). 
• To enroll patients or request slot reservations:  Be on an ETCTN Corresponding or 
Participating Organization roster with the role of Registrar. Registrars must hold a 
minimum of an AP registration type.   
• To approve slot reservations or access cohort management:  Be identified to Theradex as the 
“Client Admin” for the study.  
• Have regulatory approval for the conduct of the study at their site. 
 
Prior to accessing OPEN/IWRS, site staff should verify the following: 
• All eligibility criteria have been met within the protocol stated timeframes.   
• If applicable, all patients have signed an appropriate consent form and HIPAA authorization 
form. 
 
 
OPEN/IWRS Questions? 
 
Further instructional information on OPEN is provided on the OPEN tab of the CTSU website at 
https://www.ctsu.org or at https://open.ctsu.org.  For any additional questions contact the CTSU 
Help Desk at 1-888-823-5923 or ctsucontact@westat.com.  
 
Theradex has developed a Slot Reservations and Cohort Management User Guide, which is 
available on the Theradex website:  http://www.theradex.com/clinicalTechnologies/?National-
Cancer-Institute-NCI-11. This link to the Theradex website is also on the CTSU website OPEN 


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tab. For questions about the use of IWRS for slot reservations, contact the Theradex Helpdesk at 
609-619-7862 or Theradex main number 609-799-7580; CTMSSupport@theradex.com. 
 
 
4.4   General Guidelines 
 
Following registration, patients should begin protocol treatment within 10 days.* Issues that 
would cause treatment delays should be discussed with the Principal Investigator.  If a patient 
does not receive protocol therapy following registration, the patient’s registration on the study 
may be canceled.  The Study Coordinator should be notified of cancellations as soon as possible. 
 
Except in very unusual circumstances, each participating institution will order DCTD-supplied 
agents directly from CTEP.  Agents may be ordered by a participating site only after the initial 
IRB approval for the site has been forwarded by the Coordinating Center to the CTEP PIO 
(PIO@ctep.nci.nih.gov) except for Group studies. 
 
 
 


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5.   TREATMENT PLAN 
 
Cycles are 21 days long. 5.1 Agent Administration 
Treatment will be administered on an outpatient basis.  Reported adverse events and potential 
risks are described in Section 7.  Appropriate dose modifications are described in Section 6.  No 
investigational or commercial agents or therapies other than those described below may be 
administered with the intent to treat the patient's malignancy. 
 
AZD1775 Administration 
Oral AZD1775 will be given 300 mg once daily for 5 days a week for 2 weeks every cycle (QD 
days 1 to 5, and 8 to 12). One cycle of therapy will be 21 days: therefore, 10 doses each cycle. 
Dose escalation will not be permitted. Appropriate dose modifications and interruptions will be 
allowed. Other investigational or commercially available agents specifically for cancer control 
will be prohibited. Patients will continue the treatment until they experience tumor progression, 
prohibitive toxicity or withdraw from the study by themselves. 
 
Table3.  Regimen Description 
Agent 
Premedications; Precautions* 
Schedule 
Cycle Length 
AZD1775 5-HT3 antagonist (ondansetron 8 mg PO or 
granisetron 1 mg PO) prior to each dose of 
AZD1775;  
 
taken with 8 ounces of water approximately 
2 hours before or 2 hours after food 
Days 1-5,  
week 1 
Days 8-12, 
week 2 
21 days 
(3 weeks) 
 
 
 
*Dexamethasone (oral or intravenous) or the 5-HT3 antagonist (intravenous) may be given for 
better symptom control as indicated. Prophylactic anti-emetic therapy with aprepitant or 
fosaprepitant is prohibited. 
 
The patient will be requested to maintain a medication diary of each dose of medication.  The 
medication diary will be returned to clinic staff at the end of each course. 
 
AZD1775 should be taken with 8 ounces of water approximately 2 hours before or 2 hours after 
food. If a patient misses the daily dose according to the schedule, the dose should be taken as 
soon as possible, but not more than 12 hours after the missed dose was scheduled. If greater than 
12 hours, the missed dose should be skipped and the patient should take the next dose when 
scheduled. If vomiting occurs after a patient takes the AZD1775 dose, the patient should be 
instructed not to retake the dose, but to wait until the next scheduled dose of AZD1775.  If no 
dose is scheduled for the following day, the dose will not be ‘made up’.  If vomiting persists, the 
patient should contact the Investigator. 
 
2.7 5.2 Study Procedures 
 
All patients will be evaluated and study procedures will be performed in Figure 1. Dosing, 
laboratory tests, imaging studies, and office visits will occur per protocol (within ± 4 days) 
unless patients’ medical or logistical issues necessitate adjustment. 
 


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Pretreatment Evaluations 
To be completed within 4 weeks before initiation of therapy: 
• Medical history, including list of current medications 
• Tumor markers, if applicable 
• Imaging studies, as appropriate for specific types of cancers  
• ECG and ECHO or MUGA, and then as clinically indicated 
• Mandatory biopsy  
• Urinalysis, to be repeated as clinically indicated 
• For women of childbearing potential, blood or urine pregnancy test at baseline and then 
periodically as clinically indicated 
• Obtainment of the informed consent document 
 
To be completed within 7 days before initiation of therapy: 
• Physical examination  
• ECOG performance status determination 
• Complete blood count with leukocyte differential and platelet count 
• Serum chemistry analysis, including sodium, potassium, chloride, carbon dioxide, BUN, 
creatinine, glucose, calcium, magnesium, albumin, alkaline phosphatase, total bilirubin, 
ALT, TSH, and free T4 
 
 
On-Study Evaluations  
To be obtained weekly during the first cycle and then as clinically indicated: 
• Complete blood count and leukocyte differential and platelet count 
• Serum chemistry, including sodium, potassium, chloride, carbon dioxide, BUN, creatinine, 
glucose, calcium, magnesium, albumin, alkaline phosphatase, total bilirubin, AST, and ALT 
 
To be obtained once in every cycle of therapy (once every 3 weeks): 
• Medical history and physical examination  
• ECOG performance status 
• Assessment of treatment-related and treatment-unrelated toxic effects 
• Complete blood count with leukocyte differential and platelet count 
• Serum chemistry, including sodium, potassium, chloride, carbon dioxide, BUN, creatinine, 
glucose, calcium, magnesium, albumin, alkaline phosphatase, total bilirubin, AST and ALT 
 
To be obtained for tumor responses: 
• Evaluation of the tumor: the same imaging technique used during the initial evaluation or 
more sophisticated studies will be performed about once every 9 weeks ± 7 days for six 
months, then once every 12 weeks ± 7 days, or sooner if clinically indicated 
• Tumor markers, if applicable, once with each imaging study, or more frequently as indicated 
 
 
Sample Collection for Correlative Studies 
Correlative studies will be performed by Dr. Robert Kinders at the NCI-Frederick National 
Laboratory for Cancer Research, Dr. K. Keyomarsi, in the Department of Experimental 


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Radiation Oncology, and Dr. F. Meric-Bernstam in the Department of Investigational Cancer 
Therapeutics. Patients who experience CR/PR will be encouraged to enroll into the Unusual 
Responders protocol (F. Meric-Bernstam) at MD Anderson. 
 
Blood Sample Collections:  
After the informed consent is obtained, blood samples (~10 mL each) will be obtained at each of 
the following time-points: 
 
1. Prior to the initiation of treatment (pre-study), and cycle 1 day 8 (C1D8) 
2. Prior to the initiation of cycle 2 (C2D1) and cycle 2 day 8 (C2D8) 
3. At the end of the study, if available 
 
 
Tumor Sample Collections: 
After informed consent is obtained, tumor samples will be collected as follows: 
1. Ten 5 µm-thick FFPE slides, or tumor block – all patients if available 
2. Pre-treatment core biopsy – mandatory for all patients 
3. Tumor core biopsy on C1D10 at 4 hours (± 2 hours) post dose– mandatory for all patients if 
clinically feasible 
4. Tumor core biopsy at relapse – optional for all patients.  
The time of drug administration and the time of biopsy will be documented. 
 
 Please refer to Section 9 and Appendix D for more details. 
 
 
 
 
 
 


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2.8 5.3 General Concomitant Medication and Supportive Care Guidelines 
 
Because there is a potential for interaction of AZD1775 with other concomitantly administered 
drugs, the case report form must capture the concurrent use of all other drugs, over-the-counter 
medications, or alternative therapies.  The Principal Investigator should be alerted if the patient is 
taking any agent known to affect or with the potential for drug interactions. 
  
Concomitant treatment with aprepitant and fosaprepitant is not allowable per protocol until 
further evaluation.  
 
All concomitant medications received within 14 days before the first dose of study medication 
and 30 days after the last dose of study medication should be recorded. Concomitant medications 
must be recorded in the appropriate sections of the CRF. 
 
Mandatory prophylactic anti-emetics will be required for all patients. Treatment with the anti-
emetics aprepitant [Emend] and fosaprepitant are excluded because of known drug-drug 
interactions. In addition, loperamide (Imodium) is required at the first onset of diarrhea 
according to ASCO guidelines. Oral loperamide (Imodium) 4mg should be administered at the 
first onset of diarrhea and then 2mg every 2 hours until diarrhea-free for at least 12 hours. The 
first dose of loperamide could be lowered to 2mg if the diarrhea is recurrent and if, in the opinion 
of the treating physician, the diarrhea is not severe. 
Medications may be administered for maintenance of existing conditions prior to study 
enrollment or for a new condition that develops while on study, including but not limited to the 
following: 
 
•   Bisphosphonates and receptor activator of nuclear factor kappa-B ligand (RANKL) 
inhibitors (e.g. denosumab). 
•   Patients requiring therapeutic warfarin or coumarin-derivative anticoagulants will be 
monitored with international normalized ratio (INR) and prothrombin time (PT) as clinically 
indicated. 
•   Low molecular weight heparin (LMWH), rivaroxaban, or equivalent anticoagulant therapy is 
permitted where clinically indicated. 
•   Patients may receive treatment with megestrol acetate when prescribed for appetite 
stimulation. 
  
 
 


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2.9 5.4 Restrictions during the study 
 
The following restrictions are considered the minimum standards for each study within the 
project.  Additional restrictions may be included depending on individual study requirements. 
 
• AZD1775 should be taken with 8 ounces of water approximately 2 hours before or 2 hours 
after food.  
• In once a day dosing if a patient misses the daily dose according to the schedule, the dose 
should be taken as soon as possible, but not more than 12 hours after the missed dose was 
scheduled. If greater than 12 hours, the missed dose should be skipped and the patient should 
take the next dose when scheduled 
• If vomiting occurs after a patient takes the AZD1775 dose, the patient should be instructed 
not to retake the dose, but to wait until the next scheduled dose of AZD1775.  If no dose is 
scheduled for the following day, the dose will not be ‘made up’.  If vomiting persists, the 
patient should contact the Investigator. 
• Women of childbearing potential (WoCBP) may be included only if acceptable contraception 
is in place for two weeks before study entry, for the duration of the treatment with the study 
drug and for 1 month after the last dose of AZD1775  
• WoCBP defined as: Women between menarche and menopause who have not been 
permanently or surgically sterilized and are capable of procreation. 
• All WoCBP must have a negative pregnancy test within 3 days prior to study entry and prior 
to starting each treatment cycle. 
• Male patients who are involved in the study must agree to avoid procreative and unprotected 
sex and must not donate sperm during the study and for 3 months after the last dose of 
AZD1775.  Where the female partner is pregnant or not using effective birth control, men 
should be advised to abstain while in the study and for 3 months after the last dose of 
AZD1775. 
• Female partners, who are of child-bearing potential, of men participating in clinical studies of 
AZD1775 will also be required to use effective contraceptive measures (detailed in Appendix 
D) while their partner is on study drug and for 3 months thereafter. 
• Male patients will be advised to arrange for the freezing of sperm samples prior to the start of 
the study should they wish to father children while on AZD1775 or during the 3 months after 
stopping AZD1775.  
 


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Prohibited concomitant medications 
 
The following treatments and the medications listed in Appendix D are prohibited or should be 
used with caution while in AZD1775 studies.  Any further questions regarding concomitant 
treatments should be referred to a Medical Monitor. 
• No formal clinical drug interaction studies have been performed with AZD1775. An 
exploratory assessment of the effect of aprepitant on AZD1775 exposure in oncology 
patients suggests that there is a drug interaction between AZD1775 and aprepitant, as 
exposure to AZD1775 increased by ~60% when aprepitant was co-administered with 
AZD1775. The observed increase in AZD1775 exposure is likely the result of CYP3A4 
inhibition by aprepitant. This increase in exposure is statistically significant. At the 
selected MTDs, this increase may also be of clinical importance. Therefore, concomitant 
treatment with aprepitant and fosaprepitant is not allowable per protocol until further 
evaluation.   
 
Potent or moderate inhibitors or inducers of CYP3A4, sensitive CYP3A4 substrates, and 
CYP3A4 substrates with a narrow therapeutic window should be avoided until additional 
data on drug-drug interactions (DDI) becomes available. The use of sensitive substrates 
of CYP3A4, such as atorvastatin, simvastatin and lovastatin, is prohibited in this study. 
As grapefruit and Seville oranges are known to contain moderate inhibitors of CYP3A4, 
these fruits or their products (including marmalade, juice, etc.) should be avoided while 
taking AZD1775. 
 
• In vitro data suggests that AZD1775 may also be a weak reversible inhibitor of 
CYP2C19.  Caution should be exercised with concomitant administration of AZD1775 
and agents that are sensitive substrates of CYP2C19, or substrates of this enzyme with 
narrow therapeutic range; refer to Appendix D for a list of sensitive substrates of 
CYP2C19, or substrates of this enzyme with narrow therapeutic range. 
 
• AZD1775 has been shown to be a weak inducer of CYP1A2 in vitro with a maximum 
measured response between donors of 39.9% to 93.1% (at 10 μM) and 18.6% to 32.5% 
(at 5 μM) of the positive control omeprazole (50 μM). Given the nature of the AZD1775 
dosing schedule, however, the risk of induction in the clinic is considered low. No 
specific precautions are recommended at this time, except to be initially vigilant when 
using substrates of CYP1A2 with a narrow therapeutic range. 
 
• In vitro studies have shown that AZD1775 may be a substrate and inhibitor for human P-
glycoprotein (P-gp). Caution should be exercised when agents that are inhibitors or 
substrates of P-gp are administered concomitantly with AZD1775 (see Appendix D). 
 
• in vitro transporter studies have shown AZD1775 to be an inhibitor of BCRP (IC50 5.1 
μM). This finding is particularly relevant for drugs administered orally where exposure is 
normally limited by BCRP-mediated efflux, in particular some statins, such as 
rosuvastatin. Other drugs where the disposition is mediated via BCRP should be 


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administered with caution, dose modification considered or substituted by an alternative 
drug. 
 
• Metformin should be used with caution. AZD1775 has been shown to be an inhibitor of 
MATE1 and MATE2K transporters. A drug interaction with substrates of either 
transporter cannot be ruled out, the most important substrate known to date being 
metformin. 
 
• Herbal preparations/medications are not allowed throughout the study. These herbal 
medications include, but are not limited to: St. John’s wort, kava, ephedra (ma huang), 
gingko biloba, dehydroepiandrosterone (DHEA), yohimbe, saw palmetto, and ginseng. 
Patients should stop using these herbal medications 7 days prior to first dose of 
AZD1775. 
 
2.10 5.5 Duration of Therapy 
 
In the absence of treatment delays due to adverse event(s), treatment may continue or until one 
of the following criteria applies: 
 
•  Disease progression 
 
•  Intercurrent illness that prevents further administration of treatment 
 
•  Unacceptable adverse event(s) 
 
•  Patient decides to withdraw from the study 
 
•  General or specific changes in the patient's condition render the patient unacceptable for 
further treatment in the judgment of the investigator 
 
•  Clinical progression 
 
•  Patient non-compliance 
 
•  Pregnancy 
 
All women of child bearing potential should be instructed to contact the investigator 
immediately if they suspect they might be pregnant (e.g., missed or late menstrual period) at 
any time during study participation. 
 
The investigator must immediately notify CTEP in the event of a confirmed pregnancy in a 
patient participating in the study. 
 
• Termination of the study by sponsor 
 


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• The drug manufacturer can no longer provide the study agent 
 
The reason(s) for protocol therapy discontinuation, the reason(s) for study removal, and the 
corresponding dates must be documented in the Case Report Form (CRF). A patient that decides 
to discontinue investigational product (IP) should be asked about the reason(s) and the 
occurrence of any AEs. 
   
 
 


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2.11 5.6 Duration of Follow Up 
 
•   Patients will be followed for 4 weeks after discontinuing study treatment or until death, 
whichever occurs first. 
 
•   Patients who discontinue the study treatment for reasons other than disease progression, will 
continue to undergo radiographic tumor assessment until radiographic documentation of disease 
progression every 12 weeks, or earlier clinically indicated, or start of new therapy, or until 
overall study completion, whichever occurs first. 
 
•   Patients who discontinue the study treatment for unacceptable adverse event(s) will be 
followed until resolution or stabilization of the adverse event, or until radiographic 
documentation 
 
  
 
 


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2.12 5.7 Criteria for Removal from Study 
 
Patients will be removed from study when any of the criteria listed in Section 5.6 applies. The 
reason for study removal and the date the patient was removed must be documented in the Case 
Report Form. 
 
In the event of unusual or life-threatening complications, participating investigators must 
immediately notify the Principal Investigator, Siqing Fu, MD, PhD at 713-404-1141 (Pager). 
 
 
 
 


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6.  DOSING DELAYS/DOSE MODIFICATIONS 
 
The principal investigator is responsible for ensuring that all staff involved in the study is 
familiar with the content of this section.6.1  Definition of adverse events  
 
An adverse event is the development of an undesirable medical condition or the deterioration of 
a pre-existing medical condition following or during exposure to a pharmaceutical product, 
whether or not considered causally related to the product.  An undesirable medical condition can 
be symptoms (e.g., nausea, chest pain), signs (e.g., tachycardia, enlarged liver) or the abnormal 
results of an investigation (e.g., laboratory findings, electrocardiogram).  In clinical studies, an 
AE can include an undesirable medical condition occurring at any time, including run-in or 
washout periods, even if no study treatment has been administered. 
 
The term AE is used to include both serious and non-serious AEs. 
 
