Treatment of early-stage mycosis fungoides: results from the PROspective Cutaneous Lymphoma International Prognostic Index (PROCLIPI) study.

BACKGROUND: The PROspective Cutaneous Lymphoma International Prognostic Index (PROCLIPI) study is a prospective analysis of an international database. Here we examine front-line treatments and quality of life (QoL) in patients with newly diagnosed mycosis fungoides (MF). OBJECTIVES: To identify (i) differences in first-line approaches according to tumour-nodes-metastasis-blood (TNMB) staging; (ii) parameters related to a first-line systemic approach and (iii) response rates and QoL measures. METHODS: In total, 395 newly diagnosed patients with early-stage MF (stage IA-IIA) were recruited from 41 centres in 17 countries between 1 January 2015 and 31 December 2018 following central clinicopathological review. RESULTS: The most common first-line therapy was skin-directed therapy (SDT) (322 cases, 81·5%), while a smaller percentage (44 cases, 11·1%) received systemic therapy. Expectant observation was used in 7·3%. In univariate analysis, the use of systemic therapy was significantly associated with higher clinical stage (IA, 6%; IB, 14%; IIA, 20%; IA-IB vs. IIA, P < 0·001), presence of plaques (T1a/T2a, 5%; T1b/T2b, 17%; P < 0·001), higher modified Severity Weighted Assessment Tool (> 10, 15%; ≤ 10, 7%; P = 0·01) and folliculotropic MF (FMF) (24% vs. 12%, P = 0·001). Multivariate analysis demonstrated significant associations with the presence of plaques (T1b/T2b vs. T1a/T2a, odds ratio 3·07) and FMF (odds ratio 2·83). The overall response rate (ORR) to first-line SDT was 73%, while the ORR to first-line systemic treatments was lower (57%) (P = 0·027). Health-related QoL improved significantly both in patients with responsive disease and in those with stable disease. CONCLUSIONS: Disease characteristics such as presence of plaques and FMF influence physician treatment choices, and SDT was superior to systemic therapy even in patients with such disease characteristics. Consequently, future treatment guidelines for early-stage MF need to address these issues.

Should we be imaging lymph nodes at initial diagnosis of early-stage mycosis fungoides? Results from the PROspective Cutaneous Lymphoma International Prognostic Index (PROCLIPI) international study.

BACKGROUND: Early-stage mycosis fungoides (MF) includes involvement of dermatopathic lymph nodes (LNs) or early lymphomatous LNs. There is a lack of unanimity among current guidelines regarding the indications for initial staging imaging in early-stage presentation of MF in the absence of enlarged palpable LNs. OBJECTIVES: To investigate how often imaging is performed in patients with early-stage presentation of MF, to assess the yield of LN imaging, and to determine what disease characteristics promoted imaging. METHODS: A review of clinicopathologically confirmed newly diagnosed patients with cutaneous patch/plaque (T1/T2) MF from PROspective Cutaneous Lymphoma International Prognostic Index (PROCLIPI) data. RESULTS: PROCLIPI enrolled 375 patients with stage T1/T2 MF: 304 with classical MF and 71 with folliculotropic MF. Imaging was performed in 169 patients (45%): 83 with computed tomography, 18 with positron emission tomography-computed tomography and 68 with ultrasound. Only nine of these (5%) had palpable enlarged (≥ 15 mm) LNs, with an over-representation of plaques, irrespectively of the 10% body surface area cutoff that distinguishes T1 from T2. Folliculotropic MF was not more frequently imaged than classical MF. Radiologically enlarged LNs (≥ 15 mm) were detected in 30 patients (18%); only seven had clinical lymphadenopathy. On multivariate analysis, plaque presentation was the sole parameter significantly associated with radiologically enlarged LNs. Imaging of only clinically enlarged LNs upstaged 4% of patients (seven of 169) to at least IIA, whereas nonselective imaging upstaged another 14% (24 of 169). LN biopsy, performed in eight of 30 patients, identified N3 (extensive lymphomatous involvement) in two and N1 (dermatopathic changes) in six. CONCLUSIONS: Physical examination was a poor determinant of LN enlargement or involvement. Presence of plaques was associated with a significant increase in identification of enlarged or involved LNs in patients with early-stage presentation of MF, which may be important when deciding who to image. Imaging increases the detection rate of stage IIA MF, and identifies rare cases of extensive lymphomatous nodes, upstaging them to advanced-stage IVA2.

