Late-onset and long-lasting immune-related adverse events from immune checkpoint-inhibitors: An overlooked aspect in immunotherapy.

BACKGROUND: Immune checkpoint inhibitors (ICIs) have revolutionised cancer therapy but frequently cause immune-related adverse events (irAEs). Description of late-onset and duration of irAEs in the literature is often incomplete. METHODS: To investigate reporting and incidence of late-onset and long-lasting irAEs, we reviewed all registration trials leading to ICI's approval by the US FDA and/or EMA up to December 2019. We analysed real-world data from all lung cancer (LC) and melanoma (Mel) patients treated with approved ICIs at the University Hospital of Lausanne (CHUV) from 2011 to 2019. To account for the immortal time bias, we used a time-dependent analysis to assess the potential association between irAEs and overall survival (OS). RESULTS: Duration of irAEs and proportion of patients with ongoing toxicities at data cut-off were not specified in 56/62 (90%) publications of ICIs registration trials. In our real-world analysis, including 437 patients (217 LC, 220 Mel), 229 (52.4%) experienced at least one grade ≥2 toxicity, for a total of 318 reported irAEs, of which 112 (35.2%) were long-lasting (≥6 months) and about 40% were ongoing at a median follow-up of 369 days [194-695] or patient death. The cumulative probability of irAE onset from treatment initiation was 42.8%, 51.0% and 57.3% at 6, 12 and 24 months, respectively. The rate of ongoing toxicity from the time of first toxicity onset was 42.8%, 38.4% and 35.7% at 6, 12 and 24 months. Time-dependent analysis showed no significant association between the incidence of irAEs and OS in both cohorts (log Rank p = 0.67 and 0.19 for LC and Mel, respectively). CONCLUSIONS: Late-onset and long-lasting irAEs are underreported but common events during ICIs therapy. Time-dependent survival analysis is advocated to assess their impact on OS. Real-world evidence is warranted to fully capture and characterise late-onset and long-lasting irAEs in order to implement appropriate strategies for patient surveillance and follow-up.

Arming embolic beads with anti-VEGF antibodies and controlling their release using LbL technology.

Transarterial chemoembolization (TACE) is used to treat various types of hypervascular tumors such as hepatocellular carcinoma and renal cancer. However, embolization and blocking of blood vessels nourishing a tumor mass evokes an angiogenic response due to the secretion of vascular endothelial growth factor (VEGF), which results in the formation of new blood vessels and eventually limitation in therapeutic efficacy. The presented work investigates the feasibility of loading the clinically used embolic beads (DC Bead®) with Bevacizumab (BEV), an anti-VEGF antibody, and control its release kinetics via Layer-by-Layer (LbL) coating. This strategy has the aim to achieve high, localized and sustained concentrations of BEV at the tumor site and reduce drug exposure in the systemic circulation. High loading of BEV on lyophilized beads of about 76mg BEV/bead vial was achieved. LbL coating was carried out by depositing alternating layers of the biocompatible polymers alginate and poly-L-lysine. Coating was proven successful by monitoring the reversal of zeta potential after addition of each layer. Morphological changes of the bead surface before and after coating were illustrated using SEM imaging. Moreover, release profiles from different formulations were studied and results showed that optimizing the number of deposited layers effectively slows the release of BEV for three days. Activity of released BEV was studied in different 2D and 3D cell based assays. Released BEV fractions showed comparable activity to fresh BEV solution used as control after 3days. In conclusion, our results suggest the opportunity for loading anti-VEGF antibodies on commercially available embolic beads to increase the efficacy of TACE of hypervascular tumors.

Structure-Based, Rational Design of T Cell Receptors.

Adoptive cell transfer using engineered T cells is emerging as a promising treatment for metastatic melanoma. Such an approach allows one to introduce T cell receptor (TCR) modifications that, while maintaining the specificity for the targeted antigen, can enhance the binding and kinetic parameters for the interaction with peptides (p) bound to major histocompatibility complexes (MHC). Using the well-characterized 2C TCR/SIYR/H-2K(b) structure as a model system, we demonstrated that a binding free energy decomposition based on the MM-GBSA approach provides a detailed and reliable description of the TCR/pMHC interactions at the structural and thermodynamic levels. Starting from this result, we developed a new structure-based approach, to rationally design new TCR sequences, and applied it to the BC1 TCR targeting the HLA-A2 restricted NY-ESO-1157-165 cancer-testis epitope. Fifty-four percent of the designed sequence replacements exhibited improved pMHC binding as compared to the native TCR, with up to 150-fold increase in affinity, while preserving specificity. Genetically engineered CD8(+) T cells expressing these modified TCRs showed an improved functional activity compared to those expressing BC1 TCR. We measured maximum levels of activities for TCRs within the upper limit of natural affinity, K D = ∼1 - 5 µM. Beyond the affinity threshold at K D < 1 µM we observed an attenuation in cellular function, in line with the "half-life" model of T cell activation. Our computer-aided protein-engineering approach requires the 3D-structure of the TCR-pMHC complex of interest, which can be obtained from X-ray crystallography. We have also developed a homology modeling-based approach, TCRep 3D, to obtain accurate structural models of any TCR-pMHC complexes when experimental data is not available. Since the accuracy of the models depends on the prediction of the TCR orientation over pMHC, we have complemented the approach with a simplified rigid method to predict this orientation and successfully assessed it using all non-redundant TCR-pMHC crystal structures available. These methods potentially extend the use of our TCR engineering method to entire TCR repertoires for which no X-ray structure is available. We have also performed a steered molecular dynamics study of the unbinding of the TCR-pMHC complex to get a better understanding of how TCRs interact with pMHCs. This entire rational TCR design pipeline is now being used to produce rationally optimized TCRs for adoptive cell therapies of stage IV melanoma.