 
 Definitions of serious adverse event 
 
A serious adverse event is an AE occurring during any study phase (i.e., run-in, treatment, 
washout, follow-up), that fulfils one or more of the following criteria: 
• Results in death 
• Is immediately life-threatening 
• Requires in-patient hospitalization or prolongation of existing hospitalization  
• Results in persistent or significant disability/incapacity or substantial disruption of the ability 
to conduct normal life functions 
• Is a congenital abnormality or birth defect 
• Is an important medical event that may jeopardize the subject or may require medical 
intervention to prevent one of the outcomes listed above. 
 
The causality of SAEs (their relationship to all study treatment/procedures) will be assessed by 
the investigator(s) and communicated to AstraZeneca. 
 
 


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 Dose Modification 
 
Toxicity will be assessed utilizing the NCI CTCAE v5.0 
(http://evs.nci.nih.gov/ftp1/CTCAE/CTCAE), unless otherwise specified. 
 
Dose adjustments will be based on the organ system exhibiting the greatest degree of toxicity. 
Dose reductions or holds and initiation of supportive care are allowed as clinically indicated by 
the treating physician. A maximum of 2 dose reductions for AZD1775 will be allowed. Patients 
requiring >2 dose reductions will be discontinued from the study drug. 
 
Table 5 AZD1775 Dose Level Reductions for Toxicity 
 
Dose Level 
AZD1775 
Schedule 21-day cycle 
Starting Dose 
Dose Level -1 
Dose Level -2 
300 mg QD PO  
250 mg QD PO 
200 mg QD PO 
Days 1-5 and 8-12 
Days 1-5 and 8-12 
Days 1-5 and 8-12 
 
 
 
Any patient requiring a toxicity-related dose delay of more than 21 days from the intended day of 
the next scheduled dose must be discontinued from the study unless there is approval from the 
Medical Monitor for the patient to continue. 
 
 
 
 


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2.13 Dose modifications due to hematologic toxicity 
 
Complete blood counts (CBC) will be obtained for all patients at the beginning of each treatment 
cycle (Day 1).  If hematologic toxicity occurs (see Table6 and Table 7), treatment should be held 
and ANC and platelets should be monitored weekly (or more often as clinically indicated) until 
recovery. 
 
Table 6:   Day 1 Hematologic Dose Modifications 
Treatment Day Blood Counts and Toxicity 
ANC 
 
Platelets 
Action 
≥1000/μL 
And 
≥75,000/μL 
No dose modification or interruption 
<1000/μL 
Or 
<75,000/μL 
Delay by 1 week intervals until recovery 
 
If hematologic toxicity parameters do not recover within 21 days, the patient should be 
removed from the study treatment.  
 
 
Table 7:   Neutropenia, Infection, Febrile Neutropenia Dose Modifications and Management 
Any Day 
 
Grade 3 neutropenic fever (ANC <1000/µL + Temperature 
≥101°F [38.5°C]) or neutropenic infection 
Documented infection with Grade 3 neutropenia (ANC 
<1000/µL)  
Grade 4 neutropenia 
(ANC <500/μL >7 days 
Grade 4 thrombocytopenia (platelet count <25,000/μL >7 days) 
Hold dose until recovery.   
Then, upon resuming dosing, 
reduce AZD1775 to the next 
lower dose levela.   
 
Grade 4 febrile neutropenia or Grade 4 infection with 
neutropenia (both defined as septic shock) 
Thrombocytopenic haemorrhage (gross occult bleeding) 
associated with a platelet count <50,000/μL 
Discontinue treatment is 
recommended, and follow for 
disease progression. 
However patient may continue 
after dose reductions only when 
treating physician, the study 
chair and the patient concur. 
 
a No more than two dose reductions will be allowed for any patient.  Patients requiring additional 
dose modifications due to toxicity will discontinue study treatment. 
 
 
 
 


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2.14 Dose modifications due to non-hematologic toxicity  
 
Substantial acute toxicities should be managed as medically indicated and with temporary 
suspension of investigational product, as appropriate. Dose reductions or holds and initiation of 
supportive care are allowed as clinically indicated by the treatment physician.  
 
Dose reductions of AZD1775 should be considered only if toxicity is considered to be related to 
AZD1775. Dose re-escalation is not permitted.  
 
In general, if a patient experiences a Grade 1 / Grade 2 non-hematological toxicity, no dose 
modification is required (except QTc prolongation, see Table 8in ). If a patient experiences a G3 
or G4 toxicity which is not attributable to the disease or disease related processes under 
investigation, dosing will be interrupted and/or the dose reduced and supportive therapy 
administered as required.  Any patient who develops a Grade 3 or 4 non-hematologic toxicity 
that does not resolve to ≤ Grade 1 within 21 days should be removed from the study treatment 
unless approved by the Medical Monitor.   
 
 
Dose modification for QTc interval prolongation 
Table 8:  AZD1775 dose modifications for QTc interval prolongation 
Electrocardiogram QT corrected interval prolonged 
QTc Value 
AZD1775 
QTc 450-480 ms (males) or 
470-480 (females) 
Hold.  Once QTc interval has returned to pre-treatment 
status and correction of possible electrolyte imbalance has 
been made, resume at next lower dose level. 
QTc 481-500 ms 
Hold. Seek cardiologist advice. 
QTc ≥ 501 ms  
Discontinue treatment 
Shift from baseline of ≥ 60ms 
Discontinue treatment 
 
 
Diarrhea 
Due to frequent reports of diarrhea with AZD1775 administration, vigorous anti-diarrheal 
treatment loperamide (Imodium) is required at the first onset of diarrhea according to American 
Society of Clinical Oncology (ASCO) guidelines. Oral loperamide (Imodium) 4 mg should be 
administered every 2 hours until diarrhea-free for at least 12 hours. The first dose of loperamide 
could be lowered to 2 mg if the diarrhea is recurrent and if, in the opinion of the treating 
physician, the diarrhea is not severe.   
 
Patients should be instructed to notify the Investigator or research staff of the occurrence of 
bloody or black stools, symptoms of dehydration, fever, inability to take liquids by mouth, and 
inability to control diarrhea within 24 hours of using loperamide or other prescribed antidiarrheal 
medications. 
 


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If diarrhea is severe (i.e., requiring intravenous [IV] rehydration) and/or associated with fever or 
severe neutropenia (Grade 3 or 4), broad-spectrum antibiotics must be prescribed. Patients with 
severe diarrhea or any diarrhea associated with severe nausea or vomiting should be hospitalized 
for IV hydration and correction of electrolyte imbalances. Guidelines for dose modification of 
AZD1775 will be considered as seen in Table 8. 
 
Table 9:  AZD1775 dose modifications for diarrhea 
Diarrhea 
Management/Next Dose for AZD1775 
≤ Grade 1 
No change in dose 
Grade 2 
Hold until ≤ Grade 1.  Resume at same dose level. 
Grade 3 
Hold* until ≤ Grade 1.  Resume at one dose level lower, if 
indicated.** 
Grade 4 
Off protocol therapy 
*Patients requiring a delay of >2 weeks should go off protocol therapy. 
**Patients requiring > two dose reductions should go off protocol therapy. 
Recommended management:  Loperamide antidiarrheal therapy 
Dosage schedule:  4 mg at first onset, followed by 2 mg with each loose motion until 
diarrhea-free for 12 hours (maximum dosage:  16 mg/24 hours) 
Adjunct anti-diarrheal therapy is permitted and should be recorded when used. 
 
 
Nausea and vomiting (mandatory antiemetic prophylaxis) 
All patients must receive a 5-HT3 antagonist, ondansetron (Zofran) 8 mg PO QD or granisetron 
(Kytril) 1 mg PO QD prior to each dose of AZD1775.  If nausea and vomiting continue, a second 
dose of antiemetics can be taken 8 hours later if necessary. In addition, dexamethasone 4 mg PO 
will be given with each AZD1775 dose unless contraindicated or not well-tolerated.  
Dexamethasone or the 5-HT3 antagonist may be given by IV as needed..   
 
Promethazine (Phenergan), prochlorperazine (Compazine), and benzodiazepine may still be used 
as additional adjunctive treatments during AZD1775 therapy.   
 
Please note: aprepitant [Emend] and fosaprepitant are not permitted due to known DDIs. 
Patients should be strongly encouraged to maintain liberal oral fluid intake. 
 
 
Suitable alternative medications may be used, with adequate justification, in those studies where 
the use of any of the above medications might interfere with other study procedures or are 
deemed insufficient.  Guidelines for dose modification of AZD1775 will be considered for 
nausea and vomiting as seen in Table 9 and Table 10, respectively. 
 
Table 10:  AZD1775 dose modifications for nausea 
Nausea 
Management/Next Dose for AZD1775 
≤ Grade 1 
No change in dose 
Grade 2 
Hold until ≤ Grade 1.  Resume at same dose level. 
Grade 3 
Hold* until ≤Grade 1.  Resume at one dose level lower, if indicated.** 


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Table 10:  AZD1775 dose modifications for nausea 
Nausea 
Management/Next Dose for AZD1775 
Grade 4 
Off protocol therapy 
*Patients requiring a delay of >2 weeks should go off protocol therapy. 
**Patients requiring > two dose reductions should go off protocol therapy. 
Recommended management:  antiemetics. 
 
 
Table 11:  AZD1775 dose modifications for vomiting 
Vomiting 
Management/Next Dose for AZD1775 
≤ Grade 1 
No change in dose 
Grade 2 
Hold until ≤ Grade 1.  Resume at same dose level. 
Grade 3 
Hold* until ≤ Grade 1.  Resume at one dose level lower, if indicated.** 
Grade 4 
Off protocol therapy 
*Patients requiring a delay of >2 weeks should go off protocol therapy. 
**Patients requiring > two dose reductions should go off protocol therapy. 
Recommended management:  antiemetics. 
 
 
Febrile neutropenia 
Patients experiencing febrile neutropenia with significant symptoms should be managed in a 
hospital setting according to standard procedures, with the urgent initiation of IV antibiotic 
therapy. Patients with febrile neutropenia without symptoms should be managed according to 
standard guidelines. 
 
 


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2.15 Recording of adverse events 
 
Time period for collection of adverse events 
Adverse events will be collected throughout the study, from informed consent until 30 days after 
the end of the last investigational product administration. 
 
SAEs will be recorded from the time of informed consent and should be reported to CTEP.  
 
 
Follow-up of unresolved adverse events 
Any AEs that are unresolved at the patient’s last visit in the study are followed up by the 
Investigator for as long as medically indicated, but without further recording in the eCRF. 
 
 
If an Investigator learns of any SAEs, including death, at any time after a patient has completed 
the study and he/she considers there is a reasonable possibility that the event is related to 
AZD1775, the Investigator should notify CTEP. 
 
The following variables will be collected for each AE: 
• AE (verbatim) 
• The date and time when the AE started and stopped 
• The maximum intensity or intensity or changes in intensity 
• CTCAE grade/max CTAE grade/changes in CTCAE grade 
• Whether the AE is serious or not 
• Investigator causality rating against the Investigational Product (yes or no) 
• Action taken with regard to investigational product 
• AE caused subject’s withdrawal from study (yes or no) 
• Outcome. 
 
In addition, the following variables will be collected for SAEs: 
• Date AE met criteria for serious AE 
• Date Investigator became aware of serious AE 
• Reason AE is serious  
• Date of hospitalization 
• Date of discharge 
• Probable cause of death 
• Date of death 
• Autopsy performed 
• Causality assessment in relation to Study procedure(s) 
• Causality assessment in relation to Other medication 
• Causality assessment in relation to module-specific combination treatments 
• Description of AE. 
 
The grading scales found in the revised CTCAE Version 5.0 will be utilized for all events with 


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an assigned CTCAE grading. For those events without assigned CTCAE grades, the 
recommendation in the CTCAE criteria should be utilized that converts mild, moderate, and 
severe events. A copy of the current CTCAE version can be downloaded from the Cancer 
Therapy Evaluation Program website (http://ctep.cancer.gov). 
 
It is important to distinguish between serious and severe AEs.  Severity is a measure of intensity 
whereas seriousness is defined by the criteria in Section 7.3.  An AE of severe intensity need not 
necessarily be considered serious.  For example, nausea that persists for several hours may be 
considered severe nausea, but not a SAE. On the other hand, a stroke that results in only a 
limited degree of disability may be considered a mild stroke but would be a SAE. 
 
 
Adverse Events based on signs and symptoms 
All AEs spontaneously reported by the patient, reported in response to the open question from 
the study personnel, or revealed by observation will be collected and recorded in the eCRF. 
 
When collecting AEs, the recording of a diagnosis is preferred (when possible) to the recording 
of a list of signs and symptoms. However, if a diagnosis is known and there are other signs or 
symptoms that are not generally part of the diagnosis, the diagnosis and each sign or symptom 
will be recorded separately. 
 
They do not include metastases of the original cancer. 
 
 
Adverse Events based on examinations and tests 
The results from protocol mandated laboratory tests and vital signs will be summarised in the 
CSR.  Deterioration as compared to baseline in protocol-mandated laboratory values, vital signs 
and ECG should therefore only be reported as AEs if they fulfil any of the SAE criteria or are the 
reason for discontinuation of the study treatment unless clearly due to the progression of disease 
under study. 
 
If deterioration in a laboratory value/vital sign/ECG is associated with clinical signs and 
symptoms, the sign or symptom will be reported as an AE and the associated laboratory 
result/vital sign/ECG will be considered as additional information. Wherever possible the 
reporting Investigator uses the clinical, rather than the laboratory term (e.g., anemia versus low 
hemoglobin value). In the absence of clinical signs or symptoms, clinically relevant 
deteriorations in non-mandated parameters should be reported as AE(s). 
Deterioration of a laboratory value, which is unequivocally due to disease progression, should 
not be reported as an AE/SAE. 
 
Any new or aggravated clinically relevant abnormal medical finding at a physical examination as 
compared with the baseline assessment will be reported as an AE. 
 
Examples of these are marked hematological and other laboratory abnormalities, and certain 
events that lead to intervention (other than those already classified as serious), dose reduction or 
significant additional treatment. 


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Hy’s Law 
Cases where a patient shows elevations in liver biochemistry may require further evaluation and 
occurrences of AST or ALT ≥3 x ULN together with total bilirubin ≥2 x ULN may need to be 
reported as SAEs.  
 
 
Disease progression 
Disease progression can be considered as a worsening of a subject’s condition attributable to the 
disease for which the investigational product is being studied.  It may be an increase in the 
severity of the disease under study and/or increases in the symptoms of the disease.  (For 
example, the progression of existing metastasis to the primary cancer under study) should be 
considered as disease progression and not an AE.  Events, which are unequivocally due to 
disease progression, should not be reported as an AE during the study. 
 
 
New cancers 
The development of a new cancer should be regarded as an AE and will generally meet at least 
one of the serious criteria. New cancers are those that are not the primary reason for the 
administration of the study treatment and have been identified after the patient’s inclusion in this 
study. They do not include metastases of the original cancer. 
 
Investigators must report SAEs and follow-up information to their responsible IRB according to 
the policies of the responsible IRB. For fatal or life-threatening AEs where important or relevant 
information is missing, active follow-up is undertaken immediately. 
 
 


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Laboratory safety assessment 
Blood and samples for determination of clinical chemistry, haematology, coagulation, and PD 
will be performed at the times indicated in the Study Plan (Table 12). 
 
Additional safety samples may be collected as clinically indicated and at the discretion of the 
Investigator. The date, time of collection and results (values, units and reference ranges) will be 
recorded on the appropriate eCRF. 
 
The clinical chemistry and haematology will be performed at a CLIA-certified laboratory. 
Sample tubes and sample sizes may vary depending on laboratory method used and routine 
practice at the study centre.  
 
The following laboratory variables will be measured as a minimum (some of these variables may 
be measured at baseline only): 
 
  Table 12:  Laboratory Safety Variables 
Hematology (2.7 mL whole blood 
sample) 
Clinical chemistry (2.7 mL serum or 
plasma sample) 
B-Hemoglobin 
S/P-Albumin 
B-Leukocyte  
S/P-Alanine transaminase (ALT) 
B-Hematocrit 
S/P-Aspartate transaminase (AST) 
B-Red blood cell count 
S/P-Alkaline phosphatase (ALP) 
B-Absolute leukocyte differential count 
S/P-Bilirubin, total 
Neutrophils 
S/P-Calcium, total 
Lymphocytes 
S/P-Creatinine 
Monocytes 
S/P-Chloride  
S/P-Glucose 
S/P-Magnesium 
Basophils 
S/P-Potassium  
Eosinophils 
S/P-Sodium  
B-Platelet count 
S/P-Urea nitrogen or blood urea nitrogen 
S/P-TSH 
Coagulation (1.8 mL sample) 
S/P-Free T4 
S/P-CO2 
S/P-BUN 
B-PT or INR with PTT  
Pregnancy test (Blood or urine) 
 
Pre-menopausal women of childbearing potential must have a negative urine or serum pregnancy 
test within 3 days prior to starting study treatment, and a confirmatory test before treatment 
before each treatment period and each subsequent check-in prior to administration of study 
treatment in the next period. In the event of suspected pregnancy during the study, the test should 
be repeated and, if positive, the patient discontinued from study treatment immediately. 
The Investigator should make an assessment of the available results with regard to clinically 
relevant abnormalities. Any clinically significant abnormal laboratory values should be repeated 
as clinically indicated and recorded on the eCRF. The laboratory results should be signed and 
dated and retained at centre as source data for laboratory variables.  


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NB.  In case a patient shows an AST or ALT ≥3 x ULN (upper limit of normal) and total 
bilirubin ≥2 x ULN please refer to Appendix D ‘Actions required in cases of combined increase 
of Aminotransferase and Total Bilirubin – Hy’s Law’, for further instructions. 
 
 
Physical examination 
A complete physical examination will be performed and include an assessment of the following: 
general appearance, respiratory, cardiovascular, abdomen, skin, head and neck (including ears, 
eyes, nose and throat), lymph nodes, thyroid, musculo-skeletal (including spine and extremities) 
and neurological systems.  
 
If new or aggravated physical findings imply deterioration compared with baseline, the finding 
should be reported as an AE. Performance status will be assessed using the ECOG performance 
status criteria. 
 
 
ECG 
Resting 12-lead ECG 
A triplicate 12-lead safety ECG (paper ECG printout of 10 seconds for Investigator review) will 
be taken at screening and prior to dosing of each study cycle, and as clinically indicated.  
 
At any other time the Investigator deems necessary for safety during the administration period, 
triplicate ECG recordings should be taken within an approximate 5-minute period.  Additional 
ECGs may be taken.  The patients will rest for at least 10 minutes before the start of each 
recording and they must be in the same supine body position (maximum 30 degrees flexion in 
the hip and feet not in contact with the footboard) at the recording time point.  
 
