Spiral.imperial.ac.uk



Pembrolizumab induces long term complete remission in patients with chemotherapy resistant gestational trophoblastic neoplasia despite classical HLA deficiencyEhsan Ghorani MRCP1,2, Baljeet Kaur FRCPath3, Rosemary A. Fisher PhD1, Dee Short1, Ulrika Joneborg PhD4, Joseph W. Carlson PhD5, Ayse Akarca BSc6, Teresa Marafioti PhD6, Sergio A. Quezada PhD2, Naveed Sarwar PhD1 and Michael J. Seckl PhD1Department of Medical Oncology, Charing Cross Gestational Trophoblastic Disease Centre, Charing Cross Hospital Campus of Imperial College London, UK.Cancer Immunology Unit, University College London (UCL) Cancer Institute, London UK.Department of Histopathology, Charing Cross Gestational Trophoblastic Disease Centre, Charing Cross Hospital Campus of Imperial College London, London, UKDepartment of Women's and Children's Health, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden.Department of Oncology-Pathology, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden. Department of Cellular Pathology, UCL Hospital, London, UKCorrespondence Prof Michael Seckl, Department of Medical Oncology, Charing Cross Gestational Trophoblastic Disease Centre, Charing Cross Hospital Campus of Imperial College London, London W6 8RF, UK. E-mail: m.seckl@imperial.ac.uk.Telephone: +44 (0)20 3311 1421Fax: +44 (0)20 8383 5577IntroductionGestational trophoblastic disease (GTD) is a spectrum of pregnancy related disorders ranging from pre-malignant hydatidiform mole through to malignant forms collectively referred to as gestational trophoblastic neoplasia (GTN). GTN includes the malignant invasive mole, choriocarcinoma and rare placental site trophoblastic and epithelioid trophoblastic tumours (PSTT/ETT).ADDIN CSL_CITATION { "citationItems" : [ { "id" : "ITEM-1", "itemData" : { "DOI" : "10.1016/S0140-6736(10)60280-2", "ISSN" : "1474-547X", "PMID" : "20673583", "abstract" : "Gestational trophoblastic disease encompasses a range of pregnancy-related disorders, consisting of the premalignant disorders of complete and partial hydatidiform mole, and the malignant disorders of invasive mole, choriocarcinoma, and the rare placental-site trophoblastic tumour. These malignant forms are termed gestational trophoblastic tumours or neoplasia. Improvements in management and follow-up protocols mean that overall cure rates can exceed 98% with fertility retention, whereas most women would have died from malignant disease 60 years ago. This success can be explained by the development of effective treatments, the use of human chorionic gonadotropin as a biomarker, and centralisation of care. We summarise strategies for management of gestational trophoblastic disease and address some of the controversies and future research directions.", "author" : [ { "dropping-particle" : "", "family" : "Seckl", "given" : "Michael J", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Sebire", "given" : "Neil J", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Berkowitz", "given" : "Ross S", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" } ], "container-title" : "Lancet (London, England)", "id" : "ITEM-1", "issue" : "9742", "issued" : { "date-parts" : [ [ "2010", "8", "28" ] ] }, "page" : "717-29", "title" : "Gestational trophoblastic disease.", "type" : "article-journal", "volume" : "376" }, "uris" : [ "" ] } ], "mendeley" : { "formattedCitation" : "¹", "plainTextFormattedCitation" : "1", "previouslyFormattedCitation" : "¹" }, "properties" : { "noteIndex" : 0 }, "schema" : "" }1 Globally, approximately 18,000 women are diagnosed annually with GTN, most of whom are cured with chemotherapy guided by a sensitive disease response biomarker, human chorionic gonadotrophin (hCG). However, 0.5-5% of women die from multi-drug resistance, necessitating novel approaches. Risk factors for poor survival include drug refractory diseaseADDIN CSL_CITATION { "citationItems" : [ { "id" : "ITEM-1", "itemData" : { "DOI" : "10.1038/sj.bjc.6603608", "ISSN" : "0007-0920", "PMID" : "17299394", "abstract" : "The majority of women requiring chemotherapy for gestational trophoblastic disease (GTN) are cured with their initial chemotherapy treatment. However, a small percentage either become refractory to treatment, or relapse after the completion of treatment. This study investigates the characteristics and outcome of these patients. Patients were identified from the Charing Cross Hospital GTD database. The outcome of these patients with relapsed disease was compared to those with refractory disease. Between 1980 and 2004, 1708 patients were treated with chemotherapy for GTN. Sixty (3.5%) patents relapsed following completion of initial therapy. The overall 5-year survival for patients with relapsed GTN was 93% (95% CI 86-100%). The overall survival for patients with low-risk and high-risk disease at presentation, who subsequently relapsed was 100% (n=35), and 84% (n=25) (95% CI: 66-96%: P<0.05), respectively. Eleven patients were identified who failed to enter remission and had refractory disease. These patients had a worse outcome compared to patients with relapsed disease (5-year survival 43% (95% CI:12-73% P<0.01)). The outcome of patients with relapsed GTN is good. However, patients with primary chemo-refractory disease do poorly and novel therapies are required for this group of patients.", "author" : [ { "dropping-particle" : "", "family" : "Powles", "given" : "T", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Savage", "given" : "P M", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Stebbing", "given" : "J", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Short", "given" : "D", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Young", "given" : "A", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Bower", "given" : "M", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Pappin", "given" : "C", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Schmid", "given" : "P", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Seckl", "given" : "M J", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" } ], "container-title" : "British journal of cancer", "id" : "ITEM-1", "issue" : "5", "issued" : { "date-parts" : [ [ "2007" ] ] }, "page" : "732-7", "title" : "A comparison of patients with relapsed and chemo-refractory gestational trophoblastic neoplasia.", "type" : "article-journal", "volume" : "96" }, "uris" : [ "" ] } ], "mendeley" : { "formattedCitation" : "²", "plainTextFormattedCitation" : "2", "previouslyFormattedCitation" : "²" }, "properties" : { "noteIndex" : 0 }, "schema" : "" }2, liver and/or brain metastasesADDIN CSL_CITATION { "citationItems" : [ { "id" : "ITEM-1", "itemData" : { "ISSN" : "0024-7758", "PMID" : "22696824", "abstract" : "OBJECTIVE To determine whether survival outcomes of women with liver metastases from gestational trophoblastic neoplasia (GTN) have improved from the previous finding of 27% at 5 years. STUDY DESIGN The Charing Cross GTN database was searched for patients with liver metastases treated between 1975 and 2007. Prognostic variables were recorded and analyzed for effect on survival. RESULTS Thirty-eight (1.8%) of 2,100 GTN patients had liver metastases. One patient with placental site trophoblastic tumor was excluded. In the remaining 37 cases the overall survival was 48% at 5 years. Seven patients with very advanced disease died <4 weeks after admission, and 12 late deaths occurred, 5 due to non-GTN causes (1 stroke and 4 second cancers). After exclusion of the early deaths and censoring for the non-GTN related deaths, the cause-specific survival was 68%. No prognostic variable was significant on univariate analysis. However, patients presenting >2.8 years and <2.8 years from the antecedent pregnancy had a 32% and 75% (p = 0.08) chance of long-term survival, respectively. CONCLUSION The prognosis of patients with liver metastases from GTN has improved. Outcome may be best in those patients presenting within 2.8 years of the causative pregnancy and without very large volumes of disease.", "author" : [ { "dropping-particle" : "", "family" : "Ahamed", "given" : "Eliyaz", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Short", "given" : "Dee", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "North", "given" : "Bernard", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Savage", "given" : "Philip M", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Seckl", "given" : "Michael J", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" } ], "container-title" : "The Journal of reproductive