ORCID Profile
0000-0002-1137-1726
Current Organisations
University of New South Wales
,
Broad Institute
,
Garvan Institute of Medical Research
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Publisher: Elsevier BV
Date: 12-2022
DOI: 10.1016/J.IMMUNI.2022.11.001
Abstract: The association between cancer and autoimmune disease is unexplained, exemplified by T cell large granular lymphocytic leukemia (T-LGL) where gain-of-function (GOF) somatic STAT3 mutations correlate with co-existing autoimmunity. To investigate whether these mutations are the cause or consequence of CD8
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/1078-0432.22551385.V1
Abstract: Supplementary Table S1. ADT + PI3Ki + PD-1 antibody leads to TAM activation within TME of PTEN 53-deficient prostate tumors.
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/1078-0432.22551388.V1
Abstract: Supplementary Figure S1. The majority of Pb-Cre PTENfl/fl Trp53fl/fl mice are de novo resistant to ADT. Supplementary Figure S2. ADT/PI3Ki combination therapy halts prostate tumor growth up to 14 days, followed by development of resistance in majority of Pb-Cre PTENfl/fl Trp53fl/fl mice. Supplementary Figure S3. PI3Ki treatment with concurrent androgen depletion does not alter proliferation and survival of PTEN 53-deficient murine PC cells in vitro. Supplementary Figure S4. ADT/PI3K inhibitor combination increases MHC-II and PD-1 expression on TAM within the TME of PTEN 53-deficient murine PC. Supplementary Figure S5. PD-1 upregulation suppresses phagocytic capacity of activated TAM. Supplementary Figure S6. Ex vivo AD + PI3Ki + PD-1 antibody treatment activates MHCIIlo TAM when co-cultured with PTEN 53-deficient murine prostate tumor cells. Supplementary Figure S7. The addition of PD-1 blockade to androgen depletion/PI3Ki therapy does not alter phagocytic capacity of PD-1 lo macrophages. Supplementary Figure S8. The combination of androgen depletion, PI3Ki and aPD-1 blockade does not alter phagocytic checkpoint expression on PTEN 53-deficient prostate tumor cells. Supplementary Figure S9. Androgen depletion, singly and in combination with aPD-1, did not alter phagocytosis activity of inactivated MHC-IIlo/PD-1 lo and MHC-IIlo/PD-1 hi TAM subsets. Supplementary Figure S10. Androgen depletion, not PI3Ki or aPD1, directly enhances TAM activation within the TME of PTEN 53-deficient PC. Supplementary Figure S11. PI3Ki does not alter phagocytosis/histone lactylation status of MHC-IIlo/PD-1 lo TAM and MHC-IIlo/PD-1 hi TAM. Supplementary Figure S12. PI3Ki inhibits lactate secretion from PTEN 53-deficient prostate tumor cells within TME. Supplementary Figure S13. Direct ex vivo treatment of TAM with PI3Ki, singly and in combination with PD-1 antibody and/or androgen depletion does not alter their histone lactylation profile. Supplementary Figure S14. ADT + PI3Ki + aPD-1 induces tumor control in 60% of Pb-Cre PTENfl/fl TP53fl/fl mice. Supplementary Figure S15. Depletion of activated TAM abrogates anti-cancer response elicited by ADT + PI3Ki + PD-1 antibody treatment in the PTEN 53-deficient murine prostate GEMM tumors. Supplementary Figure S16. Long-term treatment of ADT + PI3Ki + aPD-1 activates Wnt/βcatenin pathway in murine PTEN 53-deficient GEMM-derived SC1 cells. Supplementary Figure S17. Feedback Wnt/β-catenin-pathway activation within murine PTEN 53-deficient GEMM-derived PC cells following long-term ADT + copanlisib + aPD1 treatment suppresses phagocytosis via increased histone lactylation within bone marrow derived macrophages (BMDM).
Publisher: Springer Science and Business Media LLC
Date: 16-09-2023
Publisher: American Association for Cancer Research (AACR)
Date: 15-05-2023
DOI: 10.1158/1078-0432.22820227.V1
Abstract: Supplementary Table S1. ADT + PI3Ki + PD-1 antibody leads to TAM activation within TME of PTEN 53-deficient prostate tumors.
