Article Text
PDF
Abstract
Introduction The PRaG regimen, which consists of hypofractionated radiotherapy combined with a programmed cell death-1/programmed cell death ligand-1 (PD-1/PD-L1) inhibitor and granulocyte-macrophage colony stimulating factor (GM-CSF), has been demonstrated to have a survival benefit in patients with advanced solid tumours who have failed at least two lines of treatment. Nonetheless, lymphopenia poses an impediment to the enduring efficacy of PD-1/PD-L1 inhibitor therapy. Adequate lymphocyte reserves are essential for the efficacy of immunotherapy. Coupling the PRaG regimen with immunomodulatory agents that augment the number and functionality of lymphocytes may yield further survival benefits in this cohort of patients.
Objective The aim of this study is to investigate the effectiveness and safety of a meticulously thymalfasin-controlled PRaG regimen in patients with advanced and chemotherapy-resistant solid tumours.
Methods and analysis The study has a prospective, single-arm, open-label, multicentre design and aims to recruit up to 60 patients with histologically confirmed advanced solid tumours that have relapsed or metastasised. All eligible patients will receive a minimum of two cycles of the PRaG regimen comprising thymalfasin followed by maintenance treatment with a PD-1/PD-L1 inhibitor and thymalfasin for 1 year or until disease progression. Patients will be monitored according to the predetermined protocol for a year or until disease progression after initiation of radiotherapy.
Ethics and dissemination The study protocol was approved by the Ethics Committee of the Second Affiliated Hospital of Soochow University, on 25 November 2022 (JD-LK-2022-151-01) and all other participating hospitals. Findings will be disseminated through national and international conferences. We also plan to publish our findings in high-impact peer-reviewed journal.
Trial registration number NCT05790447.
- IMMUNOLOGY
- RADIOTHERAPY
- Protocols & guidelines
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, appropriate credit is given, any changes made indicated, and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/.
Statistics from Altmetric.com
Strengths and limitations of this study
To our knowledge, this is the first study to try to evaluate the efficacy and safety of thymalfasin in combination with PRaG therapy (PD-1/PD-L1 inhibitor, Radiotherapy and GM-CSF) for patients with metastatic solid tumours.
The strength of this study is the dynamic adjustment of thymalfasin based on the number of lymphocyte subpopulations.
The study setting, criteria, inclusion, interventions and outcomes are based on a pragmatic approach to ensure external validity.
The limitations of the study include its single-arm design, which is a lack of comparator, making it difficult to evaluate internal validity.
Other limitations include a small number of patients and a single-country design.
Introduction
Use of programmed cell death-1 (PD-1)/programmed cell death ligand-1 (PD-L1) inhibitors for tumour immunotherapy has had a significant impact on the treatment of various cancers.1 However, for the most of solid tumours, the response rate to PD-1/PD-L1 inhibitors such as nivolumab or pembrolizumab as monotherapy was around 15–25%. Poor T cell and cytotoxic T cell infiltration in the tumour microenvironment was considered one of the reasons for primary resistance to PD-1/PD-L1 inhibitors.2 3 Furthermore, as a result of factors such as negative PD-L1 expression, microsatellite stability, low tumour mutational burden (TMB), PD-1/PD-L1 monotherapy are less effective in treating metastatic malignant tumours that have failed second-line and subsequent treatments. Therefore, developing novel therapeutic strategies is critical to improving the prognosis of patients with advanced, recurrent or metastatic malignancy.
Hypofractionated radiotherapy (HFRT) is increasing in clinical practice owing to the advent of precision radiation therapy technology. HFRT can activate the immune system by inducing in situ tumour vaccine effects, causing DNA damage that leads to tumour cell death, and altering the tumour microenvironment.4 5 Furthermore, radiation therapy can upregulate the expression of PD-L1 in tumour cells, potentially heightening their sensitivity to PD-1/PD-L1 inhibitor therapy. Clinical research has demonstrated that combining radiotherapy with PD-1/PD-L1 inhibitors can improve response rates and prolong survival time in patients with metastatic malignant tumours.4 6 7 In a Phase I clinical trial, treatment consisting of 10–15 Gy×3 f and PD-1 inhibitor therapy was confirmed to be safe and effective with an objective response rate of 13.6% and an incidence of grade 3 or higher toxicity of less than 10%.8 In the PEMBRO-RT trial, a treatment regimen consisting of 8 Gy×3 f combined with PD-1 inhibitor therapy was used to treat advanced metastatic non-small cell lung cancer.7 The objective response rate was 36%, with progression-free survival (PFS) being more than double that in the control group.