Characteristics associated with significantly worse quality of life in mycosis fungoides/Sézary syndrome from the Prospective Cutaneous Lymphoma International Prognostic Index (PROCLIPI) study.

BACKGROUND: Mycosis fungoides (MF) and Sézary Syndrome (SS) are the most common cutaneous T-cell lymphomas. MF/SS is accompanied by considerable morbidity from pain, itching and disfigurement. AIM: To identify factors associated with poorer health-related quality of life (HRQoL) in patients newly diagnosed with MF/SS. METHODS: Patients enrolled into Prospective Cutaneous Lymphoma International Prognostic Index (PROCLIPI; an international observational study in MF/SS) had their HRQoL assessed using the Skindex-29 questionnaire. Skindex-29 scores were analysed in relation to patient- and disease-specific characteristics. RESULTS: The study population consisted of 237 patients [60·3% male; median age 60 years, (interquartile range 49-70)], of whom 179 had early MF and 58 had advanced MF/SS. In univariate analysis, HRQoL, as measured by Skindex-29, was worse in women, SS, late-stage MF, those with elevated lactate dehydrogenase, alopecia, high modified Severity Weighted Assessment Tool and confluent erythema. Linear regression models only identified female gender (ß = 8·61; P = 0·003) and alopecia (ß = 9·71, P = 0·02) as independent predictors of worse global HRQoL. Item-level analysis showed that the severe impairment in symptoms [odds ratio (OR) 2·14, 95% confidence interval (CI) 1·19-3·89] and emotions (OR 1·88, 95% CI 1·09-3·27) subscale scores seen in women was caused by more burning/stinging, pruritus, irritation and greater feelings of depression, shame, embarrassment and annoyance with their diagnosis of MF/SS. CONCLUSIONS: HRQoL is significantly more impaired in newly diagnosed women with MF/SS and in those with alopecia. As Skindex-29 does not include existential questions on cancer, which may cause additional worry and distress, a comprehensive validated cutaneous T-cell lymphoma-specific questionnaire is urgently needed to more accurately assess disease-specific HRQoL in these patients. What's already known about this topic? Cross-sectional studies of mixed populations of known and newly diagnosed patients with mycosis fungoides (MF)/Sézary syndrome (SS) have shown significant impairment in health-related quality of life (HRQoL). Previous studies on assessing gender-specific differences in HRQoL in MF/SS are conflicting. More advanced-stage disease and pruritus is associated with poorer HRQoL in patients with MF/SS. What does this study add? This is the first prospective study to investigate HRQoL in a homogenous group of newly diagnosed patients with MF/SS. In patients newly diagnosed with MF/SS, HRQoL is worse in women and in those with alopecia and confluent erythema. MF/SS diagnosis has a multidimensional impact on patient HRQoL, including a large burden of cutaneous symptoms, as well as a negative impact on emotional well-being.

Recommendations for Radiotherapy Technique and Dose in Extra-nodal Lymphoma.

Extra-nodal sites may be involved in around 40% of patients with non-Hodgkin lymphoma. The general principles for target volume delineation in this setting are presented, together with specific examples. In general, the entire organ affected should be encompassed in the clinical target volume with an expansion of at least 10 mm, increased in some instances to account for patterns of potential lymphatic flow. Adjacent lymph nodes may be treated using standard techniques for nodal irradiation. Doses for extra-nodal lymphoma follow the same principles as nodal lymphoma, delivering 30 Gy in 15 fractions for Hodgkin and aggressive non-Hodgkin lymphoma and 24 Gy in 12 fractions for indolent lymphomas, with the exception of certain palliative situations, mycosis fungoides, central nervous system lymphoma and natural killer/T-cell lymphoma.

Recommendations for the use of radiotherapy in nodal lymphoma.