The Investigator will judge the overall interpretation as normal or abnormal. If abnormal, it will 
be decided as to whether or not the abnormality is clinically significant or not clinically 
significant. The paper copy of each ECG reading will be retained with the patient’s completed 
source documents. Only overall evaluation (normal/abnormal) will be recorded in the eCRF. If 
there is a clinically significant abnormal unscheduled ECG finding during the treatment period, 
this should be recorded on the AE eCRF, according to standard AE collection and reporting 
processes.  
 
Attention should be paid to any detected increases in QTc interval. Patients who develop a single 
resting value of QTc interval of >450 msec/male and >470 msec/female or a shift from baseline 
of 60ms should stop taking AZD1775. Dosing can be resumed at a reduced dose (see section 
3.4.2.1) after return of the resting QTc interval to pre-treatment status has been confirmed and 
correction of possible electrolyte imbalance has been made. 
 
Monitoring of QTc, checking and correction of abnormal electrolyte levels and renal function are 
advised, especially in case of severe/prolonged diarrhoea.  If QTc increases markedly from 
baseline, but stays below the above limits, a cardiologist’s advice should be sought. 
 
The concomitant use of ondansetron (known to prolong the QTc interval in rare cases, per 
labelling) should be taken into account when interpreting QTc changes. 


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7.   ADVERSE EVENTS:  LIST AND REPORTING REQUIREMENTS  
 
Adverse event (AE) monitoring and reporting is a routine part of every clinical trial.  The 
following list of AEs (Section 7.1) and the characteristics of an observed AE (Section 7.2 and 
7.3) will determine whether the event requires expedited reporting via the CTEP Adverse Event 
Reporting System (CTEP-AERS) in addition to routine reporting. 
 
 
7.1   Comprehensive Adverse Events and Potential Risks List (CAEPRs) 
 
The Comprehensive Adverse Event and Potential Risks list (CAEPR) provides a single list of 
reported and/or potential adverse events (AE) associated with an agent using a uniform 
presentation of events by body system.  In addition to the comprehensive list, a subset of AEs, 
the Specific Protocol Exceptions to Expedited Reporting (SPEER), appears in a separate column 
and is identified with bold and italicized text.  The SPEER is a list of events that are protocol-
specific exceptions to expedited reporting to NCI (except as noted below).  Refer to the 'CTEP, 
NCI Guidelines: Adverse Event Reporting Requirements' 
http://ctep.cancer.gov/protocolDevelopment/adverse_effects.htm for further clarification.  
 
NOTE:  The highest grade currently reported is noted in parentheses next to the AE in the 
SPEER.  Report ONLY AEs higher than this grade expeditiously.  If this CAEPR is part of a 
combination protocol using multiple investigational agents and has an AE listed on different 
SPEERs, use the lower of the grades to determine if expedited reporting is required. 
 
7.1.1 CAEPR for AZD1775 
 
Comprehensive Adverse Events and Potential Risks list (CAEPR) for AZD1775 
(adavosertib, NSC 751084) 
 
 
The Comprehensive Adverse Events and Potential Risks list (CAEPR) provides a single list of reported and/or 
potential adverse events (AE) associated with an agent using a uniform presentation of events by body system. In 
addition to the comprehensive list, a subset, the Specific Protocol Exceptions to Expedited Reporting (SPEER), 
appears in a separate column and is identified with bold and italicized text. This subset of AEs (SPEER) is a list of 
events that are protocol specific exceptions to expedited reporting to NCI (except as noted below).  Refer to the 
'CTEP, NCI Guidelines: Adverse Event Reporting Requirements' 
http://ctep.cancer.gov/protocolDevelopment/electronic_applications/docs/aeguidelines.pdf for further clarification. 
Frequency is provided based on 323 patients. Below is the CAEPR for AZD1775 (adavosertib). 
NOTE: Report AEs on the SPEER ONLY IF they exceed the grade noted in parentheses next to the AE in the 
SPEER.  If this CAEPR is part of a combination protocol using multiple investigational agents and has an AE 
listed on different SPEERs, use the lower of the grades to determine if expedited reporting is required. 
 
 
 


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Table 13: CAEPR for AZD1775 (Adavosertib, NSC 751084) 
 
Version 2.7, April 27, 20201 
 
 Adverse Events with Possible  
 Relationship to AZD1775 (adavosertib) 
 (CTCAE 5.0 Term) 
[n= 323] 
 
 
 Specific Protocol Exceptions 
to Expedited Reporting  
Likely (>20%) 
Less Likely (<=20%) 
Rare but Serious (<3%) 
 
 
BLOOD AND LYMPHATIC SYSTEM DISORDERS 
  
 
Anemia 
 
 Anemia (Gr 3) 
 
 
Febrile neutropenia 
  
CARDIAC DISORDERS 
  
 
 
Atrial fibrillation 
  
 
 
Supraventricular tachycardia 
  
GASTROINTESTINAL DISORDERS 
  
 
Abdominal pain 
 
 Abdominal pain (Gr 2) 
 
Constipation 
 
 Constipation (Gr 2) 
Diarrhea 
 
 
 Diarrhea (Gr 3) 
 
Dyspepsia 
 
  
 
 
Gastrointestinal hemorrhage2 
  
 
Mucositis oral 
 
 Mucositis oral (Gr 2) 
Nausea 
 
 
 Nausea (Gr 3) 
Vomiting 
 
 
 Vomiting (Gr 3) 
GENERAL DISORDERS AND ADMINISTRATION SITE CONDITIONS 
  
 
Edema limbs 
 
 Edema limbs (Gr 2) 
Fatigue 
 
 
 Fatigue (Gr 3) 
 
Fever 
 
 Fever (Gr 2) 
HEPATOBILIARY DISORDERS 
  
 
 
Hepatobiliary disorders - Other 
(hepatitis) 
  
INFECTIONS AND INFESTATIONS 
  
 
Infection3 
 
 Infection3 (Gr 3) 
INVESTIGATIONS 
  
 
Alanine aminotransferase increased  
 Alanine aminotransferase 
increased (Gr 3) 
 
 
Electrocardiogram QT corrected 
interval prolonged 
  
 
Lymphocyte count decreased 
 
  
 
Neutrophil count decreased 
 
 Neutrophil count decreased (Gr 4) 
 
Platelet count decreased 
 
 Platelet count decreased (Gr 4) 
 
Weight loss 
 
  
 
White blood cell decreased 
 
 White blood cell decreased (Gr 4) 
METABOLISM AND NUTRITION DISORDERS 
  
 
Anorexia 
 
 Anorexia (Gr 2) 
 
Dehydration 
 
  
 
Hypokalemia 
 
 Hypokalemia (Gr 2) 
 
Hypomagnesemia 
 
 Hypomagnesemia (Gr 2) 
MUSCULOSKELETAL AND CONNECTIVE TISSUE DISORDERS 
  
 
Back pain 
 
 Back pain (Gr 2) 
 
Muscle cramp 
 
  
 
Myalgia 
 
 Myalgia (Gr 2) 
NERVOUS SYSTEM DISORDERS 
  
` 
Dizziness 
 
 Dizziness (Gr 2) 


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 Adverse Events with Possible  
 Relationship to AZD1775 (adavosertib) 
 (CTCAE 5.0 Term) 
[n= 323] 
 
 
 Specific Protocol Exceptions 
to Expedited Reporting  
Likely (>20%) 
Less Likely (<=20%) 
Rare but Serious (<3%) 
 
 
 
Headache 
 
 Headache (Gr 2) 
 
 
Intracranial hemorrhage 
  
PSYCHIATRIC DISORDERS 
  
 
Insomnia 
 
  
RESPIRATORY, THORACIC AND MEDIASTINAL DISORDERS 
  
 
Cough 
 
 Cough (Gr 2) 
 
Dyspnea 
 
 Dyspnea (Gr 2) 
 
 
Hypoxia 
  
SKIN AND SUBCUTANEOUS TISSUE DISORDERS 
  
 
Rash4 
 
 Rash4 (Gr 2) 
VASCULAR DISORDERS 
  
 
 
Phlebitis 
  
 
 
1This table will be updated as the toxicity profile of the agent is revised.  Updates will be distributed to all Principal 
Investigators at the time of revision.  The current version can be obtained by contacting 
PIO@CTEP.NCI.NIH.GOV.  Your name, the name of the investigator, the protocol and the agent should be 
included in the e-mail. 
 
2Gastrointestinal hemorrhage includes Anal hemorrhage, Cecal hemorrhage, Colonic hemorrhage, Duodenal 
hemorrhage, Esophageal hemorrhage, Esophageal varices hemorrhage, Gastric hemorrhage, Hemorrhoidal 
hemorrhage, Ileal hemorrhage, Intra-abdominal hemorrhage, Jejunal hemorrhage, Lower gastrointestinal 
hemorrhage, Oral hemorrhage, Pancreatic hemorrhage, Rectal hemorrhage, Retroperitoneal hemorrhage, and Upper 
gastrointestinal hemorrhage under the GASTROINTESTINAL DISORDERS SOC. 
 
3Infection includes all 75 sites of infection under the INFECTIONS AND INFESTATIONS SOC. 
 
4Rash may include rash, erythema, eczema, and rash maculo-papular. 
 
5Peripheral neuropathy includes both peripheral motor neuropathy and peripheral sensory neuropathy. 
 
6Acute kidney injury includes renal impairment and acute renal insufficiency. 
 
 
 
Adverse events reported on AZD1775 (adavosertib) trials, but for which there is insufficient evidence to 
suggest that there was a reasonable possibility that AZD1775 (adavosertib) caused the adverse event: 
 
BLOOD AND LYMPHATIC SYSTEM DISORDERS - Blood and lymphatic system disorders - Other 
(pancytopenia); Blood and lymphatic system disorders - Other (thrombocytosis); Blood and lymphatic system 
disorders - Other (right leg deep vein thrombosis); Leukocytosis 
Cardiac disorders - Cardiac disorders - Other (cardiomegaly); Chest pain - cardiac; Myocardial infarction; 
Palpitations; Sinus bradycardia; Sinus tachycardia 
EAR AND LABYRINTH DISORDERS - Ear pain; Hearing impaired; Tinnitus 
EYE DISORDERS - Blurred vision; Cataract; Eye disorders - Other (eye swelling); Eye pain; Keratitis; 
Photophobia; Scleral disorder; Vision decreased; Watering eyes 
GASTROINTESTINAL DISORDERS - Abdominal distension; Anal pain; Ascites; Belching; Bloating; Cheilitis; 
Colitis; Colonic obstruction; Dry mouth; Duodenal ulcer; Dysphagia; Enterocolitis; Flatulence; Gastric ulcer; 
Gastritis; Hemorrhoids; Oral pain; Rectal pain; Small intestinal obstruction 


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GENERAL DISORDERS AND ADMINISTRATION SITE CONDITIONS - Chills; Death NOS; Edema trunk; 
Flu like symptoms; Gait disturbance; General disorders and administration site conditions - Other (catheter site 
pain); Infusion site extravasation; Malaise; Non-cardiac chest pain; Pain 
IMMUNE SYSTEM DISORDERS - Allergic reaction; Anaphylaxis; Cytokine release syndrome 
INJURY, POISONING AND PROCEDURAL COMPLICATIONS - Fall; Injury, poisoning and procedural 
complications - Other (excoriation); Injury, poisoning and procedural complications - Other (ligament sprain) 
INVESTIGATIONS - Alkaline phosphatase increased; Aspartate aminotransferase increased; Blood bilirubin 
increased; Creatinine increased; GGT increased; Investigations - Other (blood urea increased); Lymphocyte count 
increased 
METABOLISM AND NUTRITION DISORDERS - Alkalosis; Hypercalcemia; Hyperglycemia; Hyperkalemia; 
Hyperuricemia; Hypoalbuminemia; Hypocalcemia; Hyponatremia; Hypophosphatemia; Tumor lysis syndrome 
MUSCULOSKELETAL AND CONNECTIVE TISSUE DISORDERS - Arthralgia; Arthritis; Bone pain; Flank 
pain; Generalized muscle weakness; Muscle weakness lower limb; Musculoskeletal and connective tissue disorder - 
Other (groin pain); Neck pain; Pain in extremity 
NEOPLASMS BENIGN, MALIGNANT AND UNSPECIFIED (INCL CYSTS AND POLYPS) - Neoplasms 
benign, malignant and unspecified (incl cysts and polyps) - Other (carcinoid tumor); Tumor pain 
NERVOUS SYSTEM DISORDERS - Central nervous system necrosis; Cognitive disturbance; Depressed level of 
consciousness; Dysesthesia; Dysgeusia; Encephalopathy; Lethargy; Nervous system disorders - Other (hemiparesis); 
Paresthesia; Peripheral neuropathy5; Presyncope; Somnolence; Syncope 
PSYCHIATRIC DISORDERS - Agitation; Anxiety; Confusion; Depression 
RENAL AND URINARY DISORDERS - Acute kidney injury6; Hematuria; Urinary frequency; Urinary 
incontinence; Urinary retention; Urinary tract pain 
REPRODUCTIVE SYSTEM AND BREAST DISORDERS - Genital edema; Reproductive system and breast 
disorders - Other (female genital tract fistula) 
RESPIRATORY, THORACIC AND MEDIASTINAL DISORDERS - Allergic rhinitis; Apnea; 
Bronchopulmonary hemorrhage; Epistaxis; Hiccups; Nasal congestion; Pleural effusion; Pneumonitis; Pulmonary 
hypertension; Respiratory, thoracic and mediastinal disorders - Other (diaphragmalgia); Voice alteration; Wheezing 
SKIN AND SUBCUTANEOUS TISSUE DISORDERS - Alopecia; Bullous dermatitis; Dry skin; Hyperhidrosis; 
Pain of skin; Palmar-plantar erythrodysesthesia syndrome; Pruritus; Purpura; Rash acneiform; Skin ulceration; 
Urticaria 
VASCULAR DISORDERS - Flushing; Hematoma; Hot flashes; Hypertension; Hypotension; Thromboembolic 
event 
 
 
 
Note: AZD1775 (adavosertib) in combination with other agents could cause an exacerbation of any adverse event 
currently known to be caused by the other agent, or the combination may result in events never previously 
associated with either agent. 
 
7.2   Adverse Event Characteristics  
 
•  CTCAE term (AE description) and grade:  The descriptions and grading scales found in the 
revised NCI Common Terminology Criteria for Adverse Events (CTCAE) version 5.0 will be 
utilized for AE reporting.  All appropriate treatment areas should have access to a copy of the 
CTCAE version 5.0.  A copy of the CTCAE version 5.0 can be downloaded from the CTEP web 
site http://ctep.cancer.gov/protocolDevelopment/electronic_applications/ctc.htm. 
 
•  For expedited reporting purposes only:   
− 
AEs for the AZD1775 that are bold and italicized in the CAEPR (i.e., those listed in the 
SPEER column, Section 7.1) should be reported through CTEP-AERS only if the grade is 
above the grade provided in the SPEER. 
− 
Other AEs for the NCI Protocol #: 10136 that do not require expedited reporting are 
outlined in section 7.3. 


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64 
 
 
•  Attribution of the AE: 
− 
Definite – The AE is clearly related to the study treatment. 
− 
Probable – The AE is likely related to the study treatment. 
− 
Possible – The AE may be related to the study treatment. 
− 
Unlikely – The AE is doubtfully related to the study treatment. 
− 
Unrelated – The AE is clearly NOT related to the study treatment. 
 
 
 


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7.3   Expedited Adverse Event Reporting 
 
Expedited AE reporting for this study must use CTEP-AERS (CTEP Adverse Event Reporting 
System), accessed via the CTEP Web site (https://eapps-ctep.nci.nih.gov/ctepaers).  The 
reporting procedures to be followed are presented in the “NCI Guidelines for Investigators: 
Adverse Event Reporting Requirements for DCTD (CTEP and CIP) and DCP INDs and IDEs” 
which can be downloaded from the CTEP Web site 
(http://ctep.cancer.gov/protocolDevelopment/electronic_applications/adverse_events.htm).  
These requirements are briefly outlined in the tables below (Table 14). 
 
In the rare occurrence when Internet connectivity is lost, a 24-hour notification is to be made to 
CTEP by telephone at 301-897-7497.  Once Internet connectivity is restored, the 24-hour 
notification phoned in must be entered electronically into CTEP-AERS by the original submitter 
at the site. 
 
 
Distribution of Adverse Event Reports 
 
CTEP-AERS is programmed for automatic electronic distribution of reports to the following 
individuals:  Principal Investigator and Adverse Event Coordinator(s) (if applicable) of the 
Corresponding Organization or Lead Organization, the local treating physician, and the Reporter 
and Submitter.  CTEP-AERS provides a copy feature for other e-mail recipients.  
 
The Coordinating Center of the Corresponding Organization is responsible for submitting to the 
CTSU documentation of AEs that they deem reportable for posting on the CTSU protocol web page 
and inclusion on the CTSU bi-monthly broadcast. 
 
 
Expedited Reporting Guidelines 
 
Use the NCI protocol number and the protocol-specific patient ID assigned during trial 
registration on all reports (Table 14). 
 
Note:  A death on study requires both routine and expedited reporting, regardless of 
causality.  Attribution to treatment or other cause must be provided. 
 
Death due to progressive disease should be reported as Grade 5 “Neoplasms benign, malignant 
and unspecified (incl cysts and polyps) - Other (Progressive Disease)” under the system 
organ class (SOC) of the same name.  Evidence that the death was a manifestation of underlying 
disease (e.g., radiological changes suggesting tumor growth or progression: clinical deterioration 
associated with a disease process) should be submitted.  
 
Table 14: Expedited Reporting Requirements for Adverse Events that Occur on Studies under an 
IND/IDE within 30 Days of the Last Administration of the Investigational Agent/Intervention 1, 2 


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FDA REPORTING REQUIREMENTS FOR SERIOUS ADVERSE EVENTS (21 CFR Part 312) 
 
NOTE:  Investigators MUST immediately report to the sponsor (NCI) ANY Serious Adverse Events, whether or not 
they are considered related to the investigational agent(s)/intervention (21 CFR 312.64) 
An adverse event is considered serious if it results in ANY of the following outcomes:   
1) Death 
2) A life-threatening adverse event  
3) An adverse event that results in inpatient hospitalization or prolongation of existing hospitalization for ≥ 24 hours  
4) A persistent or significant incapacity or substantial disruption of the ability to conduct normal life functions  
5) A congenital anomaly/birth defect.  
6) Important Medical Events (IME) that may not result in death, be life threatening, or require hospitalization may 
be considered serious when, based upon medical judgment, they may jeopardize the patient or subject and may 
require medical or surgical intervention to prevent one of the outcomes listed in this definition. (FDA, 21 CFR 
312.32; ICH E2A and ICH E6). 
 
