Macrophage colony-stimulating factor receptor (M-CSFR) is found in cells of the mononuclear phago... more Macrophage colony-stimulating factor receptor (M-CSFR) is found in cells of the mononuclear phagocyte lineage and is aberrantly expressed in a range of tumours, in addition to tumour-associated macrophages. Consequently, a variety of cancer therapies directed against M-CSFR are under development. We set out to engineer chimeric antigen receptors (CARs) that employ the natural ligands of this receptor, namely M-CSF or interleukin (IL)-34, to achieve specificity for M-CSFR-expressing target cells. Both M-CSF and IL-34 bind to overlapping regions of M-CSFR, although affinity of IL-34 is significantly greater than that of M-CSF. Matched second- and third-generation CARs targeted using M-CSF or IL-34 were expressed in human T-cells using the SFG retroviral vector. We found that both M-CSF- and IL-34-containing CARs enable T-cells to mediate selective destruction of tumour cells that express enforced or endogenous M-CSFR, accompanied by production of both IL-2 and interferon (IFN)-γ. Alth...
Co-stimulation is critical to the function of chimeric antigen receptor (CAR) T-cells. Previously... more Co-stimulation is critical to the function of chimeric antigen receptor (CAR) T-cells. Previously, we demonstrated that dual co-stimulation can be effectively harnessed by a parallel (p)CAR architecture in which a CD28-containing second generation CAR is co-expressed with a 4-1BB containing chimeric co-stimulatory receptor (CCR). When compared to linear CARs, pCAR-engineered T-cells elicit superior anti-tumor activity in a range of pre-clinical models. Since CD19 is the best validated clinical target for cellular immunotherapy, we evaluated a panel of CD19-specific CAR and pCAR T-cells in this study. First, we generated a panel of single chain antibody fragments (scFvs) by alanine scanning mutagenesis of the CD19-specific FMC63 scFv (VH domain) and these were incorporated into second generation CD28+CD3ζ CARs. The resulting panel of CAR T-cells demonstrated a broad range of CD19 binding ability and avidity for CD19-expressing tumor cells. Each scFv-modified CAR was then converted in...
C himeric antigen receptors are fusion molecules that couple the direct (antibody-like) binding o... more C himeric antigen receptors are fusion molecules that couple the direct (antibody-like) binding of a cell surface target to the delivery of a tailored T-cell activating signal. These molecules are delivered to peripheral blood T or NK cells, using retroviral, lentiviral or a number of nonviral vector systems. Three generations of CARs have been described in which signalling is provided by CD3ζ alone (first generation), with either CD28 or 4-1BB co-stimulation (second generation) or with a combination of both of these co-stimulatory signals (third generation). Immunotherapy using autologous CAR T-cells has made a profound impact on the management of refractory B-cell malignancy. In that setting, second generation CAR T-cells are re-targeted against the ubiquitous B-cell antigen, CD19, enabling them to eliminate both malignant and healthy B-cells. Complete response rates approaching 90% have been achieved in patients with refractory acute lymphoblastic leukaemia. Emphasizing robustness, these data have been achieved in a number of distinct centres, using CARs that vary in their antibody targeting moiety or co-stimulatory domain (either CD28 or 4-1BB). However, successful implementation of CAR T-cell immunotherapy for solid tumours remains an elusive goal. Mindful of the role played by ErbB dysre-gulation in many solid tumours, we set out to target this family using a CAR-based approach. The ErbB family of receptor tyrosine kinases comprises four members (ErbB1-4) which form a series of 9 possible homo-or heterodimeric pairs. Several highly successful anti-cancer pharmaceuticals have been developed to target one or more members of the family, including herceptin, cetuximab and erlotinib. To achieve broad targeting specificity across the network, we used a chimeric peptide named T1E. T1E is derived from transforming growth factor-α (TGF-α) and epidermal growth factor (EGF). While TGF-α and EGF are both unique ErbB1 ligands, the T1E chimera binds to all ErbB1 and ErbB4 homo-and heterodimers, in addition to the ErbB2/3 heterodimer. A second generation CAR named T1E28z was engineered by fusing T1E to a chimeric endodomain comprising CD28 and CD3ζ. Pre-clinical evaluation of