The presentation of tumour antigens by dendritic cells to T-cells to activate adaptive immunity is an essential event in the tumour immune cycle. Granulocyte-macrophage colony stimulating factor (GM-CSF) is an immune-sensitising cytokine that is commonly used in clinical practice. GM-CSF can promote the differentiation of monocytes/M1 macrophages and dendritic cells, enhance their activity, increase antigen presentation and amplify immunity. The preliminary results for PD-1 inhibitor therapy combined with GM-CSF in patients with advanced cholangiocarcinoma showed that PFS reached 35% after 6 months of enrolment and the proportion of adverse reactions grade 3 or higher was 7%, suggesting that a PD-1 inhibitor combined with GM-CSF was safe and had a good short-term effect.9 In addition, GM-CSF can also amplify the immune activation effects caused by radiotherapy. A prospective clinical study has shown that local radiotherapy combined with GM-CSF can induce abscopal effects and improve the prognosis in patients with advanced solid tumours.10
The ‘PRaG’ regimen was initially proposed as radiotherapy combined with PD-1 inhibitor and GM-CSF therapy for patients with advanced solid tumours who have progressed after at least first-line chemotherapy. The PRaG 1.0 study included 54 patients who were followed up for a median of 16.4 months.11 The objective response rate was 16.7% with a disease control rate of 46.3% in intention to treat patients (ITT). Median PFS was 4.0 months, and median overall survival (OS) was 10.5 months. Patients who received the PRaG regimen tolerated it well overall, with only five experiencing grade 3 treatment-related adverse events and a single patient experiencing grade 4 treatment-related adverse events. These results indicate that the PRaG regimen is effective for patients with advanced recurrent solid tumours. Adequate lymphocyte reserves are critical for effective immunotherapy, and depletion of T lymphocytes poses a challenge in terms of the long-term efficacy of PD-1/PD-L1 inhibitor therapy. Furthermore, both radiotherapy and chemotherapy can cause severe or persistent depletion of lymphocytes, thereby adversely affecting the implementation and effectiveness of tumour treatment. Moreover, the findings of the exploratory PRaG 1.0 study revealed a significant relationship between lymphocyte subsets and clinical efficacy, with a low CD4+/CD8+ ratio indicating a poorer therapeutic effect (p=0.026), high CD3+T cells, CD3+CD4+T cells, CD3+CD8+T cells and NK cell levels after one cycle of the PRaG regimen reveals better effects.12 Therefore, we suggest that combining the PRaG regimen with immunomodulatory agents that augment the number and functionality of lymphocytes may expand the patient population that can benefit from this treatment and improve its efficacy.
Thymalfasin (thymosin α−1 [Tα−1]) is a classic immunomodulator with a polypeptide structure comprising 28 amino acids that is conserved across different species. It has been demonstrated to be clinically effective and safe in the treatment of several diseases, including lung cancer, liver cancer, melanoma, infectious diseases and chronic hepatitis B. Rare and mild adverse events of Tα−1 occurred during the clinical practice, even in the elderly, immuno-compromised individuals and paediatric subjects since Tα−1 possesses an exceptional safety profile.13 Thymalfasin has been found to participate in multiple steps in the tumour immune cycle, acting directly on precursor T-cells to promote their proliferation and maturation.14 It can also activate Toll-like receptors 2 and 9 on dendritic cells, thereby activating these cells, increasing the number of cytotoxic T-cells and enhancing the effectiveness of tumour cell killing.14 15 Recent studies have shown that thymalfasin can reverse the immunosuppressive tumour microenvironment by transforming M2-type macrophages into the M1-type in combination with chemotherapy, promoting infiltration of T-cells into tumour tissues and synergizing with chemotherapy to improve antitumor efficacy.16 Previous clinical trials have also demonstrated that thymalfasin can increase the number and function of lymphocytes and show synergistic efficacy when combined with radiotherapy and chemotherapy. In patients with non-small cell lung cancer who received radiotherapy, a loading dose of thymalfasin upregulated the number and function of lymphocytes.17 In another study, patients with recurrent or metastatic oesophageal squamous cell carcinoma after multiple lines of treatment received thymalfasin in combination with stereotactic body radiation therapy, resulting in a significant abscopal effect. After treatment, patients in the control group with abscopal metastasis showed a significant increase in the number of CD8+T lymphocytes and an increase in the ratio of lymphocytes to monocytes, indicating a better clinical prognosis.18 In a recent study, thymalfasin was administered in combination with concurrent chemoradiotherapy in patients with unresectable stage IIIA–IIIC non-small cell lung cancer. Thymalfasin significantly reduced the incidence of grade 2 or higher radiation pneumonitis, and the number of lymphocytes in the group receiving thymalfasin combination therapy was noticeably protected.19 These findings indicate that thymalfasin in combination with chemotherapy, radiotherapy might synergistically enhance the efficacy of immunotherapy and be of benefit to patients.
