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Objective:To prepare 99Tc m-hydrazinonicotinamide(HYNIC)-αCD8/Fab ( 99Tc m-αCD8/Fab), and explore the predictive value of 99Tc m-αCD8/Fab SPECT/CT imaging for the efficacy of anti-programmed death-1 (PD-1) immunotherapy. Methods:The αCD8/Fab was modified with HYNIC- N-hydroxysuccinimide (NHS) and IRDye800-NHS to form HYNIC-αCD8/Fab and IRDye800-αCD8/Fab (Dye-αCD8/Fab), respectively. 99Tc m-αCD8/Fab was prepared in sodium bicarbonate buffer (pH=8.5), with SnCl 2 being used as the reducing agent. The labeling yield and radiochemical purity of 99Tc m-αCD8/Fab and its stability in PBS and fetal bovine serum (FBS) were tested in vitro. The mouse spleen and human peripheral blood lymphocytes were isolated for cell-specific binding and blocking experiments of 99Tc m-αCD8/Fab in vitro. SPECT/CT imaging was used to analyze the specific binding ability of the 99Tc m-αCD8/Fab probe in CT26 colon cancer mouse models (BALB/c). The near infrared fluorescence imaging and SPECT/CT imaging were performed to detect the intra-tumoral CD8 + T cell infiltration after anti-PD-1 therapy in tumor bearing mice, and the results were further verified by HE and immunofluorescence staining. CD8 + T cell depletion study was performed to determine the role of CD8 + T cells in the tumor responses to anti-PD-1 therapy. Two-way analysis of variance was used to compare the data difference. Results:The labeling yield of 99Tc m-αCD8/Fab was 90% with a high radiochemical purity (95%) and good stability in vitro (radiochemical purity still more than 80% after 720 min in PBS and FBS). 99Tc m-αCD8/Fab could specifically bind to mouse CD8 + T cells ((10.30±0.81) percent added radioactive dose (%AD)/10 6 cells), compared with the binding ability in human peripheral blood lymphocytes group and CD8 antibody blocking group ((1.78±0.61) and (1.59±0.25) %AD/10 6 cells; F=10.07, P<0.001). SPECT/CT imaging showed that 99Tc m-αCD8/Fab had markedly higher tumor uptake in the CT26 colon cancer mouse models. Near-infrared fluorescence imaging showed that the tumor uptake of 99Tc m-αCD8/Fab in the responsive group was significantly higher than in the nonresponsive group after anti-PD-1 treatment ((8.9±1.1)% vs (7.1±0.8)%; F=4.69, P=0.024), and SPECT/CT imaging found the similar result. HE and immunofluorescence staining of tumor and lymph nodes showed that the proportion of lymphocyte infiltration was higher in the responsive group. Furthermore, CD8 + T cell depletion significantly reversed the therapeutic effect of anti-PD-1 immunotherapy in tumor-bearing mice. Conclusions:In this study, 99Tc m-αCD8/Fab was successfully obtained. CD8-specific SPECT imaging could sensitively visualize the tumor-infiltrating CD8 + T cells, suggesting the potential application value to predict and evaluate the efficacy of immunotherapy in the clinical settings.
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Objective To investigate the efficacy and toxicity of stereotactic ablative radiotherapy for stage Ⅰ peripheral non-small cell lung cancers.Methods Thirty six patients of stage Ⅰ peripheral non-small cell lung cancers were treated with stereotactic ablative radiotherapy.The prescription dose was 12 Gy per fraction ×4 fraction in one to two weeks.The 100％ planning target volume (PTV) was covered by the isodose curve of 95％ prescription dose.Organs at risk and their respective tolerance doses used during treatment planning were developed from the research scheme of the Radiation Therapy Oncology Group 0236.Before the radiation delivery,all patients were scanned by the fan beam CT or the cone beam CT for image guidance and registration.The follow-up for the patients was given to observe the toxicity and efficacy of stereotactic ablative radiotherapy (SABR).Results The median follow-up time was 18.7 months (range of 4 to 36 months).After treatment,the overall response rate was 88.9％,with complete response (CR) 17 cases(47.2％),partial response (PR) 15 cases(41.7％),and stable disease (SD) 4 cases(11.1％).The estimated overall survival rate at 1 and 3 years was 92.3％ (95％ confidence interval [CI],86.3％ ～97.1％) and 85.3％ (95％ CI,80.5％ ～90.6％).The estimated local control rate at 3 years was 90.2％ (95％ CI,85.7％ ～94.8％).There was no gradeⅢ or above toxicity related to treatment.Conclusions The stereotactic ablative radiotherapy attains good local control and survival efficacy for the stage Ⅰ peripheral non-small lung cancer patients.It is well tolerated owing to low toxicity.