medicine", "id" : "ITEM-1", "issue" : "5-6", "issued" : { "date-parts" : [ [ "0" ] ] }, "page" : "262-9", "title" : "Survival of women with gestational trophoblastic neoplasia and liver metastases: is it improving?", "type" : "article-journal", "volume" : "57" }, "uris" : [ "" ] }, { "id" : "ITEM-2", "itemData" : { "DOI" : "10.1016/j.ygyno.2015.01.530", "ISSN" : "1095-6859", "PMID" : "25598530", "abstract" : "OBJECTIVE To update the demographic data, treatment details and outcomes for GTN patients with brain metastases managed with the modern treatment protocols at the UK centre for gestational trophoblast neoplasia at Charing Cross Hospital in London. METHODS The hospital database and pharmacy records were reviewed to identify GTN patients treated with brain metastases. Data was assembled on the specific GTN diagnosis, staging, prognostic scores, chemotherapy regimens, additional interventions and outcomes. RESULTS During the 22 year study period, 27 GTN patients with brain metastases were treated. One case clearly resulted from a prior molar pregnancy, 3 were of uncertain aetiology and 23 cases had no prior molar pregnancy. The standard chemotherapy regimens were EMA-CO or EMA-EP given with an enhanced CNS methotrexate dose combined with intrathecal methotrexate. Five patients required emergency neurosurgery and routine radiotherapy was not employed. Twenty three (85%) patients are long term survivors and four patients died. Of the patients who died, all four had chemotherapy refractive disease and two had extended intervals of 18 and 30 years from their antecedent pregnancy. CONCLUSION The incidence of brain metastases in postmolar pregnancy GTN is extremely low. Patients with non-molar choriocarcinoma have an approximate 20% risk of CNS disease and should have routine CNS imaging. Treatment with CNS doses of EMA-CO or EMA-EP appears curative for most patients.", "author" : [ { "dropping-particle" : "", "family" : "Savage", "given" : "Philip", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Kelpanides", "given" : "Inga", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Tuthill", "given" : "Mark", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Short", "given" : "Dee", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Seckl", "given" : "Michael J", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" } ], "container-title" : "Gynecologic oncology", "id" : "ITEM-2", "issue" : "1", "issued" : { "date-parts" : [ [ "2015", "4" ] ] }, "page" : "73-6", "title" : "Brain metastases in gestational trophoblast neoplasia: an update on incidence, management and outcome.", "type" : "article-journal", "volume" : "137" }, "uris" : [ "" ] } ], "mendeley" : { "formattedCitation" : "^3,4", "plainTextFormattedCitation" : "3,4" }, "properties" : { "noteIndex" : 0 }, "schema" : "" }3,4 and PSTT/ETT that develops four or more years after the antecedent pregnancy.ADDIN CSL_CITATION { "citationItems" : [ { "id" : "ITEM-1", "itemData" : { "DOI" : "10.1016/S0140-6736(09)60618-8", "ISSN" : "1474-547X", "PMID" : "19552948", "abstract" : "BACKGROUND: Placental-site trophoblastic tumours are a rare form of gestational trophoblastic disease and consequently information about optimum management or prognostic factors is restricted. We aimed to assess the long-term outcome of stage-adapted management by surgery, chemotherapy, or both for patients with the disorder.\n\nMETHODS: 35 550 women were registered with gestational trophoblastic disease in the UK (1976-2006), of whom 62 were diagnosed with placental-site trophoblastic tumours and included, retrospectively, in the study. Patients were treated by surgery, chemotherapy, or both. We estimated the probabilities of overall survival and survival without recurrence of disease 5 and 10 years after the date of first treatment, and calculated the association of these endpoints with prognostic factors, including time since antecedent pregnancy, serum concentration of beta-human chorionic gonadotropin, and stage of disease, with both univariate and multivariate analyses.\n\nFINDINGS: Probabilities of overall and recurrence-free survival 10 years after first treatment were 70% (95% CI 54-82) and 73% (54-85), respectively. Patients with stage I disease had a 10-year probability of overall survival of 90% (77-100) and did not benefit from postoperative chemotherapy. By contrast, patients with stage II, III, and IV disease required combined treatment with surgery and chemotherapy; probability of overall survival at 10 years was 52% (3-100) for patients with stage II disease and 49% (26-72) for stage III or IV disease. Outcome for patients who had recurrent or refractory disease was poor: only four (22%) patients achieved long-term survival beyond 60 months. Multivariate analysis showed that the only significant independent predictor of overall and recurrence-free survival was time since antecedent pregnancy. A cutoff point of 48 months since antecedent pregnancy could differentiate between patients' probability of survival (<48 months) or death (>/=48 months) with 93% specificity and 100% sensitivity, and with a positive predictive value of 100% and a negative predictive value of 98%.\n\nINTERPRETATION: Stage-adapted management with surgery for stage I disease, and combined surgery and chemotherapy for stage II, III, and IV disease could improve the effectiveness of treatment for placental-site trophoblastic tumours. Use of 48 months since antecedent pregnancy as a prognostic indicator of survival could help select patients for risk-adapted treatment.\n\nFUNDI\u2026", "author" : [ { "dropping-particle" : "", "family" : "Schmid", "given" : "Peter", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Nagai", "given" : "Yutaka", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Agarwal", "given" : "Roshan", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Hancock", "given" : "Barry", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Savage", "given" : "Philip M", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Sebire", "given" : "Neil J", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Lindsay", "given" : "Iain", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Wells", "given" : "Michael", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Fisher", "given" : "Rosemary A", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Short", "given" : "Delia", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Newlands", "given" : "Edward S", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Wischnewsky", "given" : "Manfred B", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Seckl", "given" : "Michael J", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" } ], "container-title" : "Lancet (London, England)", "id" : "ITEM-1", "issue" : "9683", "issued" : { "date-parts" : [ [ "2009", "7" ] ] }, "page" : "48-55", "title" : "Prognostic markers and long-term outcome of placental-site trophoblastic tumours: a retrospective observational study.", "type" : "article-journal", "volume" : "374" }, "uris" : [ "" ] } ], "mendeley" : { "formattedCitation" : "⁵", "plainTextFormattedCitation" : "5", "previouslyFormattedCitation" : "³" }, "properties" : { "noteIndex" : 0 }, "schema" : "" }5Anti-cancer T-cell activity is regulated by multiple suppressive mechanisms, including tumour-expressed programmed death ligand 1 (PDL1) signalling to the T-cell inhibitory receptor PD1. Monoclonal antibodies to block this pathway, such as the anti-PD1 agent pembrolizumab, have shown impressive clinical activity in several cancer types.ADDIN CSL_CITATION { "citationItems" : [ { "id" : "ITEM-1", "itemData" : { "DOI" : "10.1016/S0140-6736(15)01281-7", "ISSN" : "1474-547X", "PMID" : "26712084", "abstract" : "BACKGROUND Despite recent advances in the treatment of advanced non-small-cell lung cancer, there remains a need for effective treatments for progressive disease. We assessed the efficacy of pembrolizumab for patients with previously treated, PD-L1-positive, advanced non-small-cell lung cancer. METHODS We did this randomised, open-label, phase 2/3 study at 202 academic medical centres in 24 countries. Patients with previously treated non-small-cell lung cancer with PD-L1 expression on at least 1% of tumour cells were randomly assigned (1:1:1) in blocks of six per stratum with an interactive voice-response system to receive pembrolizumab 2 mg/kg, pembrolizumab 10 mg/kg, or docetaxel 75 mg/m(2) every 3 weeks. The primary endpoints were overall survival and progression-free survival both in the total population and in patients with PD-L1 