ALL SERIOUS adverse events that meet the above criteria MUST be immediately reported to the NCI via electronic 
submission within the timeframes detailed in the table below. 
Hospitalization 
Grade 1 and Grade 2 Timeframes 
Grade 3-5 
Timeframes 
Resulting in Hospitalization ≥ 24 hrs 
10 Calendar Days 
24-Hour 5 Calendar Days 
Not resulting in Hospitalization ≥ 24 hrs 
Not required 
NOTE:  Protocol specific exceptions to expedited reporting of serious adverse events are found in the Specific 
Protocol Exceptions to Expedited Reporting (SPEER) portion of the CAEPR. 
 
Expedited AE reporting timelines are defined as: 
o “24-Hour; 5 Calendar Days” - The AE must initially be submitted electronically within 24 hours of learning of the 
AE, followed by a complete expedited report within 5 calendar days of the initial 24-hour report. 
o “10 Calendar Days” - A complete expedited report on the AE must be submitted electronically within 10 calendar 
days of learning of the AE. 
1Serious adverse events that occur more than 30 days after the last administration of investigational 
agent/intervention and have an attribution of possible, probable, or definite require reporting as follows:  
Expedited 24-hour notification followed by complete report within 5 calendar days for: 
• All Grade 3, 4, and Grade 5 AEs 
Expedited 10 calendar day reports for: 
Grade 2 AEs resulting in hospitalization or prolongation of hospitalization 
 
2For studies using PET or SPECT IND agents, the AE reporting period is limited to 10 radioactive half-lives, 
rounded UP to the nearest whole day, after the agent/intervention was last administered.  Footnote “1” above 
applies after this reporting period. 
 
Effective Date:  May 5, 2011 
 
 
 
 


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7.4   Routine Adverse Event Reporting 
 
All Adverse Events must be reported in routine study data submissions.  AEs reported 
expeditiously through CTEP-AERS must also be reported in routine study data 
submissions. 
 
Adverse event data collection and reporting, which are required as part of every clinical trial, are 
done to ensure the safety of patients enrolled in the studies as well as those who will enroll in 
future studies using similar agents.  AEs are reported in a routine manner at scheduled times 
during the trial using Medidata Rave.  For this trial the Adverse Event CRF is used for routine 
AE reporting in Rave. 
 
 
7.5   Secondary Malignancy 
 
A secondary malignancy is a cancer caused by treatment for a previous malignancy (e.g., 
treatment with investigational agent/intervention, radiation or chemotherapy). A secondary 
malignancy is not considered a metastasis of the initial neoplasm. 
 
CTEP requires all secondary malignancies that occur following treatment with an agent under an 
NCI IND/IDE be reported expeditiously via CTEP-AERS.  Three options are available to 
describe the event:  
 
•  Leukemia secondary to oncology chemotherapy (e.g., acute myelocytic leukemia [AML])  
•  Myelodysplastic syndrome (MDS)  
•  Treatment-related secondary malignancy  
 
Any malignancy possibly related to cancer treatment (including AML/MDS) should also be 
reported via the routine reporting mechanisms outlined in each protocol.  
 
Indicate form for reporting in Rave, timeframes, and if loading of the pathology report is 
required. 
 
7.6   Second Malignancy 
 
A second malignancy is one unrelated to the treatment of a prior malignancy (and is NOT a 
metastasis from the initial malignancy).  Second malignancies require ONLY routine AE 
reporting unless otherwise specified. 
 
 
 
 


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8. PHARMACEUTICAL INFORMATION 
 
A list of the adverse events and potential risks associated with the investigational agent 
administered in this study can be found in Section 7. 
 
8.1   AZD1775 (NSC #751084) 
 
Chemical Name: 2-allyl-1-[6-(1-hydroxy-1-methylethyl)pyridin-2-yl]-6-([4-(4-methylpiperazin-
1-yl)phenyl]amino)-1,2-dihydro-3H-pyrazolo[3,4-d]pyrimidin-3- one 
 
Classification: Inhibitor of Wee1-kinase 
 
CAS: 955365-80-7 
 
Molecular Formula:  C27H32N8O2·H2O 
M.W.: 518.623 
 
 
Mode of Action: AZD1775 is an inhibitor of the Wee1-kinase. Wee1 is a tyrosine kinase 
upstream of CDC2 thereby involved in regulation of cell cycle checkpoints, particularly the G2 
checkpoint. As the majority of human cancers harbor abnormalities in the p53 pathway they 
become more dependent on S- and G2-phase checkpoints. In preclinical models, AZD1775 
selectively enhanced chemotherapy-induced death of cells deficient in p53 signaling. 
Description: AZD1775 is a crystalline, non-hygroscopic, monohydrate of the neutral drug.  It 
dehydrates upon heating leading to formation of a crystalline anhydrate. 
 
How Supplied: AZD1775 is supplied by AstraZeneca and distributed by the Pharmaceutical 
Management Branch, CTEP/DCTD/NCI as capsules available in 25 mg (yellow color, size 2 
gelatin capsule) and 100 mg (orange color, size 2 gelatin capsule) strengths.  The dry-filled 
capsules consist of a roller-compacted granule of drug substance, lactose monohydrate, 
microcrystalline cellulose, croscarmellose sodium, and magnesium stearate.  Each high-density 
polyethylene (HDPE) bottle contains 20 capsules. 
 
The pharmaceutical collaborator does not have stability data to support repackaging AZD1775 
capsules in any container other than what is provided. 
 
Storage:   
Store at 2 to 30˚C (36 to 86˚F).  Do not freeze. 
 
If a storage temperature excursion is identified, promptly return AZD1775 to between 2-30°C and 
quarantine the supplies.  Provide a detailed report of the excursion (including documentation of 
temperature monitoring and duration of the excursion) to PMBAfterHours@mail.nih.gov for 
determination of suitability. 
Stability:  Shelf life studies of AZD1775 are on-going. 


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Route of Administration: Oral administration. Take AZD1775 at least two hours before or two 
hours after a meal. 
 
Potential Drug Interactions: AZD1775 is primarily metabolized by CYP3A4 and is a weak, 
time-dependent inhibitor of CYP3A4. Avoid concomitant CYP3A4 moderate or strong 
inhibitors/inducers, and sensitive substrates with a narrow therapeutic index. AZD1775 is also a 
weak inhibitor of CYP2C19. Caution should be exercised with concomitant administration of 
sensitive substrates or substrates with a narrow therapeutic index. 
 
In vitro transporter studies have shown that AZD1775 was an inhibitor of OATP1B1, OATP1B3, 
MATE1, MATE2K, P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP), and a 
substrate for P-gp and BCRP. The PK parameters of AZD1775 could be altered if AZD1775 is 
co-administered with P- gp and BCRP inhibitors/inducers, and there is potential for drug-drug 
interactions when co-administered with OATP1B1, OATP1B3, MATE1, MATE2K, P-gp and 
BCRP substrates. This finding is particularly relevant for drugs administered orally where 
exposure is normally limited by BCRP-mediated efflux, in particular some statins. Modelling has 
predicted a substantial increase in the exposure of atorvastatin when co- administered with 
AZD1775 and the use of atorvastatin is therefore prohibited. 
 
Contraindications: Treatment with AZD1775 is contraindicated in subjects with hypersensitivity 
to any component of the drug. Developmental and reproductive toxicity studies of AZD1775 
have not been performed. AZD1775 is not to be given to women who are pregnant or breast 
feeding. 
 
Availability AZD1775 is an investigational agent supplied to investigators by the Division of 
Cancer Treatment and Diagnosis (DCTD), NCI. AZD1775 is provided to the NCI under a 
Collaborative Agreement between the Pharmaceutical Collaborator and DCTD, NCI. 
 
 
 


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8.2   Agent Ordering and Agent Accountability 
 
NCI-supplied agents may be requested by eligible participating Investigators (or their authorized 
designee) at each participating institution.  The CTEP-assigned protocol number must be used 
for ordering all CTEP-supplied investigational agents.  The eligible participating investigators at 
each participating institution must be registered with CTEP, DCTD through an annual 
submission of FDA Form 1572 (Statement of Investigator), NCI Biosketch, Agent Shipment 
Form, and Financial Disclosure Form (FDF).  If there are several participating investigators at 
one institution, CTEP-supplied investigational agents for the study should be ordered under the 
name of one lead participating investigator at that institution. 
 
In general, sites may order initial agent supplies when a subject is being screened for enrollment 
onto the study. 
 
Active CTEP-registered investigators and investigator-designated shipping designees and 
ordering designees can submit agent requests through the PMB Online Agent Order Processing 
(OAOP) application.  Access to OAOP requires the establishment of a CTEP Identity and Access 
Management (IAM) account and the maintenance of an “active” account status and a “current” 
password.  For questions about drug orders, transfers, returns, or accountability, call or email 
PMB any time.  Refer to the PMB’s website for specific policies and guidelines related to agent 
management.   
 
Agent Inventory Records – The investigator, or a responsible party designated by the 
investigator, must maintain a careful record of the receipt, dispensing and final disposition of all 
agents received from the PMB using the appropriate NCI Investigational Agent (Drug) 
Accountability Record (DARF) available on the CTEP forms page.  Store and maintain separate 
NCI Investigational Agent Accountability Records for each agent, strength, formulation and 
ordering investigator on this protocol. 
 
8.3   Investigator Brochure Availability  
The current versions of the IB for the agent will be accessible to site investigators and research 
staff through the PMB Online Agent Order Processing (OAOP) application.  Access to OAOP 
requires the establishment of a CTEP Identity and Access Management (IAM) account and the 
maintenance of an “active” account status, a “current” password, and active person registration 
status.  Questions about IB access may be directed to the PMB IB coordinator via email. 
 
8.4   Useful Links and Contacts  
Useful Links and Contacts:  
CTEP Forms, Templates, Documents:  http://ctep.cancer.gov/forms/ 
NCI CTEP Investigator Registration:   RCRHelpDesk@nih.gov 
PMB policies and guidelines: http://ctep.cancer.gov/branches/pmb/agent_management.htm  
PMB Online Agent Order Processing (OAOP) application: 
https://ctepcore.nci.nih.gov/OAOP/ CTEP Identity and Access Management (IAM) account: 


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https://ctepcore.nci.nih.gov/iam/index.jsp 
CTEP IAM account help:  ctepreghelp@ctep.nci.nih.gov  
PMB email:  PMBAfterHours@mail.nih.gov 
IB Coordinator: IBCoordinator@mail.nih.gov 
PMB phone and hours of service: (240) 276-6575 Monday through Friday between 8:30 am 
and 4:30 pm (ET) 
 
 


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9. BIOMARKER, CORRELATIVES AND SPECIAL STUDIES 
 
9.1   Collection of Specimens 
 
Table 15: Summary of Specimen Requirements 
 
Time Point 
Specimen and Quantity 
Send Specimens to: 
Baseline 
 
• 
2 tissue cores snap-frozen  
PADIS Laboratory, NCI-
Frederick 
 
• 
Archival FFPE tumor block or 10 5-
micron unstained slides 
• 
1-2 cores snap-frozen 
• 
10 mL blood in purple top EDTA tube 
• 
10 mL blood in green top (sodium 
heparin) tube 
Keyomarsi Laboratory 
 
 
o  
 
 
o  
 
C1D8 
 
o 10 mL blood in purple top EDTA tube 
o 10 ml  blood in green top (sodium 
heparin) tube 
Keyomarsi Laboratory 
C1D10 – 4h post dose (+/- 1 h) 
 
o 2 tissue cores, snap frozen 
PADIS Laboratory, NCI 
Frederick 
 
o 1-2 tissue cores, snap forzen 
Keyomarsi Laboratory 
C2D1 
 
o 10 mL blood in purple top EDTA tube 
o 10 ml  blood in green top (sodium 
heparin) tube 
Keyomarsi Laboratory 
C2D8 
 
o 10 mL blood in purple top EDTA tube 
o 10 ml blood in green top (sodium heparin) 
tube 
Keyomarsi Laboratory 
Progression/Relapse 
 
o 2 tissue cores, snap frozen 
o 10 mL blood in purple top EDTA tube 
o 10 ml blood in green top (sodium heparin) 
tube 
Keyomarsi Laboratory 


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Due to the nature of the correlative analysis, fresh biopsies are necessary. A minimum of four 
core biopsy samples (18 gauge cores) should be attained. Please see Appendix D for detailed 
instructions and request sample shipment kits directly from NCI. Biopsy costs are covered for all 
ETCTN sites. 
 
Note that if any site is unable to send flash frozen biopsy tissue, please contact Dr. Siqing Fu 
and Dr. Robert Kinders for a case by case discussion prior to collection of the samples.  
Time Point 
Specimen and Quantity 
Send Specimens to: 
Baseline 
 
• 
2 tissue cores snap-frozen  
PADIS Laboratory, NCI-Frederick 
 
• 
Archival FFPE tumor block or 10 5-micron 
unstained slides 
• 
1-2 cores snap-frozen 
• 
10 mL blood in purple top EDTA tube 
• 
10 mL blood in green top (sodium heparin) 
tube 
Keyomarsi Laboratory 
04h post dose 5 
 
• 
2 tissue cores, snap-frozen 
PADIS Laboratory, NCI-Frederick  
 
• 
1-2 tissue cores, snap-frozen  
Keyomarsi Laboratory 
C1D8 
 
• 
10 mL blood in purple top EDTA tube 
• 
10 ml  blood in green top (sodium heparin) 
tube 
Keyomarsi Laboratory 
C1D10 – 4h post dose (+/- 1 h) 
 
• 
2 tissue cores, snap frozen 
PADIS Laboratory, NCI Frederick 
 
• 
1-2 tissue cores, snap forzen 
Keyomarsi Laboratory 
C2D1 
 
• 
10 mL blood in purple top EDTA tube 
• 
10 ml  blood in green top (sodium heparin) 
tube 
Keyomarsi Laboratory 
C2D8 
 
• 
10 mL blood in purple top EDTA tube 
• 
10 ml blood in green top (sodium heparin) 
tube 
Keyomarsi Laboratory 
Progression/Relapse 
 
• 
2 tissue cores, snap frozen 
• 
10 mL blood in purple top EDTA tube 
• 
10 ml blood in green top (sodium heparin) 
tube 
Keyomarsi Laboratory 
  


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Collaborating labs should collect 10ml of whole blood in lavender/purple top Vacutainer 
tube (EDTA) and 10ml in green top Vacutainer tube (sodium heparin). After sample 
collection, gently invert tubes 5-10 times to mix the blood with the tube additive. All tubes 
should be labelled with the patient’s study ID. Ship whole blood to the address below for 
Dr. Keyomarsi’s laboratory on the day the biospecimen is collected. If the whole blood 
absolutely cannot be shipped the day it is collected, the tube(s) should be refrigerated (4°C) 
and shipped within 24 hours. 
 
Dr. Keyomarsi’s labs will isolate DNA for NGS sequencing and PBMCs and plasma. 
PBMCs and plasma will be isolated using Ficoll density gradient and cryopreserved in 
liquid nitrogen in 10% DMSO and 90% human AB serum at 10 × 106 cells/vial. Plasma will 
be stored at -70°C (0.5-1 mL/vial) until later analysis. 
 
 
2.16 9.2   Shipping Information 
1. Two cores per time point should be sent to the Pharmacodynamics Assay Development & 
Implementation Section (PADIS) laboratory at NCI, where they will be embedded. Monthly 
batch shipment is allowed.  
 
The shipping address is listed in the SOP under sections 8.3 and 8.4 
 
Attention: Dan Danner 
NCI-F/FNLCR 
1073 Beasley Street, Building 1073 
Fort Detrick 
Frederick, MD 21701 
Phone: 301-846-5748 
 
Please notify Rachel Andrews, Dr. Robert Kinders and Dr. Siqing Fu by email before sending a 
shipment. A separate email with FedEx shipping information will be sent to the research nurse 
for shipment. 
 
Note that FedEx packages should not be shipped any later than Thursday mornings for overnight 
to the facility, and should not be shipped to arrive on Federal holidays because the receiving 
docks are closed on weekends and holidays. 
 
 
Robert Kinders: kindersr@mail.nih.gov  
TEL: 301-846-6410 
Sr. Principal Scientist 
Head, Pharmacodynamics Section 
Clinical Pharmacodynamics Program 
Rachel Andrews:  Rachel.andrews@nih.gov 
TEL: 240-344-5697. 
 
 


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75 
 
Applied/Developmental Research Directorate 
Frederick National Laboratory/Leidos 
 
2. Two additional cores, along with pharmacodynamics blood samples should be sent to Dr. 
Khandan Keyomarsi’s laboratory at MD Anderson Cancer Center. Please contact Dr. Siqing 
Fu should you have any questions.  
 
Khandan Keyomarsi 
Zayed Building for Personalize (Z6.3012) 
1515 Holcombe Blvd. 
Unit 0066 
Houston, TX 77030 
kkeyomar@mdanderson.org 
 
 
10.  STUDY CALENDAR 
 
Baseline evaluations are to be conducted within 1 week prior to start of protocol therapy.  Scans 
and x-rays must be done <4 weeks prior to the start of therapy.  In the event that the patient’s 
condition is deteriorating, laboratory evaluations should be repeated within 48 hours prior to 
initiation of the next cycle of therapy (Table 15). 
 