Anti-inflammatory, antimicrobial and antioxidant activities of Diospyros bipindensis (Gürke) extracts and its main constituents.

ETHNOPHARMACOLOGICAL RELEVANCE: Diospyros bipindensis (Gürke) stem bark is used in Cameroon by Baka Pygmies for the treatment of respiratory disorders. AIM OF THE STUDY: To assess the anti-inflammatory, antibacterial and antioxidant properties of constituents from the bark extracts through bioassay-guided fractionation. MATERIALS AND METHODS: The anti-inflammatory activity of extracts, fractions and pure compounds was assessed through the inhibition of the pro-inflammatory mediator nuclear factor-kappa B (NF-κB) transcriptional activity and nitric oxide (NO) production. DPPH, ABTS and ORAC assays were used for determining the antioxidant properties. The activity against Streptococcus pneumoniae, Staphylococcus aureus, Streptococcus pyogenes, Escherichia coli and Klebsiella pneumoniae, was evaluated on the basis of the minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) by the macrodilution method. RESULTS: The water extract showed antimicrobial activity against S. pneumoniae (MIC: 300 µg/ml) and S. pyogenes (MIC: 300 µg/ml). The dichloromethane extract efficiently inhibited NF-κB transcriptional activity and NO production and exhibited significant antioxidant activity in the ORAC assay. An interesting activity was also found against S. pneumoniae (MIC: 200 µg/ml), S. aureus (MIC: 400 µg/ml) and S. pyogenes (MIC: 200 µg/ml). The phytochemical investigation of the dichloromethane extract afforded plumbagin, canaliculatin, ismailin, betulinic acid and 4-hydroxy-5-methyl-coumarin as the main constituents. Plumbagin and ismailin were found to be responsible for the main biological activities observed. CONCLUSIONS: These results may provide a rational support for the traditional use of Diospyros bipindensis stem bark in the treatment of respiratory disorders, since the anti-inflammatory, antimicrobial and antioxidant compounds isolated from the dichloromethane extract were also present in the traditional water extract.

Brusatol-mediated induction of leukemic cell differentiation and G(1) arrest is associated with down-regulation of c-myc.

Employing the natural product quassinoid brusatol, we currently report cellular and molecular events leading to cell death or terminal differentiation in a panel of leukemic cells. Brusatol and bruceantin exerted significant cytotoxic effects with several leukemic cell lines, but not with K562 or normal lymphocytic cells. Cell lines that were less sensitive to the cytotoxic effects of brusatol responded primarily through induction of terminal differentiation. The differentiated phenotype in cell lines derived from acute or chronic myeloid leukemias (HL-60, K562, Kasumi-1, NB4, U937, BV173) was characterized for producing superoxide and non-specific esterase, and some with up-regulation of CD13 (cluster of differentiation) and down-regulation of CD15. Chronic myeloid leukemic cell lines, K562 and BV173, and acute lymphoblastic cell lines, SUPB13 and RS4;11, were induced to differentiate along the erythrocytic pathway. Withdrawal studies showed that brusatol treatment for 48 h was sufficient to induce commitment towards terminal differentiation in HL-60, K562 and SUPB13. Reh cells did not undergo maturation. Analysis of c-MYC protein expression revealed that brusatol or bruceantin down-regulated expression to undetectable levels in cell lines that were most sensitive, based on cell death or terminal differentiation. Generally, c-myc RNA was reduced, but to a lower extent than c-MYC protein levels, indicating c-myc expression was regulated by quassinoids at the post-transcriptional level. Thus, regulation of c-myc expression may represent a critical event that leads to terminal differentiation. Since these responses are facilitated at clinically achievable concentrations, quassinoids may be of value for the management of hematological malignancies.