These guidelines have been developed to define the use of radiotherapy for lymphoma in the current era of combined modality treatment taking into account increasing concern over the late side-effects associated with previous radiotherapy. The role of reduced volume and reduced doses is addressed, integrating modern imaging with three-dimensional planning and advanced techniques of treatment delivery. Both wide-field and involved-field techniques have now been supplanted by the use of defined volumes based on node involvement shown on computed tomography (CT) and positron emission tomography (PET) imaging and applying the International Commission on Radiation Units and Measurements concepts of gross tumour volume (GTV), clinical target volume (CTV) and planning target volume (PTV). The planning of lymphoma patients for radical radiotherapy should now be based upon contrast enhanced 3 mm contiguous CT with three-dimensional definition of volumes using the convention of GTV, CTV and PTV. The involved-site radiotherapy concept defines the CTV based on the PET-defined pre-chemotherapy sites of involvement with an expansion in the cranio-caudal direction of lymphatic spread by 1.5 cm, constrained to tissue planes such as bone, muscle and air cavities. The margin allows for uncertainties in PET resolution, image registration and changes in patient positioning and shape. There is increasing evidence in both Hodgkin and non-Hodgkin lymphoma that traditional doses are higher than necessary for disease control and related to the incidence of late effects. No more than 30 Gy for Hodgkin and aggressive non-Hodgkin lymphoma and 24 Gy for indolent lymphomas is recommended; lower doses of 20 Gy in combination therapy for early-stage low-risk Hodgkin lymphoma may be sufficient. As yet there are no large datasets validating the use of involved-site radiotherapy; these will emerge from the current generation of clinical trials. Radiotherapy remains the most effective single modality in the treatment of lymphoma. A reduction in both treatment volume and overall treatment dose should now be considered to minimise the risks of late sequelae. However, it is important that this is not at the expense of the excellent disease control currently achieved.

Validation of an ELISA for the determination of rituximab pharmacokinetics in clinical trials subjects.

Rituximab is a chimeric anti-CD20 monoclonal antibody that has revolutionised the treatment of many B-cell malignancies, and is now increasingly being used in non-malignant conditions such as auto-immune disorders. Serum rituximab levels are highly variable in patients receiving similar 'standard' approved doses. Little is known regarding the factors that affect serum rituximab concentration and that in turn may influence clinical outcome. In order to provide a tool that may ultimately enable patient specific dosing of rituximab therapy, we have validated a reliable, robust ELISA for the quantitation of serum rituximab levels to provide accurate pharmacokinetic (PK) data that will guide the optimisation of rituximab dosing regimes. Extensive validation of the assay was performed in order to utilise the assay for clinical applications. The within and between day plate coating reproducibility was tested and proved a robust starting platform for the assay. The within day precision for the assay was determined using spiked serum samples and was shown to have a coefficient of variation (CV) of <10% with an accuracy between 91 and 125%. The between day precision (CV) was <25% with an accuracy between 95 and 109%. Dilution linearity and parallelism were demonstrated. Spike recovery for all concentrations and donors was shown to be within +/-15% on average, with a CV below 10%. This assay is highly accurate and reproducible in determining the levels of rituximab in spiked serum samples. It meets stringent acceptance criteria, is fit for purpose, and is currently being applied to several clinical trials incorporating rituximab in the treatment of lymphoma. This assay represents a useful tool for clinical application of this widely used therapeutic.

Novel CDH3 mutations in hypotrichosis with juvenile macular dystrophy.

BACKGROUND: Hypotrichosis with juvenile macular dystrophy (HJMD) is a rare autosomal recessive disorder characterized by sparse and short hair, heralding progressive degeneration of the retinal pigment epithelium, which leads to blindness by the second decade of life. The disorder is caused by mutations in CDH3, a gene encoding P-cadherin, a major component of adherens junctions. Most HJMD cases reported to date have been shown to be caused by homozygous CDH3 mutations segregating in consanguineous families. AIM AND METHODS: To elucidate the genetic basis of HJMD in two nonconsanguineous families, we established the coding sequence of CDH3 in four patients and their healthy siblings. RESULTS: The four patients demonstrated markedly variable degrees of visual acuity impairment. Novel biallelic recessive mutations were identified in all affected individuals. One patient in the first family was found to carry two heterozygous mutations, IVS2 + 1G-->A and p.E504K; the other three patients in the second family were compound heterozygous for a missense mutation, p.H575R, and a nonsense mutation, p.R221X. CONCLUSION: This paper expands the spectrum of known mutations in CDH3 and points to the existence of clinical heterogeneity in this syndrome.

