A Novel C-C Chemoattractant Cytokine (Chemokine) Receptor 6 (CCR6) Antagonist (PF-07054894) Distinguishes between Homologous Chemokine Receptors, Increases Basal Circulating CCR6+ T Cells, and Ameliorates Interleukin-23-Induced Skin Inflammation.

Blocking chemokine receptor C-C chemoattractant cytokine (chemokine) receptor (CCR) 6-dependent T cell migration has therapeutic promise in inflammatory diseases. PF-07054894 is a novel CCR6 antagonist that blocked only CCR6, CCR7, and C-X-C chemoattractant cytokine (chemokine) receptor (CXCR) 2 in a ß-arrestin assay panel of 168 G protein-coupled receptors. Inhibition of CCR6-mediated human T cell chemotaxis by (R)-4-((2-(((1,4-Dimethyl-1H-pyrazol-3-yl)(1-methylcyclopentyl)methyl)amino)-3,4-dioxocyclobut-1-en-1-yl)amino)-3-hydroxy-N,N-dimethylpicolinamide (PF-07054894) was insurmountable by CCR6 ligand, C-C motif ligand (CCL) 20. In contrast, blockade of CCR7-dependent chemotaxis in human T cells and CXCR2-dependent chemotaxis in human neutrophils by PF-07054894 were surmountable by CCL19 and C-X-C motif ligand 1, respectively. [3H]-PF-07054894 showed a slower dissociation rate for CCR6 than for CCR7 and CXCR2 suggesting that differences in chemotaxis patterns of inhibition could be attributable to offset kinetics. Consistent with this notion, an analog of PF-07054894 with fast dissociation rate showed surmountable inhibition of CCL20/CCR6 chemotaxis. Furthermore, pre-equilibration of T cells with PF-07054894 increased its inhibitory potency in CCL20/CCR6 chemotaxis by 10-fold. The functional selectivity of PF-07054894 for inhibition of CCR6 relative to CCR7 and CXCR2 is estimated to be at least 50- and 150-fold, respectively. When administered orally to naïve cynomolgus monkeys, PF-07054894 increased the frequency of CCR6+ peripheral blood T cells, suggesting that blockade of CCR6 inhibited homeostatic migration of T cells from blood to tissues. PF-07054894 inhibited interleukin-23-induced mouse skin ear swelling to a similar extent as genetic ablation of CCR6. PF-07054894 caused an increase in cell surface CCR6 in mouse and monkey B cells, which was recapitulated in mouse splenocytes in vitro. In conclusion, PF-07054894 is a potent and functionally selective CCR6 antagonist that blocks CCR6-mediated chemotaxis in vitro and in vivo. SIGNIFICANCE STATEMENT: The chemokine receptor, C-C chemoattractant cytokine (chemokine) receptor 6 (CCR6) plays a key role in the migration of pathogenic lymphocytes and dendritic cells into sites of inflammation. (R)-4-((2-(((1,4-Dimethyl-1H-pyrazol-3-yl)(1-methylcyclopentyl)methyl)amino)-3,4-dioxocyclobut-1-en-1-yl)amino)-3-hydroxy-N,N-dimethylpicolinamide (PF-07054894) is a novel CCR6 small molecule antagonist that illustrates the importance of binding kinetics in achieving pharmacological potency and selectivity. Orally administered PF-07054894 blocks homeostatic and pathogenic functions of CCR6, suggesting that it is a promising therapeutic agent for the treatment of a variety of autoimmune and inflammatory diseases.

Detecting expression of 5T4 in CTCs and tumor samples from NSCLC patients.

The fetal oncogene 5T4 is a cell surface protein, with overexpression observed in a variety of cancers as compared to normal adult tissue. The ability to select patients with tumors that express high levels of 5T4 may enrich a clinical trial cohort with patients most likely to respond to 5T4 targeted therapy. To that end, we developed assays to measure 5T4 in both tumors and in circulating tumor cells (CTCs). We identified the presence of 5T4 in both adenocarcinoma and squamous cell carcinoma of lung, in all clinical stages and grades of disease. CTCs were identified in peripheral blood from the majority of patients with NSCLC, and 5T4 was detectable in most samples. Although 5T4 was present in both CTCs and tumors in most patients, there was no concordance between relative amount in either sample type. Clinical response rates of patients treated with the therapies directed against 5T4 in early stage clinical trials, as determined by these assays, may provide important insights into the biology of 5T4 in tumors and the mechanisms of action of 5T4-targeting therapy.

