title: Ratios of T-cell immune effectors and checkpoint molecules as prognostic biomarkers in diffuse large B-cell lymphoma: a population-based study.
abstract: BACKGROUND
  Risk-stratification of diffuse large B-cell lymphoma (DLBCL) requires identification of patients with disease that is not cured, despite initial treatment with R-CHOP. The prognostic importance of the revised International Prognostic Index (R-IPI) and cell of origin of the malignant B cell are established in DLBCL. We aimed to develop a novel, easily applicable, tissue-based prognostic biomarker based on quantification of the tumour microenvironment that is independent of and additive to the R-IPI and cell of origin.
  
  
  METHODS
  We performed digital hybridisation on the NanoString platform to assess the relation between immune effector and inhibitory (checkpoint) genes in 252 formalin-fixed, paraffin-embedded DLBCL tissue specimens obtained from patients treated with R-CHOP. We used a tree-based survival model to quantify net antitumoral immunity (using ratios of immune effector to checkpoint genes) and to generate a cutoff as an outcome predictor in 158 of the 252 patients. We validated this model in tissue (n=233) and blood (n=140) samples from two independent cohorts treated with R-CHOP.
  
  
  RESULTS
  T-cell and NK-cell immune effector molecule expression correlated with tumour-associated macrophage and PD-1/PD-L1 axis markers, consistent with malignant B cells triggering a dynamic checkpoint response to adapt to and evade immune surveillance. The ratio of CD4*CD8 to (CD163:CD68[M2])*PD-L1 was better able to stratify overall survival than was any one immune marker or combination, distinguishing groups with disparate 4-year overall survival. 94 (59%) of 158 patients had a score above the cutoff and 4-year overall survival of 92·1% (95% CI 82·9-96·7), and the remaining 64 (41%) patients had a score below the cutoff and 4-year overall survival of 47·0% (32·8-60·5; hazard ratio [HR] 8·3, 95% CI 4·3-17·3; p<0·0001). The CD4*CD8:M2*PD-L1 immune ratio was independent of and added to the R-IPI and cell of origin. Tissue findings in the independent tissue cohort accorded with those in our initial tissue cohort. 139 (60%) of 233 patients had a score above the cutoff and 4-year overall survival of 75·6% (95% CI 64·6-83·6), with the remaining 94 (40%) patients having a score below the cutoff (63·5% [52·5-72·7]; HR 1·9, 95% CI 1·1-3·3; p=0·0067).
  
  
  CONCLUSIONS
  Ratios of immune effectors to checkpoints augment the cell of origin and R-IPI in DLBCL and are applicable to paraffin-embedded biopsy specimens. These findings might have potential implications for selection of patients for checkpoint blockade within clinical trials.
  