Publisher: EMBO
Date: 13-08-2020
Publisher: American Association for Cancer Research (AACR)
Date: 15-05-2023
DOI: 10.1158/1078-0432.22820230.V1
Abstract: Supplementary Figure S1. The majority of Pb-Cre PTENfl/fl Trp53fl/fl mice are de novo resistant to ADT.Supplementary Figure S2. ADT/PI3Ki combination therapy halts prostate tumor growth up to 14 days, followed by development of resistance in majority of Pb-Cre PTENfl/fl Trp53fl/fl mice.Supplementary Figure S3. PI3Ki treatment with concurrent androgen depletion does not alter proliferation and survival of PTEN 53-deficient murine PC cells in vitro.Supplementary Figure S4. ADT/PI3K inhibitor combination increases MHC-II and PD-1 expression on TAM within the TME of PTEN 53-deficient murine PC.Supplementary Figure S5. PD-1 upregulation suppresses phagocytic capacity of activated TAM.Supplementary Figure S6. Ex vivo AD + PI3Ki + PD-1 antibody treatment activates MHCIIlo TAM when co-cultured with PTEN 53-deficient murine prostate tumor cells.Supplementary Figure S7. The addition of PD-1 blockade to androgen depletion/PI3Ki therapy does not alter phagocytic capacity of PD-1 lo macrophages.Supplementary Figure S8. The combination of androgen depletion, PI3Ki and aPD-1 blockade does not alter phagocytic checkpoint expression on PTEN 53-deficient prostate tumor cells.Supplementary Figure S9. Androgen depletion, singly and in combination with aPD-1, did not alter phagocytosis activity of inactivated MHC-IIlo/PD-1 lo and MHC-IIlo/PD-1 hi TAM subsets.Supplementary Figure S10. Androgen depletion, not PI3Ki or aPD1, directly enhances TAM activation within the TME of PTEN 53-deficient PC.Supplementary Figure S11. PI3Ki does not alter phagocytosis/histone lactylation status of MHC-IIlo/PD-1 lo TAM and MHC-IIlo/PD-1 hi TAM.Supplementary Figure S12. PI3Ki inhibits lactate secretion from PTEN 53-deficient prostate tumor cells within TME.Supplementary Figure S13. Direct ex vivo treatment of TAM with PI3Ki, singly and in combination with PD-1 antibody and/or androgen depletion does not alter their histone lactylation profile.Supplementary Figure S14. ADT + PI3Ki + aPD-1 induces tumor control in 60% of Pb-Cre PTENfl/fl TP53fl/fl mice.Supplementary Figure S15. Depletion of activated TAM abrogates anti-cancer response elicited by ADT + PI3Ki + PD-1 antibody treatment in the PTEN 53-deficient murine prostate GEMM tumors.Supplementary Figure S16. Long-term treatment of ADT + PI3Ki + aPD-1 activates Wnt/βcatenin pathway in murine PTEN 53-deficient GEMM-derived SC1 cells.Supplementary Figure S17. Feedback Wnt/β-catenin-pathway activation within murine PTEN 53-deficient GEMM-derived PC cells following long-term ADT + copanlisib + aPD1 treatment suppresses phagocytosis via increased histone lactylation within bone marrow derived macrophages (BMDM).