Table 16:  Phase II Study of AZD1775 in CCNE1 Amplified Patients  
Assessment Tool 
Screeninga Baselineb 
Cycle 1 p 
Cycle 2, 3, 4, … p 
End of Studyc 
Week 
Week 
1 
2 
3 
1 
2 
3 
AZD1775 d 
X 
X 
 
X 
X 
 
Medical history 
X 
X 
Physical exam  
X 
X 
X 
X 
ECOG performance status e 
X 
X 
 
 
X 
X 
CBC with differential 
X 
X 
X 
X 
X 
X 
Serum chemistries (electrolytes, 
hepatic and renal function tests) 
X 
X 
X 
X 
X 
X 
Pregnancy test f 
X 
X 
 
 
X 
X 
Triplicate ECG g 
X 
X 
 
 
X 
 
ECHO/MUGAh 
X 
 
 
 
 
 
Urinalysis i 
X 
 
 
X 
Tumor markers, if available j 
X 
X 
Radiological evaluations (CT, MRI 
or PET as appropriate) k 
X 
X 
Pharmacodynamics (blood) l 
 
X 
X 
 
X 
X 
X 
Tumor specimens m 
X 
 
X 
 
 
 
 
Adverse event assessment n 
X 


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Concurrent medications 
X 
DLT assessment o 
X 
a) 
Screening visit to occur within 4 weeks prior to initiation of therapy 
b) 
Baseline visit to occur within 7 days prior to initiation of therapy 
c) 
End of study visit to occur within 30 days of the last dose of study agents, if feasible 
d) 
This agent is administered orally daily from D1 to D5 (Week 1) and D8-D12 (Week 2), there will not be 
agent administration for Week 3 of each cycle. One cycle is 21 days in length 
e) 
To be obtained at the screening, on Day 1 of each cycle of therapy and at EOT 
f) 
To be tested in women of child-bearing potential prior to the enrollment, on Day 1 of each cycle of 
therapy and at EOT  
g) 
Triplicate 12-lead ECGs at screening and during the study. Triplicate ECGs will be performed on Day 1 of 
each treatment cycle. The mean QTcF interval for all 3 ECGs must be <450 msec for male patients and <470 
msec for female patients. Triplicate 12-lead ECGs will be conducted approximately 2-5 minutes apart prior to 
dosing 
h) 
To be obtained at screening and then as clinically indicated 
i) 
To be obtained at screening then periodically as indicated clinically 
j) 
To be tested at screening and with each restaging if abnormal at baseline, or more frequently as 
appropriate 
k) 
To be performed at screening, once approximately at 9 weeks, another at 18 weeks and also at 27 
weeks +/- 7 days or sooner if clinically indicated, and then every 12 weeks or sooner if clinically 
indicated 
l) 
To collect pharmacodynamics blood samples (20 mL each collection: plasma and peripheral blood 
monocytes): mandatory on cycle 1 day 1 pre-dose and optional on cycle 1 day 8, cycle 2 day 1, cycle 2 
day 8, and at the end of study, if feasible. Optional blood (about 10 mL) will be collected with optional 
tumor biopsy on C1D10 or at relapse. 
m) Mandatory tumor biopsy pre-study and archived tumor tissues (ten 5 µm-thick slides), if available. 
Additional mandatory tumor biopsy on C1D10 at 4 hours (± 2 hours) post dose. Optional biopsies will be 
collected at relapse if a patient has achieved a partial remission or complete remission.  
n) 
Adverse events to be assessed at all study visits and/or over the phone as clinically indicated 
o) 
DLT assessment to occur with 21 days of the first dose 
p) 
Assessments should occur within +/- 4 days of each scheduled visit 
 
 
11.  MEASUREMENT OF EFFECT 
 
Although the clinical benefit of this drug has not yet been established, the intent of offering this 
treatment is to provide a possible therapeutic benefit, and thus the patient will be carefully 
monitored for tumor response and symptom relief in addition to safety and tolerability.  Patients 
with measurable disease will be assessed by standard criteria.  For the purposes of this study, 
patients should be re-evaluated every 9 weeks or sooner as clinically indicated.  In addition to a 
baseline scan, confirmatory scans will also be obtained 4 weeks following initial documentation 
of an objective response. 
 
Antitumor Effect – Solid Tumors 
For the purposes of this study, patients should be re-evaluated for response every 9 weeks.  In 
addition to a baseline scan, confirmatory scans should also be obtained 4 (not less than 4) weeks 
following initial documentation of objective response. 
 
Response and progression will be evaluated in this study using the new international criteria 
proposed by the revised Response Evaluation Criteria in Solid Tumors (RECIST) guideline 
(version 1.1) [51].  Changes in the largest diameter (unidimensional measurement) of the tumor 
lesions and the shortest diameter in the case of malignant lymph nodes are used in the RECIST 
criteria. 
 
 


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11.1  Definitions 
 
Evaluable for toxicity.  All patients will be evaluable for toxicity from the time of their first 
treatment with AZD1775. 
 
Evaluable for objective response.  Only those patients who have measurable disease present at 
baseline, have received at least one cycle of therapy, and have had their disease re-evaluated will 
be considered evaluable for response.  These patients will have their response classified 
according to the definitions stated below.  (Note:  Patients who exhibit objective disease 
progression prior to the end of cycle 1 will also be considered evaluable.) 
 
11.2  Disease Parameters 
 
Measurable disease.  Measurable lesions are defined as those that can be accurately measured in 
at least one dimension (longest diameter to be recorded) as ≥20 mm (≥2 cm) by chest x-ray or as 
≥10 mm (≥1 cm) with CT scan, MRI, or calipers by clinical exam.  All tumor measurements 
must be recorded in millimeters (or decimal fractions of centimeters). 
 
Note:  Tumor lesions that are situated in a previously irradiated area might or might not be 
considered measurable.   
 
Malignant lymph nodes.  To be considered pathologically enlarged and measurable, a lymph 
node must be ≥15 mm (≥1.5 cm) in short axis when assessed by CT scan (CT scan slice 
thickness recommended to be no greater than 5 mm [0.5 cm]).  At baseline and in follow-up, 
only the short axis will be measured and followed. 
 
Non-measurable disease.  All other lesions (or sites of disease), including small lesions (longest 
diameter <10 mm [<1 cm] or pathological lymph nodes with ≥10 to <15 mm [≥1 to <1.5 cm] 
short axis), are considered non-measurable disease.  Bone lesions, leptomeningeal disease, 
ascites, pleural/pericardial effusions, lymphangitis cutis/pulmonitis, inflammatory breast disease, 
and abdominal masses (not followed by CT or MRI), are considered as non-measurable. 
 
Note:  Cystic lesions that meet the criteria for radiographically defined simple cysts should not 
be considered as malignant lesions (neither measurable nor non-measurable) since they are, by 
definition, simple cysts. 
 
‘Cystic lesions’ thought to represent cystic metastases can be considered as measurable lesions, 
if they meet the definition of measurability described above. However, if non-cystic lesions are 
present in the same patient, these are preferred for selection as target lesions. 
 
Target lesions.  All measurable lesions up to a maximum of 2 lesions per organ and 5 lesions in 
total, representative of all involved organs, should be identified as target lesions and recorded 
and measured at baseline.  Target lesions should be selected on the basis of their size (lesions 
with the longest diameter), be representative of all involved organs, but in addition should be 
those that lend themselves to reproducible repeated measurements.  It may be the case that, on 
occasion, the largest lesion does not lend itself to reproducible measurement in which 


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circumstance the next largest lesion which can be measured reproducibly should be selected.  A 
sum of the diameters (longest for non-nodal lesions, short axis for nodal lesions) for all target 
lesions will be calculated and reported as the baseline sum diameters.  If lymph nodes are to be 
included in the sum, then only the short axis is added into the sum.  The baseline sum diameters 
will be used as reference to further characterize any objective tumor regression in the measurable 
dimension of the disease. 
 
Non-target lesions.  All other lesions (or sites of disease) including any measurable lesions over 
and above the 5 target lesions should be identified as non-target lesions and should also be 
recorded at baseline.  Measurements of these lesions are not required, but the presence, absence, 
or in rare cases unequivocal progression of each should be noted throughout follow-up.  
 
 
 


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11.3  Methods for Evaluation of Measurable Disease 
 
All measurements should be taken and recorded in metric notation using a ruler or calipers.  All 
baseline evaluations should be performed as closely as possible to the beginning of treatment and 
never more than 4 weeks before the beginning of the treatment. 
 
The same method of assessment and the same technique should be used to characterize each 
identified and reported lesion at baseline and during follow-up. Imaging-based evaluation is 
preferred to evaluation by clinical examination unless the lesion(s) being followed cannot be 
imaged but are assessable by clinical exam. 
 
Clinical lesions  Clinical lesions will only be considered measurable when they are superficial 
(e.g., skin nodules and palpable lymph nodes) and ≥10 mm (≥1 cm) diameter as assessed using 
calipers (e.g., skin nodules).  In the case of skin lesions, documentation by color photography, 
including a ruler to estimate the size of the lesion, is recommended.  
 
Chest x-ray  Lesions on chest x-ray are acceptable as measurable lesions when they are clearly 
defined and surrounded by aerated lung.  However, CT is preferable.  
 
Conventional CT and MRI  This guideline has defined measurability of lesions on CT scan based 
on the assumption that CT slice thickness is 5 mm (0.5 cm) or less.  If CT scans have slice 
thickness greater than 5 mm (0.5 cm), the minimum size for a measurable lesion should be twice 
the slice thickness.  MRI is also acceptable in certain situations (e.g. for body scans).   
 
Use of MRI remains a complex issue.  MRI has excellent contrast, spatial, and temporal 
resolution; however, there are many image acquisition variables involved in MRI, which greatly 
impact image quality, lesion conspicuity, and measurement.  Furthermore, the availability of 
MRI is variable globally.  As with CT, if an MRI is performed, the technical specifications of the 
scanning sequences used should be optimized for the evaluation of the type and site of disease.  
Furthermore, as with CT, the modality used at follow-up should be the same as was used at 
baseline and the lesions should be measured/assessed on the same pulse sequence.  It is beyond 
the scope of the RECIST guidelines to prescribe specific MRI pulse sequence parameters for all 
scanners, body parts, and diseases.  Ideally, the same type of scanner should be used and the 
image acquisition protocol should be followed as closely as possible to prior scans.  Body scans 
should be performed with breath-hold scanning techniques, if possible. 
 
PET-CT  At present, the low dose or attenuation correction CT portion of a combined PET-CT is 
not always of optimal diagnostic CT quality for use with RECIST measurements.  However, if 
the site can document that the CT performed as part of a PET-CT is of identical diagnostic 
quality to a diagnostic CT (with IV and oral contrast), then the CT portion of the PET-CT can be 
used for RECIST measurements and can be used interchangeably with conventional CT in 
accurately measuring cancer lesions over time.  Note, however, that the PET portion of the CT 
introduces additional data which may bias an investigator if it is not routinely or serially 
performed.   
 
Ultrasound  Ultrasound is not useful in assessment of lesion size and should not be used as a 


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method of measurement.  Ultrasound examinations cannot be reproduced in their entirety for 
independent review at a later date and, because they are operator dependent, it cannot be 
guaranteed that the same technique and measurements will be taken from one assessment to the 
next.  If new lesions are identified by ultrasound in the course of the study, confirmation by CT 
or MRI is advised.  If there is concern about radiation exposure at CT, MRI may be used instead 
of CT in selected instances. 
 
Endoscopy, Laparoscopy  The utilization of these techniques for objective tumor evaluation is 
not advised.  However, such techniques may be useful to confirm complete pathological 
response when biopsies are obtained or to determine relapse in trials where recurrence following 
complete response (CR) or surgical resection is an endpoint. 
 
Tumor markers  Tumor markers alone cannot be used to assess response.  If markers are initially 
above the upper normal limit, they must normalize for a patient to be considered in complete 
clinical response.  Specific guidelines for both CA-125 response (in recurrent ovarian cancer) 
and PSA response (in recurrent prostate cancer) have been published [JNCI 96:487-488, 2004; J 
Clin Oncol 17, 3461-3467, 1999; J Clin Oncol 26:1148-1159, 2008].  In addition, the 
Gynecologic Cancer Intergroup has developed CA-125 progression criteria which are to be 
integrated with objective tumor assessment for use in first-line trials in ovarian cancer [JNCI 
92:1534-1535, 2000]. 
 
Cytology, Histology  These techniques can be used to differentiate between partial responses 
(PR) and complete responses (CR) in rare cases (e.g., residual lesions in tumor types, such as 
germ cell tumors, where known residual benign tumors can remain). 
 
The cytological confirmation of the neoplastic origin of any effusion that appears or worsens 
during treatment when the measurable tumor has met criteria for response or stable disease is 
mandatory to differentiate between response or stable disease (an effusion may be a side effect of 
the treatment) and progressive disease. 
 
FDG-PET  While FDG-PET response assessments need additional study, it is sometimes 
reasonable to incorporate the use of FDG-PET scanning to complement CT scanning in 
assessment of progression (particularly possible 'new' disease).  New lesions on the basis of 
FDG-PET imaging can be identified according to the following algorithm:  
− 
Negative FDG-PET at baseline, with a positive FDG-PET at follow-up is a sign of PD based 
on a new lesion. 
− 
No FDG-PET at baseline and a positive FDG-PET at follow-up:  If the positive FDG-PET at 
follow-up corresponds to a new site of disease confirmed by CT, this is PD.  If the positive 
FDG-PET at follow-up is not confirmed as a new site of disease on CT, additional follow-up 
CT  scans are needed to determine if there is truly progression occurring at that site (if so, the 
date of PD will be the date of the initial abnormal FDG-PET scan).  If the positive FDG-PET 
at follow-up corresponds to a pre-existing site of disease on CT that is not progressing on the 
basis of the anatomic images, this is not PD. 
− 
FDG-PET may be used to upgrade a response to a CR in a manner similar to a biopsy in 
cases where a residual radiographic abnormality is thought to represent fibrosis or scarring.  
The use of FDG-PET in this circumstance should be prospectively described in the protocol 


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and supported by disease-specific medical literature for the indication.  However, it must be 
acknowledged that both approaches may lead to false positive CR due to limitations of FDG-
PET and biopsy resolution/sensitivity. 
 
Note:  A ‘positive’ FDG-PET scan lesion means one which is FDG avid with an uptake greater 
than twice that of the surrounding tissue on the attenuation corrected image. 
 
 
 


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11.4  Response Criteria 
 
Evaluation of Target Lesions 
 
Complete Response (CR):  Disappearance of all target lesions.  Any pathological lymph nodes 
(whether target or non-target) must have reduction in short axis to <10 mm (<1 cm). 
 
Partial Response (PR):  At least a 30% decrease in the sum of the diameters of target lesions, 
taking as reference the baseline sum diameters. 
 
Progressive Disease (PD):  At least a 20% increase in the sum of the diameters of target lesions, 
taking as reference the smallest sum on study (this includes the baseline sum if that is the 
smallest on study).  In addition to the relative increase of 20%, the sum must also demonstrate an 
absolute increase of at least 5 mm (0.5 cm).  (Note:  the appearance of one or more new lesions is 
also considered progressions). 
 
Stable Disease (SD):  Neither sufficient shrinkage to qualify for PR nor sufficient increase to 
qualify for PD, taking as reference the smallest sum diameters while on study. 
 
 
Evaluation of Non-Target Lesions 
 
Complete Response (CR):  Disappearance of all non-target lesions and normalization of tumor 
marker level.  All lymph nodes must be non-pathological in size (<10 mm [<1 cm] short axis). 
 
Note:  If tumor markers are initially above the upper normal limit, they must normalize for a 
patient to be considered in complete clinical response. 
 
Non-CR/Non-PD:  Persistence of one or more non-target lesion(s) and/or maintenance of tumor 
marker level above the normal limits. 
 
Progressive Disease (PD):  Appearance of one or more new lesions and/or unequivocal 
progression of existing non-target lesions.  Unequivocal progression should not normally trump 
target lesion status.  It must be representative of overall disease status change, not a single lesion 
increase. 
 
Although a clear progression of “non-target” lesions only is exceptional, the opinion of the 
treating physician should prevail in such circumstances, and the progression status should be 
confirmed at a later time by the review panel (or Principal Investigator). 
 
 
Evaluation of Best Overall Response 
 
The best overall response is the best response recorded from the start of the treatment until 
disease progression/recurrence (taking as reference for progressive disease the smallest 
measurements recorded since the treatment started).  The patient's best response assignment will 


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depend on the achievement of both measurement and confirmation criteria (Tables 15 and 16). 
 
  Table 17:  For Patients with Measurable Disease (i.e., Target Disease) 
Target 
Lesions 
Non-Target 
Lesions 
New 
Lesions 
Overall 
Response 
Best Overall Response when 
Confirmation is Required* 
CR 
CR 
No 
CR 
≥4 wks. Confirmation** 
CR 
Non-CR/Non-
PD 
No 
PR 
≥4 wks. Confirmation** 
CR 
Not evaluated 
No 
PR 
PR 
Non-CR/Non-
PD/not 
evaluated 
No 
PR 
SD 
Non-CR/Non-
PD/not 
evaluated 
No 
SD 
Documented at least once ≥4 
wks. from baseline** 
PD 
Any 
Yes or No 
PD 
no prior SD, PR or CR 
Any 
PD*** 
Yes or No 
PD 
Any 
Any 
Yes 
PD 
 See RECIST 1.1 manuscript for further details on what is evidence of a new lesion. 
**    Only for non-randomized trials with response as primary endpoint. 
***  In exceptional circumstances, unequivocal progression in non-target lesions may be 
accepted as disease progression. 
 
Note:  Patients with a global deterioration of health status requiring discontinuation of 
treatment without objective evidence of disease progression at that time should be reported as 
“symptomatic deterioration.”  Every effort should be made to document the objective 
progression even after discontinuation of treatment. 
 
  Table 17:  For Patients with Non-Measurable Disease (i.e., Non-Target Disease) 
Non-Target Lesions 
New Lesions 
Overall Response 
CR 
No 
CR 
Non-CR/non-PD 
No 
Non-CR/non-PD* 
Not all evaluated 
No 
not evaluated 
Unequivocal PD 
Yes or No 
PD 
Any 
Yes 
PD 
 ‘Non-CR/non-PD’ is preferred over ‘stable disease’ for non-target disease since SD is 
increasingly used as an endpoint for assessment of efficacy in some trials so to assign this 
category when no lesions can be measured is not advised 
 
 
Duration of Response 
 
Duration of overall response:  The duration of overall response is measured from the time 
measurement criteria are met for CR or PR (whichever is first recorded) until the first date that 


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recurrent or progressive disease is objectively documented (taking as reference for progressive 
disease the smallest measurements recorded since the treatment started). 
 
The duration of overall CR is measured from the time measurement criteria are first met for CR 
until the first date that progressive disease is objectively documented.  
 
Duration of stable disease:  Stable disease is measured from the start of the treatment until the 
criteria for progression are met, taking as reference the smallest measurements recorded since the 
treatment started, including the baseline measurements.  
 
 
 
 


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12.   STUDY OVERSIGHT AND DATA REPORTING / REGULATORY 
REQUIREMENTS 
 
Adverse event lists, guidelines, and instructions for AE reporting can be found in Section 7.0 
(Adverse Events: List and Reporting Requirements). 
 