Hypercalcaemia, parathyroid hormone-related protein and malignancy.

Hypercalcaemia is the most common serious metabolic complication of malignancy. Recent advances have significantly increased our understanding of the pathophysiology of hypercalcaemia of malignancy and revealed the importance of parathyroid hormone-related protein (PTHrP) in a wide range of physiological and pathological processes. This review examines the pathophysiology of hypercalcaemia of malignancy, focusing on the role of PTHrP before discussing further pathological and physiological processes in which PTHrP may be implicated, and the impact of this knowledge on the management of malignant disease.

Antibody therapy of lymphoma.

The availability of rituximab and the possible imminent availability of two new radiolabelled monoclonal anti-CD20 antibodies (Yttrium-90 (90Y)-ibritumomab and Iodine-131(131I)-tositumomab) have captured much attention in the treatment of lymphoma. The chimeric monoclonal anti-CD20 antibody, rituximab has truly heralded a new era for the treatment of lymphoma and human malignancies. The full potential of antibody-based therapy to improve the outcome in patients with B-cell non-Hodgkin's lymphoma has yet to be defined, but recent data suggests that the combination of chemotherapy plus rituximab may significantly improve outcome for patients with aggressive lymphoma over chemotherapy alone. Highly promising data are also emerging for the use of rituximab in combination with chemotherapy in other types of lymphoma. New advances in antibody therapy, driven by new technologies and defining novel antigen targets, offer the promise of more effective tumour specific therapies. Combinations of antibodies, either conjugated with radioisotopes or unlabelled, used with chemotherapy are likely to provide definitive advances in the treatment of lymphoma in the immediate future.

Identification and pharmacological characterization of a novel human melanin-concentrating hormone receptor, mch-r2.

Melanin-concentrating hormone (MCH) is a neuropeptide highly expressed in the brain that regulates several physiological functions mediated by receptors in the G protein-coupled receptor family. Recently an orphan receptor, SLC-1, has been identified as an MCH receptor (MCH-R1). Herein we identify and characterize a novel receptor for human MCH (MCH-R2). The receptor is composed of 340 amino acids encoded by a 1023-base pair cDNA and is 35% homologous to SLC-1. (125)I-MCH specifically bound to Chinese hamster ovary cells stably expressing MCH-R2. MCH stimulated dose-dependent increases in intracellular free Ca(2+) and inositol phosphate production in these cells but did not affect cAMP production. The pharmacological profile for mammalian MCH, [Phe(13),Tyr(19)]MCH, and salmon MCH at MCH-R2 differed compared with MCH-R1 as assessed by intracellular signaling and radioligand binding assays. The EC(50) in signaling assays and the IC(50) in radioligand binding assays of salmon MCH was an order of magnitude higher than mammalian MCH at MCH-R2. By comparison, the EC(50) and IC(50) values of salmon MCH and mammalian MCH at MCH-R1 were relatively similar. Blot hybridization revealed exclusive expression of MCH-R2 mRNA in several distinct brain regions, particularly in the cortical area, suggesting the involvement of MCH-R2 in the central regulation of MCH-mediated functions.

Molecular identification and characterization of the platelet ADP receptor targeted by thienopyridine antithrombotic drugs.