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/1078-0432.C.6563227.V1
Abstract: AbstractPurpose: Phosphatase and tensin homolog (PTEN) loss of function occurs in approximately 50% of patients with metastatic castrate-resistant prostate cancer (mCRPC), and is associated with poor prognosis and responsiveness to standard-of-care therapies and immune checkpoint inhibitors. While PTEN loss of function hyperactivates PI3K signaling, combinatorial PI3K/AKT pathway and androgen deprivation therapy (ADT) has demonstrated limited anticancer efficacy in clinical trials. Here, we aimed to elucidate mechanism(s) of resistance to ADT/PI3K-AKT axis blockade, and to develop rational combinatorial strategies to effectively treat this molecular subset of mCRPC. Experimental Design: Prostate-specific PTEN 53-deficient genetically engineered mice (GEM) with established 150–200 mm sup /sup tumors, as assessed by ultrasound, were treated with either ADT (degarelix), PI3K inhibitor (copanlisib), or anti–PD-1 antibody (aPD-1), as single agents or their combinations, and tumors were monitored by MRI and harvested for immune, transcriptomic, and proteomic profiling, or i ex vivo /i co-culture studies. Single-cell RNA sequencing on human mCRPC s les was performed using 10X Genomics platform. Results: Coclinical trials in PTEN 53-deficient GEM revealed that recruitment of PD-1–expressing tumor-associated macrophages (TAM) thwarts ADT/PI3Ki combination–induced tumor control. The addition of aPD-1 to ADT/PI3Ki combination led to TAM-dependent approximately 3-fold increase in anticancer responses. Mechanistically, decreased lactate production from PI3Ki-treated tumor cells suppressed histone lactylation within TAM, resulting in their anticancer phagocytic activation, which was augmented by ADT/aPD-1 treatment and abrogated by feedback activation of Wnt/β-catenin pathway. Single-cell RNA-sequencing analysis in mCRPC patient biopsy s les revealed a direct correlation between high glycolytic activity and TAM phagocytosis suppression. Conclusions: Immunometabolic strategies that reverse lactate and PD-1–mediated TAM immunosuppression, in combination with ADT, warrant further investigation in patients with PTEN-deficient mCRPC. /
Publisher: American Association for Cancer Research (AACR)
Date: 15-05-2023
DOI: 10.1158/1078-0432.C.6563227.V2
Abstract: AbstractPurpose: Phosphatase and tensin homolog (PTEN) loss of function occurs in approximately 50% of patients with metastatic castrate-resistant prostate cancer (mCRPC), and is associated with poor prognosis and responsiveness to standard-of-care therapies and immune checkpoint inhibitors. While PTEN loss of function hyperactivates PI3K signaling, combinatorial PI3K/AKT pathway and androgen deprivation therapy (ADT) has demonstrated limited anticancer efficacy in clinical trials. Here, we aimed to elucidate mechanism(s) of resistance to ADT/PI3K-AKT axis blockade, and to develop rational combinatorial strategies to effectively treat this molecular subset of mCRPC. Experimental Design: Prostate-specific PTEN 53-deficient genetically engineered mice (GEM) with established 150–200 mm sup /sup tumors, as assessed by ultrasound, were treated with either ADT (degarelix), PI3K inhibitor (copanlisib), or anti–PD-1 antibody (aPD-1), as single agents or their combinations, and tumors were monitored by MRI and harvested for immune, transcriptomic, and proteomic profiling, or i ex vivo /i co-culture studies. Single-cell RNA sequencing on human mCRPC s les was performed using 10X Genomics platform. Results: Coclinical trials in PTEN 53-deficient GEM revealed that recruitment of PD-1–expressing tumor-associated macrophages (TAM) thwarts ADT/PI3Ki combination–induced tumor control. The addition of aPD-1 to ADT/PI3Ki combination led to TAM-dependent approximately 3-fold increase in anticancer responses. Mechanistically, decreased lactate production from PI3Ki-treated tumor cells suppressed histone lactylation within TAM, resulting in their anticancer phagocytic activation, which was augmented by ADT/aPD-1 treatment and abrogated by feedback activation of Wnt/β-catenin pathway. Single-cell RNA-sequencing analysis in mCRPC patient biopsy s les revealed a direct correlation between high glycolytic activity and TAM phagocytosis suppression. Conclusions: Immunometabolic strategies that reverse lactate and PD-1–mediated TAM immunosuppression, in combination with ADT, warrant further investigation in patients with PTEN-deficient mCRPC. /
Publisher: Elsevier BV
Date: 2022
DOI: 10.1016/J.GIM.2021.09.001
Abstract: Genetic variants causing aberrant premessenger RNA splicing are increasingly being recognized as causal variants in genetic disorders. In this study, we devise standardized practices for polymerase chain reaction (PCR)-based RNA diagnostics using clinically accessible specimens (blood, fibroblasts, urothelia, biopsy). A total of 74 families with erse monogenic conditions (31% prenatal-congenital onset, 47% early childhood, and 22% teenage-adult onset) were triaged into PCR-based RNA testing, with comparative RNA sequencing for 19 cases. Informative RNA assay data were obtained for 96% of cases, enabling variant reclassification for 75% variants that can be used for genetic counseling (71%), to inform clinical care (32%) and prenatal counseling (41%). Variant-associated mis-splicing was highly reproducible for 28 cases with s les from ≥2 affected in iduals or heterozygotes and 10 cases with ≥2 biospecimens. PCR licons encompassing another segregated heterozygous variant was vital for clinical interpretation of 22 of 79 variants to phase RNA splicing events and discern complete from partial mis-splicing. RNA diagnostics enabled provision of a genetic diagnosis for 64% of recruited cases. PCR-based RNA diagnostics has capacity to analyze 81.3% of clinically significant genes, with long licons providing an advantage over RNA sequencing to phase RNA splicing events. The Australasian Consortium for RNA Diagnostics (SpliceACORD) provide clinically-endorsed, standardized protocols and recommendations for interpreting RNA assay data.