12.1  Study Oversight 
 
This protocol is monitored at several levels, as described in this section.  The Protocol Principal 
Investigator is responsible for monitoring the conduct and progress of the clinical trial, including 
the ongoing review of accrual, patient-specific clinical and laboratory data, and routine and 
serious adverse events; reporting of expedited adverse events; and accumulation of reported 
adverse events from other trials testing the same drug(s).  The Protocol Principal Investigator and 
statistician have access to the data at all times through the CTMS web-based reporting portal.   
 
During the Phase 2 study, the Protocol Principal Investigator will have, at a minimum, quarterly 
conference calls with the Study Investigators and the CTEP Medical Officer(s) to review accrual, 
progress, and pharmacovigilance.  Decisions to proceed to the second stage of a Phase 2 trial will 
require sign-off by the Protocol Principal Investigator and the Protocol Statistician. 
   
All Study Investigators at participating sites who register/enroll patients on a given protocol are 
responsible for timely submission of data via Medidata Rave and timely reporting of adverse 
events for that particular study.  This includes timely review of data collected on the electronic 
CRFs submitted via Medidata Rave. 
 
All studies are also reviewed in accordance with the enrolling institution’s data safety monitoring 
plan.  
 
 
 


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12.2  Data Reporting 
 
Data collection for this study will be done exclusively through Medidata Rave.  Access to the 
trial in Rave is granted through the iMedidata application to all persons with the appropriate 
roles assigned in the Regulatory Support System (RSS).  To access Rave via iMedidata, the site 
user must have an active CTEP IAM account (check at < https://ctepcore.nci.nih.gov/iam  >) and 
the appropriate Rave role (Rave CRA, Read-Only, CRA, Lab Admin, SLA, or Site Investigator) 
on either the LPO or participating organization roster at the enrolling site. To the hold Rave CRA 
role or CRA Lab Admin role, the user must hold a minimum of an AP registration type.  To hold 
the Rave Site Investigator role, the individual must be registered as an NPIVR or IVR.  
Associates can hold read-only roles in Rave. Upon initial site registration approval for the study 
in RSS, all persons with Rave roles assigned on the appropriate roster will be sent a study 
invitation e-mail from iMedidata.  To accept the invitation, site users must log into the Select 
Login (https://login.imedidata.com/selectlogin) using their CTEP-IAM user name and password, 
and click on the “accept” link in the upper right-corner of the iMedidata page.  Please note, site 
users will not be able to access the study in Rave until all required Medidata and study specific 
trainings are completed.  Trainings will be in the form of electronic learnings (eLearnings), and can 
be accessed by clicking on the link in the upper right pane of the iMedidata screen.   
 
Users that have not previously activated their iMedidata/Rave account at the time of initial site 
registration approval for the study in RSS will also receive a separate invitation from iMedidata 
to activate their account. Account activation instructions are located on the CTSU website, Rave 
tab under the Rave resource materials (Medidata Account Activation and Study Invitation 
Acceptance).  Additional information on iMedidata/Rave is available on the CTSU members’ 
website under the Rave tab or by contacting the CTSU Help Desk at 1-888-823-5923 or by e-
mail at ctsucontact@westat.com. 
 
Method 
 
This study will be monitored by the Clinical Trials Monitoring Service (CTMS).  Data will be 
submitted to CTMS at least once every two weeks via Medidata Rave (or other modality if 
approved by CTEP).  Information on CTMS reporting is available at:  
http://www.theradex.com/clinicalTechnologies/?National-Cancer-Institute-NCI-11.  On-site 
audits will be conducted on an 18-36 month basis as part of routine cancer center site visits. 
More frequent audits may be conducted if warranted by accrual or due to concerns regarding 
data quality or timely submission.  For CTMS monitored studies, after users have activated their 
accounts, please contact the Theradex Help Desk at (609) 799-7580 or by email at 
CTMSSupport@theradex.com for additional support with Rave and completion of CRFs. 
 
 
Responsibility for Data Submission 
 
For ETCTN trials, it is the responsibility of the PI(s) at the site to ensure that all investigators at 
the ETCTN Sites understand the procedures for data submission for each ETCTN protocol and 
that protocol specified data are submitted accurately and in a timely manner to the CTMS via the 
electronic data capture system, Medidata Rave.   


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Data are to be submitted via Medidata Rave to CTMS on a real-time basis, but no less than once 
every 2 weeks.  The timeliness of data submissions and timeliness in resolving data queries will 
be tracked by CTMS.  Metrics for timeliness will be followed and assessed on a quarterly basis.  
For the purpose of Institutional Performance Monitoring, data will be considered delinquent if it 
is greater than 4 weeks past due. 
 
Data from Medidata Rave and CTEP-AERS is reviewed by the CTMS on an ongoing basis as 
data is received.  Queries will be issued by CTMS directly within Rave.  The queries will appear 
on the Task Summary Tab within Rave for the CRA at the ETCTN to resolve.  Monthly web-
based reports are posted for review by the Drug Monitors in the IDB, CTEP.  Onsite audits will 
be conducted by the CTMS to ensure compliance with regulatory requirements, GCP, and NCI 
policies and procedures with the overarching goal of ensuring the integrity of data generated 
from NCI-sponsored clinical trials, as described in the ETCTN Program Guidelines, which may 
be found on the CTEP 
(http://ctep.cancer.gov/protocolDevelopment/electronic_applications/adverse_events.htm)  and 
CTSU websites. 
 
An End of Study CRF is to be completed by the PI, and is to include a summary of study 
endpoints not otherwise captured in the database, such as (for phase I trials) the recommended 
phase 2 dose (RP2D), and a description of any dose-limiting toxicities (DLTs).  CTMS will 
utilize a core set of eCRFs that are Cancer Data Standards Registry and Repository (caDSR) 
compliant (http://cbiit.nci.nih.gov/ncip/biomedical-informatics-resources/interoperability-and-
semantics/metadata-and-models).  Customized eCRFs will be included when appropriate to meet 
unique study requirements.  The PI is encouraged to review the eCRFs, working closely with 
CTMS to ensure prospectively that all required items are appropriately captured in the eCRFs 
prior to study activation.  CTMS will prepare the eCRFs with built-in edit checks to the extent 
possible to promote data integrity. 
 
CDUS data submissions for ETCTN trials activated after 03/01/2014, will be carried out by the 
CTMS contractor, Theradex.  CDUS submissions are performed by Theradex on a monthly 
basis.  The trial’s lead institution is responsible for timely submission to CTMS via Rave. 
 
Further information on data submission procedures can be found in the ETCTN Program 
Guidelines 
(http://ctep.cancer.gov/protocolDevelopment/electronic_applications/adverse_events.htm). 
 
 
12.3  CTEP Multicenter Guidelines 
 
N/A 
 
 
12.4  Collaborative Agreements Language 
The agent(s) supplied by CTEP, DCTD, NCI used in this protocol is/are provided to the NCI 
under a Collaborative Agreement (CRADA, CTA, CSA) between the Pharmaceutical 


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Company(ies) (hereinafter referred to as “Collaborator(s)”) and the NCI Division of Cancer 
Treatment and Diagnosis.  Therefore, the following obligations/guidelines, in addition to the 
provisions in the “Intellectual Property Option to Collaborator”  
(http://ctep.cancer.gov/industryCollaborations2/intellectual_property.htm) contained within the 
terms of award, apply to the use of the Agent(s) in this study: 
 
1. Agent(s) may not be used for any purpose outside the scope of this protocol, nor can Agent(s) 
be transferred or licensed to any party not participating in the clinical study.  Collaborator(s) data 
for Agent(s) are confidential and proprietary to Collaborator(s) and shall be maintained as such 
by the investigators.  The protocol documents for studies utilizing Agents contain confidential 
information and should not be shared or distributed without the permission of the NCI.  If a copy 
of this protocol is requested by a patient or patient’s family member participating on the study, 
the individual should sign a confidentiality agreement.  A suitable model agreement can be 
downloaded from: http://ctep.cancer.gov. 
 
2. For a clinical protocol where there is an investigational Agent used in combination with 
(an)other Agent(s), each the subject of different Collaborative Agreements, the access to and use 
of data by each Collaborator shall be as follows (data pertaining to such combination use shall 
hereinafter be referred to as "Multi-Party Data”): 
 
a. NCI will provide all Collaborators with prior written notice regarding the existence and nature 
of any agreements governing their collaboration with NCI, the design of the proposed 
combination protocol, and the existence of any obligations that would tend to restrict NCI's 
participation in the proposed combination protocol. 
 
b. Each Collaborator shall agree to permit use of the Multi-Party Data from the clinical trial by 
any other Collaborator solely to the extent necessary to allow said other Collaborator to develop, 
obtain regulatory approval or commercialize its own Agent. 
 
c. Any Collaborator having the right to use the Multi-Party Data from these trials must agree in 
writing prior to the commencement of the trials that it will use the Multi-Party Data solely for 
development, regulatory approval, and commercialization of its own Agent. 
 
3. Clinical Trial Data and Results and Raw Data developed under a Collaborative Agreement 
will be made available to Collaborator(s), the NCI, and the FDA, as appropriate and unless 
additional disclosure is required by law or court order as described in the IP Option to 
Collaborator (http://ctep.cancer.gov/industryCollaborations2/intellectual_property.htm).  
Additionally, all Clinical Data and Results and Raw Data will be collected, used and disclosed 
consistent with all applicable federal statutes and regulations for the protection of human 
subjects, including, if applicable, the Standards for Privacy of Individually Identifiable Health 
Information set forth in 45 C.F.R. Part 164. 
 
4. When a Collaborator wishes to initiate a data request, the request should first be sent to the 
NCI, who will then notify the appropriate investigators (Group Chair for Cooperative Group 
studies, or PI for other studies) of Collaborator's wish to contact them. 
 


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5. Any data provided to Collaborator(s) for Phase 3 studies must be in accordance with the 
guidelines and policies of the responsible Data Monitoring Committee (DMC), if there is a DMC 
for this clinical trial. 
 
6. Any manuscripts reporting the results of this clinical trial must be provided to CTEP by the 
Group office for Cooperative Group studies or by the principal investigator for non-Cooperative 
Group studies for immediate delivery to Collaborator(s) for advisory review and comment prior 
to submission for publication.  Collaborator(s) will have 30 days from the date of receipt for 
review.  Collaborator shall have the right to request that publication be delayed for up to an 
additional 30 days in order to ensure that Collaborator’s confidential and proprietary data, in 
addition to Collaborator(s)’s intellectual property rights, are protected.  Copies of abstracts must 
be provided to CTEP for forwarding to Collaborator(s) for courtesy review as soon as possible 
and preferably at least three (3) days prior to submission, but in any case, prior to presentation at 
the meeting or publication in the proceedings.  Press releases and other media presentations must 
also be forwarded to CTEP prior to release.  Copies of any manuscript, abstract and/or press 
release/ media presentation should be sent to: 
 
Email:  ncicteppubs@mail.nih.gov 
 
The Regulatory Affairs Branch will then distribute them to Collaborator(s).  No publication, 
manuscript or other form of public disclosure shall contain any of Collaborator’s confidential/ 
proprietary information. 
 
12.5  Genomic Data Sharing Plan 
N/A 
 
 


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12.6  Safety Monitoring  
 
The site investigators will meet by teleconference at a weekly frequency for approximately 30 
minutes. The site investigator(s) and research staff involved with the conduct of the protocol are 
required to attend. During the weekly teleconference, the investigators will discuss:  
•  Safety of protocol participants (AEs and reporting)  
•  Validity and integrity of the data (data completeness on case report forms and complete 
source documentation)  
•  Enrollment rate relative to expectation of target accrual (eligible and ineligible participants)  
•  Retention of participants, adherence to the protocol, and protocol violations  
•  Protocol amendments  
 
 
12.7  Data Safety Monitoring Plan  
 
The MD Anderson Cancer Center Data and Safety Monitoring Committee (DSMC) will provide 
the primary oversight of data and safety monitoring. The MD Anderson DSMC will review and 
monitor compliance, toxicity and deviations from this study. The committee is composed of 
clinical specialists with experience in oncology. Information that raises any questions about 
participant safety will be addressed with the Principal Investigator.  
 
The DSMC will review this protocol at a minimum of once every six months. Information to be 
provided to the committee includes: a study narrative by the PI, a summary DSMC report 
produced by RAVE/Theradex which includes participant accrual, response, trial status history, 
SAEs, Adverse Events, Deviations and survival); audit results, and monitoring reports as 
applicable. Other information (e.g. scans, laboratory values) will be provided upon request.  
 
The MD Anderson Cancer Center Office of Investigational New Drug/Device (the MD Anderson 
IND Office) will audit the trial at least annually or as determined by the MD Anderson DSMC. 
The overall principal investigator, study coordinator and/or data manager may request access to 
all source documents and other study documentation for on-site or remote monitoring, audit or 
inspection. 
 
 
 
 


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13  STATISTICAL CONSIDERATIONS 
 
13.1  Study Design/Endpoints 
 
The primary endpoint is objective response rate (ORR), defined as a CR or PR within 6 months 
of start of treatment, which is defined as consistent with RECIST version 1.1 criteria for solid 
tumors. The ORR rate will be compared against a null benchmark value of 5%. A maximum of 
29 patients will be treated. If at least 4 patients have a response, it will then be concluded that the 
agent is promising and worthy of further testing. Allowing for 10% ineligibility or dropout rate, 
we will accrue 32 patients to obtain 29 evaluable patients.  
 
 
 


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13.2  Sample Size/Accrual Rate 
 
The ORR rate will be compared against a null benchmark value of 5%. If the observed objective 
response rate is ≥ 4/29 (14%), it will then be concluded that the agent is promising and worthy of 
further testing. Allowing for 10% ineligibility or dropout rate, a total of 32 patients will need to 
be accrued in order to obtain 29 evaluable patients (Table 17). If a response has been observed in 
a particular tumor type, then the study may be expanded to include a total of 14 participants with 
that tumor type. 
 
In order to accrue 29 evaluable patients, we will use a 2-stage design with 10 patients in the first 
stage. If at least one of these 10 patients has a response, we will continue to a second stage of 19 
patients. The proposed design has the following operating characteristics: power is 80% to 
conclude an agent is promising if the true ORR is 0.2; type 1 error rate (one-sided) is 6% (this is 
the probability to conclude that a given agent is promising if the true ORR is 0.05). If the true 
ORR is 0.05, there is a 49% chance of stopping after the first stage. 
 
Table 18:  PLANNED ENROLLMENT REPORT 
Racial Categories 
Ethnic Categories 
Total 
Not Hispanic or Latino 
Hispanic or Latino 
 
Female 
Male 
Female 
Male 
 
American Indian/ 
Alaska Native 
1 
1 
0 
1 
3 
Asian 
1 
1 
1 
0 
3 
Native Hawaiian or 
Other Pacific Islander 
1 
0 
1 
1 
3 
Black or African 
American 
1 
1 
1 
1 
4 
White 
4 
3 
3 
3 
13 
More Than One Race 
2 
1 
1 
2 
6 
Total 
 
 
 
 
32 
PHS 398 / PHS 2590 (Rev. 08/12 Approved Through 8/31/2015)  OMB No. 0925-0001/0002 
 
 
13.3  Stratification Factors 
 
No stratification factor is applied in this study. 
 
 
13.4  Analysis of Secondary Endpoints 
All patients will be evaluated for toxicity and efficacy from the time of their first treatment. All 
patients must be assessed for response to treatment to be assigned one of the following 
categories, even if there are major protocol treatment deviations or if they are ineligible. 
1) Complete response                                              


NCI Protocol #: 10136 
Version Date:  2 June 2020 
93 
 
2) Partial response 
3) Stable disease                                                      
4) Progressive disease 
5) Early death from malignant disease                     
6) Early death from toxicity 
7) Early death because of other cause                     
8) Unknown (not assessable, insufficient data) 
 
Categorization of response will be based on RECIST 1.1. All of the patients who meet the 
eligibility criteria (with the possible exception of those who received no study medication) will 
be included in the main analysis of the response rate. Patients in response categories 4-8 as above 
should be considered as failing to respond to therapy (disease progression). A patient will be 
determined as having a response if he/she has CR or PR; a patient will be determined as a non-
response if there is no evidence of response by 6 months during this study. Thus, an incorrect 
treatment schedule or drug administration does not result in exclusion from the analysis of the 
response rate.  
 
The method of Thall, Simon, and Estey [52] will be employed to perform interim safety 
monitoring. We will assume a Beta (.3, .7) prior distribution for the DLT rate, and target a 30% 
DLT rate and we will terminate enrollment into the trial if Pr{DLT  rate > 30%| data} > 0.9. That 
is, if at any time during the study we determine that there is more than a 90% chance that the 
DLT rate is more than 30% we will stop enrollment. We will monitor at 10 and 20 patients with 
a possible maximum of 32 patients. Stopping boundaries corresponding to this probability 
criterion are to terminate the trial if [# of patients with DLT/ # of patients evaluated] ≥5/10, 8/20. 
The operating characteristics of this rule in the trial are shown in Table 18.  
 
Table 19:  Operating Characteristics for Safety Monitoring Rule 
True DLT Rate 
Probability of Early Termination (PET) 
Mean number of patients 
10% 
0% 
32.0 
20% 
5% 
31.0 
30% 
28% 
27.1 
40% 
63% 
20.7 
50% 
89% 
15.1 
Calculations performed using MultcLean Desktop Version 2.1.0. 
 
Descriptive summary statistics will be used to assess demographics, safety, and antitumor 
activity. Categorical data will be summarized as percentages. Continuous data will be 
summarized by means, medians, ranges, coefficients of variation, and standard deviations. 
Differences in categorical variables will be assessed using the Fisher exact test. PFS and OS will 
be estimated using the Kaplan-Meier method. In correlative mechanism studies, we will focus on 
the association between clinical responses and potential biomarkers and on comparison of 
biomarkers before and after treatment. Statistical inferences will be based on 2-sided tests at a 
significance level of P<0.05. In 20 patients with paired blood samples or biopsies for comparing 
biomarker levels before and after treatment, the study will have 80% power for a paired t-test to 


NCI Protocol #: 10136 
Version Date:  2 June 2020 
94 
 
detect a 0.7 standard deviation mean difference, assuming a 2-sided alpha 5% level. To compare 
biomarker levels between responders and non-responders, the study will have 80% power for a 
2-sample t-test to detect a 1.5 standard deviation mean difference, using a 2-sided 5% alpha level 
if the response rate is 25% and N = 20. Non-parametric analogs of the t-tests will be used if the 
data violate the assumptions of the t-test. 
 
