Trial Outcomes & Findings for Window of Opportunity Study of Pembrolizumab Alone and in Combinations in Bladder Cancer (NCT NCT03978624)

Last Updated: 2025-10-15

Results Overview

Gene expression levels with the CD8 T cell gene signature were measured using RNA sequencing. The expression level of each gene in the signature were log2 transformed and the mean of the signature genes was calculated in each sample. Then the mean expression of signature in each sample was compared to the average expression and variability across other sample to center the expression values (mean = 0) and scale them by standard deviation and generate a Z-score. Change in z-score was then calculated between pre- and post-treatments for each individual patient and then changes in z-score were compared between treatment arms. An increase in Z-score with treatment denotes an increase in expression of the immune gene signature (IGS).

Recruitment status

ACTIVE_NOT_RECRUITING

Study phase

PHASE2

Target enrollment

20 participants

Primary outcome timeframe

Pre-treatment and at time of definitive treatment within 10 weeks after starting neoadjuvant treatment

Results posted on

2025-10-15

Participant Flow

Participants were enrolled in the study between 09/23/2020 and 04/18/2023 at two cancer centers in the United States.

Participant milestones

Participant milestones
Measure	Pembrolizumab Patients will receive an infusion of pembrolizumab (200mg IV) on Day 1 and Day 22.	Pembrolizumab Plus Entinostat Patients will receive an infusion of pembrolizumab (200mg intravenous) on Day 1 and Day 22, as well as entinostat (50mg given orally) on Day 1, Day 8, and Day 15.
Overall Study STARTED	10	10
Overall Study COMPLETED	9	10
Overall Study NOT COMPLETED	1	0

Reasons for withdrawal

Reasons for withdrawal
Measure	Pembrolizumab Patients will receive an infusion of pembrolizumab (200mg IV) on Day 1 and Day 22.	Pembrolizumab Plus Entinostat Patients will receive an infusion of pembrolizumab (200mg intravenous) on Day 1 and Day 22, as well as entinostat (50mg given orally) on Day 1, Day 8, and Day 15.
Overall Study Protocol Violation	1	0

Baseline Characteristics

Window of Opportunity Study of Pembrolizumab Alone and in Combinations in Bladder Cancer

Baseline characteristics by cohort

Baseline characteristics by cohort
Measure	Pembrolizumab n=10 Participants Patients will receive an infusion of pembrolizumab (200mg IV) on Day 1 and Day 22.	Pembrolizumab Plus Entinostat n=10 Participants Patients will receive an infusion of pembrolizumab (200mg intravenous) on Day 1 and Day 22, as well as entinostat (50mg given orally) on Day 1, Day 8, and Day 15.	Total n=20 Participants Total of all reporting groups
Age, Continuous	73.5 years n=5 Participants	74.9 years n=7 Participants	74.2 years n=5 Participants
Sex: Female, Male Female	1 Participants n=5 Participants	2 Participants n=7 Participants	3 Participants n=5 Participants
Sex: Female, Male Male	9 Participants n=5 Participants	8 Participants n=7 Participants	17 Participants n=5 Participants
Ethnicity (NIH/OMB) Hispanic or Latino	0 Participants n=5 Participants	0 Participants n=7 Participants	0 Participants n=5 Participants
Ethnicity (NIH/OMB) Not Hispanic or Latino	10 Participants n=5 Participants	10 Participants n=7 Participants	20 Participants n=5 Participants
Ethnicity (NIH/OMB) Unknown or Not Reported	0 Participants n=5 Participants	0 Participants n=7 Participants	0 Participants n=5 Participants
Race (NIH/OMB) American Indian or Alaska Native	0 Participants n=5 Participants	0 Participants n=7 Participants	0 Participants n=5 Participants
Race (NIH/OMB) Asian	0 Participants n=5 Participants	0 Participants n=7 Participants	0 Participants n=5 Participants
Race (NIH/OMB) Native Hawaiian or Other Pacific Islander	0 Participants n=5 Participants	0 Participants n=7 Participants	0 Participants n=5 Participants
Race (NIH/OMB) Black or African American	1 Participants n=5 Participants	4 Participants n=7 Participants	5 Participants n=5 Participants
Race (NIH/OMB) White	9 Participants n=5 Participants	6 Participants n=7 Participants	15 Participants n=5 Participants
Race (NIH/OMB) More than one race	0 Participants n=5 Participants	0 Participants n=7 Participants	0 Participants n=5 Participants
Race (NIH/OMB) Unknown or Not Reported	0 Participants n=5 Participants	0 Participants n=7 Participants	0 Participants n=5 Participants
Region of Enrollment United States	10 participants n=5 Participants	10 participants n=7 Participants	20 participants n=5 Participants

PRIMARY outcome

Timeframe: Pre-treatment and at time of definitive treatment within 10 weeks after starting neoadjuvant treatment

Population: Participants started the study, and tissue analysis was completed.

