TORC1 is an essential regulator of nutrient-controlled proliferation and differentiation in Leishmania.

Leishmania parasites undergo differentiation between various proliferating and non-dividing forms to adapt to changing host environments. The mechanisms that link environmental cues with the parasite's developmental changes remain elusive. Here, we report that Leishmania TORC1 is a key environmental sensor for parasite proliferation and differentiation in the sand fly-stage promastigotes and for replication of mammalian-stage amastigotes. We show that Leishmania RPTOR1, interacts with TOR1 and LST8, and identify new parasite-specific proteins that interact in this complex. We investigate TORC1 function by conditional deletion of RPTOR1, where under nutrient-rich conditions RPTOR1 depletion results in decreased protein synthesis and growth, G1 cell cycle arrest and premature differentiation from proliferative promastigotes to non-dividing mammalian-infective metacyclic forms. These parasites are unable to respond to nutrients to differentiate into proliferative retroleptomonads, which are required for their blood-meal induced amplification in sand flies and enhanced mammalian infectivity. We additionally show that RPTOR1-/- metacyclic promastigotes develop into amastigotes but do not proliferate in the mammalian host to cause pathology. RPTOR1-dependent TORC1 functionality represents a critical mechanism for driving parasite growth and proliferation.

Phase II trial of CV301 vaccine combined with atezolizumab in advanced urothelial carcinoma.

CV301 comprises recombinant poxviruses, Modified Vaccinia Ankara (MVA) and Fowlpox (FPV), encoding CEA, MUC-1, and co-stimulatory Molecules (TRICOM) ICAM-1, LFA-3, and B7-1. MVA-BN-CV301 is used for priming and FPV-CV301 is used for boosting. A Phase 2, single-arm trial was designed to evaluate CV301 plus atezolizumab as first-line treatment for cisplatin-ineligible advanced urothelial carcinoma (aUC) (Cohort 1) or progressing after platinum chemotherapy (Cohort 2). MVA-CV301 was given subcutaneously (SC) on Days 1 and 22 and FPV-CV301 SC from day 43 every 21 days for 4 doses, then tapered gradually over up to 2 years. Atezolizumab 1200 mg IV was given every 21 days. The primary endpoint was objective response rate (ORR). Overall, 43 evaluable patients received therapy: 19 in Cohort 1; 24 in Cohort 2; nine experienced ≥ Grade 3 therapy-related adverse events. In Cohort 1, one had partial response (PR) (ORR 5.3%, 90% CI 0.3, 22.6). In Cohort 2, 1 complete response and 1 PR were noted (ORR 8.3%, 90% CI 1.5, 24.0). The trial was halted for futility. Patients exhibiting benefit demonstrated T-cell response to CEA and MUC-1. The trial illustrates the challenges in the development of vaccines, which should be guided by robust preclinical data.

Crystal Structure and Activity Studies of the C11 Cysteine Peptidase from Parabacteroides merdae in the Human Gut Microbiome.

Clan CD cysteine peptidases, a structurally related group of peptidases that include mammalian caspases, exhibit a wide range of important functions, along with a variety of specificities and activation mechanisms. However, for the clostripain family (denoted C11), little is currently known. Here, we describe the first crystal structure of a C11 protein from the human gut bacterium, Parabacteroides merdae (PmC11), determined to 1.7-Å resolution. PmC11 is a monomeric cysteine peptidase that comprises an extended caspase-like α/ß/α sandwich and an unusual C-terminal domain. It shares core structural elements with clan CD cysteine peptidases but otherwise structurally differs from the other families in the clan. These studies also revealed a well ordered break in the polypeptide chain at Lys(147), resulting in a large conformational rearrangement close to the active site. Biochemical and kinetic analysis revealed Lys(147) to be an intramolecular processing site at which cleavage is required for full activation of the enzyme, suggesting an autoinhibitory mechanism for self-preservation. PmC11 has an acidic binding pocket and a preference for basic substrates, and accepts substrates with Arg and Lys in P1 and does not require Ca(2+) for activity. Collectively, these data provide insights into the mechanism and activity of PmC11 and a detailed framework for studies on C11 peptidases from other phylogenetic kingdoms.

PNT1 Is a C11 Cysteine Peptidase Essential for Replication of the Trypanosome Kinetoplast.