 
13.5  Stopping Rules 
 
An interim futility monitoring analysis will be conducted after the enrollment of 10 evaluable 
patients. If no objective response is observed, the trial should be terminated for futility. If one or 
more objective responses are observed, this study will continue the enrollment of additional 19 
evaluable patients. 
 
Patients who meet a stop rule may remain on study only in their own best medical interest. 
Otherwise patients will be removed from the study for one of the following stop rules: 
 
•  Any uncontrolled side effect affecting quality of life. 
•  Intercurrent illness that prevents further administration of treatment. 
•  At least 20% increase in tumor burden compared with nadir. 
•  Patient chooses to withdraw. Patients can stop participating at any time.  
•  The researcher may decide to take a patient off under several circumstances: discontinuing is 
in the participant's medical best interest, funding is stopped, agent supply is insufficient, patient's 
condition worsens, or new information becomes available 
 
 
 
 
 
 
 


NCI Protocol #: 10136 
Version Date:  2 June 2020 
95 
 
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23. Wang Y, Klijn JG, Zhang Y et al. Gene-expression profiles to predict distant metastasis of 
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25. Sehdev AS, Kurman RJ, Kuhn E, Shih Ie M. Serous tubal intraepithelial carcinoma 
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cyclin E and fatty acid synthase. Mod Pathol 2010; 23: 844-855. 
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biologic properties of ovarian cancer cells. Oncogene 2004; 23: 2648-2657. 
27. Loden M, Stighall M, Nielsen NH et al. The cyclin D1 high and cyclin E high subgroups of 
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28. Keyomarsi K, Tucker SL, Buchholz TA et al. Cyclin E and survival in patients with breast 
cancer. N Engl J Med 2002; 347: 1566-1575. 
29. Pils D, Bachmayr-Heyda A, Auer K et al. Cyclin E1 (CCNE1) as independent positive 
prognostic factor in advanced stage serous ovarian cancer patients - a study of the OVCAD 
consortium. Eur J Cancer 2014; 50: 99-110. 
30. Santala S, Talvensaari-Mattila A, Soini Y, Santala M. Cyclin E Expression Correlates with 
Cancer-specific Survival in Endometrial Endometrioid Adenocarcinoma. Anticancer Res 2015; 
35: 3393-3397. 
31. Cassia R, Moreno-Bueno G, Rodriguez-Perales S et al. Cyclin E gene (CCNE) amplification 
and hCDC4 mutations in endometrial carcinoma. J Pathol 2003; 201: 589-595. 
32. Huang LN, Wang DS, Chen YQ et al. Meta-analysis for cyclin E in lung cancer survival. 
Clin Chim Acta 2012; 413: 663-668. 
33. Zhou Z, Bandla S, Ye J et al. Cyclin E involved in early stage carcinogenesis of esophageal 
adenocarcinoma by SNP DNA microarray and immunohistochemical studies. BMC 
Gastroenterol 2014; 14: 78. 
34. Alsina M, Landolfi S, Aura C et al. Cyclin E amplification/overexpression is associated with 
poor prognosis in gastric cancer. Ann Oncol 2015; 26: 438-439. 
35. Geng Y, Lee YM, Welcker M et al. Kinase-independent function of cyclin E. Mol Cell 
2007; 25: 127-139. 
36. Bagheri-Yarmand R, Biernacka A, Hunt KK, Keyomarsi K. Low molecular weight cyclin E 
overexpression shortens mitosis, leading to chromosome missegregation and centrosome 
amplification. Cancer Res 2010; 70: 5074-5084. 
37. Bagheri-Yarmand R, Nanos-Webb A, Biernacka A et al. Cyclin E deregulation impairs 
mitotic progression through premature activation of Cdc25C. Cancer Res 2010; 70: 5085-5095. 
38. Beck H, Nahse-Kumpf V, Larsen MS et al. Cyclin-dependent kinase suppression by WEE1 


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kinase protects the genome through control of replication initiation and nucleotide consumption. 
Mol Cell Biol 2012; 32: 4226-4236. 
39. Aarts M, Sharpe R, Garcia-Murillas I et al. Forced mitotic entry of S-phase cells as a 
therapeutic strategy induced by inhibition of WEE1. Cancer Discov 2012; 2: 524-539. 
40. Mahajan K, Mahajan NP. WEE1 tyrosine kinase, a novel epigenetic modifier. Trends Genet 
2013; 29: 394-402. 
41. Heijink AM, Blomen VA, Bisteau X et al. A haploid genetic screen identifies the G1/S 
regulatory machinery as a determinant of Wee1 inhibitor sensitivity. Proc Natl Acad Sci U S A 
2015; 112: 15160-15165. 
42. Karakas C, Biernacka A, Bui T et al. Cytoplasmic Cyclin E and Phospho-Cyclin-Dependent 
Kinase 2 Are Biomarkers of Aggressive Breast Cancer. Am J Pathol 2016. 
43. Bauer TM, Jones SF, Greenlees C et al. A phase 1b, open-label, multi-center study to assess 
the safety, tolerability, pharmacokinetics and anti-tumor activity of AZD1775 monotherapy in 
patients with advanced solid tumors: expansion cohorts. J Clin Oncol 2016; 34. 
44. Wingate H, Puskas A, Duong M et al. Low molecular weight cyclin E is specific in breast 
cancer and is associated with mechanisms of tumor progression. Cell Cycle 2009; 8: 1062-1068. 
45. Duong MT, Akli S, Wei C et al. LMW-E/CDK2 deregulates acinar morphogenesis, induces 
tumorigenesis, and associates with the activated b-Raf-ERK1/2-mTOR pathway in breast cancer 
patients. PLoS Genet 2012; 8: e1002538. 
46. Schellens JHM, Marabelle A, Zeigenfuss S et al. Pembrolizumab for previously treated 
advanced cervical squamous cell cancer: Preliminary results from the phase 2 KEYNOTE-158 
study. J Clin Oncol 35, 2017 (suppl; abstr 5514) 2017. 
47. Infection includes all 75 sites of infection under the INFECTIONS AND INFESTATIONS 
SOC. In. 
48. Acute kidney injury includes renal impairment and acute renal insufficiency. In. 
49. Rash may include rash, erythema, eczema, and rash maculo-papular. In. 
50. Peripheral neuropathy includes both peripheral motor neuropathy and peripheral sensory 
neuropathy. In. 
51. Eisenhauer EA, Therasse P, Bogaerts J et al. New response evaluation criteria in solid 
tumours: revised RECIST guideline (version 1.1). Eur J Cancer 2009; 45: 228-247. 
52. Thall PF, Simon RM, Estey EH. Bayesian sequential monitoring designs for single-arm 
clinical trials with multiple outcomes. Stat Med 1995; 14: 357-379. 
53.   Do, K, Wilsker D, Ji J et al. Phase I study of single-agent AZD1775 (MK-1775), a Wee1 
kinase inhibitor, in patients with refractory solid tumors. J Clin Oncol 2015; 33:3409-15.  
 
 


DCTD Standard Operating Procedures (SOP) 
 
98 
 
 
 
APPENDIX A: PERFORMANCE STATUS CRITERIA 
 
ECOG Performance Status Scale 
Karnofsky Performance Scale 
Grade 
Descriptions 
Percent 
Description 
0 
Normal activity.  Fully active, able to 
carry on all pre-disease performance 
without restriction. 
100 
Normal, no complaints, no evidence 
of disease. 
90 
Able to carry on normal activity; 
minor signs or symptoms of disease. 
1 
Symptoms, but ambulatory.  
Restricted in physically strenuous 
activity, but ambulatory and able to 
carry out work of a light or sedentary 
nature (e.g., light housework, office 
work). 
80 
Normal activity with effort; some 
signs or symptoms of disease. 
70 
Cares for self, unable to carry on 
normal activity or to do active work. 
2 
In bed <50% of the time.  Ambulatory 
and capable of all self-care, but 
unable to carry out any work 
activities.  Up and about more than 
50% of waking hours. 
60 
Requires occasional assistance, but 
is able to care for most of his/her 
needs. 
50 
Requires considerable assistance and 
frequent medical care. 
3 
In bed >50% of the time.  Capable of 
only limited self-care, confined to bed 
or chair more than 50% of waking 
hours. 
40 
Disabled, requires special care and 
assistance. 
30 
Severely disabled, hospitalization 
indicated.  Death not imminent. 
4 
100% bedridden.  Completely 
disabled.  Cannot carry on any self-
care.  Totally confined to bed or chair. 
20 
Very sick, hospitalization indicated. 
Death not imminent. 
10 
Moribund, fatal processes 
progressing rapidly. 
5 
Dead. 
0 
Dead. 
 
 
 


DCTD Standard Operating Procedures (SOP) 
 
99 
 
 
 
APPENDIX B: PATIENT DRUG INFORMATION HANDOUT AND WALLET CARD 
Information for Patients, Their Caregivers, and Non-Study Healthcare Team on Possible 
Interactions with Other Drugs and Herbal Supplements  
 
The patient ____________________________ is enrolled on a clinical trial using the experimental study 
drug, AZD1775.  This clinical trial is sponsored by the National Cancer Institute.  This form is addressed 
to the patient, but includes important information for others who care for this patient.  
 
These are the things that you as a healthcare provider need to know: 
AZD1775 interacts with certain specific enzyme(s) in your liver* and certain transport proteins that help 
move drugs in and out of cells**. 
 
*The enzyme(s) in question is/are CYP3A4, CYP3A, CYP2C19, CYP1A2 and AZD1775 is broken down 
by CYP3A4, CYP3A and is affected by strong and moderate inhibitors and inducers of CYP3A4. AZD1775 
is also a possible weak inhibitor of CYP2C19 and inhibits CYP3A4 and may increase levels of other drugs 
that are cleared by these enzymes. AZD1775 induces CYP1A2 enzyme that may result in decreased level of 
other drugs that are cleared by these enzymes. 
 
 **The protein(s) in question is/are P-glycoprotein (P-gp), multi-drug and toxin extrusion proteins 
(MATE1 and MATE2K) and breast cancer resistance protein (BCRP). AZD1775 requires P-gp to move 
out of cells and concomitant administration of strong P-gp inhibitors and inducers should be avoided. 
AZD1775 inhibits BCRP, MATE1 and MATE2K transporters and has the possibility of inducing P-gp and 
that may affect the transport of other drugs that depend on these proteins to move out of cells. Use caution 
when taking substrates of these transporters, such as statins. 
 
To the patient: Take this paper with you to your medical appointments and keep the attached 
information card in your wallet.   
AZD1775 may interact with other drugs which can cause side effects. For this reason, it is very important 
to tell your study doctors of any medicines you are taking before you enroll onto this clinical trial.  It is 
also very important to tell your doctors if you stop taking any regular medicines, or if you start taking a 
new medicine while you take part in this study.  When you talk about your current medications with your 
doctors, include medicine you buy without a prescription (over-the-counter remedy), or any herbal 
supplements such as St. John’s Wort.  It is helpful to bring your medication bottles or an updated 
medication list with you. 
 
Many health care providers can write prescriptions.  You must tell all of your health care providers 
(doctors, physician assistants, nurse practitioners, pharmacists) you are taking part in a clinical trial.  
 
These are the things that you and they need to know:  
 
AZD1775 must be used very carefully with other medicines that use certain liver enzymes or transport 
proteins to be effective or to be cleared from your system or that may affect your heart’s electrical activity.  
Before you enroll onto the clinical trial, your study doctor will work with your regular health care 
providers to review any medicines and herbal supplements  that are considered strong inducers/inhibitors 
or substrates of CYP3A4, CYP3A, CYP2C19, CYP1A2, P-gp, MATE1, MATE2K and BCRP 
 
• Please be very careful!  Over-the-counter drugs (including herbal supplements) may contain ingredients 


DCTD Standard Operating Procedures (SOP) 
 
100 
 
 
 
that could interact with your study drug.  Speak to your doctors or pharmacist to determine if there could 
be any side effects.  
• Avoid ingesting grapefruit, grapefruit juice and Seville oranges while taking AZD1775. 
• You may need to be monitored more frequently if you are taking any drugs that have narrow therapeutic 
ranges.  
•  Your regular health care provider should check a frequently updated medical reference or call your study 
doctor before prescribing any new medicine or discontinuing any medicine.  Your study doctor’s name is 
_____________________________________ and he or she can be contacted at 
_____________________________________. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
STUDY DRUG INFORMATION WALLET CARD   
You are enrolled on a clinical trial using the experimental study  
drug AZD1775.  This clinical trial is sponsored by the NCI.  
__________________ may interact with drugs that are processed by 
your liver, or use certain transport proteins in your body.  Because of 
this, it is very important to: 
➢ Tell your doctors if you stop taking any medicines or if you start 
taking any new medicines. 
➢ Tell all of your health care providers (doctors, physician assistants, 
nurse practitioners, or pharmacists) that you are taking part in a 
clinical trial. 
➢ Check with your doctor or pharmacist whenever you need to use 
an over-the-counter medicine or herbal supplement. 
 
AZD1775 interacts with specific liver enzymes called CYP3A4, CYP3A, 
CYP2C19, CYP1A2, transport protein, P-gp, MATE1, MATE2K and 
BCRP and must be used very carefully with other medicines that 
interact with these enzymes and  transporter proteins.  
➢ 
Before you enroll onto the clinical trial, your study doctor will 
work with your regular health care providers to review any 
medicines and herbal supplements that are prohibited. 
➢ Before prescribing new medicines, your regular health care 
providers should go to a frequently-updated medical reference for 
a list of drugs to avoid, or contact your study doctor.   
➢ Your study doctor’s name is _____________________________ 
and can be contacted at _________________________________.  


DCTD Standard Operating Procedures (SOP) 
 
101 
 
 
 
APPENDIX C: MEDICATION DIARY 
Study Name: A Phase II Study of AZD1775, a WEE1 Inhibitor, in Patients with CCNE1 Amplification. 
 
 
CTEP Study ID: 10136 
 
Patient Initials: ______________________________ 
 
Patient Identification Number: ______________________________ 
 
 
Instructions to the patient:  
 
PLEASE complete and return this dosing diary along with any unused AZD1775 and the 
container(s). You will receive a new dosing diary for each treatment cycle. Thank you for your 
participation in the very important study! 
 
You will take: AZD1775  
 
Each time you take the AZD1775 study medication, your dose is _________ mg and is made up of:            
 _____ 100mg capsules              
                              
              _____ 25mg capsules              
  
How to store your AZD1775 study medication 
The AZD1775 study medication must be stored at room temperature (below 30ºC/86ºF) at all times.  
Please read the medication label for additional storage instructions. 
 
How to take your AZD1775 study medication 
• The AZD1775 study medication dose indicated above is to be taken once daily for five (5) 
consecutive days at the start of Week 1 (Days 1, 2, 3, 4 and 5) and Week 2 (Days 8, 9, 10, 11 and 
12) of each cycle.   
• Each dose is taken orally (by mouth) 2 hours before or 2 hours after food and at approximately the 
same time each morning or evening.  
• You will take 10 doses in each cycle of 21 days. 
 
If you miss the daily dose according to the schedule,  
• the dose should be taken as soon as possible, but not more than 12 hours after the missed dose was 
scheduled.  
• If greater than 12 hours, the missed dose should be skipped and you should take the next dose when 
scheduled. 
 
How to fill-in your AZD1775 dosing diary 
It is very important that you complete this dosing diary as accurately as possible and bring it completed 
along with all remaining study medication (used and unused containers) to your next clinic visit. 
 
Please fill-in this dosing diary each day that you are scheduled to take study medication.   


DCTD Standard Operating Procedures (SOP) 
 
102 
 
 
 
Write the date in the box marked Date. Please record the exact time you took your study medication and 
how many capsules taken. ( number of 100mg and/or number of 25mg) 
 
It is important that you do not miss any of your study medication, but if you do please provide a reason in 
the Comments box along with any other comments you would like the doctor to know. 
 