Outcome measures

Outcome measures
Measure	Pembrolizumab n=6 Participants Patients will receive an infusion of pembrolizumab (200mg IV) on Day 1 and Day 22.	Pembrolizumab Plus Entinostat n=8 Participants Patients will receive an infusion of pembrolizumab (200mg intravenous) on Day 1 and Day 22, as well as entinostat (50mg given orally) on Day 1, Day 8, and Day 15
Changes in CD8 T Cell Immune Gene Signature in the Pembrolizumab + Entinostat Group and the Pembrolizumab Group	1.11 Z-score Standard Deviation 1.39	1.01 Z-score Standard Deviation 1.12

SECONDARY outcome

Timeframe: Less than 10 weeks

Neoantigens will be predicted based on whole exome sequencing data using mRNAseq-based filtering. The number of predicted neoantigens will be calculated in the pre-treatment biopsy specimen and the post-treatment cystectomy specimen for each patient. The change from baseline in number and character of neoantigens will be described and compared for each treatment group. Similarly, the T cell receptor (TCR) repertoire will be sequenced and clonality in blood and tumor will be compared between pre- and post-treatment samples for each patient, and change in clonality will be compared between treatment groups.

Outcome measures

Outcome data not reported

SECONDARY outcome

Timeframe: 10 weeks

Compare changes in STAT and NF-κB gene signatures and histone acetylation (H3K9Ac, H3K27Ac, and others) levels after combination treatment with pembrolizumab and entinostat as compared to pembrolizumab alone (subset of subjects with frozen pre- and post-treatment tumor tissue available) Gene expression for each gene in representative STAT, NF-κB, and histone acetylation gene signatures (from MSigDB) will be quantified based on mRNA sequencing. Change in Z-score of these gene signatures will be calculated from pre- and post-treatment tissue samples for patients in both cohorts (pembrolizumab alone or pembrolizumab plus entinostat), and then changes in Z-scores will be compared between cohorts.

Outcome measures

Outcome data not reported

SECONDARY outcome

Timeframe: 10 weeks

Population: Subjects started the treatment.

The analysis of toxicity and safety will be based on the frequency and severity of adverse events attributed to the study treatment, occurring from Day 1 of treatment through 30 days after its completion. The worst toxicity grades per subject will be tabulated for adverse events and laboratory measurements using the National Cancer Institute Common Terminology Criteria for Adverse Events (NCI-CTCAE v5.0) and reported in frequency tables.

Outcome measures

Outcome measures
Measure	Pembrolizumab n=10 Participants Patients will receive an infusion of pembrolizumab (200mg IV) on Day 1 and Day 22.	Pembrolizumab Plus Entinostat n=10 Participants Patients will receive an infusion of pembrolizumab (200mg intravenous) on Day 1 and Day 22, as well as entinostat (50mg given orally) on Day 1, Day 8, and Day 15
Frequency and Severity of Attributed Adverse Events as Assessed by CTCAE v5.0 Grade 1 Diarrhea	2 Participants	2 Participants
Frequency and Severity of Attributed Adverse Events as Assessed by CTCAE v5.0 Grade 1 Dry mouth	1 Participants	0 Participants
Frequency and Severity of Attributed Adverse Events as Assessed by CTCAE v5.0 Grade 1 Nausea	1 Participants	0 Participants
Frequency and Severity of Attributed Adverse Events as Assessed by CTCAE v5.0 Grade 1 Fatigue	1 Participants	1 Participants
Frequency and Severity of Attributed Adverse Events as Assessed by CTCAE v5.0 Grade 1 Alanine aminotransferase increased	1 Participants	0 Participants
Frequency and Severity of Attributed Adverse Events as Assessed by CTCAE v5.0 Grade 1 Alkaline phosphatase increased	1 Participants	0 Participants
Frequency and Severity of Attributed Adverse Events as Assessed by CTCAE v5.0 Grade 1 Aspartate aminotransferase increased	2 Participants	0 Participants
Frequency and Severity of Attributed Adverse Events as Assessed by CTCAE v5.0 Grade 1 Arthralgia	1 Participants	0 Participants
Frequency and Severity of Attributed Adverse Events as Assessed by CTCAE v5.0 Grade 1 Myalgia	0 Participants	1 Participants
Frequency and Severity of Attributed Adverse Events as Assessed by CTCAE v5.0 Grade 3 Myalgia	1 Participants	0 Participants
Frequency and Severity of Attributed Adverse Events as Assessed by CTCAE v5.0 Grade 1 Pruritus	1 Participants	0 Participants
Frequency and Severity of Attributed Adverse Events as Assessed by CTCAE v5.0 Grade 1 Rash maculo-papular	1 Participants	0 Participants
Frequency and Severity of Attributed Adverse Events as Assessed by CTCAE v5.0 Grade 1 SFacial erythema/bilateral axilla rash	1 Participants	0 Participants
Frequency and Severity of Attributed Adverse Events as Assessed by CTCAE v5.0 Grade 2 Skin tag	0 Participants	1 Participants
Frequency and Severity of Attributed Adverse Events as Assessed by CTCAE v5.0 Grade 2 Hyperthyroidism	1 Participants	0 Participants