ADP plays a critical role in modulating thrombosis and hemostasis. ADP initiates platelet aggregation by simultaneous activation of two G protein-coupled receptors, P2Y1 and P2Y12. Activation of P2Y1 activates phospholipase C and triggers shape change, while P2Y12 couples to Gi to reduce adenylyl cyclase activity. P2Y12 has been shown to be the target of the thienopyridine drugs, ticlopidine and clopidogrel. Recently, we cloned a human orphan receptor, SP1999, highly expressed in brain and platelets, which responded to ADP and had a pharmacological profile similar to that of P2Y12. To determine whether SP1999 is P2Y12, we generated SP1999-null mice. These mice appear normal, but they exhibit highly prolonged bleeding times, and their platelets aggregate poorly in responses to ADP and display a reduced sensitivity to thrombin and collagen. These platelets retain normal shape change and calcium flux in response to ADP but fail to inhibit adenylyl cyclase. In addition, oral clopidogrel does not inhibit aggregation responses to ADP in these mice. These results demonstrate that SP1999 is indeed the elusive receptor, P2Y12. Identification of the target receptor of the thienopyridine drugs affords us a better understanding of platelet function and provides tools that may lead to the discovery of more effective antithrombotic therapies.

Cloning and characterization of a novel human histamine receptor.

Histamine exerts its numerous physiological functions through interaction with G protein-coupled receptors. Three such receptors have been defined at both the pharmacological and molecular level, while pharmacological evidence hints at the existence of further subtypes. We report here the cloning and characterization of a fourth histamine receptor subtype. Initially discovered in an expressed-sequence tag database, the full coding sequence (SP9144) was subsequently identified in chromosome 18 genomic sequence. This virtual coding sequence exhibited highest homology to the H(3) histamine receptor and was used to generate a full-length clone by polymerase chain reaction (PCR). The distribution of mRNA encoding SP9144 was restricted to cells of the immune system as determined by quantitative PCR. HEK-293 cells transiently transfected with SP9144 and a chimeric G protein alpha-subunit (Galpha(q/i1,2)) exhibited increases in intracellular [Ca(2+)] in response to histamine but not other biogenic amines. SP9144-transfected cells exhibited saturable, specific, high-affinity binding of [(3)H]histamine, which was potently inhibited by H(3) receptor-selective compounds. The rank order and potency of these compounds at SP9144 differed from the rank order at the H(3) receptor. Although SP9144 apparently coupled to Galpha(i), HEK-293 cells stably transfected with SP9144 did not exhibit histamine-mediated inhibition of forskolin-stimulated cAMP levels. However, both [(35)S]GTPgammaS binding and phosphorylation of mitogen-activated protein kinase were stimulated by histamine via SP9144 activation. In both of these assays, SP9144 exhibited evidence of constitutive activation. Taken together, these data demonstrate that SP9144 is a unique, fourth histamine receptor subtype.

ADP is the cognate ligand for the orphan G protein-coupled receptor SP1999.

P2Y receptors are a class of G protein-coupled receptors activated primarily by ATP, UTP, and UDP. Five mammalian P2Y receptors have been cloned so far including P2Y1, P2Y2, P2Y4, P2Y6, and P2Y11. P2Y1, P2Y2, and P2Y6 couple to the activation of phospholipase C, whereas P2Y4 and P2Y11 couple to the activation of both phospholipase C and the adenylyl cyclase pathways. Additional ADP receptors linked to Galpha(i) have been described but have not yet been cloned. SP1999 is an orphan G protein-coupled receptor, which is highly expressed in brain, spinal cord, and blood platelets. In the present study, we demonstrate that SP1999 is a Galpha(i)-coupled receptor that is potently activated by ADP. In an effort to identify ligands for SP1999, fractionated rat spinal cord extracts were assayed for Ca(2+) mobilization activity against Chinese hamster ovary cells transiently transfected with SP1999 and chimeric Galpha subunits (Galpha(q/i)). A substance that selectively activated SP1999-transfected cells was identified and purified through a series of chromatographic steps. Mass spectral analysis of the purified material definitively identified it as ADP. ADP was subsequently shown to inhibit forskolin-stimulated adenylyl cyclase activity through selective activation of SP1999 with an EC(50) of 60 nM. Other nucleotides were able to activate SP1999 with a rank order of potency 2-MeS-ATP = 2-MeS-ADP > ADP = adenosine 5'-O-2-(thio)diphosphate > 2-Cl-ATP > adenosine 5'-O-(thiotriphosphate). Thus, SP1999 is a novel, Galpha(i)-linked receptor for ADP.