expression on at least 50% of tumour cells. We used a threshold for significance of p<0.00825 (one-sided) for the analysis of overall survival and a threshold of p<0.001 for progression-free survival. This trial is registered at , number NCT01905657. FINDINGS Between Aug 28, 2013, and Feb 27, 2015, we enrolled 1034 patients: 345 allocated to pembrolizumab 2 mg/kg, 346 allocated to pembrolizumab 10 mg/kg, and 343 allocated to docetaxel. By Sept 30, 2015, 521 patients had died. In the total population, median overall survival was 10.4 months with pembrolizumab 2 mg/kg, 12.7 months with pembrolizumab 10 mg/kg, and 8.5 months with docetaxel. Overall survival was significantly longer for pembrolizumab 2 mg/kg versus docetaxel (hazard ratio [HR] 0.71, 95% CI 0.58-0.88; p=0.0008) and for pembrolizumab 10 mg/kg versus docetaxel (0.61, 0.49-0.75; p<0.0001). Median progression-free survival was 3.9 months with pembrolizumab 2 mg/kg, 4.0 months with pembrolizumab 10 mg/kg, and 4.0 months with docetaxel, with no significant difference for pembrolizumab 2 mg/kg versus docetaxel (0.88, 0.74-1.05; p=0.07) or for pembrolizumab 10 mg/kg versus docetaxel (HR 0.79, 95% CI 0.66-0.94; p=0.004). Among patients with at least 50% of tumour cells expressing PD-L1, overall survival was significantly longer with pembrolizumab 2 mg/kg than with docetaxel (median 14.9 months vs 8.2 months; HR 0.54, 95% CI 0.38-0.77; p=0.0002) and with pembrolizumab 10 mg/kg than with docetaxel (17.3 months vs 8.2 months; 0.50, 0.36-0.70; p<0.0001). Likewise, for this patient population, progression-free survival was significantly longer with pembroli\u2026", "author" : [ { "dropping-particle" : "", "family" : "Herbst", "given" : "Roy S", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Baas", "given" : "Paul", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Kim", "given" : "Dong-Wan", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Felip", "given" : "Enriqueta", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "P\u00e9rez-Gracia", "given" : "Jos\u00e9 L", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Han", "given" : "Ji-Youn", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Molina", "given" : "Julian", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Kim", "given" : "Joo-Hang", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Arvis", "given" : "Catherine Dubos", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Ahn", "given" : "Myung-Ju", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Majem", "given" : "Margarita", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Fidler", "given" : "Mary J", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Castro", "given" : "Gilberto", "non-dropping-particle" : "de", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Garrido", "given" : "Marcelo", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Lubiniecki", "given" : "Gregory M", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Shentu", "given" : "Yue", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Im", "given" : "Ellie", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Dolled-Filhart", "given" : "Marisa", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Garon", "given" : "Edward B", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" } ], "container-title" : "Lancet (London, England)", "id" : "ITEM-1", "issue" : "10027", "issued" : { "date-parts" : [ [ "2016", "4", "9" ] ] }, "page" : "1540-50", "title" : "Pembrolizumab versus docetaxel for previously treated, PD-L1-positive, advanced non-small-cell lung cancer (KEYNOTE-010): a randomised controlled trial.", "type" : "article-journal", "volume" : "387" }, "uris" : [ "" ] } ], "mendeley" : { "formattedCitation" : "⁶", "plainTextFormattedCitation" : "6", "previouslyFormattedCitation" : "⁴" }, "properties" : { "noteIndex" : 0 }, "schema" : "" }6 Placental expression of paternal antigens make this organ a target for maternal immune recognition during pregnancy and PDL1 expression maintains gestational tolerance. In pre-clinical models, loss of placental PDL1/PD1 signalling results in foetal rejection.ADDIN CSL_CITATION { "citationItems" : [ { "id" : "ITEM-1", "itemData" : { "DOI" : "10.1084/jem.20050019", "PMID" : "16027236", "abstract" : "Fetal survival during gestation implies that tolerance mechanisms suppress the maternal immune response to paternally inherited alloantigens. Here we show that the inhibitory T cell costimulatory molecule, programmed death ligand 1 (PDL1), has an important role in conferring fetomaternal tolerance in an allogeneic pregnancy model. Blockade of PDL1 signaling during murine pregnancy resulted in increased rejection rates of allogeneic concepti but not syngeneic concepti. Fetal rejection was T cell- but not B cell-dependent because PDL1-specific antibody treatment caused fetal rejection in B cell-deficient but not in RAG-1-deficient females. Blockade of PDL1 also resulted in a significant increase in the frequency of IFN-gamma-producing lymphocytes in response to alloantigen in an ELISPOT assay and higher IFN-gamma levels in placental homogenates by ELISA. Finally, PDL1-deficient females exhibited decreased allogeneic fetal survival rates as compared with littermate and heterozygote controls and showed evidence of expansion of T helper type 1 immune responses in vivo. These results provide the first evidence that PDL1 is involved in fetomaternal tolerance.", "author" : [ { "dropping-particle" : "", "family" : "Guleria", "given" : "Indira", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Khosroshahi", "given" : "Arezou", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Ansari", "given" : "Mohammed Javeed", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Habicht", "given" : "Antje", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Azuma", "given" : "Miyuki", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Yagita", "given" : "Hideo", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Noelle", "given" : "Randolph J", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Coyle", "given" : "Anthony", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Mellor", "given" : "Andrew L", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Khoury", "given" : "Samia J", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Sayegh", "given" : "Mohamed H", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" } ], "container-title" : "The Journal of experimental medicine", "id" : "ITEM-1", "issue" : "2", "issued" : { "date-parts" : [ [ "2005", "7", "18" ] ] }, "page" : "231-7", "publisher" : "The Rockefeller University Press", "title" : "A critical role for the programmed death ligand 1 in fetomaternal tolerance.", "type" : "article-journal", "volume" : "202" }, "uris" : [ "" ] } ], "mendeley" : { "formattedCitation" : "⁷", "plainTextFormattedCitation" : "7", "previouslyFormattedCitation" : "⁵" }, "properties" : { "noteIndex" : 0 }, "schema" : "" }7 PDL1 is strongly expressed by GTNADDIN CSL_CITATION { "citationItems" : [ { "id" : "ITEM-1", "itemData" : { "DOI" : "10.1097/PGP.0000000000000305", "PMID" : "27362903", "abstract" : "One of the major immune checkpoints responsible for immune evasion in cancer cells is the interaction between programmed cell death-1 (PD-1) and its ligand (PD-L1). As human trophoblastic cells display many of the features of malignant cells such as the ability to invade normal tissue including blood vessels and are apparently not eradicated by the host immune system, we undertook the present study to determine whether PD-L1 was upregulated in different types of trophoblastic cells during normal pregnancy and in gestational trophoblastic diseases. Immunohistochemistry using an anti-PD-L1-specific antibody demonstrated that in early and term normal placentas, PD-L1 was highly expressed in syncytiotrophoblast and to a much lower extent in intermediate trophoblastic cells located in the chorion laeve and implantation site. PD-L1 immunoreactivity was undetectable in cytotrophoblastic cells. This staining pattern in normal placenta was recapitulated in various types of gestational trophoblastic disease. PD-L1 was highly expressed by syncytiotrophoblast in complete moles and choriocarcinomas. The intermediate trophoblastic neoplasms, placental site trophoblastic tumors, and epithelioid trophoblastic tumors showed variable PD-L1 immunoreactivity but at a lower intensity than in the neoplastic syncytiotrophoblast in choriocarcinoma. In addition, we observed PD-1-positive lymphocytes located within the implantation site and