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Abstract
Background Vulvar squamous cell carcinoma (VSCC) is a rare cancer for which the cornerstone of treatment is surgery with high complication rates. The unmet need is a less radical and more effective treatment for VSCC.
Primary Objectives To investigate the impact of mono-immunotherapy pembrolizumab as neoadjuvant treatment for primary resectable VSCC patients.
Study Hypothesis Some primary VSCC patients display a specific immune profile which is associated with better survival. In other tumors, this profile is associated with a better response to programmed cell death protein 1 (PD-1) checkpoint blockade which may reinvigorate tumor-specific T cells. This potentially results in a reduced tumor load and less radical surgery and/or adjuvant treatment in patients with this immune profile.
Trial Design This is an investigator-initiated, prospective, single arm, multicenter, phase II clinical trial.
Inclusion Criteria Patients with VSCC clinical stage International Federation of Gynecology and Obstetrics (FIGO) I-III (2021) eligible for primary surgery, with at least one measurable lesion of at least one dimension ≥10 mm in the largest diameter, are included in this study.
Main Exclusion Criteria Patients not suitable for surgery and/or previously treated with immunomodulatory agents, and/or who suffer from comorbidities that may interfere with PD-1 blockade, are excluded from the study.
Endpoints The clinical efficacy of neoadjuvant pembrolizumab in VSCC is measured by an objective change in tumor size according to the Response Evaluation Criteria In Solid Tumors version 1.1 (RECIST 1.1) and documented by calipers using standardized digital photography with a reference ruler. In addition, the activation, proliferation, and migration of T cells in the tumor will be studied. The secondary endpoints are pathological complete responses at the time of surgery, feasibility, and safety.
Sample Size 40 patients with FIGO I-III (2021) primary VSCC will be enrolled.
Estimated Dates for Completing Accrual and Presenting Results The intervention phase started in July 2023 and will continue until July 2025. The expected completion of the entire study is July 2026.
Trial Registration Number NCT05761132
- Vulvar and Vaginal Cancer
- Immunotherapy
Data availability statement
There are no data in this work.
This is an open access article distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited, an indication of whether changes were made, and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/.
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INTRODUCTION
Vulvar squamous cell carcinoma (VSCC) is a relatively rare gynecological cancer type that accounts for 10% of all gynecological cancers, with an annual incidence of 2–3 per 100 000 women. The majority of these patients (90%) have a T1 (restricted to the vulva, not involving the urethra, vagina, or anus), N0-1, M0 tumor. Patients with International Federation of Gynecology and Obstetrics (FIGO) stage (2009) I/II VSCC have approximately a 70% 5-year survival, which decreases drastically to 40% in stage III disease when lymph nodes are involved.1 Surgical resection of the primary vulvar tumor and excision of sentinel inguinal lymph nodes or full inguinal lymph node dissection depending on clinicopathological features remains the standard-of-care for VSCC, and approximately 20% of the patients receive adjuvant radiotherapy.2 The prognosis of VSCC has not improved over the past decades and post-operative morbidity remains a lingering burden, consisting of wound complications, infections, positive margins, vital tissue damage to clitoris, urethra, and/or anus, and associated psycho-social issues and sexual dysfunction.3
In the end, up to 40% of VSCC patients will develop recurrent disease and eventually die of their disease and/or accompanied complications.4
To improve outcome, as well as reduce short-term and late morbidity after cancer treatment, there is a need to study potential therapeutic agents that may result in less (surgical or treatment-related) morbidity as well as decreased recurrences.
We recently showed that patients treated by surgery and if needed adjuvant radiotherapy displayed better overall survival and recurrence-free survival when their VSCC was strongly infiltrated by T cells and showed signs of active immune signaling.5–7 In other solid tumors, immunotherapy in the form of checkpoint blockade works best in patients with such an inflamed/hot tumor,8 especially when it is applied in a neoadjuvant setting where it is associated with partial and complete responses in substantial numbers of patients.9–13 Based on these data, we hypothesized that programmed cell death protein 1 (PD-1) blockade may activate and boost T cell immunity when patients have an inflamed/hot VSCC, and consequently show a reduction in tumor size prior to surgery. This may lead to less radical surgery, thereby reducing the deleterious post-surgical morbidity and meanwhile limiting the recurrence rate of VSCC. Therefore, we initiated APOLLO, a prospective, multicenter, phase II, non-controlled clinical trial with pembrolizumab, to evaluate the efficacy and anti-tumor response in VSCC in a neoadjuvant setting.
METHODS
Trial Design
Patients with primary VSCC who present at the outpatient clinic of participating centers will be informed of the study by their treating physician. For those patients who are potentially eligible and interested in the study, further information and counseling will be done by the gynecological oncologist and medical oncologist. Written informed consent will be obtained after a reflection period (week 2). At baseline, the tumor size will be recorded by calipers using digital photography. Pre-treatment biopsies will be obtained for confirmation of VSCC and for translational studies.