Publisher: Cold Spring Harbor Laboratory
Date: 31-05-2022
DOI: 10.1101/2022.05.31.494081
Abstract: Immune cells are critical determinants of solid tumour aetiology, but the erse phenotypes of intra-tumoural immune cells remain incompletely characterised. We applied integrated single cell RNA sequencing (scRNA-Seq) and highly multiplexed protein epitope analysis to a cohort of breast cancer s les to resolve cell states within the tumour microenvironment. We reveal novel protein markers for resting and activated tumour infiltrating lymphocytes, and show that high expression of CD103 primarily marks exhausted CD8 rather than tissue resident CD8 T-cells in human breast cancers. We identify two distinct states of activated CD4+ T follicular helper (Tfh) cells. A population resembling conventional Tfh (cTfh) cells were localised primarily to lymphoid aggregates by spatial transcriptomics. In contrast, cancer associated Tfh (caTfh) cells expressing markers of tissue residency and exhaustion co-localized with cancer foci and signalled to macrophages. Importantly, increased caTfh : cTfh ratio associated with improved disease outcome and response to checkpoint immunotherapy.
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/1078-0432.22551388
Abstract: Supplementary Figure S1. The majority of Pb-Cre PTENfl/fl Trp53fl/fl mice are de novo resistant to ADT. Supplementary Figure S2. ADT/PI3Ki combination therapy halts prostate tumor growth up to 14 days, followed by development of resistance in majority of Pb-Cre PTENfl/fl Trp53fl/fl mice. Supplementary Figure S3. PI3Ki treatment with concurrent androgen depletion does not alter proliferation and survival of PTEN 53-deficient murine PC cells in vitro. Supplementary Figure S4. ADT/PI3K inhibitor combination increases MHC-II and PD-1 expression on TAM within the TME of PTEN 53-deficient murine PC. Supplementary Figure S5. PD-1 upregulation suppresses phagocytic capacity of activated TAM. Supplementary Figure S6. Ex vivo AD + PI3Ki + PD-1 antibody treatment activates MHCIIlo TAM when co-cultured with PTEN 53-deficient murine prostate tumor cells. Supplementary Figure S7. The addition of PD-1 blockade to androgen depletion/PI3Ki therapy does not alter phagocytic capacity of PD-1 lo macrophages. Supplementary Figure S8. The combination of androgen depletion, PI3Ki and aPD-1 blockade does not alter phagocytic checkpoint expression on PTEN 53-deficient prostate tumor cells. Supplementary Figure S9. Androgen depletion, singly and in combination with aPD-1, did not alter phagocytosis activity of inactivated MHC-IIlo/PD-1 lo and MHC-IIlo/PD-1 hi TAM subsets. Supplementary Figure S10. Androgen depletion, not PI3Ki or aPD1, directly enhances TAM activation within the TME of PTEN 53-deficient PC. Supplementary Figure S11. PI3Ki does not alter phagocytosis/histone lactylation status of MHC-IIlo/PD-1 lo TAM and MHC-IIlo/PD-1 hi TAM. Supplementary Figure S12. PI3Ki inhibits lactate secretion from PTEN 53-deficient prostate tumor cells within TME. Supplementary Figure S13. Direct ex vivo treatment of TAM with PI3Ki, singly and in combination with PD-1 antibody and/or androgen depletion does not alter their histone lactylation profile. Supplementary Figure S14. ADT + PI3Ki + aPD-1 induces tumor control in 60% of Pb-Cre PTENfl/fl TP53fl/fl mice. Supplementary Figure S15. Depletion of activated TAM abrogates anti-cancer response elicited by ADT + PI3Ki + PD-1 antibody treatment in the PTEN 53-deficient murine prostate GEMM tumors. Supplementary Figure S16. Long-term treatment of ADT + PI3Ki + aPD-1 activates Wnt/βcatenin pathway in murine PTEN 53-deficient GEMM-derived SC1 cells. Supplementary Figure S17. Feedback Wnt/β-catenin-pathway activation within murine PTEN 53-deficient GEMM-derived PC cells following long-term ADT + copanlisib + aPD1 treatment suppresses phagocytosis via increased histone lactylation within bone marrow derived macrophages (BMDM).