in trophoblastic tumors. In summary, this study describes a novel mechanism for trophoblastic cells to create a tolerogenic feto-maternal interface by upregulating PD-L1 in syncytiotrophoblast and in intermediate trophoblast. Trophoblastic tumors may also use PD-L1 expression to evade the host immune response thereby promoting their survival.", "author" : [ { "dropping-particle" : "", "family" : "Veras", "given" : "Emanuela", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Kurman", "given" : "Robert J", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Wang", "given" : "Tian-Li", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Shih", "given" : "Ie-Ming", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" } ], "container-title" : "International journal of gynecological pathology : official journal of the International Society of Gynecological Pathologists", "id" : "ITEM-1", "issued" : { "date-parts" : [ [ "2016", "6", "29" ] ] }, "title" : "PD-L1 Expression in Human Placentas and Gestational Trophoblastic Diseases.", "type" : "article-journal" }, "uris" : [ "" ] }, { "id" : "ITEM-2", "itemData" : { "DOI" : "10.1097/IGC.0000000000000892", "PMID" : "28060141", "abstract" : "OBJECTIVE Recently reported expression of programmed cell death 1 ligand 1 (PD-L1) in gestational trophoblastic diseases (GTDs) suggests that the immune tolerance of pregnancy might be hijacked during neoplastic process. We assessed PD-L1 protein expression in premalignant and malignant GTD lesions and analyzed associations with disease severity and chemotherapy outcomes. METHODS We included 83 GTD whole-tissue sections from 76 patients in different treatment settings. PD-L1 protein expression was assessed with immunohistochemistry in each trophoblast subtype with the Allred total score (ATS), which combines intensity and proportion expression on a 0- to 8-point scale. Peritumoral immune infiltrate was scored on hematoxylin-eosin-safran-stained slides. RESULTS PD-L1 expression was ubiquitous and strong in all GTD trophoblast subtypes. For invasive moles, ATS scores were maximal at 8 in 100%, 100%, and 75% of syncytiotrophoblast, villous cytotrophoblast, and extravillous cytotrophoblast specimens, respectively. For choriocarcinomas, ATS was 8 in 80% of specimens. Immune infiltrates were moderate to severe in 61%, 100%, and 100% of peritumoral zones of choriocarcinoma, epithelioid trophoblastic tumor, and invasive moles, respectively. Because of the homogeneous pathological findings, no significant differences in PD-L1 expression profiles or peritumoral immune infiltrates were found regarding FIGO (International Federation of Gynecology Obstetrics) prognostic score, fatal outcome, or chemosensitivity. CONCLUSIONS We confirm that PD-L1 is constitutively expressed in all GTD premalignant and malignant trophoblast subtypes, independently from FIGO score, chemoresistance, or fatal outcomes, thereby suggesting a crucial role for PD-L1 in the development and tolerance of GTD. Assessment of anti-PD-L1 drug in GTD patients has been activated.", "author" : [ { "dropping-particle" : "", "family" : "Bolze", "given" : "Pierre-Adrien", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Patrier", "given" : "Sophie", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Massardier", "given" : "J\u00e9r\u00f4me", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Hajri", "given" : "Touria", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Abbas", "given" : "Fatima", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Schott", "given" : "Anne Marie", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Allias", "given" : "Fabienne", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Devouassoux-Shisheboran", "given" : "Mojgan", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Freyer", "given" : "Gilles", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Golfier", "given" : "Fran\u00e7ois", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "You", "given" : "Beno\u00eet", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" } ], "container-title" : "International journal of gynecological cancer : official journal of the International Gynecological Cancer Society", "id" : "ITEM-2", "issue" : "3", "issued" : { "date-parts" : [ [ "2017", "3" ] ] }, "page" : "554-561", "title" : "PD-L1 Expression in Premalignant and Malignant Trophoblasts From Gestational Trophoblastic Diseases Is Ubiquitous and Independent of Clinical Outcomes.", "type" : "article-journal", "volume" : "27" }, "uris" : [ "" ] }, { "id" : "ITEM-3", "itemData" : { "DOI" : "10.1097/PAS.0000000000000653", "PMID" : "27158757", "abstract" : "Programmed cell death 1/programmed cell death ligand (PD-1/PD-Ls) axis is crucial for the modulation of immune responses and self-tolerance. Also, aberrant PD-L1 expression on the tumor cells or tumor-associated inflammatory cells accelerates immune evasion of tumor cells. In the past decade, PD-1/PD-L immune checkpoint inhibitors were introduced to cancer treatment trials and, in some cases, showed significant anticancer effects. PD-L1 immunohistochemical staining is considered a potential predictor of clinical response to PD-1/PD-L immune checkpoint inhibitor treatment. However, immunohistochemical data on PD-L1 expression in different types of cancer especially rare entities remain incomplete. In this study, PD-L1 expression was immunohistochemically analyzed in 5536 tumors including germ cell, epithelial, mesenchymal, melanocytic/neuroectodermal, and lymphohematopoietic tumors, as well as in a set of human normal tissues including a fetus. Immunohistochemical analysis was performed with E1L3N rabbit monoclonal antibody and Leica Bond Max automation using multitumor blocks containing up to 70 tumor samples. PD-L1 was constitutively and strongly expressed in placental trophoblasts as well as choriocarcinomas and trophoblastic components of germ cell tumors. Also, the neoplastic cells of classical Hodgkin lymphoma, anaplastic large cell lymphoma, schwannoma, thymoma, and squamous cell carcinoma of various sites frequently expressed PD-L1. In gastrointestinal adenocarcinomas, PD-L1-expression was associated with EBER positivity and mismatch-repair deficiency. In addition, PD-L1 was variably expressed in non-neoplastic macrophages and dendritic cells. PD-L1 immunohistochemistry may have some role in the immunophenotypic differential diagnosis of tumors and pinpointing potential candidates for anti-PD-1/PD-L immune checkpoint therapy.", "author" : [ { "dropping-particle" : "", "family" : "Inaguma", "given" : "Shingo", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Wang", "given" : "Zengfeng", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Lasota", "given" : "Jerzy", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Sarlomo-Rikala", "given" : "Maarit", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "McCue", "given" : "Peter A.", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Ikeda", "given" : "Hiroshi", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Miettinen", "given" : "Markku", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" } ], "container-title" : "The American Journal of Surgical Pathology", "id" : "ITEM-3", "issue" : "8", "issued" : { "date-parts" : [ [ "2016", "8" ] ] }, "page" : "1133-1142", "title" : "Comprehensive Immunohistochemical Study of Programmed Cell Death Ligand 1 (PD-L1)", "type" : "article-journal", "volume" : "40" }, "uris" : [ "" ] } ], "mendeley" : { "formattedCitation" : "^8\u201310", "plainTextFormattedCitation" : "8\u201310", "previouslyFormattedCitation" : "^6\u20138" }, "properties" : { "noteIndex" : 0 }, "schema" : "" }8–10, suggesting its role in tumour immune-evasion. We therefore hypothesised that targeting PD-1 inhibitory signalling with pembrolizumab may be effective in drug resistant GTN. Here we report the outcomes of four patients treated across two European centres.CasesAll patients had genetically verified gestational tumours (Table S1 and supplementary methods) with demographics summarised in Table S2. Patient 1 presented aged 39 with choriocarcinoma (Figure 1A) and multiple high risk factors including liver and brain metastases. At a second relapse three years later, she progressed through a fifth line of treatment including previous high dose chemotherapy (Figure 1B). Multiplex immunohistochemistry (IHC) on a chest wall biopsy revealed close to 100% tumour PDL1 expression, rich peritumoural and focal densities of tumour infiltrating lymphocytes (TILs; Figure 1C and D) predominantly