  
  BACKGROUND
  Leukaemia Foundation of Queensland, Kasey-Anne Oklobdzijato Memorial Fund, the Australasian Leukaemia and Lymphoma Group (Malcolm Broomhead Bequest), the Australian Cancer Research Foundation, and the Cancer Council of Queensland.
authors.affiliation: University of Queensland Diamantina Institute, Translational Research Institute, University of Queensland, Brisbane, QLD, Australia; Princess Alexandra Hospital, Brisbane, QLD, Australia; Genomics Research Centre, Griffith University, Southport, QLD, Australia.
authors.forename: Colm
authors.initials: C
authors.lastname: Keane
authors.affiliation: University of Queensland Diamantina Institute, Translational Research Institute, University of Queensland, Brisbane, QLD, Australia.
authors.forename: Frank
authors.initials: F
authors.lastname: Vari
authors.affiliation: Department of Haematology, Prince of Wales Hospital, Sydney, NSW, Australia.
authors.forename: Mark
authors.initials: M
authors.lastname: Hertzberg
authors.affiliation: University of Queensland Diamantina Institute, Translational Research Institute, University of Queensland, Brisbane, QLD, Australia.
authors.forename: Kim-Anh Lê
authors.initials: KA
authors.lastname: Cao
authors.affiliation: Eppley Institute for Research in Cancer and Allied Diseases, University of Nebraska Medical Center, Omaha, NE, USA.
authors.forename: Michael R
authors.initials: MR
authors.lastname: Green
authors.affiliation: University of Queensland Diamantina Institute, Translational Research Institute, University of Queensland, Brisbane, QLD, Australia.
authors.forename: Erica
authors.initials: E
authors.lastname: Han
authors.affiliation: Peter MacCallum Cancer Centre and University of Melbourne, Melbourne, VIC, Australia.
authors.forename: John F
authors.initials: JF
authors.lastname: Seymour
authors.affiliation: Peter MacCallum Cancer Centre and University of Melbourne, Melbourne, VIC, Australia.
authors.forename: Rodney J
authors.initials: RJ
authors.lastname: Hicks
authors.affiliation: University of Queensland Diamantina Institute, Translational Research Institute, University of Queensland, Brisbane, QLD, Australia; Princess Alexandra Hospital, Brisbane, QLD, Australia.
authors.forename: Devinder
authors.initials: D
authors.lastname: Gill
authors.affiliation: University of Queensland Diamantina Institute, Translational Research Institute, University of Queensland, Brisbane, QLD, Australia.
authors.forename: Pauline
authors.initials: P
authors.lastname: Crooks
authors.affiliation: University of Queensland Diamantina Institute, Translational Research Institute, University of Queensland, Brisbane, QLD, Australia; Princess Alexandra Hospital, Brisbane, QLD, Australia.
authors.forename: Clare
authors.initials: C
authors.lastname: Gould
authors.affiliation: University of Queensland Diamantina Institute, Translational Research Institute, University of Queensland, Brisbane, QLD, Australia.
authors.forename: Kimberley
authors.initials: K
authors.lastname: Jones
authors.affiliation: Genomics Research Centre, Griffith University, Southport, QLD, Australia.
authors.forename: Lyn R
authors.initials: LR
authors.lastname: Griffiths
authors.affiliation: Canberra Hospital, Canberra, ACT, Australia; Australian National University Medical School, Acton, ACT, Australia.
authors.forename: Dipti
authors.initials: D
authors.lastname: Talaulikar
authors.affiliation: Canberra Hospital, Canberra, ACT, Australia.
authors.forename: Sanjiv
authors.initials: S
authors.lastname: Jain
authors.affiliation: Australian National University Medical School, Acton, ACT, Australia.
authors.forename: Josh
authors.initials: J
authors.lastname: Tobin
authors.affiliation: University of Queensland Diamantina Institute, Translational Research Institute, University of Queensland, Brisbane, QLD, Australia; Princess Alexandra Hospital, Brisbane, QLD, Australia. Electronic address: M.Gandhi@uq.edu.au.
authors.forename: Maher K
authors.initials: MK
authors.lastname: Gandhi
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D058846:Antibodies, Monoclonal, Murine-Derived / Q000627:therapeutic use; D000971:Antineoplastic Combined Chemotherapy Protocols / Q000627:therapeutic use; D001315:Australia; D060890:B7-H1 Antigen / Q000276:immunology; D015415:Biomarkers / Q000378:metabolism; D016516:CD4-CD8 Ratio; D003520:Cyclophosphamide / Q000627:therapeutic use; D004317:Doxorubicin / Q000627:therapeutic use; D005260:Female; D006801:Humans; D007694:Killer Cells, Natural / Q000276:immunology; D016403:Lymphoma, Large B-Cell, Diffuse / Q000188:drug therapy / Q000276:immunology*; D008297:Male; D008875:Middle Aged; D011241:Prednisone / Q000627:therapeutic use; D011379:Prognosis; D061026:Programmed Cell Death 1 Receptor / Q000276:immunology; D011793:Queensland; D000069283:Rituximab; D015996:Survival Rate; D013601:T-Lymphocytes / Q000276:immunology*; D016896:Treatment Outcome; D014750:Vincristine / Q000627:therapeutic use; D055815:Young Adult
chemical_list: D058846:Antibodies, Monoclonal, Murine-Derived; D060890:B7-H1 Antigen; D015415:Biomarkers; C423236:CD274 protein, human; C105992:PDCD1 protein, human; D061026:Programmed Cell Death 1 Receptor; C571759:R-CHOP protocol; D000069283:Rituximab; D014750:Vincristine; D004317:Doxorubicin; D003520:Cyclophosphamide; D011241:Prednisone