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Objective To measure the volumetric changes precisely during stereotactic ablative radiotherapy for early-stage and oligo-metastatic lung tumors and optimize the treatment plan timely.Methods From October 2011 to October 2014, 66 patients with 71 early-stage or oligo-metastatic lung tumors received SABR.Median age was 66 years.To measure the volume of tumors, the verification images were registered before each treatment fraction with stimulation images by reference to bone structure.Tumors volume was measured by the first verification images, and were defined as the reference when evaluating the trend of tumors volume change during SABR treatment.Generalized estimated equations were used to analyze the trend of the change of tumors volume over time with several possible predictors.The primary plan (P-plan) was modified when the biological effective dose (BED) of a tumor reached 60 Gy and volume change reached 25％.The modified plan was named as M-plan.Paired t-test was used to compare the dose of organs at risk (OAR) between M-plan and P-plan.Results In 71 tumors, 49 (69％) tumors showed volumetric shrinkage, 21 (30％) tumors showed enlargement and 1 tumor showed invariance.Generalized estimated equation showed no statistical significance (P =0.281) for the volumetric shrinkage of lung tumors.M-plan was made in 26 tumors.Of these tumors, 21 tumors decreased over 25 ％ and the result of paired t-test showed V5 of lung, Dmax and D1.2 cm3 of spinal cord, Dmax and D5 cm3 of esophagus and D30 cm3 of chest wall were statistically different between two plans(t =3.139 ～11.939 ,P＜0.05).5 tumors enlarged over 25％ and the result of paired t-test showed V5 and V20 of lung,Dmax and D1.2cm3 of spinal cord, Dmax of esophagus and D30cm3 of chest wall were statistically different between the two plans(t =-10.436--2.518, P ＜ 0.05).Conclusions Size of lung tumors changed dynamically during SABR, but it is unnecessary to modify treatment plans for all tumors.The tumors which showed obvious volumetric change may benefit from modifying treatment plans.
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Objective: To determine the potency of QN-2013, a derivative of quinoxaline 1,4-N-oxide, as a hypoxia-selective cytotoxin or a radiosensitizer. Methods: In vitro cytotoxicity and radiosensitization, as well as in vivo antitumor activity were determined by colony formation and tumor growth delay respectively. The changes in the cell cycle, DNA damage and repair of damaged DNA were assayed by FCM and “comet” assay, separately. Results: ICN250 and ICair50 of QN-2013 for HeLa-S3 cells were 0.08 and 1.7 mmol*L－1 respectively, namely, HCR=21. This suggested that QN-2013 was a fairly hypoxic cytotoxin, but inferior to SR-4233. QN-2013 had an evident radiosensitization either in vitro or in vivo. It was noted, however, that the value of in vitro SERs increased exponentially with increasing concentration of the drug, but the in situ antitumor activity seemed to be independent of doses of the drug. The systemic toxicity of QN-2013 was superior to an LD50 of 265 mg*kg－1 compared with 80 mg*kg－1 for SR-4233. In hypoxic condition QN-2013 induced S retension effect and G2M block in HeLa-S3 cells, caused DNA double strand break, and inhibited the repair of radiation-induced DNA damages. All of these reactivenesses might be involved in the action mechanism of QN-2013. Conclusion: QN-2013 is a fair hypoxia-selective cytotoxin, and has shown improved antitumor activity in vivo in combination with radiation. In general, These results suggest that the series of quinoxaline di-N-oxide derivatives hold out bright prospect for the development of novel bioreductive antitumor drugs.