CD8+ cytotoxic T-cells, half of which were PD1+ (Figure 1H). Tumour cells were negative for the class I major histocompatibility complex (MHC-I) antigen HLA-A and MHC-II (Figure 1F-G) but positive for the immuno-inhibitory, non-classical MHC-I antigen HLA-G (Figure 1E). The serum hCG of 80 IU/L normalised to <5 IU/L after four cycles of pembrolizumab (3 mg/kg, every three weeks), that was discontinued after five further consolidation cycles. Residual liver lesions regressed on serial imaging and she remains in complete remission over 24 months later. Patient 2 presented aged 44, 16 years after her last known pregnancy with uterine PSTT/ETT. Having failed multiple therapies over seven years (Figure S1A), tumour cells in the original hysterectomy sample and a lung metastasis biopsy were >90% PDL1 positive but TILs were absent (Figure S1B and C). Tumour cells were negative for HLA-A, pan-MHC-II and HLA-G (Figure S2D, E and data not shown). Reimaging after five cycles of pembrolizumab revealed disease progression in keeping with a rising serum hCG and she died four months later.Patient 3 presented aged 47 with metastatic PSTT to the lung, liver and brain. Her brain metastases progressed during third line therapy (Figure S2A). Significant treatment related neuropathy precluded further chemotherapy. IHC on tissue fragments from a uterine biopsy revealed > 90% PDL1 staining and the presence of TILs. Tumour cells were negative for HLA-A and pan-MHC-II but positive for HLA-G. Upon commencing pembrolizumab, the serum hCG rose from 73 to 4476 IU/L before falling after four cycles. Re-imaging showed a partial response of the pulmonary metastases but multiple new brain lesions (Figure S2B and C). Her serum and cerebrospinal fluid hCG normalised after eight cycles. After five consolidation cycles, reimaging demonstrated residual uterine necrotic tissue only, confirmed pathologically. She remains in remission over 15 months following marker normalisation. Patient 4 presented aged 37 with lung metastatic choriocarcinoma. She achieved remission after five lines of chemotherapy including high dose. IHC on a lung deposit revealed close to 100% tumour PDL1 expression with dense peri- and intratumoural foci of TILs, composed of approximately equal numbers of predominantly PD1 negative CD8+ and CD4+ helper T-cells with a high density of CD56+ natural killer (NK) cells. Tumour cells were HLA-A/MHC-II negative and HLA-G positive (Figure S1B-F and data not shown). On relapse five months later with lung and pelvic nodal disease, her hCG was 118 IU/L and pembrolizumab was commenced with marker normalisation after two cycles (Figure S3A). She received five consolidation courses and remains in remission over 5 months later. Treatment was well tolerated in all cases with mild (grade 1-2) toxicities managed conservatively and details presented in Table S1.DiscussionPatients with unresectable drug resistant GTN have a fatal outcome and anti-PD1 immunotherapy may be a life-saving treatment. The efficacy and favourable toxicity profile of pembrolizumab make its earlier use an attractive alternative to high dose chemotherapy. However, lasting fertility impairment due to persistent anti-trophoblast immunity is a concern that requires further study before offering therapy earlier in the treatment sequence. As previously reported, we found GTN to strongly express PD-L1 that in this small sample was not a biomarker of response to immunotherapy. Features of the TIL landscape correlate with anti-PD1 response,ADDIN CSL_CITATION { "citationItems" : [ { "id" : "ITEM-1", "itemData" : { "DOI" : "10.1038/nature13954", "ISBN" : "1476-4687 (Electronic)\\r0028-0836 (Linking)", "ISSN" : "0028-0836", "PMID" : "25428505", "abstract" : "Therapies that target the programmed death-1 (PD-1) receptor have shown unprecedented rates of durable clinical responses in patients with various cancer types. One mechanism by which cancer tissues limit the host immune response is via upregulation of PD-1 ligand (PD-L1) and its ligation to PD-1 on antigen-specific CD8(+) T cells (termed adaptive immune resistance). Here we show that pre-existing CD8(+) T cells distinctly located at the invasive tumour margin are associated with expression of the PD-1/PD-L1 immune inhibitory axis and may predict response to therapy. We analysed samples from 46 patients with metastatic melanoma obtained before and during anti-PD-1 therapy (pembrolizumab) using quantitative immunohistochemistry, quantitative multiplex immunofluorescence, and next-generation sequencing for T-cell antigen receptors (TCRs). In serially sampled tumours, patients responding to treatment showed proliferation of intratumoral CD8(+) T cells that directly correlated with radiographic reduction in tumour size. Pre-treatment samples obtained from responding patients showed higher numbers of CD8-, PD-1- and PD-L1-expressing cells at the invasive tumour margin and inside tumours, with close proximity between PD-1 and PD-L1, and a more clonal TCR repertoire. Using multivariate analysis, we established a predictive model based on CD8 expression at the invasive margin and validated the model in an independent cohort of 15 patients. Our findings indicate that tumour regression after therapeutic PD-1 blockade requires pre-existing CD8(+) T cells that are negatively regulated by PD-1/PD-L1-mediated adaptive immune resistance.", "author" : [ { "dropping-particle" : "", "family" : "Tumeh", "given" : "Paul C.", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Harview", "given" : "Christina L.", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Yearley", "given" : "Jennifer H.", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Shintaku", "given" : "I. Peter", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Taylor", "given" : "Emma J. M.", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Robert", "given" : "Lidia", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Chmielowski", "given" : "Bartosz", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Spasic", "given" : "Marko", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Henry", "given" : "Gina", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Ciobanu", "given" : "Voicu", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "West", "given" : "Alisha N.", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Carmona", "given" : "Manuel", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Kivork", "given" : "Christine", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Seja", "given" : "Elizabeth", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Cherry", "given" : "Grace", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Gutierrez", "given" : "Antonio J.", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Grogan", "given" : "Tristan R.", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Mateus", "given" : "Christine", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Tomasic", "given" : "Gorana", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Glaspy", "given" : "John a.", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Emerson", "given" : "Ryan O.", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Robins", "given" : "Harlan", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Pierce", "given" : "Robert H.", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Elashoff", "given" : "David a.", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Robert", "given" : "Caroline", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Ribas", "given" : "Antoni", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" } ], "container-title" : "Nature", "id" : "ITEM-1", "issue" : "7528", "issued" : { "date-parts" : [ [ "2014" ] ] }, "page" : "568-571", "title" : "PD-1 blockade induces responses by inhibiting adaptive immune resistance", "type" : "article-journal", "volume" : "515" }, "uris" : [ "" ] } ], "mendeley" : { "formattedCitation" : "¹¹", "plainTextFormattedCitation" : "11", "previouslyFormattedCitation" : "⁹" }, "properties" : { "noteIndex" : 0 }, "schema" : "" }11 suggesting a causal link between TIL absence and outcome in the single non-responder. Several cell types may mediate the effect of pembrolizumab on GTN rejection. Trophoblasts do not express the classical MHC-I molecules HLA-A/B, nor MHC–II, offering protection from T-cell mediated placental destruction.ADDIN CSL_CITATION { "citationItems" : [ { "id" : "ITEM-1", "itemData" : { "DOI" : "10.1016/j.fertnstert.2017.05.001", "abstract" : "Immunological adjustments are needed to accommodate the close