T1E28z-engineered human T-cells demonstrated antitumour activity against diverse solid tumour cell lines, provided that one or more of the indicated ErbB dimer species was present. Furthermore, it was anticipated that evasion of T1E28z + CAR T-cells by tumour cells through antigen loss was unlikely since the CAR recognized multiple targets, coupled with the fact that ErbB dysregulation is an intrinsic driver of transformation. However, the major risk anticipated with clinical development of this CAR was potential for on-target toxicity. Underlining this, a fatal SUSAR (suspected unexpected severe adverse reaction) was described recently following the intravenous administration of 10 billion ErbB2 re-targeted CAR T-cells to a patient with metastatic ErbB2+ colorectal cancer. To mitigate risk, we identified a clinical niche whereby localized rather than systemic delivery of CAR T-cells could be justified. Refractory head and neck squamous cell cancer (HNSCC) provides such an indication since locally advanced or recurrent tumour formation represents
Summary Immunotherapy of cancer using chimeric antigen receptor-engineered T-cells has transforme... more Summary Immunotherapy of cancer using chimeric antigen receptor-engineered T-cells has transformed the management of selected haematological malignancies, triggering intense clinical trial activity in this arena. This article summarises trial activity that has been published to date across the spectrum of haematological and solid tumour types.
CAR-engineered T cell immunotherapy has proven transformative in selected hematological malignanc... more CAR-engineered T cell immunotherapy has proven transformative in selected hematological malignancies. However, solid tumors largely remain impervious to these approaches. In addressing this challenge, Srivastava et al. in this issue demonstrate that oxaliplatin-based lymphodepleting chemotherapy promotes enhanced CAR T cell recruitment to lung tumors, boosting therapeutic impact in combination with anti-PD-L1.
If citing, it is advised that you check and use the publisher's definitive version for pagination... more If citing, it is advised that you check and use the publisher's definitive version for pagination, volume/issue, and date of publication details. And where the final published version is provided on the Research Portal, if citing you are again advised to check the publisher's website for any subsequent corrections.
Immunotherapy using CAR-T-cells is acquiring an expanding role in the treatment of malignant dise... more Immunotherapy using CAR-T-cells is acquiring an expanding role in the treatment of malignant disease. However, efficacy of solid tumour CAR-T-immunotherapy has proven inconsistent. Limitations include poor T-cell trafficking to tumour deposits, insufficient T-cell longevity in-vivo and the occurrence of both predicted and unanticipated on-target and off-target toxicity. To refine this therapeutic approach, it is desirable to develop systems that allow the monitoring of T-cell location and persistence in-vivo. A retroviral vector named PiN-4 was constructed, which co-expresses: (i) an interleukin (IL)-4-responsive chimeric cytokine receptor (4αβ); (ii) a prostate-specific membrane antigen (PSMA)-targeted CAR (P28z) and (iii) hNIS, which promotes the uptake of technetium-99m pertechnetate (99mTcO4−) in viable cells. IL-4 enriched human 4P28zN+ T-cells were administered intravenously to Nod Scid Gamma (NSG) mice bearing prostate tumour xenografts. Measurement of the tumour xenografts by bioluminescent imaging (BLI) found that only PSMA-expressing tumours responded to 4P28zN+ T-cell treatment. Longitudinal SPECT-CT imaging further confirmed this with the preferential accumulation of 4P28zN+ T-cells in PSMA-expressing tumours compared to PSMA-negative tumours. Use of NIS as a T-cell imaging reporter brings several potential advantages. It promotes receptor-mediated uptake of the inexpensive, low toxicity and clinically useful SPECT tracer, technetium-99m pertechnetate (99mTcO4−). In addition, the ectopic expression of NIS is well tolerated by host cells and hNIS functions only in viable cells. These data demonstrate proof of concept for the utility of hNIS as an imaging reporter in genetically engineered T-cells. Citation Format: Nia Emami-Shahri, Julie Foster, Jane Sosabowski, John Maher, Sophie Papa. Dynamic SPECT imaging of PSMA-specific CAR T cells in mice bearing prostate cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2315.