The structure of a C11 peptidase PmC11 from the gut bacterium, Parabacteroides merdae, has recently been determined, enabling the identification and characterization of a C11 orthologue, PNT1, in the parasitic protozoon Trypanosoma brucei. A phylogenetic analysis identified PmC11 orthologues in bacteria, archaea, Chromerids, Coccidia, and Kinetoplastida, the latter being the most divergent. A primary sequence alignment of PNT1 with clostripain and PmC11 revealed the position of the characteristic His-Cys catalytic dyad (His(99) and Cys(136)), and an Asp (Asp(134)) in the potential S1 binding site. Immunofluorescence and cryoelectron microscopy revealed that PNT1 localizes to the kinetoplast, an organelle containing the mitochondrial genome of the parasite (kDNA), with an accumulation of the protein at or near the antipodal sites. Depletion of PNT1 by RNAi in the T. brucei bloodstream form was lethal both in in vitro culture and in vivo in mice and the induced population accumulated cells lacking a kinetoplast. In contrast, overexpression of PNT1 led to cells having mislocated kinetoplasts. RNAi depletion of PNT1 in a kDNA independent cell line resulted in kinetoplast loss but was viable, indicating that PNT1 is required exclusively for kinetoplast maintenance. Expression of a recoded wild-type PNT1 allele, but not of an active site mutant restored parasite viability after induction in vitro and in vivo confirming that the peptidase activity of PNT1 is essential for parasite survival. These data provide evidence that PNT1 is a cysteine peptidase that is required exclusively for maintenance of the trypanosome kinetoplast.

Tuning the photoactivated anticancer activity of Pt(iv) compounds via distant ferrocene conjugation.

Photoactive prodrugs offer potential for spatially-selective antitumour activity with minimal effects on normal tissues. Excited-state chemistry can induce novel effects on biochemical pathways and combat resistance to conventional drugs. Photoactive metal complexes in particular, have a rich and relatively unexplored photochemistry, especially an ability to undergo facile intersystem crossing and populate triplet states. We have conjugated the photoactive octahedral Pt(iv) complex trans, trans, trans-[Pt(N3)2(OH)2(py)2] to ferrocene to introduce novel features into a candidate photochemotherapeutic drug. The X-ray crystal structure of the conjugate Pt-Fe confirmed the axial coordination of a ferrocene carboxylate, with Pt(iv) and Fe(ii) 6.07 Å apart. The conjugation of ferrocene red-shifted the absorption spectrum and ferrocene behaves as a light antenna allowing charge transfer from iron to platinum, promoting the photoactivation of Pt-Fe with light of longer wavelength. Cancer cellular accumulation is enhanced, and generation of reactive species is catalysed after photoirradiation, introducing ferroptosis as a contribution towards the cell-death mechanism. TDDFT calculations were performed to shed light on the behaviour of Pt-Fe when it is irradiated. Intersystem spin-crossing allows the formation of triplet states centred on both metal atoms. The dissociative nature of triplet states confirms that they can be involved in ligand detachment due to irradiation. The Pt(ii) photoproducts mainly retain the trans-{Pt(py)2}2+fragment. Visible light irradiation gives rise to micromolar activity for Pt-Fe towards ovarian, lung, prostate and bladder cancer cells under both normoxia and hypoxia, and some photoproducts appear to retain Pt(iv)-Fe(ii) conjugation.

Phase I studies of davoceticept (ALPN-202), a PD-L1-dependent CD28 co-stimulator and dual PD-L1/CTLA-4 inhibitor, as monotherapy and in combination with pembrolizumab in advanced solid tumors (NEON-1 and NEON-2).

BACKGROUND: Davoceticept (ALPN-202) is an Fc fusion of a CD80 variant immunoglobulin domain designed to mediate programmed death-ligand 1 (PD-L1)-dependent CD28 co-stimulation while inhibiting the PD-L1 and cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) checkpoints. The safety and efficacy of davoceticept monotherapy and davoceticept and pembrolizumab combination therapy in adult patients with advanced solid tumors were explored in NEON-1 and NEON-2, respectively. METHODS: In NEON-1 (n=58), davoceticept 0.001-10 mg/kg was administered intravenous either once weekly (Q1W) or once every 3 weeks (Q3W). In NEON-2 (n=29), davoceticept was administered intravenously at 2 dose levels (0.1 or 0.3 mg/kg) Q1W or Q3W with pembrolizumab (400 mg once every 6 weeks). In both studies, primary endpoints included incidence of dose-limiting toxicities (DLT); type, incidence, and severity of adverse events (AEs) and laboratory abnormalities; and seriousness of AEs. Secondary endpoints included antitumor efficacy assessed using RECIST v1.1, pharmacokinetics, anti-drug antibodies, and pharmacodynamic biomarkers. RESULTS: The incidence of treatment-related AEs (TRAEs) and immune-related adverse events (irAEs) was 67% (39/58) and 36% (21/58) with davoceticept monotherapy, and 62% (18/29) and 31% (9/29) with davoceticept and pembrolizumab combination, respectively. The incidence of ≥grade (Gr)3 TRAEs and ≥Gr3 irAEs was 12% (7/58) and 5% (3/58) with davoceticept monotherapy, and 24% (7/29) and 10% (3/29) with davoceticept and pembrolizumab combination, respectively. One DLT of Gr3 immune-related gastritis occurred during davoceticept monotherapy 3 mg/kg Q3W. During davoceticept combination with pembrolizumab, two Gr5 cardiac DLTs occurred; one instance each of cardiogenic shock (0.3 mg/kg Q3W, choroidal melanoma metastatic to the liver) and immune-mediated myocarditis (0.1 mg/kg Q3W, microsatellite stable metastatic colorectal adenocarcinoma), prompting early termination of both studies. Across both studies, five patients with renal cell carcinoma (RCC) exhibited evidence of clinical benefit (two partial response, three stable disease). CONCLUSIONS: Davoceticept was generally well tolerated as monotherapy at intravenous doses up to 10 mg/kg. Evidence of clinical activity was observed with davoceticept monotherapy and davoceticept in combination with pembrolizumab, notably in RCC. However, two fatal cardiac events occurred with the combination of low-dose davoceticept and pembrolizumab. Future clinical investigation with davoceticept should not consider combination with programmed death-1-inhibitor anticancer mechanisms, until its safety profile is more fully elucidated. TRIAL REGISTRATION NUMBER: NEON-1 (NCT04186637) and NEON-2 (NCT04920383).