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/1078-0432.22551385
Abstract: Supplementary Table S1. ADT + PI3Ki + PD-1 antibody leads to TAM activation within TME of PTEN 53-deficient prostate tumors.
Publisher: Springer Science and Business Media LLC
Date: 07-11-2022
DOI: 10.1038/S41467-022-34041-X
Abstract: Cancers evade the immune system through the process of cancer immunoediting. While immune checkpoint inhibitors are effective for reactivating tumour immunity in some cancer types, many other solid cancers, including breast cancer, remain largely non-responsive. Understanding how non-responsive cancers evade immunity and whether this occurs at the clonal level will improve immunotherapeutic design. Here we use DNA barcoding to track murine mammary cancer cell clones during immunoediting and determine clonal transcriptional profiles that allow immune evasion following anti-PD1 plus anti-CTLA4 immunotherapy. Clonal ersity is significantly restricted by immunotherapy treatment in both primary tumours and metastases, demonstrating selection for pre-existing breast cancer cell populations and ongoing immunoediting during metastasis and treatment. Immunotherapy resistant clones express a common gene signature associated with poor survival of basal-like breast cancer patient cohorts. At least one of these genes has an existing small molecule that can potentially be used to improve immunotherapy response.
Publisher: Springer Science and Business Media LLC
Date: 17-02-2019
DOI: 10.1038/S41467-019-11049-4
Abstract: High-throughput single-cell RNA sequencing is a powerful technique but only generates short reads from one end of a cDNA template, limiting the reconstruction of highly erse sequences such as antigen receptors. To overcome this limitation, we combined targeted capture and long-read sequencing of T-cell-receptor (TCR) and B-cell-receptor (BCR) mRNA transcripts with short-read transcriptome profiling of barcoded single-cell libraries generated by droplet-based partitioning. We show that Repertoire and Gene Expression by Sequencing (RAGE-Seq) can generate accurate full-length antigen receptor sequences at nucleotide resolution, infer B-cell clonal evolution and identify alternatively spliced BCR transcripts. We apply RAGE-Seq to 7138 cells s led from the primary tumor and draining lymph node of a breast cancer patient to track transcriptome profiles of expanded lymphocyte clones across tissues. Our results demonstrate that RAGE-Seq is a powerful method for tracking the clonal evolution from large numbers of lymphocytes applicable to the study of immunity, autoimmunity and cancer.
Publisher: Springer Science and Business Media LLC
Date: 09-11-2022
Publisher: American Association for Cancer Research (AACR)
Date: 15-05-2023
DOI: 10.1158/1078-0432.22820227
Abstract: Supplementary Table S1. ADT + PI3Ki + PD-1 antibody leads to TAM activation within TME of PTEN 53-deficient prostate tumors.