Immunotherapy of cancer using chimeric antigen receptor (CAR) T-cells is a rapidly expanding fiel... more Immunotherapy of cancer using chimeric antigen receptor (CAR) T-cells is a rapidly expanding field. CARs are fusion molecules that couple the binding of a tumour-associated cell surface target to the delivery of a tailored T-cell activating signal. Re-infusion of such genetically engineered T-cells to patients with haematological disease has demonstrated unprecedented response rates in Phase I clinical trials. However, such successes have not yet been observed using CAR T-cells against solid malignancies and this is, in part, due to a lack of safe tumour-specific targets. The αvβ6 integrin is strongly up-regulated in multiple solid tumours including those derived from colon, lung, breast, cervix, ovaries/fallopian tube, pancreas and head and neck. It is associated with poorer prognosis in several cancers and exerts pro-tumorigenic activities including promotion of tumour growth, migration and invasion. By contrast, physiologic expression of αvβ6 is largely restricted to wound healin...
Chimeric antigen receptor (CAR) based immunotherapy has been under development for the last 25 ye... more Chimeric antigen receptor (CAR) based immunotherapy has been under development for the last 25 years and is now a promising new treatment modality in the field of cancer immunotherapy. The approach involves genetically engineering T cells to target malignant cells through expression of a bespoke fusion receptor that couples an HLA-independent antigen recognition domain to one or more intracellular T-cell activating modules. Multiple clinical trials are now underway in several centers to investigate CAR T-cell immunotherapy of diverse hematologic and solid tumor types. The most successful results have been achieved in the treatment of patients with B-cell malignancies, in whom several complete and durable responses have been achieved. This review focuses on the preclinical and clinical development of CAR T-cell immunotherapy of solid cancers, targeted against members of the ErbB family.
Chimeric antigen receptor- (CAR-) based immunotherapy has been under development for almost 25 ye... more Chimeric antigen receptor- (CAR-) based immunotherapy has been under development for almost 25 years, over which period it has progressed from a new but cumbersome technology to an emerging therapeutic modality for malignant disease. The approach involves the genetic engineering of fusion receptors (CARs) that couple the HLA-independent binding of cell surface target molecules to the delivery of a tailored activating signal to host immune cells. Engineered CARs are delivered most commonly to peripheral blood T cells using a range of vector systems, most commonly integrating viral vectors. Preclinical refinement of this approach has proceeded over several years to the point that clinical testing is now being undertaken at several centres, using increasingly sophisticated and therapeutically successful genetic payloads. This paper considers several aspects of the pre-clinical and clinical development of CAR-based immunotherapy and how this technology is acquiring an increasing niche i...
Sensitive and specific detection of nodal status, sites of metastases and low-volume recurrent di... more Sensitive and specific detection of nodal status, sites of metastases and low-volume recurrent disease could greatly improve management of patients with advanced prostate cancer. Prostate-specific membrane antigen (PSMA) is a well-established marker for prostate carcinoma with increased levels of expression in high-grade, hormone-refractory and metastatic disease. The monoclonal antibody (mAb) J591 is directed against an extracellular epitope of PSMA and has been shown to efficiently target disseminated disease including metastases in lymph nodes and bone. Its use as a diagnostic imaging agent however is limited due to its slow pharmacokinetics. In this study a diabody derived from mAb J591 was developed as a single photon emission computed tomography (SPECT) tracer with improved pharmacokinetics for the detection of PSMA expression in prostate cancer. A diabody in VH-VL orientation and with a C-terminal cysteine was expressed in HEK293T cells and purified by a combination of metal ...