The planned dose of pembrolizumab for this study is 200 mg intravenously (IV) 3-weekly, based on safety and feasibility data generated from the Keytruda development program. Patients will receive standard-of-care surgery after two cycles of neoadjuvant pembrolizumab approximately 3–4 weeks after the second dose. The duration of pre-operative treatment of approximately 6 weeks has been chosen as this is a short time period unlikely to result in significant disease progression which might preclude complete surgical resection.
Clinical responses are categorized according to the Response Evaluation Criteria In Solid Tumors version 1.1 (RECIST 1.1)14 and documented by calipers using standardized digital photography with a reference ruler as either a complete response with disappearance of all target lesions, in which any pathological lymph nodes (whether target or non-target) must have a reduction in short axis to <10 mm, or a partial response with at least a 30% decrease in the sum of diameters of target lesions, taking as reference the baseline sum diameters. Progressive disease is defined as at least a 20% increase in the sum of diameters of target vulvar 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. The appearance of one or more new lesions is also considered as progression. Finally, stable disease in which neither sufficient shrinkage to qualify for partial response nor sufficient increase to qualify for progressive disease, taking as reference the smallest sum diameters while on study, is observed. Responders have the option to fall into an extension cohort and, after consultation with the multidisciplinary team, will be treated with adjuvant pembrolizumab from week 16 until week 58 (400 mg IV, 6-weekly, seven times). These patients will be monitored for long-term effects, including blood collections for safety values and digital photography of the vulva. Non-responders and responders who will not have extended pembrolizumab will visit the hospital at week 13 and 16 to monitor potential long-term effects of pembrolizumab, including blood collections for safety values and for oncological follow-up in accordance with standard-of-care guidelines.
Baseline immunological parameters are measured from two fresh pre-treatment 5 mm wide biopsies (one diagnostic and one translational biopsy) that will be taken at week 3 and paraffin embedded using the standard workflow for diagnostic pathology including a p16-, p53-, and programmed death-ligand 1 (PD-L1) immunohistochemistry. At surgical resection, diagnostic and translational material of the tumor will be paraffin embedded. Part of the tumor at surgical resection (if the tumor size allows for this) will be used to make a fresh tumor digest. Peripheral blood mononuclear cells are isolated, washed, and cryopreserved according to standard operating procedures. A graphical representation of the study is provided in (Figure 1).
Overview of study design. After the enrollment of patients in the study, the tumor is measured according to RECIST 1.1 criteria and documented by calipers using standardized digital photography with a reference ruler. Both vulvar biopsies and blood samples are taken. In short, pembrolizumab is given in week 4 and week 7 following the collection of the blood samples. In week 10 surgery is planned with a second measurement of the tumor, together with the obtainment of pre-surgery blood samples and resection material (see trial design). RECIST 1.1, Response Evaluation Criteria In Solid Tumors, version 1.1.
Participants
Patients who are clinically diagnosed with FIGO I-III (2021) will be accrued from the Leiden University Medical Center, the University Medical Center Groningen, and the Erasmus University Medical Center. Subjects will be recruited by the gynecologist-oncologist. Included are women with a histologically confirmed primary diagnosis of VSCC with at least one lesion that can be measured in at least one dimension with ≥10 mm in largest diameter and clinical stage FIGO I-III (2021) who are eligible for primary surgery. Women should not be pregnant or breastfeeding and will be excluded when they have received prior therapy with an anti-PDL-1 or anti-PD-L2 agent, or other systemic anti-cancer therapies within 4 weeks prior to allocation. Also prior radiotherapy or major surgery within 2 weeks of the start of the study treatment, and a live vaccine within 30 days before the first dose of pembrolizumab, are exclusion criteria. In addition, an immunodeficiency or chronic systemic steroid therapy (≥10 mg daily of prednisone equivalent) are excluded (see full list of inclusion/exclusion criteria in (Table 1).
Inclusion and exclusion criteria of the APOLLO trial
Outcomes
Primary Endpoints
The primary endpoint of the study is: (1) the clinical efficacy of neoadjuvant pembrolizumab in VSCC, measured by an objective change in tumor size (according to RECIST 1.1) and documented by calipers using standardized digital photography with reference ruler at the time of surgery; and (2) the activation, proliferation, and migration of tumor-specific T cells on PD-1 blockade.
Secondary Endpoints
In addition to the clinical response, the pathological response in tumor tissue derived from surgery (week 10) will be assessed by a gynecological pathologist, blinded to other study data. The pathological tumor response (pTR) is defined as the presence of tumor cell necrosis and keratinous debris with giant cell/histiocytic reaction, quantified as a percentage of the overall tumor bed (area pathologic response/area pathologic response plus viable tumor): pTR-0 (<10%), pTR-1 (10–49%), pTR-2 (≥50%), pTR-3 (100%, complete response). Moreover, the feasibility (defined as delay in planned surgery and surgical outcome) and safety according to the National Cancer Institute Common Terminology Criteria for Adverse Events (NCI-CTAE) version 5.0 of pembrolizumab in a neoadjuvant setting will be assessed. Moreover, we will study the activation, proliferation, and migration of other cell populations on PD-1 blockade.