Publisher: Springer Science and Business Media LLC
Date: 09-2021
Publisher: American Association for Cancer Research (AACR)
Date: 04-04-2023
DOI: 10.1158/1078-0432.CCR-22-3350
Abstract: Phosphatase and tensin homolog (PTEN) loss of function occurs in approximately 50% of patients with metastatic castrate-resistant prostate cancer (mCRPC), and is associated with poor prognosis and responsiveness to standard-of-care therapies and immune checkpoint inhibitors. While PTEN loss of function hyperactivates PI3K signaling, combinatorial PI3K/AKT pathway and androgen deprivation therapy (ADT) has demonstrated limited anticancer efficacy in clinical trials. Here, we aimed to elucidate mechanism(s) of resistance to ADT/PI3K-AKT axis blockade, and to develop rational combinatorial strategies to effectively treat this molecular subset of mCRPC. Prostate-specific PTEN 53-deficient genetically engineered mice (GEM) with established 150–200 mm3 tumors, as assessed by ultrasound, were treated with either ADT (degarelix), PI3K inhibitor (copanlisib), or anti–PD-1 antibody (aPD-1), as single agents or their combinations, and tumors were monitored by MRI and harvested for immune, transcriptomic, and proteomic profiling, or ex vivo co-culture studies. Single-cell RNA sequencing on human mCRPC s les was performed using 10X Genomics platform. Coclinical trials in PTEN 53-deficient GEM revealed that recruitment of PD-1–expressing tumor-associated macrophages (TAM) thwarts ADT/PI3Ki combination–induced tumor control. The addition of aPD-1 to ADT/PI3Ki combination led to TAM-dependent approximately 3-fold increase in anticancer responses. Mechanistically, decreased lactate production from PI3Ki-treated tumor cells suppressed histone lactylation within TAM, resulting in their anticancer phagocytic activation, which was augmented by ADT/aPD-1 treatment and abrogated by feedback activation of Wnt/β-catenin pathway. Single-cell RNA-sequencing analysis in mCRPC patient biopsy s les revealed a direct correlation between high glycolytic activity and TAM phagocytosis suppression. Immunometabolic strategies that reverse lactate and PD-1–mediated TAM immunosuppression, in combination with ADT, warrant further investigation in patients with PTEN-deficient mCRPC.
Publisher: American Association for Cancer Research (AACR)
Date: 15-05-2023
DOI: 10.1158/1078-0432.C.6563227
Abstract: AbstractPurpose: Phosphatase and tensin homolog (PTEN) loss of function occurs in approximately 50% of patients with metastatic castrate-resistant prostate cancer (mCRPC), and is associated with poor prognosis and responsiveness to standard-of-care therapies and immune checkpoint inhibitors. While PTEN loss of function hyperactivates PI3K signaling, combinatorial PI3K/AKT pathway and androgen deprivation therapy (ADT) has demonstrated limited anticancer efficacy in clinical trials. Here, we aimed to elucidate mechanism(s) of resistance to ADT/PI3K-AKT axis blockade, and to develop rational combinatorial strategies to effectively treat this molecular subset of mCRPC. Experimental Design: Prostate-specific PTEN 53-deficient genetically engineered mice (GEM) with established 150–200 mm sup /sup tumors, as assessed by ultrasound, were treated with either ADT (degarelix), PI3K inhibitor (copanlisib), or anti–PD-1 antibody (aPD-1), as single agents or their combinations, and tumors were monitored by MRI and harvested for immune, transcriptomic, and proteomic profiling, or i ex vivo /i co-culture studies. Single-cell RNA sequencing on human mCRPC s les was performed using 10X Genomics platform. Results: Coclinical trials in PTEN 53-deficient GEM revealed that recruitment of PD-1–expressing tumor-associated macrophages (TAM) thwarts ADT/PI3Ki combination–induced tumor control. The addition of aPD-1 to ADT/PI3Ki combination led to TAM-dependent approximately 3-fold increase in anticancer responses. Mechanistically, decreased lactate production from PI3Ki-treated tumor cells suppressed histone lactylation within TAM, resulting in their anticancer phagocytic activation, which was augmented by ADT/aPD-1 treatment and abrogated by feedback activation of Wnt/β-catenin pathway. Single-cell RNA-sequencing analysis in mCRPC patient biopsy s les revealed a direct correlation between high glycolytic activity and TAM phagocytosis suppression. Conclusions: Immunometabolic strategies that reverse lactate and PD-1–mediated TAM immunosuppression, in combination with ADT, warrant further investigation in patients with PTEN-deficient mCRPC. /
Publisher: Springer Science and Business Media LLC
Date: 14-10-2021
DOI: 10.1038/S41467-021-26271-2