Chinese hamster ovary (CHO) cells are widely used for the production of recombinant proteins for ... more Chinese hamster ovary (CHO) cells are widely used for the production of recombinant proteins for clinical use as well as academic research. They are particularly important for the production of glycoproteins where bacteria cannot be used. TGFβ1 is a potent cytokine highly conserved across species with multiple immunological and non-immunological effects. We have discovered that CHOK1, the CHO clone most commonly used by the pharmaceutical industry, constitutively secretes latent TGFβ1 and that this hamster TGFβ1 is active on human cells inducing profound immunological effects. As far as we are aware, the production of TGFβ1 by CHOK1 cells has not been reported before in the literature. As TGFβ1 exerts powerful and pleiotropic effects on diverse cell types, and as CHO cells are used to produce a large number of clinical and non-clinical products, our findings are highly relevant to studies that rely on recombinant proteins.
www.jbmethods.org 1 PROTOCOLJournal of Biological Methods | 2014 | Vol. 1(2) | e7 DOI: 10.14440/j... more www.jbmethods.org 1 PROTOCOLJournal of Biological Methods | 2014 | Vol. 1(2) | e7 DOI: 10.14440/jbm.2014.30 POL Scientific Use of retroviral-mediated gene transfer to deliver and test function of chimeric antigen receptors in human
Prostate cancer is the most common cancer in men, both in the USA and Europe. Although incurable,... more Prostate cancer is the most common cancer in men, both in the USA and Europe. Although incurable, metastatic disease can often be controlled for years with anti-androgen therapy. Once the disease becomes castrate resistant, the median survival is 18 months. There is growing evidence that the immune system, and in particular cytokines, play an important role in prostate cancer immunosurveillance and progression. Here, we have undertaken a clinical investigation of the role of two closely related cytokines, IL-4 and IL-13 in prostate cancer. In the largest series studied to date, we show that serum IL-4, but not IL-13 is significantly elevated in castrate resistant, compared to androgen sensitive disease. Notably however, serum IL-4 levels are also raised in patients with benign prostatic OPEN ACCESS Cancers 2011, 3 4282 disease. Analysis of benign and malignant prostate tissue demonstrates that the source of IL-4 is epithelial cells rather than infiltrating leukocytes. Together, our data are consistent with a dual role for IL-4 in prostate cancer development. In benign disease, our data add to the evidence that IL-4 serves a protective role. By contrast, the data support a direct role for IL-4 in the progression of prostate cancer from androgen responsive, to advanced castrate-resistant disease.
Macrophage colony-stimulating factor receptor (M-CSFR) is found in cells of the mononuclear phago... more Macrophage colony-stimulating factor receptor (M-CSFR) is found in cells of the mononuclear phagocyte lineage and is aberrantly expressed in a range of tumours, in addition to tumour-associated macrophages. Consequently, a variety of cancer therapies directed against M-CSFR are under development. We set out to engineer chimeric antigen receptors (CARs) that employ the natural ligands of this receptor, namely M-CSF or interleukin (IL)-34, to achieve specificity for M-CSFR-expressing target cells. Both M-CSF and IL-34 bind to overlapping regions of M-CSFR, although affinity of IL-34 is significantly greater than that of M-CSF. Matched second- and third-generation CARs targeted using M-CSF or IL-34 were expressed in human T-cells using the SFG retroviral vector. We found that both M-CSF- and IL-34-containing CARs enable T-cells to mediate selective destruction of tumour cells that express enforced or endogenous M-CSFR, accompanied by production of both IL-2 and interferon (IFN)-γ. Alth...
Co-stimulation is critical to the function of chimeric antigen receptor (CAR) T-cells. Previously... more Co-stimulation is critical to the function of chimeric antigen receptor (CAR) T-cells. Previously, we demonstrated that dual co-stimulation can be effectively harnessed by a parallel (p)CAR architecture in which a CD28-containing second generation CAR is co-expressed with a 4-1BB containing chimeric co-stimulatory receptor (CCR). When compared to linear CARs, pCAR-engineered T-cells elicit superior anti-tumor activity in a range of pre-clinical models. Since CD19 is the best validated clinical target for cellular immunotherapy, we evaluated a panel of CD19-specific CAR and pCAR T-cells in this study. First, we generated a panel of single chain antibody fragments (scFvs) by alanine scanning mutagenesis of the CD19-specific FMC63 scFv (VH domain) and these were incorporated into second generation CD28+CD3ζ CARs. The resulting panel of CAR T-cells demonstrated a broad range of CD19 binding ability and avidity for CD19-expressing tumor cells. Each scFv-modified CAR was then converted in...