Recovery of Native Renal Function in Patients with Hepatorenal Syndrome Following Combined Liver and Kidney Transplant with Mercaptoacetyltriglycine-3 Renogram: Developing a Methodology.

Many patients with hepatorenal syndrome (HRS) end up receiving a combined liver and kidney transplant (CKLT) with preservation of native kidneys, specially type 1 HRS since is characterizes by a very rapid deterioration of renal function. Eventually, most of the patients regain renal function, but it is unknown if this is due to the transplanted kidney, the recovery of native renal function, or both. The aim of this study is to evaluate if there is recovery of native renal function in patients with HRS following CKLT. 22 patients (16 men; 6 women) with history of HRS and status post CKLT were studied. Mercapto-acetyltriglycine-3 renograms in the anterior and posterior views with the three kidneys in the field of view were simultaneously acquired. The renograms were analyzed by creating regions of interest around the transplanted and native kidneys. Relative contribution to the renal function, clearance, and effective renal plasma flow for the transplanted and native kidneys were obtained. 1/22 (4.5%) patients presented with a very poor functioning transplanted kidney, in 15/22 (68%) cases the combined native renal function was markedly poorer than the transplanted renal function and in 6/22 (27%) native kidneys showed a contribution to the renal function similar to the transplanted kidney. In conclusion, our series show that around 32% of the HRS patients recovered their native renal function after CKLT. Identification of common factors that affect recovery of native renal function may help to avoid unnecessary renal transplants, significantly reducing morbidity and cost, while facilitating a reallocation of scarce donor resources.

Native renal function after combined liver-kidney transplant for type 1 hepatorenal syndrome: initial report on the use of postoperative Technetium-99 m-mercaptoacetyltriglycine scans.

Type 1 hepatorenal syndrome (HRS) is characterized by rapid deterioration of renal function. We sought to assess native kidney function after combined kidney-liver transplant (CLKTx) performed for type 1 HRS. We performed a retrospective, cross-sectional, single-center study. All patients with Type 1 HRS who received a CLKTx at the University of California, San Francisco from 1997 to 2007 were screened for enrollment. Patients with a baseline estimated glomerular filtration rate (eGFR) ≥30 ml/min/1.73 m(2) were eligible. Twenty-three patients were identified and consented to receive a Technetium-99 m-mercaptoacetyltriglycine (MAG3) nuclear scan to measure the native kidney contribution to overall renal function. Only 4 of the 23 subjects (17.4%) demonstrated native renal function that consisted of a contribution ≥50% of total renal function. Several factors and comorbidities such as age, gender, race, duration of HRS, need for and duration of renal replacement therapy, need for pressors, urine sodium, proteinuria, and use of octreotide/midodrine were analyzed and not found to be significant in predicting native renal function. The assessment of post-transplant native renal function following CLKTx may allow for improved accuracy in identifying the patients in need of CLKTx, and thus allow for greater optimization of dual-organ allocation strategies in patients with concomitant liver and renal failure.

Orthotopic treatment model of prostate cancer and metastasis in the immunocompetent mouse: efficacy of flt3 ligand immunotherapy.

We established an orthotopic treatment model of prostate cancer to generate reproducible primary and metastatic carcinoma in immunocompetent C57BL/6 mice. Using an in vivo selection scheme of intraprostatic implantation of TRAMP-C1 cells, primary prostate tumors were cultured and recycled three times by intraprostatic injection resulting in the selection and establishment of the recycled cell line TRAMP-C1P3. Prostate tumors were detected approximately 30 days post-implantation with periaortic lymph node metastasis in 19/20 (95%) of mice. Tissue culture amplification, DNA ploidy and PCR amplification of the SV40 transgene were used to detect metastatic TRAMP-C1P3 in lymph node specimens. Tissue culture amplification and DNA ploidy were as sensitive as SV40 transgene amplification by PCR in detection of early metastatic disease in draining lymph nodes. To establish the use of the orthotopic model of prostate cancer for immunotherapy, mice were injected orthotopically with TRAMP-C1P3 cells and 7 days post-implantation treated daily for 28 days with either flt3L or carrier control. Carrier-treated mice had clinically detectable prostate tumors, lymph node metastasis and were moribund at 29-35 days, whereas flt3L therapy markedly suppressed primary TRAMP-C1P3 growth and lymph node metastasis, and prolonged survival. In summary, we have established a reproducible and clinically relevant orthotopic treatment model of prostate cancer in immunocompetent mice with application to a variety of therapeutic strategies. We demonstrate that flt3L treatment suppressed orthotopic prostate tumor growth and lymph node metastasis reinforcing a role for flt3L as an immunotherapeutic strategy for prostate cancer.