contact between two genetically different individuals, the mother and her baby, during mammalian pregnancy. Contact occurs between fetal somatic or placental cells that enter the maternal systemic circulation or between uterine immune cells and the invading extravillous trophoblast. Here we discuss two main types of maternal allo-recognition of the fetus. One depends on avoidance of maternal T cells recognizing and responding to paternally-derived non-self human leukocyte antigens class I and class I allotypes. The other is natural killer allo-recognition where maternally-inherited variable killer immunoglobulin-like receptors expressed by uterine natural killer cells bind to polymorphic fetal human leukocyte antigens-C molecules displayed by extravillous trophoblast. Genetic studies indicate that natural killer cell allo-recognition regulates placentation and the allocation of resources to the fetus.", "author" : [ { "dropping-particle" : "", "family" : "Moffett", "given" : "Ashley", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Chazara", "given" : "Olympe", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Colucci", "given" : "Francesco", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" } ], "container-title" : "Fertility and Sterility", "id" : "ITEM-1", "issue" : "6", "issued" : { "date-parts" : [ [ "2017" ] ] }, "page" : "1269-1272", "title" : "Maternal allo-recognition of the fetus", "type" : "article-journal", "volume" : "107" }, "uris" : [ "" ] } ], "mendeley" : { "formattedCitation" : "¹²", "plainTextFormattedCitation" : "12", "previouslyFormattedCitation" : "¹⁰" }, "properties" : { "noteIndex" : 0 }, "schema" : "" }12 We report that GTN is similarly negative for HLA-A and MHC-II. The strong presence of infiltrating T-cells in responders suggests pembrolizumab activation of HLA-C directed or indirect T-cell cytotoxicity. Effectors other than classically restricted T-cells may also be relevant. One candidate are NK cells that express PD-1, are cytotoxic towards classical MHC-I negative cells and are inhibited by HLA-G, that also acts to maintain gestational tolerance through mechanisms including T-cell suppression.ADDIN CSL_CITATION { "citationItems" : [ { "id" : "ITEM-1", "itemData" : { "DOI" : "10.1038/onc.2008.267", "author" : [ { "dropping-particle" : "", "family" : "Waldhauer", "given" : "I", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Steinle", "given" : "A", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" } ], "container-title" : "Oncogene", "id" : "ITEM-1", "issue" : "45", "issued" : { "date-parts" : [ [ "2008", "10", "6" ] ] }, "page" : "5932-5943", "publisher" : "Nature Publishing Group", "title" : "NK cells and cancer immunosurveillance", "type" : "article-journal", "volume" : "27" }, "uris" : [ "" ] } ], "mendeley" : { "formattedCitation" : "¹³", "plainTextFormattedCitation" : "13", "previouslyFormattedCitation" : "¹¹" }, "properties" : { "noteIndex" : 0 }, "schema" : "" }13 Upregulated tumour expression of HLA-G by the responders suggests a role for this molecule in inhibiting infiltrating lymphocyte effectors. Beyond T-cell activation, the immune effects of pembrolizumab are not well understood but potentially of wider relevance, especially as HLA-G expression is a recognised mediator of tumour immune evasion.ADDIN CSL_CITATION { "citationItems" : [ { "id" : "ITEM-1", "itemData" : { "DOI" : "10.1158/0008-5472.CAN-07-2704", "PMID" : "18245459", "abstract" : "Immunosurveillance is an extrinsic mechanism of cancer suppression that eliminates nascent tumors. However, the selection imposed by immunosurveillance can drive tumor evolution and the emergence of clinically apparent neoplasms. Mechanisms of immune escape acquired by less immunogenic variants during this process, termed immunoediting, may contribute significantly to malignant progression. In this review, we summarize the evidence that up-regulation of the nonclassic human leukocyte antigen (HLA) class I molecule HLA-G in tumor cells plays an important role in cancer and immune escape.", "author" : [ { "dropping-particle" : "", "family" : "Urosevic", "given" : "M.", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" }, { "dropping-particle" : "", "family" : "Dummer", "given" : "R.", "non-dropping-particle" : "", "parse-names" : false, "suffix" : "" } ], "container-title" : "Cancer Research", "id" : "ITEM-1", "issue" : "3", "issued" : { "date-parts" : [ [ "2008", "2", "1" ] ] }, "page" : "627-630", "title" : "Human Leukocyte Antigen-G and Cancer Immunoediting", "type" : "article-journal", "volume" : "68" }, "uris" : [ "" ] } ], "mendeley" : { "formattedCitation" : "¹⁴", "plainTextFormattedCitation" : "14", "previouslyFormattedCitation" : "¹²" }, "properties" : { "noteIndex" : 0 }, "schema" : "" }14Complex restrictions precluded public funding of potentially life-saving off-label pembrolizumab for the UK treated patients 1-3. This report emphasizes the need to improve mechanisms for funding rare disease treatments, for which randomised trials and established licencing pathways are not feasible. Based on the evidence presented here, UK public funding of pembrolizumab for drug resistant GTN is now under review.In summary, we provide evidence in favour of pembrolizumab as an important advance in the management of drug resistant GTN that should strongly be considered in this setting. Tumour infiltrating lymphocytes and HLA-G expression may identify responders and our analysis of MHC expression suggests potential effects of pembrolizumab on cell types other than classically restricted T-cells. Acknowledgements: Professor Seckl would like to thank the continued support of the UK Dept of Health for the funding of the Gestational Trophoblastic Disease Service. Funding: EG is funded by a Wellcome Trust Clinical Research Fellowship and an NIHR Fellowship. MJS and RAF acknowledge support from CRUK, Department of Health, Imperial College Experimental Cancer Medicine Centre and NIHR Biomedical Research Centre. The Harris and Trotter Clients Charitable Trust funded therapy for two patients.Author contributions: EG, BK, TM, SAQ and MJS designed the study. EG, UJ, NS and MJS enrolled patients and participated in clinical data collection. BK, JWC, TM and AA carried out immuno-stains and pathological review. EG, BK, JWC, TM, RF, UJ, SAQ, NS and MJS contributed to data analysis and interpretation. RF and JWC carried out genetic analyses. DS provided administrative support. EG, AA, RF, UJ and MJS wrote the report. All authors have seen and approved the final version.ReferencesADDIN Mendeley Bibliography CSL_BIBLIOGRAPHY 1Seckl MJ, Sebire NJ, Berkowitz RS. Gestational trophoblastic disease. Lancet (London, England) 2010; 376: 717–29.2Powles T, Savage PM, Stebbing J, et al. A comparison of patients with relapsed and chemo-refractory gestational trophoblastic neoplasia. Br J Cancer 2007; 96: 732–7.3Ahamed E, Short D, North B, Savage PM, Seckl MJ. Survival of women with gestational trophoblastic neoplasia and liver metastases: is it improving? J Reprod Med; 57: 262–9.4Savage P, Kelpanides I, Tuthill M, Short D, Seckl MJ. Brain metastases in gestational trophoblast neoplasia: an update on incidence, management and outcome. Gynecol Oncol 2015; 137: 73–6.5Schmid P, Nagai Y, Agarwal R, et al. Prognostic markers and long-term outcome of placental-site trophoblastic tumours: a retrospective observational study. Lancet (London, England) 2009; 374: 48–55.6Herbst RS, Baas P, Kim D-W, et al. Pembrolizumab versus docetaxel for previously treated, PD-L1-positive, advanced non-small-cell lung cancer (KEYNOTE-010): a randomised controlled trial. Lancet (London, England) 2016; 387: 1540–50.7Guleria I, Khosroshahi A, Ansari MJ, et al. A critical role for the programmed death ligand 1 in fetomaternal tolerance. J Exp Med 2005; 202: 231–7.8Veras E, Kurman RJ, Wang T-L, Shih I-M. PD-L1 Expression in Human Placentas and Gestational Trophoblastic Diseases. Int J Gynecol Pathol 2016; published online June 29. DOI:10.1097/PGP.0000000000000305.9Bolze P-A, Patrier S, Massardier J, et al. PD-L1 Expression in Premalignant and Malignant Trophoblasts From Gestational Trophoblastic Diseases Is Ubiquitous and Independent of Clinical Outcomes. Int J Gynecol Cancer 2017; 27: 554–61.10Inaguma S, Wang Z, Lasota J, et al. Comprehensive Immunohistochemical Study of Programmed Cell Death Ligand 1 (PD-L1). Am J Surg Pathol 2016; 40: 1133–42.11Tumeh PC, Harview CL, Yearley JH, et al. PD-1 blockade induces responses by inhibiting adaptive immune resistance. Nature 2014; 515: 