C himeric antigen receptors are fusion molecules that couple the direct (antibody-like) binding o... more C himeric antigen receptors are fusion molecules that couple the direct (antibody-like) binding of a cell surface target to the delivery of a tailored T-cell activating signal. These molecules are delivered to peripheral blood T or NK cells, using retroviral, lentiviral or a number of nonviral vector systems. Three generations of CARs have been described in which signalling is provided by CD3ζ alone (first generation), with either CD28 or 4-1BB co-stimulation (second generation) or with a combination of both of these co-stimulatory signals (third generation). Immunotherapy using autologous CAR T-cells has made a profound impact on the management of refractory B-cell malignancy. In that setting, second generation CAR T-cells are re-targeted against the ubiquitous B-cell antigen, CD19, enabling them to eliminate both malignant and healthy B-cells. Complete response rates approaching 90% have been achieved in patients with refractory acute lymphoblastic leukaemia. Emphasizing robustness, these data have been achieved in a number of distinct centres, using CARs that vary in their antibody targeting moiety or co-stimulatory domain (either CD28 or 4-1BB). However, successful implementation of CAR T-cell immunotherapy for solid tumours remains an elusive goal. Mindful of the role played by ErbB dysre-gulation in many solid tumours, we set out to target this family using a CAR-based approach. The ErbB family of receptor tyrosine kinases comprises four members (ErbB1-4) which form a series of 9 possible homo-or heterodimeric pairs. Several highly successful anti-cancer pharmaceuticals have been developed to target one or more members of the family, including herceptin, cetuximab and erlotinib. To achieve broad targeting specificity across the network, we used a chimeric peptide named T1E. T1E is derived from transforming growth factor-α (TGF-α) and epidermal growth factor (EGF). While TGF-α and EGF are both unique ErbB1 ligands, the T1E chimera binds to all ErbB1 and ErbB4 homo-and heterodimers, in addition to the ErbB2/3 heterodimer. A second generation CAR named T1E28z was engineered by fusing T1E to a chimeric endodomain comprising CD28 and CD3ζ. Pre-clinical evaluation of T1E28z-engineered human T-cells demonstrated antitumour activity against diverse solid tumour cell lines, provided that one or more of the indicated ErbB dimer species was present. Furthermore, it was anticipated that evasion of T1E28z + CAR T-cells by tumour cells through antigen loss was unlikely since the CAR recognized multiple targets, coupled with the fact that ErbB dysregulation is an intrinsic driver of transformation. However, the major risk anticipated with clinical development of this CAR was potential for on-target toxicity. Underlining this, a fatal SUSAR (suspected unexpected severe adverse reaction) was described recently following the intravenous administration of 10 billion ErbB2 re-targeted CAR T-cells to a patient with metastatic ErbB2+ colorectal cancer. To mitigate risk, we identified a clinical niche whereby localized rather than systemic delivery of CAR T-cells could be justified. Refractory head and neck squamous cell cancer (HNSCC) provides such an indication since locally advanced or recurrent tumour formation represents
Summary Immunotherapy of cancer using chimeric antigen receptor-engineered T-cells has transforme... more Summary Immunotherapy of cancer using chimeric antigen receptor-engineered T-cells has transformed the management of selected haematological malignancies, triggering intense clinical trial activity in this arena. This article summarises trial activity that has been published to date across the spectrum of haematological and solid tumour types.
CAR-engineered T cell immunotherapy has proven transformative in selected hematological malignanc... more CAR-engineered T cell immunotherapy has proven transformative in selected hematological malignancies. However, solid tumors largely remain impervious to these approaches. In addressing this challenge, Srivastava et al. in this issue demonstrate that oxaliplatin-based lymphodepleting chemotherapy promotes enhanced CAR T cell recruitment to lung tumors, boosting therapeutic impact in combination with anti-PD-L1.
If citing, it is advised that you check and use the publisher's definitive version for pagination... more If citing, it is advised that you check and use the publisher's definitive version for pagination, volume/issue, and date of publication details. And where the final published version is provided on the Research Portal, if citing you are again advised to check the publisher's website for any subsequent corrections.