SECONDARY outcome

Timeframe: 10 weeks

Population: Participants have started the study, completed surgery, and tumor response assessment.

The proportion of patients who have a pathologic response to less than stage 2 (\<pT2) will be reported along with a 95% confidence interval. Similarly, the proportion of patients who have a complete response (pT0) will be reported.

Outcome measures

Outcome measures
Measure	Pembrolizumab n=7 Participants Patients will receive an infusion of pembrolizumab (200mg IV) on Day 1 and Day 22.	Pembrolizumab Plus Entinostat n=7 Participants Patients will receive an infusion of pembrolizumab (200mg intravenous) on Day 1 and Day 22, as well as entinostat (50mg given orally) on Day 1, Day 8, and Day 15
Proportion of Patients Who Have no Cancer in Tissue Samples at Surgery (Pathologic Complete Response, or Only Non-invasive Cancer (Pathologic Partial Response) Complete Response	2 Participants	2 Participants
Proportion of Patients Who Have no Cancer in Tissue Samples at Surgery (Pathologic Complete Response, or Only Non-invasive Cancer (Pathologic Partial Response) Partial response / downstaging	3 Participants	3 Participants

SECONDARY outcome

Timeframe: Up to 3 years

EFS is defined as day 1 of neoadjuvant/protocol treatment to date of first documentation of disease progression (or until recurrence after surgery or radiation) or death due to any cause. Recurrence includes the development of second primary muscle-invasive urothelial malignancies.

Outcome measures

Outcome data not reported

SECONDARY outcome

Timeframe: Up to 3 years

OS is defined as day 1 of neoadjuvant/protocol treatment to date of death due to any cause. Patients will be censored at date of last follow-up.