568–71.12Moffett A, Chazara O, Colucci F. Maternal allo-recognition of the fetus. Fertil Steril 2017; 107: 1269–72.13Waldhauer I, Steinle A. NK cells and cancer immunosurveillance. Oncogene 2008; 27: 5932–43.14Urosevic M, Dummer R. Human Leukocyte Antigen-G and Cancer Immunoediting. Cancer Res 2008; 68: 627–30.Figure 1. Genetics, treatments, response and immune phenotyping for Patient 10-778510left425823Figure 1. (A) Partial electropherograms of DNA from patient 1, her partner and tumour tissue. Genotyping of the tumour for short tandem repeat loci D21S11, D7S820 (shown) demonstrates a single non-maternal allele (solid peak) representing a contribution to the tumour genome from the partner. Both maternal alleles (open peaks) are present in the tumour DNA, the lower open peak (left) representing contamination from maternal cells in the tumour sections, while the higher open peak (right) represents a maternal contribution to the tumour genome plus a small proportion of contamination from maternal cells in the tumour sections. (B) Serum hCG is plotted against time from treatment initiation. Shaded bands represent duration of chemo- and immunotherapy with; a. Low dose EP; b. EP/EMA with IT MTX; c. TE/TP; d. HDCT; e. EP; f. Gem-TIP; Pembro, pembrolizumab (arrowheads show treatment dates). See Table S2 for a summary of therapeutic regimens. (C-F) Multiplex immunohistochemistry of the tumour immune landscape. (C) Expression of PDL1 and tumour infiltrating lymphocyte (TIL) expression of the activation/exhaustion marker PD1 and CD56 expressing natural killer cells; (D) TIL expression of CD4, PD1, FOXP3 (regulatory T-cells that co-express CD4) and CD8; (E-G) tumour expression of HLA-G, HLA-A and MHC-II. The latter three markers were counterstained for PDL1 expression to identify tumour cells. Normal HLA-A/MHC-II expression is seen on non-cancer cells including endothelial cells and lymphocytes, bars represent 50 ?m. (H) TILs in three areas comprising one third of the tumour were digitally and manually counted and the mean proportions normalised to CD8 T-cells. Error bars represent one standard deviation from the mean.00Figure 1. (A) Partial electropherograms of DNA from patient 1, her partner and tumour tissue. Genotyping of the tumour for short tandem repeat loci D21S11, D7S820 (shown) demonstrates a single non-maternal allele (solid peak) representing a contribution to the tumour genome from the partner. Both maternal alleles (open peaks) are present in the tumour DNA, the lower open peak (left) representing contamination from maternal cells in the tumour sections, while the higher open peak (right) represents a maternal contribution to the tumour genome plus a small proportion of contamination from maternal cells in the tumour sections. (B) Serum hCG is plotted against time from treatment initiation. Shaded bands represent duration of chemo- and immunotherapy with; a. Low dose EP; b. EP/EMA with IT MTX; c. TE/TP; d. HDCT; e. EP; f. Gem-TIP; Pembro, pembrolizumab (arrowheads show treatment dates). See Table S2 for a summary of therapeutic regimens. (C-F) Multiplex immunohistochemistry of the tumour immune landscape. (C) Expression of PDL1 and tumour infiltrating lymphocyte (TIL) expression of the activation/exhaustion marker PD1 and CD56 expressing natural killer cells; (D) TIL expression of CD4, PD1, FOXP3 (regulatory T-cells that co-express CD4) and CD8; (E-G) tumour expression of HLA-G, HLA-A and MHC-II. The latter three markers were counterstained for PDL1 expression to identify tumour cells. Normal HLA-A/MHC-II expression is seen on non-cancer cells including endothelial cells and lymphocytes, bars represent 50 ?m. (H) TILs in three areas comprising one third of the tumour were digitally and manually counted and the mean proportions normalised to CD8 T-cells. Error bars represent one standard deviation from the mean.Supplementary dataMaterials and methodsMultiplex immunohistochemistryTumour samples and reactive human tonsil tissue were fixed in 10% buffered formalin and embedded in paraffin according to standard protocols. 2-5m tissue sections were cut and transferred onto poly-l-lysine–coated slides, dewaxed in two changes of xylene and rehydrated in a series of graded alcohols. Single immunohistochemistry was carried out using the automated platforms BenchMark Ultra (Ventana/Roche) and the Bond-III Autostainer (Leica Microsystems) according to a protocol described elsewhere.1,2 To establish optimal staining conditions each antibody was tested and optimized on sections of human reactive tonsil, used as positive control. Monoclonal antibodies used were as follows; anti-CD4 (Spring Biosciences, clone SP35), anti-CD8 (Spring Biosciences, clone SP239), anti-PD1 clone NAT 105/E3 and anti-FOXP3 clone 236/E7 (kindly gifted by Dr. G. Roncador), anti-CD56 clone CD564 (Leica Microsystems), anti-HLA-A clone EP1395Y, anti-HLA-G clone 4H84 and anti-pan-MHC-II clone CR3/43 (Abcam). Anti-PDL1 clone SP263 (Ventana) was used to score tumour PDL1 status, whereas anti-PDL1 clone QR1 (Quarttet) was used to identify tumour cells in dual staining with anti-MHC antibodies.Multiplex immunohistochemistry was carried out using a protocol described previously.3 Specificity of the staining was assessed by a haematopathologist (TM) with expertise in multiplex-immunostaining. Slides were scanned using the Hamamatsu Nanozoomer Digital scanner. Staining was independently reviewed by a second pathologist with expertise in GTD (BK). Cell counts for Patient 1 were determined in ImageJ using automatic and manual methods.Fluorescent microsatellite genotyping of tumour tissue.In each case tumour and surrounding host tissue were independently microdissected from formalin-fixed, paraffin-embedded (FFPE) pathological sections of tumour tissue, with reference to a haematoxylin and eosin stained section. DNA was prepared from tumour and host tissue in all cases using a QIAmp DNA FFPE Tissue Kit (Qiagen; cases 1-3) or a PicoPure DNA extraction kit (Arcturus; case 4) and from blood samples from the patient and her partner in case 1 and 3. DNA was amplified with primers for 15 short tandem repeat (STR) loci on 13 chromosomes, together with the amelogenin locus, using an AmpFlSTR Identifiler Plus kit (Applied Biosystems; cases 1-3) or 9 STR using an AmpFlSTR Profiler Plus. Kit (Applied Biosystems; case 4). PCR products were resolved by capillary electrophoresis using an ABI 3100 or 3500 Genetic Analyser and genotypes determined using GeneMapper version 5.0 (Applied Biosystems) or GeneMarker version 2.4 software (SoftGenetics LLC). Demonstration of the presence of paternal alleles, or non-maternal alleles in cases 2 and 4 in DNA from the tumour confirmed a diagnosis of gestational trophoblastic neoplasia.References1Marafioti T, Paterson JC, Ballabio E, et al. Novel markers of normal and neoplastic human plasmacytoid dendritic cells. Blood 2008; 111: 3778–92.2Akarca AU, Shende VH, Ramsay AD, et al. BRAF V600E mutation-specific antibody, a sensitive diagnostic marker revealing minimal residual disease in hairy cell leukaemia. Br J Haematol 2013; 162: 848–51.3Marafioti T, Jones M, Facchetti F, et al. Phenotype and genotype of interfollicular large B cells, a subpopulation of lymphocytes often with dendritic morphology. Blood 2003; 102: 2868–76.Table S1. Genotyping of patient, tumour and partner tissue confirms gestational origin of the tumourLocus:D8S1179D21S11D7S820CSF1POD3S1358THO1D13S317D16S539D2S1338D19S433vWAD18S51D5S818FGAPatient 1Patient1129 - 3011 - 129 - 1414 - 188 - 9 9 - 119 - 1118 - 2014.2 - 1617 - 1815 - 1710 - 1119 - 22Tumour1127 - 308 - 129 - 10186*9 - 1310 - 112014.2-15.217 - 1815 - 1810 - 1326*Partner10 - 1127 - 298 - 101014 - 186 - 8138 - 1019 - 2014.2-15.217 - 181811 - 1320 - 26Patient 2Patient10 - 1330 - 319 - 111215 - 186 - 711 - 169 - 1017 - 201414 - 17 12 - 17 11 - 1218 - 22Tumour10 - 1531 - 31.29 - 101215 - 176 - 910 - 169 - 111714 - 16.217 16 - 171118 - 20Patient 3Patient1231 - 32.2 1110 - 1115911 - 16920 - 2113 - 151414 - 15 1123 - 24Tumour12*30 - 318 - 11N/S15 - 17911 - 169 - 12 21 - 25N/S14 - 1614 - 15 1119 - 24Partner10 - 15308 - 1011 - 12179 - 9.311 - 1611 - 12 22 - 251316 - 1715 - 2010 - 11 19Patient 4Patient12N/S8 - 9N/D16 - 18N/D9 - 11N/DN/DN/D17 - 2018 - 18.210 - 1122 - 25Tumour11 - 12N/S8 - 11N/D16 - 19N/D9 - 12N/DN/DN/D16 - 171811 - 1221 - 25Numbers represent alleles identified for informative tandem repeat loci and the amelogenin locus. Genotypes at the TPOX locus were uninformative in all cases. All tumours had a single sex chromosome specific