Immunotherapy using CAR-T-cells is acquiring an expanding role in the treatment of malignant dise... more Immunotherapy using CAR-T-cells is acquiring an expanding role in the treatment of malignant disease. However, efficacy of solid tumour CAR-T-immunotherapy has proven inconsistent. Limitations include poor T-cell trafficking to tumour deposits, insufficient T-cell longevity in-vivo and the occurrence of both predicted and unanticipated on-target and off-target toxicity. To refine this therapeutic approach, it is desirable to develop systems that allow the monitoring of T-cell location and persistence in-vivo. A retroviral vector named PiN-4 was constructed, which co-expresses: (i) an interleukin (IL)-4-responsive chimeric cytokine receptor (4αβ); (ii) a prostate-specific membrane antigen (PSMA)-targeted CAR (P28z) and (iii) hNIS, which promotes the uptake of technetium-99m pertechnetate (99mTcO4−) in viable cells. IL-4 enriched human 4P28zN+ T-cells were administered intravenously to Nod Scid Gamma (NSG) mice bearing prostate tumour xenografts. Measurement of the tumour xenografts by bioluminescent imaging (BLI) found that only PSMA-expressing tumours responded to 4P28zN+ T-cell treatment. Longitudinal SPECT-CT imaging further confirmed this with the preferential accumulation of 4P28zN+ T-cells in PSMA-expressing tumours compared to PSMA-negative tumours. Use of NIS as a T-cell imaging reporter brings several potential advantages. It promotes receptor-mediated uptake of the inexpensive, low toxicity and clinically useful SPECT tracer, technetium-99m pertechnetate (99mTcO4−). In addition, the ectopic expression of NIS is well tolerated by host cells and hNIS functions only in viable cells. These data demonstrate proof of concept for the utility of hNIS as an imaging reporter in genetically engineered T-cells. Citation Format: Nia Emami-Shahri, Julie Foster, Jane Sosabowski, John Maher, Sophie Papa. Dynamic SPECT imaging of PSMA-specific CAR T cells in mice bearing prostate cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2315.
Immunotherapy of cancer using chimeric antigen receptor (CAR) T-cells is a rapidly expanding fiel... more Immunotherapy of cancer using chimeric antigen receptor (CAR) T-cells is a rapidly expanding field. CARs are fusion molecules that couple the binding of a tumour-associated cell surface target to the delivery of a tailored T-cell activating signal. Re-infusion of such genetically engineered T-cells to patients with haematological disease has demonstrated unprecedented response rates in Phase I clinical trials. However, such successes have not yet been observed using CAR T-cells against solid malignancies and this is, in part, due to a lack of safe tumour-specific targets. The αvβ6 integrin is strongly up-regulated in multiple solid tumours including those derived from colon, lung, breast, cervix, ovaries/fallopian tube, pancreas and head and neck. It is associated with poorer prognosis in several cancers and exerts pro-tumorigenic activities including promotion of tumour growth, migration and invasion. By contrast, physiologic expression of αvβ6 is largely restricted to wound healin...
Chimeric antigen receptor (CAR) based immunotherapy has been under development for the last 25 ye... more Chimeric antigen receptor (CAR) based immunotherapy has been under development for the last 25 years and is now a promising new treatment modality in the field of cancer immunotherapy. The approach involves genetically engineering T cells to target malignant cells through expression of a bespoke fusion receptor that couples an HLA-independent antigen recognition domain to one or more intracellular T-cell activating modules. Multiple clinical trials are now underway in several centers to investigate CAR T-cell immunotherapy of diverse hematologic and solid tumor types. The most successful results have been achieved in the treatment of patients with B-cell malignancies, in whom several complete and durable responses have been achieved. This review focuses on the preclinical and clinical development of CAR T-cell immunotherapy of solid cancers, targeted against members of the ErbB family.