Therefore, we are further exploring this updated treatment mode in a prospective study of the efficacy and safety of thymalfasin in combination with the PRaG regimen for patients with advanced solid tumours.
Methods and analysis
Objectives
The primary objective of this study is to investigate the effectiveness of a precise thymalfasin-regulated regimen that is based on a patient’s immune status and combines HFRT with sequential PD-1/PD-L1 inhibitor and GM-CSF therapy for advanced refractory solid tumours. The secondary objective is to assess the safety and toxicity of this treatment. The study also aims to explore a panel of T lymphocyte subpopulations, including tumor-associated cytotoxic T-cells, activated cytotoxic T lymphocytes, activated memory T-cells, monocytes, dendritic cells and T-cell antigen receptors.
Study design
This trial has a multicentre, prospective, open-label and single-arm design. The study was started on 1 April 2023 and aims to complete on 31 December 2025. The study protocol is shown in figure 1. Eligible patients will be treated with thymalfasin (Zadaxin, SciClone Pharmaceuticals) at three different doses based on their absolute T lymphocyte count at baseline. The thymalfasin dose is based on the commonly used dose in previous studies and the dose for severe lymphocytopenia in COVID-19.17–20 After 7 days of treatment with thymalfasin for patients with low baseline lymphocytes, patients will receive at least two cycles of the PRaG regimen. The PRaG regimen consists of HFRT delivered on day 1 followed by subcutaneous injection of GM-CSF (Molgramostim, Topleucon, Xiamen Amoytop Biotech) 200 µg daily for 1 week from day 1 to day 7. Administration of PD-1/PD-L1 inhibitor therapy will commence within 1 week following the completion of radiotherapy. The dosage of thymalfasin is adjusted in real-time based on the absolute T lymphocyte count in each PRaG treatment cycle. PRaG and thymalfasin treatments will be repeated every 21 days for at least two cycles until there are no appropriate lesions for irradiation or the tolerance dose of normal tissue is reached. Patients who complete PRaG will proceed with the maintenance of PD-1/PD-L1 inhibitor and thymalfasin therapy until disease progression or adverse events become intolerable.
Planned study protocol. GM-CSF, granulocyte-macrophage colony stimulating factor; PD-1, programmed cell death-1; PD-L1, programmed cell death ligand-1.
Inclusion criteria
The study inclusion criteria will be as follows: (1) age ≥ 18 years; (2) recurrent or metastatic advanced solid malignancy, definite pathological diagnosis or medical history, no definitely recommended standard treatment regimen in the guidelines, patient cannot tolerate or is unwilling to receive the standard treatment regimen, or presence of clear measurable metastatic lesions (>1 cm); (3) no history of congestive heart failure, unstable angina or unstable arrhythmia in the past 6 months; (4) Eastern Cooperative Oncology Group performance status score 0–3 and life expectancy ≥3 months; (5) no history of serious abnormalities in haematopoietic, heart, lung, liver or kidney function or immune deficiency; (6) aspartate transaminase and alanine transaminase levels ≤3.0 times the upper limit of normal (≤5.0 times the upper limit of normal for patients with liver cancer/metastasis liver carcinoma) or a creatinine level ≤3.0 times the upper limit of normal 1 week before enrolment and (7) ability to understand the informed consent form and willingness to sign it.
Exclusion criteria
Patients who meet any of the following criteria will be excluded: (1) pregnancy or lactation; (2) history of other malignant disease in the last 5 years, except for malignancy that could be cured after treatment (such as adequately treated thyroid cancer, cervical carcinoma in situ, basal or squamous cell skin cancer); (3) a history of uncontrolled epilepsy, disease of the central nervous system or mental disorder that could interfere with the ability to sign the informed consent form or affect compliance with drug treatment in the opinion of the investigator; (4) clinically severe (active) cardiac disease, such as symptomatic coronary heart disease, New York Heart Association class II or worse congestive heart failure, severe arrhythmias requiring medical intervention or a history of myocardial infarction within the last 12 months; (5) immunosuppressive therapy for organ transplantation; (6) known significant active infection or significant blood, renal, gastrointestinal, endocrine or metabolic disorder as determined by the investigator or other severe uncontrolled concomitant disease; (7) allergy to any ingredient of the investigational drug; (8) a medical history of immunodeficiency, including HIV positivity or other acquired or congenital immunodeficiency disease, organ transplantation or other immune-related disease requiring long-term oral hormone therapy; (9) acute or chronic tuberculosis (a positive T-spot test result with suspected tuberculous lesions on a chest radiograph) or (10) other condition deemed to render a patient unsuitable for enrolment in the opinion of the investigator.