Exploratory Endpoints
To study the on-treatment effect of neoadjuvant PD-1 blockade on the tumor microenvironment in fresh resection material and peripheral blood. To discover potential future predictors for minimal residual disease and/or relapse we will study the utility of circulating tumor DNA. The quality of life (European Organisation for Research and Treatment of Cancer Core Quality of Life Questionnaire (EORTC QLQ) C-30 and QLQ-VU34) of patients enrolled in this study will be assessed.
Sample Size
The clinical hypothesis is that the group of VSCC patients with an inflamed tumor (that is strongly T cell infiltrated tumor with ‘hot’ immune transcriptional signature) is more likely to respond to PD-1/PD-L1 blockade in a neoadjuvant setting.5–7 At this stage a conservative estimation is that 25% of primary FIGO I-III (2009) VSCC displays such an inflamed tumor.5–7 Assuming that 50% of these patients with an inflamed tumor (12.5%) should respond, a <1% spontaneous (base rate) response and an α=0.05, then n=29 (power 90%) patients are required to establish clinical outcome. For biomarker studies the power calculations (α=0.05, β=0.2, power 80%) show that with a responder to non-responder ratio of 1:3 (10 vs 30 out of 40), a positive observation in 80% of the responders and in maximally 30% of the non-responders (background) would require n=36 patients (n=9 responders and n=27 non-responders) to adequately identify potential biomarkers. Based on these calculations a total of 40 primary VSCC patients will be included in this clinical proof-of-concept study.
Randomization and Blinding
Not applicable
Statistical Methods
In order for patients to be eligible for evaluation of the primary clinical endpoint they should have received at least one cycle of pembrolizumab. All data collected until week 16 will be used for the analyses. Continuous variables will be summarized using descriptive statistics for all assessments (mean, SD, quartiles, minimum, and maximum). Categorical variables will be summarized using counts and percentages. Non-parametric (Wilcoxon signed-rank or Mann-Whitney test for two samples and Friedman or Kruskal-Wallis with Dunn’s multiple comparison test for multiple samples) and parametric (paired or unpaired t test for two samples or repeated measures one-way analysis of variance (ANOVA) or ordinary one-way ANOVA with Tukey’s multiple comparison test for multiple samples) tests are performed as appropriate. P values <0.05 are marked as significant. Objective response rates (ORRs) will be calculated using standard statistics for ORR (point estimate and confidence interval). The confidence level will be set at 95%.
DISCUSSION
This a clinical proof-of-concept study, and the first trial that studies checkpoint blockade in primary VSCC in a neoadjuvant setting. While only sporadic responses have been reported for adjuvant checkpoint therapy in metastatic VSCC, we expect that neoadjuvant treatment will be much more successful. The rationale behind this hypothesis lies in our observations that patients with an inflamed tumor (around 45% of all VSCC5) exhibit more favorable responses to standard-of-care treatments. As such, this patient population is less likely to have metastatic or recurrent VSCC5 7 and are therefore not incorporated in previous adjuvant checkpoint studies. Importantly, the literature on successful (neoadjuvant) checkpoint therapy in other cancer types indicates that patients with inflamed tumors are more likely to respond.9 10 12 13
The application of only two 3-weekly doses of checkpoint therapy in a neoadjuvant setting will likely not compromise surgical outcome in case of non-responding VSCC and may already lead to clinical responses. Importantly, recent studies, in which 1–2 doses of checkpoint therapy have been given in a neoadjuvant setting for other types of tumors, suggest that this may be enough to obtain complete and partial responses in a substantial portion of patients.9 10 12 13 By treating VSCC patients with checkpoint therapy in a neoadjuvant approach, also the patients with inflamed tumors will receive immunotherapy. If successful, neoadjuvant checkpoint therapy of VSCC is high-gain because then we would have—for the first time in decades—a new treatment option for VSCC and be able to facilitate more sparing and less morbid surgery. In addition, the study may lead to the discovery of biomarkers that could predict clinical responsiveness in this patient group.
Data availability statement
There are no data in this work.
Ethics statements
Patient consent for publication
Ethics approval
This study involves human participants and was approved by the Ethical Committee, Leiden University Medical Center NL82378.058.22. Participants gave informed consent to participate in the study before taking part.
Footnotes
MIEvP and KEK are joint first authors.
X @KimKortekaas
Contributors MvP: conceptualization, data curation, writing. KK: investigation, data curation, writing. HvD: data curation, writing. HN: data curation, writing. IB: data curation, writing. AR: data curation, writing. PEG: data curation, writing. BJ: data curation, writing. TB: data curation, writing. MW: data curation, writing. JK: conceptualization, data curation, writing. SvdB, conceptualization, writing. SvdB accepts full responsibility for the work and/or the conduct of the study, had access to the data, and controlled the decision to publish.
Funding The study is funded by a clinical proof-of-concept grant of the Oncode Institute (number 2021-0001). MSD Netherlands provided drug support for the study.
Competing interests None declared.
Provenance and peer review Commissioned; internally peer reviewed.