Cycle number: _________ 
 
Start date: _________ 
 
Patient Signature: _____________________________________ 
 
 


DCTD Standard Operating Procedures (SOP) 
 
103 
 
 
 
D
A
Y 
Date 
Time 
capsules 
taken 
Number of 
capsules 
taken 
Use the space below to make notes 
about things you would like to tell the 
doctor (including unusual symptoms 
you experience, other medicine you 
have taken and anything else you think 
would be of interest.) 
Month    Day   Year 
1 
 
 
 
 
 
 
2 
 
 
 
 
 
 
3 
 
 
 
 
 
 
4 
 
 
 
 
 
 
5 
 
 
 
 
 
 
6 
 
 
 
 
 
 
7 
 
 
 
 
 
 
8 
 
 
 
 
 
 
9 
 
 
 
 
 
 
10 
 
 
 
 
 
 
11 
 
 
 
 
 
 
12 
 
 
 
 
 
 
13 
 
 
 
 
 
 
14 
 
 
 
 
 
 
15 
 
 
 
 
 
 
16 
 
 
 
 
 
 
17 
 
 
 
 
 
 
18 
 
 
 
 
 
 
19 
 
 
 
 
 
 
20 
 
 
 
 
 
 
21 
 
 
 
 
 
 
 
 
 
 
 
 
Title: 
Appendix D: Tumor Frozen Needle Biopsy Specimen Collection and Handling 
Page 1 of 11 
Doc. #: 
SOP340507 
Revision: 
F 
Effective Date: 
2/11/2015 


DCTD Standard Operating Procedures (SOP) 
 
104 
 
 
 
 
Laboratory of Human Toxicology & Pharmacology Applied/Developmental 
Research Directorate, Leidos Biomedical Research, Inc. 
Frederick National Laboratory for Cancer Research 
 
Technical Reviewer: Yvonne A. Evrard 
 
Date:   
 
 
NCTVL Reviewer: 
Jiuping Ji 
 
Date:   
 
 
IQC Approval: 
Katherine V. Ferry-Galow 
 
Date:   
 
 
LHTP Approval: 
Ralph E. Parchment 
 
Date: 
 
DCTD OD Approval: Joseph E. Tomaszewski 
 
Date: 
 
Change History 
 
Revision Approval Date 
Description 
Originator 
Approval 
F 
2/11/2015 
Updated contact shipping address and 
process for advance notification of 
hi
KFG 
REP 
E 
7/3/2013 
Updated tube-type to 1.5-mL conical bottom 
screw cap tubes to allow for broader use in 
DCTD assays and minimize the need to 
transfer biopsies during sample extraction 
steps. Decreased maximum time from biopsy 
YAE 
REP 
D 
1/8/2013 
Update handling in surgical suite including 
details on halving of biopsy. Record total time 
elapsed from biopsy collection to freezing. 
YAE, MM 
JJ 
C 
12/29/2010 
Update sample snap freeze to dry ice/ethanol 
bath or liquid nitrogen. 
YAE 
JJ 
B 
7/24/2009 
Updated SOP format and prepared for 
publication to the DCTD Biomarkers Web site 
YAE 
JJ 
A 
10/13/2006 
Revision with New Shipping Address 
YZ 
JJ 
-- 
8/25/2006 
New Document 
YZ 
JJ 
 
Please check for revision status of the SOP at 
http://dctd.cancer.gov/ResearchResources/ResearchResources-biomarkers.htm 
and be sure to use the current version. Ralph
 E.  Parchment
  DN:
 c=US,
 o=U.S.
 Government,
 ou=HHS,
 ou=NIH,
 Digitally
 signed
 by
 Ralph
 E.
 Parchment
 -S
 (Affiliate)
 -S
 (Affiliate)
 ou=People,
 0.9.2342.19200300.100.1.1=0012315933,
 cn=Ralph
 E.
 Parchment
 -S
 (Affiliate) Date:
 2015.02.25
 10:33:40
 -05'00'
 
  
 


DCTD Standard Operating Procedures (SOP) 
 
105 
 
 
 
Title: 
Appendix D: Tumor Frozen Needle Biopsy Specimen Collection and Handling 
Page 2 of 11 
Doc. #: 
SOP340507 
Revision: 
F 
Effective Date: 
2/11/2015 
 
 
TABLE OF CONTENTS 
1.0 
PURPOSE ............................................................................................................................3 
2.0 
SCOPE. ................................................................................................................................3 
3.0 
ABBREVIATIONS .............................................................................................................3 
4.0 
INTRODUCTION ...............................................................................................................3 
5.0 
ROLES AND RESPONSIBILITIES ...................................................................................3 
6.0 
MATERIALS AND EQUIPMENT REQUIRED ................................................................4 
7.0 
OPERATING PROCEDURES ............................................................................................5 
8.0 
SHIP TO FNLCR FOR ANALYSIS (OPTIONAL)............................................................7 
APPENDIX 1: BATCH RECORD ..................................................................................................9 
APPENDIX 2: PD SAMPLE SHIPPING MANIFEST .................................................................11 


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2.17 1.0 PURPOSE 
Standardize the method for collecting and handling frozen needle tumor biopsies to enable specimen use 
for measurement of pharmacodynamic (PD) markers following treatment with anticancer agents. 
2.18 2.0 SCOPE 
This procedure applies to all personnel involved in the collection and handling of frozen needle tumor 
biopsies for use in PD marker assays during clinical trials. The goal of this SOP and associated training is 
to ensure consistency in tumor needle biopsy collection and handling between clinical sites. 
2.19 3.0 ABBREVIATIONS 
DCTD 
= 
Division of Cancer Treatment and Diagnosis 
ELISA 
= 
Enzyme-Linked ImmunoSorbent Assay 
FNLCR 
= 
Frederick National Laboratory for Cancer Research 
ID 
= 
Identification / Identifier 
IQC 
= 
Internal Quality Control 
LHTP 
= 
Laboratory of Human Toxicology and Pharmacology 
NCTVL 
= 
National Clinical Target Validation Laboratory 
PADIS 
= 
Pharmacodynamics Assay Development & Implementation Section 
PD 
= 
Pharmacodynamic 
SOP 
= 
Standard Operating Procedure 
2.20 4.0 INTRODUCTION 
Specimen handling, shipping, and storage procedures (pre-analytical variables) can have a significant 
impact on the reliability of biomarker measurements in the laboratory. Following detailed steps for 
sample collection and handling procedures and recording any deviations from this procedure allows 
retrospective identification of artifactual changes in biomarker readout and increases the reliability of the 
data and validity of the analytical results. 
5.0 ROLES AND RESPONSIBILITIES 
Laboratory Director/Supervisor 
The Laboratory Director/Supervisor, directs laboratory operations, 
supervises technical personnel and reporting of findings, and is 
responsible for the proper performance of all laboratory procedures. 
Oversees the personnel who follow the SOPs in the laboratory and is 
responsible for ensuring the personnel are certified and have sufficient 
experience to handle clinical samples. 
Certified Assay Operator and/or PK/PD Support Lab Personnel 
A Certified Assay Operator and/or PK/PD Support Lab personnel may 
be a Laboratory Technician/ Technologist, Research Associate, or 
Laboratory Scientist who has been certified through DCTD training on 
this SOP. Work under the guidance of the Laboratory 
Director/Supervisor. This person performs laboratory procedures and 
examinations in accordance with the current SOP(s), as well as any 
other procedures conducted by a laboratory, including maintaining 
equipment and records and performing quality assurance activities 
related to performance. 


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5.1 
It is the responsibility of the Laboratory Director/Supervisor to ensure that all personnel have 
documented training and qualification on this SOP prior to the actual handling and processing of 
samples from clinical trial patients. The Laboratory Director/Supervisor is responsible for 
ensuring the Certified Assay Operator running the SOP has sufficient experience to handle and 
analyze clinical samples. 
5.2 
It is the responsibility of the Certified Assay Operator and/or PK/PD Support Lab personnel to 
confirm scheduled specimen collection time points, pre-print all labels and data collection sheets 
in advance, check documentation for accuracy, and verify that the required collection tubes, 
supplies, and equipment are available for successful collection and handling of biopsy samples. 
5.3 
The Certified Assay Operator and/or PK/PD Support Lab personnel responsible for conducting 
the specimen collection and handling procedures are to follow this SOP and complete the required 
tasks  and  associated  documentation.   The  Batch  Record  (Appendix  1)  must  be  completed            
in real-time for each experimental run, with each page dated and initialed, and placed with the 
clinical sample information. 
5.4 
The responsible personnel are to check the DCTD Biomarkers Web site 
(http://dctd.cancer.gov/ResearchResources/ResearchResources-biomarkers.htm) to verify that the 
latest SOP version is being followed. 
6.0 MATERIALS AND EQUIPMENT REQUIRED 
6.1 
Stop watch, total time in minutes and seconds required 
6.2 
1.5-mL Sarstedt o-ring screw cap, conical bottomed tubes (Sarstedt, Cat#: 72.703.416) 
6.3 
Disposable, fine-tipped tweezers (e.g., VWR, Cat#: 83009-010). Tweezer tips need to easily fit 
to bottom of a 1.5-mL Sarstedt tube 
6.4 
Printable microcentrifuge tube labels or BSI labeling system 
6.5 
81-place freezer boxes (e.g., Fisher Scientific, Cat#: 12-565-182) 
6.6 
Thermoflask cooler or polystyrene foam container 
6.7 
Ice bucket 
6.8 
Liquid nitrogen or dry ice/ethanol bath 
6.9 
Wet ice 
6.10 
-80°C freezer (or lower) 


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7.0 OPERATING PROCEDURES 
7.1 
Record the name and certification number of the Certified Assay Operator and/or PK/PD Support 
Lab personnel performing the SOP, the facility/clinic collecting the specimens, the 
Patient/Sample ID, and the clinical protocol number in the Batch Record (Appendix 1). 
o 
The Batch Record for this SOP is sufficient for collection of a single patient’s biopsy 
samples; if collecting biopsy samples for more than one patient, prepare a separate Batch 
Record for each patient. 
7.2 
Prepare enough pre-printed specimen labels for each whole or halved biopsy sample to be 
collected and frozen as defined in the Pharmacodynamic/Correlative Study section of the Clinical 
Protocol; be sure to coordinate with the clinical center if they prepare the labels for sample 
collection. 
If two passes are collected from one tumor, the labels would be identical except that the specimen 
ID would be followed by a lower case a/b to designate pass number.  The specimen ID includes 
the CTEP protocol number followed by a unique patient identifier and a specimen series ID. 
NCI tumor biopsy specimen IDs for PD sampling are series 500 with consecutive numbers 
identifying the collection time point as defined in the Clinical Protocol.  Sample pre-printed label: 
 
 
7.3 
Of the pre-printed labels prepared for each sample, one label will go on each 1.5-mL Sarstedt 
tube, one on the Batch Record (Appendix 1), and the last will be given to the research nurse to 
place into the patient record sheet. 
7.4 
Tumor Needle Biopsy Collection and Handling 
7.4.1 
The research nurse is to notify the laboratory of scheduled PD sample collections, 
preferably giving at least 24-h notice. Arrive at the biopsy collection site early enough to 
allow sufficient time to set up laboratory supplies, collect relevant clinical information, 
and ensure rapid transport of specimens to the laboratory for placement at -80ºC (or 
lower) after collection. 
7.4.2 
Bring all necessary lab supplies including: disposable tweezers, a minimum of two 
1.5-mL Sarstedt tubes (one for each whole biopsy core) pre-cooled on liquid nitrogen or 
dry ice/ethanol in an insulated bucket, and one pre-printed specimen label to give to the 
research nurse for the patient record. 
Note: Pre-chill additional 1.5-mL Sarstedt tubes for specimen collection in case the 
interventional radiologist collects additional passes, or one of the other tubes is 
compromised prior to collection. 


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7.4.3 
The total time elapsed between biopsy collection and placement into the pre-chilled tube 
is of key importance to biomarker analysis; biopsies should be frozen within 2min of 
collection. The interventional radiologist will eject the biopsy onto a sterile slide (for 
optimal analyte recovery the slide should be pre-chilled). Start a stop watch (or note the 
time) at this point (Appendix 1, Section 1) and immediately walk the slide to the sample 
preparation table. 
Note: The preferred method of collection, when whole biopsies are collected, is for the 
interventional radiologist to eject the biopsy directly into the pre-chilled tube (next step). 
This minimizes the time between collection and fixation of analytes. 
7.4.4 
Indicate if a full or halved biopsy, as defined in the Pharmacodynamic/Correlative Study 
section of the Clinical Protocol, is prepared in the Batch Record (Appendix 1, Section 1). 
7.4.4.1 
For whole biopsies: Uncap an empty, prechilled 1.5-mL Sarstedt tube and 
using disposable tweezers, pick up the freshly collected needle biopsy with the 
tweezers at one end, and touch the opposite end of the biopsy to the inner 
surface of the prechilled 1.5-mL Sarstedt tube. This should attach the tissue to 
the tube, allowing it to be dropped into the tube while releasing the tissue from 
the tweezers without sticking. Dispose of the tweezers in the appropriate 
biohazardous waste container(s). 
7.4.4.2 
For halved biopsies: Use 1-2 disposable tweezers and cut/shear the biopsy in 
half cross-wise while it is on the slide (do not pull or stretch the biopsy 
longitudinally). Use the tweezers to transfer the halved biopsies to sterile pre- 
chilled tubes as indicated above. 
7.4.5 
Immediately snap freeze the biopsy by placing the tube in liquid nitrogen or a dry 
ice/ethanol bath. 
7.4.6 
Calculate the total time elapsed from biopsy collection to biopsy freezing and record the 
total number of minutes and seconds elapsed in the Batch Record (Appendix 1, 
Section 1). 
7.5 
If biopsy procedure details can be obtained from the interventional radiologist or research nurse, 
record them in the Batch Record (Appendix 1, Section 2.). Some information may not be 
available until a later time from the clinical staff. 
During first-in-human PD sample collection studies, information such as type of anesthesia and 
time-lag between biopsy needle withdrawal and sample freezing need to be tracked in order to 
determine the optimal sample collection procedure for the clinical community. 
7.6 
Return to the sample processing laboratory and transfer the frozen biopsy specimen(s) to -80°C 
(or lower) for storage until shipment to the PD processing laboratory. Record the date and time 
specimens are placed at -80ºC (or lower; Appendix 1, Section 3). 
7.7 
Review and finalize the Batch Record and document ANY and ALL deviations from this SOP in 
the Batch Record (Appendix 1, Section 5). 
7.8 
The Laboratory Director/Supervisor should review the Batch Record and sign to affirm the data 
contained within are correct (Appendix 1, Section 6). 


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8.0 SHIP TO FNLCR FOR ANALYSIS (OPTIONAL) 
If shipping to a location other than FNLCR, use the following steps as a guide. 
8.1 
FedEx return shipment labels will be provided to each approved site sending frozen shipments to 
FNLCR PD Specimen Central Receiving. 
8.1.1 
To request return shipment labels send an e-mail to NCI_PD_Support@mail.nih.gov and 
state “Protocol Name Shipment Labels Requested” in the subject line. Specify the 
address to which the shipment labels should be provided and the number of shipment 
labels requested. Shipment labels will be provided within 6 business days. 
8.2 
Once a tumor biopsy has been collected from a patient and placed at -80ºC (or lower), FNLCR 
PD Specimen Central Receiving should be notified that the specimens are ready for shipment. 
8.3 
Send an e-mail to FNLCR PD Specimen Central Receiving (NCI_PD_Support@mail.nih.gov) to 
advise that biopsy samples are being prepared for shipment.  State “Protocol Name PD 
Specimens Ready for Shipment” in the subject line. Request a confirmation e-mail that personnel 
will be available on the expected delivery date and time. Personnel are generally available to 
receive frozen shipments Tuesday through Friday, exclusive of government holidays. If needed, 
FNLCR PD Central Receiving can be contacted directly at 240-344-5697. 
8.4 
Use the PD Sample Shipping Manifest template in Appendix 2 to generate a shipping list 
containing pertinent sample information and FNLCR PD Specimen Central Receiving shipping 
address. 
Attention: Dan Danner 
NCI-F/FNLCR 
1073 Beasley Street, Building 1073 Fort Detrick 
Frederick, MD 21701 
Phone: 301-846-5748 
8.5 
Make a copy of the Shipping Manifest and specimen Batch Records so one copy can be sent to 
FNLCR with the biopsy samples and one can be maintained at the collection site for internal 
records. 
8.6 
Day of Shipment 
8.6.1 
Just prior to shipment, place specimen tubes into an 81-place freezer box and then in a 
shipping container with sufficient dry ice to maintain the samples at -20°C for at least 
72 h. All weekly processing specimens are recommended to ship out via FedEx on the 
following Monday afternoon for delivery by 10 AM Tuesday (FedEx First Overnight). 
8.6.2 
Verify the contents of the package match the Shipping Manifest and sign and date the 
bottom of both copies of the Shipping Manifest. Place one copy of the Shipping Manifest 
inside the shipping box along with copies of the completed Batch Records for all 
specimens. 
8.6.3 
Seal the box and print and attach the shipping address onto the outside of the shipping 
container; be sure the container is labeled as containing biohazardous specimens. 
8.6.4 
Record the shipping date, time, tracking number, and shipping information in the Batch 
Record (Appendix 1, Section 4). 
 


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8.6.5 
E-mail FNLCR PD Specimen Central Receiving (NCI_PD_Support@mail.nih.gov) a 
shipment notification. State “Protocol Name PD Specimen Shipment” in the subject line 
and reference the tracking number in the e-mail. 
8.6.6 
Once specimens arrive at the receiving laboratory, they should be immediately placed at - 
80ºC (or lower) pending delivery to the processing laboratory for protein extraction. 
 


DCTD Standard Operating Procedures (SOP) 
 
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BATCH RECORD 
INITIALS:  
 
DATE:   
 
 
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APPENDIX 1: BATCH RECORD 
A separate Batch Record should be started for each patient sample. 
Note: A pre-dose and post-dose sample from the same patient would have 
the same Patient ID, but different Specimen ID numbers. 
 
Note: Record times using military time (24-h designation); for 
example, specify 16:15 to indicate 4:15 PM. 
 
Certified Assay Operator:   
  
Certification Number:   
   
 
☐ Check here if PK/PD Support Lab Personnel 
 
Facility/Clinic Collecting Specimens:   
 
  
Clinical Protocol Number:   
 
        
Patient ID:    
 
 
1. 
Biopsy Collection 
 
1st Pass 
2nd 
3rd 
4th 
s 
Specimen ID 
Biopsy size prepared for PD or 
histological analysis: 
☐ Full 
☐ Halved 
☐ Full 
☐ Halved 
☐ Full 
☐ Halved 
☐ Full 
☐ Halved 
Required: 
Time elapsed from collection to 
placement in tube 
min 
sec 
min 
sec 
min 
sec 
min 
sec 
Time biopsy collected (opt) 
: 
: 
: 
: 
Time biopsy placed in tube 
(opt) 
: 
: 
: 
: 
 
 
 
 
 
 
 
 
 
 
 
 
Place 
PD Specimen 
Label Here 


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INITIALS:  
 
DATE:   
 
BATCH RECORD 
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2. 
Biopsy Procedure Details 
 
Specimen ID 
Time local anesthesia administered 
: 
Dose of local anesthetic 
mg 
Name of local anesthetic used (from Research Nurse) 
Time of skin incision 
: 
Needle Type (e.g., Temno) 
Needle diameter 
gauge 
Needle Length 
cm 
Time guide needle introduced 
: 
Time guide needle placement confirmed 
: 
Time biopsy needle introduced 
: 
 
3. 
Biopsy Storage 
 
Date/time biopsy specimen(s) placed at 
-80°C (or lower) 
 
/ 
/ 
 
 
: 
 
 
ºC   
 
4. 
Shipping to FNLCR (optional) 
 
Date and time samples shipped 
 
 
 
:  
  
Tracking information 
**Attach copy of Shipping Manifest 
5. 
Notes, including any deviations from the SOP: 
 
 
 
 
 
 
 
 
6. 
Laboratory Director/Supervisor Review of Batch Record 
Laboratory Director/Supervisor: 
(PRINT) 
  
(SIGN) 
Date: 
 
/ 
/  
 


DCTD Standard Operating Procedures (SOP) 
 
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APPENDIX 2: PD SAMPLE SHIPPING MANIFEST 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Verification of Contents                                              Signature 
 
Contents Verified Collection Laboratory 
    
 
 
 
Contents Verified FNLCR PD Central Receiving   
 
 
 
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From: 
Phone: 
E-mail: 
PD Sample 
Shipping Manifest 
In Package 
Item No 
Patient/Specimen ID 
Clinical Protocol 
Description 
Time Point Scheduled 
Example 
1234-1023-500 
12-C-0000 
Full biopsy 
Pre-dose D1 
Example 
1234-1023-501 
12-C-0000 
Half biopsy 
Cycle 1, D8 
☐  
1 
☐  
2 
☐  
3 
☐  
4 
☐  
5 
☐  
6 
☐  
7 
☐  
8 
☐  
9 
☐  
10