Abstract: In the past decades, transcriptomic studies have revolutionized cancer treatment and diagnosis. However, tumor sequencing strategies typically result in loss of spatial information, critical to understand cell interactions and their functional relevance. To address this, we investigate spatial gene expression in HER2-positive breast tumors using Spatial Transcriptomics technology. We show that expression-based clustering enables data-driven tumor annotation and assessment of intra- and interpatient heterogeneity from which we discover shared gene signatures for immune and tumor processes. By integration with single cell data, we spatially map tumor-associated cell types to find tertiary lymphoid-like structures, and a type I interferon response overlapping with regions of T-cell and macrophage subset colocalization. We construct a predictive model to infer presence of tertiary lymphoid-like structures, applicable across tissue types and technical platforms. Taken together, we combine different data modalities to define a high resolution map of cellular interactions in tumors and provide tools generalizing across tissues and diseases.
Publisher: Cold Spring Harbor Laboratory
Date: 06-04-2023
DOI: 10.1101/2023.04.06.535805
Abstract: Spatial transcriptomic technologies are powerful tools for resolving the spatial heterogeneity of gene expression in tissue s les. However, little evidence exists on relative strengths and weaknesses of the various available technologies for profiling human tumour tissue. In this study, we aimed to provide an objective assessment of two common spatial transcriptomics platforms, 10X Genomics’ Visium and Nanostring’s GeoMx DSP. The abilities of the DSP and Visium platforms to profile transcriptomic features were compared using matching cell line and primary breast cancer tissue s les. A head-to-head comparison was conducted using data generated from matching s les and synthetic tissue references. Platform specific features were also assessed according to manufacturers’ recommendations to evaluate the optimal usage of the two technologies. We identified substantial variations in assay design between the DSP and Visium assays such as transcriptomic coverage and composition of the transcripts detected. When the data was standardised according to manufacturers’ recommendations, the DSP platform was more sensitive in gene expression detection. However, its specificity was diminished by the presence of non-specific detection. Our results also confirmed the strength and weakness of each platform in characterising spatial transcriptomic features of tissue s les, in particular their application to hypothesis generation versus hypothesis testing. In this study, we share our experience on both DSP and Visium technologies as end users. We hope this can guide future users to choose the most suitable platform for their research. In addition, this dataset can be used as an important resource for the development of new analysis tools.
Publisher: Springer Science and Business Media LLC
Date: 10-05-2021
DOI: 10.1186/S13073-021-00885-Z
Abstract: High throughput single-cell RNA sequencing (scRNA-Seq) has emerged as a powerful tool for exploring cellular heterogeneity among complex human cancers. scRNA-Seq studies using fresh human surgical tissue are logistically difficult, preclude histopathological triage of s les, and limit the ability to perform batch processing. This hindrance can often introduce technical biases when integrating patient datasets and increase experimental costs. Although tissue preservation methods have been previously explored to address such issues, it is yet to be examined on complex human tissues, such as solid cancers and on high throughput scRNA-Seq platforms. Using the Chromium 10X platform, we sequenced a total of ~ 120,000 cells from fresh and cryopreserved replicates across three primary breast cancers, two primary prostate cancers and a cutaneous melanoma. We performed detailed analyses between cells from each condition to assess the effects of cryopreservation on cellular heterogeneity, cell quality, clustering and the identification of gene ontologies. In addition, we performed single-cell immunophenotyping using CITE-Seq on a single breast cancer s le cryopreserved as solid tissue fragments. Tumour heterogeneity identified from fresh tissues was largely conserved in cryopreserved replicates. We show that sequencing of single cells prepared from cryopreserved tissue fragments or from cryopreserved cell suspensions is comparable to sequenced cells prepared from fresh tissue, with cryopreserved cell suspensions displaying higher correlations with fresh tissue in gene expression. We showed that cryopreservation had minimal impacts on the results of downstream analyses such as biological pathway enrichment. For some tumours, cryopreservation modestly increased cell stress signatures compared to freshly analysed tissue. Further, we demonstrate the advantage of cryopreserving whole-cells for detecting cell-surface proteins using CITE-Seq, which is impossible using other preservation methods such as single nuclei-sequencing. We show that the viable cryopreservation of human cancers provides high-quality single-cells for multi-omics analysis. Our study guides new experimental designs for tissue biobanking for future clinical single-cell RNA sequencing studies.