Outcome measures

Outcome data not reported

Adverse Events

A: Pembrolizumab Alone

Serious events: 4 serious events

Other events: 10 other events

Deaths: 0 deaths

B: Pembrolizumab Plus Entinostat

Serious events: 2 serious events

Other events: 10 other events

Deaths: 0 deaths

Serious adverse events

Serious adverse events
Measure	A: Pembrolizumab Alone n=10 participants at risk Subjects will be administered pembrolizumab alone 200 mg IV on day 1 and day 22 Pembrolizumab: 200 mg IV on day 1 and day 22	B: Pembrolizumab Plus Entinostat n=10 participants at risk Subjects will be administered pembrolizumab on day 1 and day 22 and entinostat 5 mg given orally on day 1, day 8 and day 15 Pembrolizumab: 200 mg IV on day 1 and day 22 Entinostat: 5 mg given orally on day 1, day 8 and day 15
Gastrointestinal disorders Abdominal pain	10.0% 1/10 • Up to 142 days of the last pembrolizumab administration. Adverse events were collected from day one of the study drug administration to 30 days after the last pembrolizumab administration according to National Cancer Institute - Common Terminology Criteria for Adverse Events (NCI-CTCAE). All hospital admissions were graded as SAEs per protocol.	0.00% 0/10 • Up to 142 days of the last pembrolizumab administration. Adverse events were collected from day one of the study drug administration to 30 days after the last pembrolizumab administration according to National Cancer Institute - Common Terminology Criteria for Adverse Events (NCI-CTCAE). All hospital admissions were graded as SAEs per protocol.
Gastrointestinal disorders Upper gastrointestinal hemorrhage	10.0% 1/10 • Up to 142 days of the last pembrolizumab administration. Adverse events were collected from day one of the study drug administration to 30 days after the last pembrolizumab administration according to National Cancer Institute - Common Terminology Criteria for Adverse Events (NCI-CTCAE). All hospital admissions were graded as SAEs per protocol.	0.00% 0/10 • Up to 142 days of the last pembrolizumab administration. Adverse events were collected from day one of the study drug administration to 30 days after the last pembrolizumab administration according to National Cancer Institute - Common Terminology Criteria for Adverse Events (NCI-CTCAE). All hospital admissions were graded as SAEs per protocol.
Infections and infestations Kidney infection	10.0% 1/10 • Up to 142 days of the last pembrolizumab administration. Adverse events were collected from day one of the study drug administration to 30 days after the last pembrolizumab administration according to National Cancer Institute - Common Terminology Criteria for Adverse Events (NCI-CTCAE). All hospital admissions were graded as SAEs per protocol.	0.00% 0/10 • Up to 142 days of the last pembrolizumab administration. Adverse events were collected from day one of the study drug administration to 30 days after the last pembrolizumab administration according to National Cancer Institute - Common Terminology Criteria for Adverse Events (NCI-CTCAE). All hospital admissions were graded as SAEs per protocol.
Infections and infestations Urinary tract infection	10.0% 1/10 • Up to 142 days of the last pembrolizumab administration. Adverse events were collected from day one of the study drug administration to 30 days after the last pembrolizumab administration according to National Cancer Institute - Common Terminology Criteria for Adverse Events (NCI-CTCAE). All hospital admissions were graded as SAEs per protocol.	10.0% 1/10 • Up to 142 days of the last pembrolizumab administration. Adverse events were collected from day one of the study drug administration to 30 days after the last pembrolizumab administration according to National Cancer Institute - Common Terminology Criteria for Adverse Events (NCI-CTCAE). All hospital admissions were graded as SAEs per protocol.
Metabolism and nutrition disorders Hyponatremia	0.00% 0/10 • Up to 142 days of the last pembrolizumab administration. Adverse events were collected from day one of the study drug administration to 30 days after the last pembrolizumab administration according to National Cancer Institute - Common Terminology Criteria for Adverse Events (NCI-CTCAE). All hospital admissions were graded as SAEs per protocol.	10.0% 1/10 • Up to 142 days of the last pembrolizumab administration. Adverse events were collected from day one of the study drug administration to 30 days after the last pembrolizumab administration according to National Cancer Institute - Common Terminology Criteria for Adverse Events (NCI-CTCAE). All hospital admissions were graded as SAEs per protocol.
Psychiatric disorders Confusion	10.0% 1/10 • Up to 142 days of the last pembrolizumab administration. Adverse events were collected from day one of the study drug administration to 30 days after the last pembrolizumab administration according to National Cancer Institute - Common Terminology Criteria for Adverse Events (NCI-CTCAE). All hospital admissions were graded as SAEs per protocol.	0.00% 0/10 • Up to 142 days of the last pembrolizumab administration. Adverse events were collected from day one of the study drug administration to 30 days after the last pembrolizumab administration according to National Cancer Institute - Common Terminology Criteria for Adverse Events (NCI-CTCAE). All hospital admissions were graded as SAEs per protocol.
Renal and urinary disorders Acute kidney injury	10.0% 1/10 • Up to 142 days of the last pembrolizumab administration. Adverse events were collected from day one of the study drug administration to 30 days after the last pembrolizumab administration according to National Cancer Institute - Common Terminology Criteria for Adverse Events (NCI-CTCAE). All hospital admissions were graded as SAEs per protocol.	0.00% 0/10 • Up to 142 days of the last pembrolizumab administration. Adverse events were collected from day one of the study drug administration to 30 days after the last pembrolizumab administration according to National Cancer Institute - Common Terminology Criteria for Adverse Events (NCI-CTCAE). All hospital admissions were graded as SAEs per protocol.
Vascular disorders Thromboembolic event	0.00% 0/10 • Up to 142 days of the last pembrolizumab administration. Adverse events were collected from day one of the study drug administration to 30 days after the last pembrolizumab administration according to National Cancer Institute - Common Terminology Criteria for Adverse Events (NCI-CTCAE). All hospital admissions were graded as SAEs per protocol.	10.0% 1/10 • Up to 142 days of the last pembrolizumab administration. Adverse events were collected from day one of the study drug administration to 30 days after the last pembrolizumab administration according to National Cancer Institute - Common Terminology Criteria for Adverse Events (NCI-CTCAE). All hospital admissions were graded as SAEs per protocol.

Other adverse events

Additional Information

Melahat Canter

UNC Lineberger Comprehensive Cancer Center

Phone: 919-962-0000

Email: [email protected]

Results disclosure agreements

Principal investigator is a sponsor employee The Site shall not independently publish, present, or disclose the Study results, except as specified herein. If a joint publication is not submitted by the Institution within 18 months of the Study's conclusion, abandonment, or termination at all sites, the Site may publish. Additionally, the Site agrees to provide Syndax and Merck with a copy of any proposed publication at least 60 days before the intended publication date.
Publication restrictions are in place

Restriction type: OTHER