Chimeric antigen receptor- (CAR-) based immunotherapy has been under development for almost 25 ye... more Chimeric antigen receptor- (CAR-) based immunotherapy has been under development for almost 25 years, over which period it has progressed from a new but cumbersome technology to an emerging therapeutic modality for malignant disease. The approach involves the genetic engineering of fusion receptors (CARs) that couple the HLA-independent binding of cell surface target molecules to the delivery of a tailored activating signal to host immune cells. Engineered CARs are delivered most commonly to peripheral blood T cells using a range of vector systems, most commonly integrating viral vectors. Preclinical refinement of this approach has proceeded over several years to the point that clinical testing is now being undertaken at several centres, using increasingly sophisticated and therapeutically successful genetic payloads. This paper considers several aspects of the pre-clinical and clinical development of CAR-based immunotherapy and how this technology is acquiring an increasing niche i...
Sensitive and specific detection of nodal status, sites of metastases and low-volume recurrent di... more Sensitive and specific detection of nodal status, sites of metastases and low-volume recurrent disease could greatly improve management of patients with advanced prostate cancer. Prostate-specific membrane antigen (PSMA) is a well-established marker for prostate carcinoma with increased levels of expression in high-grade, hormone-refractory and metastatic disease. The monoclonal antibody (mAb) J591 is directed against an extracellular epitope of PSMA and has been shown to efficiently target disseminated disease including metastases in lymph nodes and bone. Its use as a diagnostic imaging agent however is limited due to its slow pharmacokinetics. In this study a diabody derived from mAb J591 was developed as a single photon emission computed tomography (SPECT) tracer with improved pharmacokinetics for the detection of PSMA expression in prostate cancer. A diabody in VH-VL orientation and with a C-terminal cysteine was expressed in HEK293T cells and purified by a combination of metal ...
Chinese hamster ovary (CHO) cells are widely used for the production of recombinant proteins for ... more Chinese hamster ovary (CHO) cells are widely used for the production of recombinant proteins for clinical use as well as academic research. They are particularly important for the production of glycoproteins where bacteria cannot be used. TGFβ1 is a potent cytokine highly conserved across species with multiple immunological and non-immunological effects. We have discovered that CHOK1, the CHO clone most commonly used by the pharmaceutical industry, constitutively secretes latent TGFβ1 and that this hamster TGFβ1 is active on human cells inducing profound immunological effects. As far as we are aware, the production of TGFβ1 by CHOK1 cells has not been reported before in the literature. As TGFβ1 exerts powerful and pleiotropic effects on diverse cell types, and as CHO cells are used to produce a large number of clinical and non-clinical products, our findings are highly relevant to studies that rely on recombinant proteins.
www.jbmethods.org 1 PROTOCOLJournal of Biological Methods | 2014 | Vol. 1(2) | e7 DOI: 10.14440/j... more www.jbmethods.org 1 PROTOCOLJournal of Biological Methods | 2014 | Vol. 1(2) | e7 DOI: 10.14440/jbm.2014.30 POL Scientific Use of retroviral-mediated gene transfer to deliver and test function of chimeric antigen receptors in human
Prostate cancer is the most common cancer in men, both in the USA and Europe. Although incurable,... more Prostate cancer is the most common cancer in men, both in the USA and Europe. Although incurable, metastatic disease can often be controlled for years with anti-androgen therapy. Once the disease becomes castrate resistant, the median survival is 18 months. There is growing evidence that the immune system, and in particular cytokines, play an important role in prostate cancer immunosurveillance and progression. Here, we have undertaken a clinical investigation of the role of two closely related cytokines, IL-4 and IL-13 in prostate cancer. In the largest series studied to date, we show that serum IL-4, but not IL-13 is significantly elevated in castrate resistant, compared to androgen sensitive disease. Notably however, serum IL-4 levels are also raised in patients with benign prostatic OPEN ACCESS Cancers 2011, 3 4282 disease. Analysis of benign and malignant prostate tissue demonstrates that the source of IL-4 is epithelial cells rather than infiltrating leukocytes. Together, our data are consistent with a dual role for IL-4 in prostate cancer development. In benign disease, our data add to the evidence that IL-4 serves a protective role. By contrast, the data support a direct role for IL-4 in the progression of prostate cancer from androgen responsive, to advanced castrate-resistant disease.
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Papers by John Maher