Identification of a human gastrointestinal tract and immune system receptor for the peptide neuromedin U.

Neuromedin U (NmU) is a 25 amino acid peptide prominently expressed in the upper gastrointestinal (GI) tract and central nervous system. It is highly conserved throughout evolution and induces smooth muscle contraction in a variety of species. Our understanding of NmU biology has been limited because the identity of its receptor was unknown. Here we demonstrate that GPR66/FM-3 is specifically stimulated by NmU, causing the mobilization of intracellular calcium. This response was dose-dependent (EC(50) = 10 nM) and specific in that none of over 1000 ligands tested, including other neuromedins (NmB, C, L, K, N), induced a calcium flux in GPR66/FM-3-transfected cells. The GPR66/FM-3 mRNA is prominently expressed in the upper GI tract of humans, as is the mRNA for NmU, consistent with role for this receptor-ligand pair in regulating the function of this organ system. In addition, we show that whereas neuromedin U is expressed by monocytes and dendritic cells, GPR66/FM-3 is expressed by T cells and NK cells. These data suggest a previously unrecognized role for NmU as an immunoregulatory molecule.

Identification of a novel neuromedin U receptor subtype expressed in the central nervous system.

Neuromedin U is a neuropeptide prominently expressed in the upper gastrointestinal tract and central nervous system. Recently, GPR66/FM-3 (NmU-R1) was identified as a specific receptor for neuromedin U. A BLAST search of the GenBank(TM) genomic database using the NmU-R1 cDNA sequence revealed a human genomic fragment encoding a G protein-coupled receptor that we designated NmU-R2 based on its homology to NmU-R1. The full-length NmU-R2 cDNA was subsequently cloned, stably expressed in 293 cells, and shown to mobilize intracellular calcium in response to neuromedin U. This response was dose-dependent (EC(50) = 5 nm) and specific in that other neuromedins did not induce a calcium flux in receptor-transfected cells. Expression analysis of human NmU-R2 demonstrated its mRNA to be most highly expressed in central nervous system tissues. Based on these data, we conclude that NmU-R2 is a novel neuromedin U receptor subtype that is likely to mediate central nervous system-specific neuromedin U effects.

A novel hepatointestinal leukotriene B4 receptor. Cloning and functional characterization.

Leukotriene B(4) (LTB(4)) is a product of eicosanoid metabolism and acts as an extremely potent chemotactic mediator for inflammation. LTB(4) exerts positive effects on the immigration and activation of leukocytes. These effects suggest an involvement of LTB(4) in several diseases: inflammatory bowel disease, psoriasis, arthritis, and asthma. LTB(4) elicits actions through interaction with one or more cell surface receptors that lead to chemotaxis and inflammation. One leukotriene B(4) receptor has been recently identified (LTB(4)-R1). In this report we describe cloning of a cDNA encoding a novel 358-amino acid receptor (LTB(4)-R2) that possesses seven membrane-spanning domains and is homologous (42%) and genetically linked to LTB(4)-R1. Expression of LTB(4)-R2 is broad but highest in liver, intestine, spleen, and kidney. In radioligand binding assays, membranes prepared from COS-7 cells transfected with LTB(4)-R2 cDNA displayed high affinity (K(d) = 0.17 nm) for [(3)H]LTB(4). Radioligand competition assays revealed high affinities of the receptor for LTB(4) and LTB(5), and 20-hydroxy-LTB(4), and intermediate affinities for 15(S)-HETE and 12-oxo-ETE. Three LTB(4) receptor antagonists, 14,15-dehydro-LTB(4), LTB(4)-3-aminopropylamide, and U-75302, had high affinity for LTB(4)-R1 but not for LTB(4)-R2. No apparent affinity binding for the receptors was detected for the CysLT1-selective antagonists montelukast and zafirlukast. LTB(4) functionally mobilized intracellular calcium and inhibited forskolin-stimulated cAMP production in 293 cells. The discovery of this new receptor should aid in further understanding the roles of LTB(4) in pathologies in these tissues and may provide a tool in identification of specific antagonists/agonists for potential therapeutic treatments.