Publisher: American Association for Cancer Research (AACR)
Date: 15-05-2023
DOI: 10.1158/1078-0432.22820230
Abstract: Supplementary Figure S1. The majority of Pb-Cre PTENfl/fl Trp53fl/fl mice are de novo resistant to ADT.Supplementary Figure S2. ADT/PI3Ki combination therapy halts prostate tumor growth up to 14 days, followed by development of resistance in majority of Pb-Cre PTENfl/fl Trp53fl/fl mice.Supplementary Figure S3. PI3Ki treatment with concurrent androgen depletion does not alter proliferation and survival of PTEN 53-deficient murine PC cells in vitro.Supplementary Figure S4. ADT/PI3K inhibitor combination increases MHC-II and PD-1 expression on TAM within the TME of PTEN 53-deficient murine PC.Supplementary Figure S5. PD-1 upregulation suppresses phagocytic capacity of activated TAM.Supplementary Figure S6. Ex vivo AD + PI3Ki + PD-1 antibody treatment activates MHCIIlo TAM when co-cultured with PTEN 53-deficient murine prostate tumor cells.Supplementary Figure S7. The addition of PD-1 blockade to androgen depletion/PI3Ki therapy does not alter phagocytic capacity of PD-1 lo macrophages.Supplementary Figure S8. The combination of androgen depletion, PI3Ki and aPD-1 blockade does not alter phagocytic checkpoint expression on PTEN 53-deficient prostate tumor cells.Supplementary Figure S9. Androgen depletion, singly and in combination with aPD-1, did not alter phagocytosis activity of inactivated MHC-IIlo/PD-1 lo and MHC-IIlo/PD-1 hi TAM subsets.Supplementary Figure S10. Androgen depletion, not PI3Ki or aPD1, directly enhances TAM activation within the TME of PTEN 53-deficient PC.Supplementary Figure S11. PI3Ki does not alter phagocytosis/histone lactylation status of MHC-IIlo/PD-1 lo TAM and MHC-IIlo/PD-1 hi TAM.Supplementary Figure S12. PI3Ki inhibits lactate secretion from PTEN 53-deficient prostate tumor cells within TME.Supplementary Figure S13. Direct ex vivo treatment of TAM with PI3Ki, singly and in combination with PD-1 antibody and/or androgen depletion does not alter their histone lactylation profile.Supplementary Figure S14. ADT + PI3Ki + aPD-1 induces tumor control in 60% of Pb-Cre PTENfl/fl TP53fl/fl mice.Supplementary Figure S15. Depletion of activated TAM abrogates anti-cancer response elicited by ADT + PI3Ki + PD-1 antibody treatment in the PTEN 53-deficient murine prostate GEMM tumors.Supplementary Figure S16. Long-term treatment of ADT + PI3Ki + aPD-1 activates Wnt/βcatenin pathway in murine PTEN 53-deficient GEMM-derived SC1 cells.Supplementary Figure S17. Feedback Wnt/β-catenin-pathway activation within murine PTEN 53-deficient GEMM-derived PC cells following long-term ADT + copanlisib + aPD1 treatment suppresses phagocytosis via increased histone lactylation within bone marrow derived macrophages (BMDM).
No related grants have been discovered for Ghamdan Al-Eryani.