Study endpoints
The primary endpoint of the study is the overall response rate, which will be evaluated according to Response Evaluation Criteria in Solid Tumors V.1.1. The secondary endpoints are the disease control rate, PFS, OS and the incidence of treatment-related adverse events. The efficacy of the PRaG and thymalfasin regimen will be evaluated every 2 months. Peripheral blood and/or tissue samples will be collected at specified time points and processed as per the study protocol for subsequent translational research.
Sample size calculation
Based on the assumption that the primary endpoint (overall response rate) would be 20%, using an exact two-sided 90% CI, a sample size of 60 cases (allowing for a 10% dropout rate) would provide acceptable precision of ±0.1.
Statistical analysis
The statistical analysis will be performed using SPSS V.18.0 software (IBM Corp., Armonk, NY, USA). The Shapiro-Wilk normality test will be conducted on the residuals, and the test level will be α>0.05. Random block analysis of variance will be conducted for indicators that are normally distributed and the rank sum test of random block design for those that are non-normally distributed, and the test level will be α>0.05. The study will compare changes in the white blood cell, granulocyte and lymphocyte counts as well as differential cells and cytokines before and after radiotherapy, and determine if any of the differences are statistically different. Cox regression analysis will also be performed, combining the changes in these counts with the patient’s survival time to determine their influence on the survival rate. The study will use the Kaplan-Meier method to analyse survival according to the presence or absence of bystander effects and to investigate the relationship between survival rate and cytokines.
Patient and public involvement
Patients and/or the public were not involved in the design, conduct, reporting or dissemination plans of this research.
Ethics and dissemination
The study protocol has been approved by the Ethics Committee of the Second Affiliated Hospital of Soochow University and all other participating hospitals and will be conducted in compliance with the Declaration of Helsinki (see online supplemental file 1). Individual consent will be sought from all trial participants (see online supplemental file 2). The outcomes of the PRaG5.0 study trial, regardless of the findings, will be planned to publish in an international peer-reviewed medical journal. Our reporting of the results will strictly follow the guidelines outlined in the Consolidated Standards of Reporting Trials statement.
Supplemental material
Supplemental material
Ethics statements
Patient consent for publication
Acknowledgments
We thank Liwen Bianji (Edanz) (www.liwenbianji.cn) for editing and drafting the English text of this manuscript.
References
Supplementary materials
Supplementary Data
This web only file has been produced by the BMJ Publishing Group from an electronic file supplied by the author(s) and has not been edited for content.
Footnotes
YK, RC and MX contributed equally.
Contributors LZ conceived the study and serves as the guarantor for the study. YK, RC, MX, and PX designed the study. JZ, GC, ZH, HZ, XD, YP and CZ were responsible for data collection and follow-up. YM and XZ contributed to statistical analysis. All authors contributed to data interpretation and the writing of this manuscript.
Funding This study was supported by Suzhou Medical Center (Szlcyxzx202103), the National Natural Science Foundation of China (82171828), the Key R&D plan of Jiangsu Province (Social Development, BE2021652), Suzhou Radiotherapy Clinical Medical Center (Szlcyxzx202103), Open project of the State Key Laboratory of Radiology and Radiation Protection of Soochow University (GZK1202014,GZK1202238), Open Project of Provincial Key Laboratory of Soochow University (KJS1961), the Subject construction support project of the Second Affiliated Hospital of Soochow University (XKTJ-RC202001, XKTJHRC20210011), the Suzhou Science and Technology Development Plan (SYS2020143,SKY2022163), Chinese Society of Clinical Oncology Research Foundation of Beijing (Y-XD202002/ZB-0015, Y-pierrefabre202102-0113), Wu Jieping Medical Foundation (320.6750.2021-01-12), The special project of “Technological Innovation” project of CNNC Medical Industry Co. Ltd (ZHYLTD2021001), Suzhou Science and Education Health Project (KJXW2021018), Postgraduate Research & Practice Innovation Program of Jiangsu Province (SJCX22_1508，KYCX22_3223),Youth Employee Pre Research Fund Project of the Second Affiliated Hospital of Soochow University (SDFEYQN1701), Key Medical Discipline Construction Unit of Jiangsu Province for the 14th Five-year plan (JSDW202236).
Competing interests None declared.
Patient and public involvement Patients and/or the public were not involved in the design, conduct, reporting or dissemination plans of this research.
Provenance and peer review Not commissioned; externally peer reviewed.
Supplemental material This content has been supplied by the author(s). It has not been vetted by BMJ Publishing Group Limited (BMJ) and may not have been peer-reviewed. Any opinions or recommendations discussed are solely those of the author(s) and are not endorsed by BMJ. BMJ disclaims all liability and responsibility arising from any reliance placed on the content. Where the content includes any translated material, BMJ does not warrant the accuracy and reliability of the translations (including but not limited to local regulations, clinical guidelines, terminology, drug names and drug dosages), and is not responsible for any error and/or omissions arising from translation and adaptation or otherwise.