peak representing DNA from the X chromosome. * Loci for which loss of heterozygosity was observed in cases 1 and 3; N/S, not scored. N/D, not done. DNA was not available for the partner of Patients 2 and 4.Table S2. Summary of patient baseline characteristics and prior therapies.Patient 1Patient 2Patient 3Patient 4Age142524837Performance status10210Obstetric statusG2P2G3P3G3P2G2P2GTN subtypeCCETT (predominant)/PSTTPSTTCCAntecedent pregnancy (years)316110.75Sites of diseaseNodal, hepaticNodal, hepatic, pulmonary, pleuralPelvic, hepatic, pulmonary, brainNodal, hepaticDisease duration (years)2.67.80.61.25FIGO score at diagnosis178206Prior chemotherapy (cycles)1Induction EP (1)EP/EMA (8)Induction EP (3)IM MTX (2)2EP/EMA CNS (8), IT MTX (2)TE/TP (4)EP/EMA CNS (5), IT MTX (3)EMA/CO (8)IT MTX (3)3TE/TP (4)-TE/TP (4)IT MTX (3)TE/TP (2)EP/EMA (1)4HDCT (2)-Escalated EP (5), IT MTX (8)HDCT (2)5Escalated EP (6)---6Gem-TIP (2)---Prior surgeryTMHBSO, TM-TMSerum hCG80246873118Biochemical response and duration (months)CR (24)PDCR (15)CR (5)Radiological responseOngoing PRPDOngoing PROngoing PRPembrolizumabCycles to hCG normalisation4-82Consolidation cycles5-55Adverse events (grade)Arthralgia (1)Pruritis (1)Synovitis (2), rash (1)Neutropaenia (2), synovitis (1)1Age or performance status at commencement of pembrolizumab. TM, thoracic metastasectomy; HBSO, hysterectomy and bilateral salpingo-oophorectomy; G (gravida); P (para). CR, complete response; PD, progressive disease; PR, partial response. Adverse events were graded according to the Common Terminology Criteria for Adverse Events (CTCAE) v4.03. Radiological responses were evaluated according to Response Evaluation Criteria In Solid Tumors (RECIST) v1.1. Details of chemotherapeutic regimens are given in Table S3.Table S3. Treatment regimensRegimen and cycle length (weeks)Agent and dosePembrolizumaba (3)3 mg/kg pembrolizumab D1IM MTX (2)50 mg methotrexate D1, 3, 5, 7EMA/CO (2)EMA0.5 mg actinomycin D1, 2100 mg/m2 etoposide D1, 2300 mg/m2 methotrexate D1CO1 g/m2 vincristine D8600 mg/m2 cyclophosphamide D8Induction EP (1)100 mg/m2 etoposide D1, 220 mg/m2 cisplatin D1, 2EP/EMA CNSb (2)EMA0.5 mg actinomycin D1, 2100 mg/m2 etoposide D1, 21000 mg/m2 methotrexate D1EP150 mg/m2 etoposide D875 mg/m2 cisplatin D8IT MTX (2)12.5 mg methotrexate D1TE/TPc (4)TE150 mg/m2 etoposide D1135 mg/m2 paclitaxel D1TP60 mg/m2 cisplatin D14135 mg/m2 paclitaxel D14Escalated EP (2)500mg/m2 etoposide D160mg/m2 cisplatin D1Gem-TIP (3)1200 mg/m2 gemcitabine D1175 mg/m2 paclitaxel D11000 mg/m2 ifosfamide D2-620 mg/m2 cisplatin D2-6HDCT75? mg/m2 paclitaxel D-7, -5, -3450? mg/m2 etoposide D-7, -5, -3AUC 10 carboplatin D-7, -5, -360? mg/kg cyclophosphamide D-5, -3Autologous haematopoietic stem cells D0IM, intramuscular; IT, intrathecal; D, day; AUC, area under the concentration-time curve. a. Patient 4 received pembrolizumab 200 mg every 2 weeks. b, For Patient 1, carboplatin AUC 4 was substituted for cisplatin. Patient 2 received 300 mg/m2 methotrexate. c, For Patient 1, carboplatin AUC 4 was substituted for cisplatin.Figure S1. Treatments, response and immune phenotyping for Patient 20389left487680 (A) Serum hCG is plotted against time from treatment initiation. Shaded bands represent duration of chemo- and immunotherapy with; a. EP/EMA; Pembro, pembrolizumab (arrowheads show treatment dates). H, hysterectomy; t, thoracic metastasectomy. TE/TP was additionally given prior to pembrolizumab at another institution (data not shown), see Table S2 for a summary of therapeutic regimens. (B-E) Multiplex immunohistochemistry on the uterine primary showing staining for PDL1, PD1 and CD56 (B); CD4, PD1, FOXP3 and CD8 (C); HLA-G (D) and HLA-A (F). The latter two were counterstained for PDL1 expression to identify tumour cells. Normal HLA-A expression is seen on non-cancer cells including endothelial cells and lymphocytes, and HLA-G expression is seen on non-tumour infiltrating cells. Bars represent 100 ?m.00 (A) Serum hCG is plotted against time from treatment initiation. Shaded bands represent duration of chemo- and immunotherapy with; a. EP/EMA; Pembro, pembrolizumab (arrowheads show treatment dates). H, hysterectomy; t, thoracic metastasectomy. TE/TP was additionally given prior to pembrolizumab at another institution (data not shown), see Table S2 for a summary of therapeutic regimens. (B-E) Multiplex immunohistochemistry on the uterine primary showing staining for PDL1, PD1 and CD56 (B); CD4, PD1, FOXP3 and CD8 (C); HLA-G (D) and HLA-A (F). The latter two were counterstained for PDL1 expression to identify tumour cells. Normal HLA-A expression is seen on non-cancer cells including endothelial cells and lymphocytes, and HLA-G expression is seen on non-tumour infiltrating cells. Bars represent 100 ?m.Figure S2. Treatments, response and imaging for Patient 3-3189985168407 (A) Serum hCG is plotted against time from treatment initiation. Shaded bands represent duration of chemo- and immunotherapy with; a. Induction EP; b. EP/EMA CNS; c. TE/TP; d. Escalated EP; Pembro, pembrolizumab (arrowheads show treatment dates). For treatments b-d, IT MTX was given concurrently. See Table S2 for a summary of therapeutic regimens. Serum (blue) and cerebrospinal fluid (CSF; purple) hCG response is shown. (B-C) Radiological response to pembrolizumab. (B) Serial contrast enhanced MRI head scans show flare and resolution of a brain metastasis. (C) Serial CT chest imaging shows resolution of a lung metastasis. Numbers represent months following initiation of pembrolizumab therapy (t=0 represents baseline imaging).00 (A) Serum hCG is plotted against time from treatment initiation. Shaded bands represent duration of chemo- and immunotherapy with; a. Induction EP; b. EP/EMA CNS; c. TE/TP; d. Escalated EP; Pembro, pembrolizumab (arrowheads show treatment dates). For treatments b-d, IT MTX was given concurrently. See Table S2 for a summary of therapeutic regimens. Serum (blue) and cerebrospinal fluid (CSF; purple) hCG response is shown. (B-C) Radiological response to pembrolizumab. (B) Serial contrast enhanced MRI head scans show flare and resolution of a brain metastasis. (C) Serial CT chest imaging shows resolution of a lung metastasis. Numbers represent months following initiation of pembrolizumab therapy (t=0 represents baseline imaging).Figure S3. Treatments, response and immunophenotyping for Patient 4-3314706891020(A) Serum hCG is plotted against time from treatment initiation. Shaded bands represent duration of chemo- and immunotherapy with; a. IM MTX; b. EMA/CO and IT MTX; c. TE/TP (and a single cycle of EP/EMA); d. HDCT; Pembro, pembrolizumab (arrowheads show treatment dates). t, thoracic metastasectomy. See Table S2 for a summary of therapeutic regimens. (B) Tumour infiltrating lymphocytes (TILs) in three areas comprising one third of the tumour were manually counted and the mean proportions normalised to CD8 T-cells. Error bars represent standard deviation. (C-F) Multiplex immunohistochemistry showing expression of PDL1, the activation/exhaustion marker PD1 and CD56 expressing natural killer cells (C); phenotype of TILs stained for CD4, PD1, FOXP3 (regulatory T-cells that co-express CD4) and CD8 (D); tumour HLA-G (E) and HLA-A (F). The latter two were counterstained for PDL1 expression to identify tumour cells. Normal HLA-A expression is seen on non-cancer cells predominantly representing stromal and inflammatory cells, bars represent 50 ?m.00(A) Serum hCG is plotted against time from treatment initiation. Shaded bands represent duration of chemo- and immunotherapy with; a. IM MTX; b. EMA/CO and IT MTX; c. TE/TP (and a single cycle of EP/EMA); d. HDCT; Pembro, pembrolizumab (arrowheads show treatment dates). t, thoracic metastasectomy. See Table S2 for a summary of therapeutic regimens. (B) Tumour infiltrating lymphocytes (TILs) in three areas comprising one third of the tumour were manually counted and the mean proportions normalised to CD8 T-cells. Error bars represent standard deviation. (C-F) Multiplex immunohistochemistry showing expression of PDL1, the activation/exhaustion marker PD1 and CD56 expressing natural killer cells (C); phenotype of TILs stained for CD4, PD1, FOXP3 (regulatory T-cells that co-express CD4) and CD8 (D); tumour HLA-G (E) and HLA-A (F). The latter two were counterstained for PDL1 expression to identify tumour cells. Normal HLA-A expression is seen on non-cancer cells predominantly representing stromal and inflammatory cells, bars represent 50 ?m. ................
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