Bing-Neel syndrome: a case report and systematic review of clinical manifestations, diagnosis, and treatment options.

BACKGROUND: Bing-Neel syndrome is an extremely rare neurologic complication of Waldenström macroglobulinemia (WM) that was first described in 1936. It is associated with central nervous system infiltration by neoplastic lymphoplasmacytoid and plasma cells with or without cerebrospinal fluid (CSF) hyperglobulinemia. CASE REPORT: We report a case of a 69-year-old white man with a 10-year history of WM. He was diagnosed with Bing-Neel syndrome based on magnetic resonance imaging and pathology studies of CSF. In addition, a comprehensive review of the reported cases of Bing-Neel syndrome in the up-to-date English-language literature was performed. RESULTS: Our patient underwent successful treatment with cranial radiation and intrathecal chemotherapy. He has been in clinical and pathologic remission for 3 years following the completion of his treatment. Based on our literature review, we also summarize and discuss clinical manifestations, diagnosis, and treatment options for Bing-Neel syndrome. CONCLUSION: Bing-Neel syndrome is a rare and potentially treatable complication of WM. Patients with a history of WM presenting with neurologic symptoms should be evaluated for possible Bing-Neel syndrome. Cranial radiation therapy alone or in combination with intrathecal chemotherapy is more likely to achieve sustainable remission than intrathecal chemotherapy alone.

Rapidly progressive metastatic multicentric epithelioid angiosarcoma of the small bowel: a case report and a review of literature.

BACKGROUND: Angiosarcoma is a rare high-grade neoplasm that frequently involves the skin and subcutaneous tissue. Rarely, angiosarcoma can occur in the gastrointestinal tract where it frequently exhibits multicentric epithelioid morphology. DESIGN: We report a case of multicentric epithelioid angiosarcoma (EAS) of the small intestine in a 73-year-old male patient who presented with weakness and melena, and was found to have bleeding lesions in the small intestine on upper gastrointestinal endoscopy. In addition to this case, we extensively reviewed the clinical and pathological features of previously reported cases of angiosarcoma of the small intestine in the English literature since 1970. RESULTS: Our patient presented with rare and aggressive EAS of the small intestine. Despite surgical resection of the lesions, the patient continued to worsen and developed rapidly progressive metastatic disease. He died within 4 months of the diagnosis. CONCLUSIONS: Angiosarcoma, especially of the deep tissues and the gastrointestinal tract, is very aggressive and rapidly metastatic. The survival rate in these patients is extremely poor, and most patients die within 6 months to 1 year of the diagnosis. Treatment usually involves surgical resection of the bleeding lesions and frequent blood transfusions for symptom alleviation.

Highly aggressive ALK-positive anaplastic large cell lymphoma with a leukemic phase and multi-organ involvement: a report of three cases and a review of the literature.

Anaplastic large cell lymphoma (ALCL) is an aggressive neoplasm of T- or null cell phenotype and is recognized as a distinct clinicopathologic subtype of non-Hodgkin lymphoma (NHL) in the revised World Health Organization (WHO) classification of hematopoietic neoplasms. It is rarely associated with leukemic phase. Most cases with leukemic involvement are the small cell variant of ALCL. These cases often lack the pleomorphism seen in the common variant of ALCL and may be misdiagnosed. We report a series of three patients who presented with leukemic phase ALCL. The patients included an 11-year-old boy, a 29-year-old man, and a 59-year-old woman. The clinical and pathologic features of these cases are reviewed. The patients in our case series with leukemic phase ALCL exhibited rare clinical features. The patients presented with massive extranodal disease involving cerebrospinal fluid (CSF), liver, spleen, lungs, and bone marrow. CSF involvement was documented morphologically as well as by flow cytometry in two patients. Two of the patients had small cell variant and the third patient had common type ALCL. The neoplastic cells in all three patients were ALK positive; however these patients died within months of diagnosis. Leukemic phase ALCL is rare, and behaves in an aggressive manner. Some, but not all, cases in the literature presenting with peripheral blood involvement had small cell variant ALCL, as seen in two of our cases. The leukemic phase of ALCL should be considered when a T-cell leukemia with unusual morphologic features is encountered.