A novel cyclooxygenase-inhibitory stilbenolignan from the seeds of Aiphanes aculeata.

[structure: see text] Aiphanol (1), a novel stilbenolignan, along with isorhapontigenin (2), piceatannol (3), and luteolin, were isolated by bioassay-guided fractionation from the seeds of Aiphanes aculeata Willd. (Arecaceae). The structure of compound 1 was elucidated by spectroscopic methods. Compound 1 is based on an unprecedented stilbenolignan skeleton in which a stilbene moiety is linked with a phenylpropane unit through a dioxane bridge. Compounds 1 and 2 exhibited significant inhibitory activities against cyclooxygenases-1 and -2.

Cyclooxygenase-inhibitory and antioxidant constituents of the aerial parts of Antirhea acutata.

Two new compounds, (6S)-hydroxy-29-nor-3,4-seco-cycloart-4(30),24-dien-3-oic acid (1) and 8-[1-(3,4-dihydroxyphenyl)-3-methoxy-3-oxopropyl]epicatechin (3), were isolated by bioassay-guided fractionation from the aerial parts of Antirhea acutata (DC.) Urb. (Rubiaceae). Compound 1 showed moderate inhibitory activities in cyclooxygenase-1 and -2 assays (IC(50) 43.7 and 4.7 microM, respectively), while compound 3 was active in 1,1-diphenyl-2-picrylhydrazyl free-radical and cytochrome c reduction antioxidant assays (IC(50) 29.1 and 16.3 microM, respectively). Additionally, one further new compound was isolated, (3S,24S)-25-trihydroxy-9,19-cycloartane-29-oic acid (2), but this was inactive in the bioassay systems used. Compound 1 is based on the unprecedented 29-nor-3,4-seco-cycloartane skeleton.

Flavonoids and phenylpropanoid derivatives from Campanula barbata.

Four new phenylpropanoid derivatives, barbatosides A-D, and a new catechin, barbatoflavan, were isolated from the whole plant of Campanula barbata L. (Campanulaceae) and identified as wahlenbergioside-3'-O-glucoside, wahlenbergioside-3'-O-(2'''-(p-methoxycinnamoyl))-glucoside, wahlenbergioside-3'-O-(4'''-(trans-p-coumaroyl))-glucoside, wahlenbergioside-3'-O-(4"'-(cis-p-coumaroyl))-glucoside and 3-acetyl-5-methoxy-7,3',4'-trihydroxy-8-O-glucoside-flavan-3-ol, respectively, by spectroscopic methods. In addition, four flavonols were isolated and identified as kaempferol-3-O-glucoside, kaempferol-3-O-rutinoside, quercetin-3-O-glucoside and quercetin-3-O-rutinoside. Barbatoflavan demonstrated scavenging properties towards the DPPH radical.

Two new stilbene dimer glucosides from grape (Vitis vinifera) cell cultures.

Two new stilbene dimer glucosides, resveratrol (E)-dehydrodimer 11-O-beta-D-glucopyranoside (1) and resveratrol (E)-dehydrodimer 11'-O-beta-D-glucopyranoside (2), were isolated together with the known resveratrol (E)-dehydrodimer (3) and pallidol (4) from Vitis vinifera cell cultures. The structures and stereochemistry of the new compounds were determined on the basis of spectroscopic data analysis. Compounds 1 and 2 are dimers that belong to a new type of oligostilbene formed from a resveratrol unit and a resveratrol glucoside unit. Compounds 1 and 3 exhibited nonspecific inhibitory activity against cyclooxygenase-1 and -2, with IC(50) values in the range of 5 microM, whereas compound 4 was approximately 10-fold less active.

Potential cancer-chemopreventive activities of wine stilbenoids and flavans extracted from grape (Vitis vinifera) cell cultures.

Moderate consumption of wine is associated with a reduced risk of cancer. Grape plant cell cultures were used to purify 12 phenols: the stilbenoids trans-astringin, trans-piceid (2), trans-resveratroloside, trans-resveratrol, trans-piceatannol, cis-resveratroloside, cis-piceid, and cis-resveratrol; the flavans (+)-catechin, (-)-epicatechin, and epicatechin 3-O-gallate; and the flavan dimer procyanidin B2 3'-O-gallate. These compounds were evaluated for potential to inhibit cyclooxygenases and preneoplastic lesion formation in carcinogen-treated mouse mammary glands in organ culture. At 10 micrograms/ml, trans-astringin and trans-piceatannol inhibited development of 7,12-dimethylbenz[a]anthracene-induced preneoplastic lesions in mouse mammary glands with 68.8% and 76.9% inhibition, respectively, compared with untreated glands. The latter compound was the most potent of the 12 compounds tested in this assay, with the exception of trans-resveratrol (87.5% inhibition). In the cyclooxygenase (COX)-1 assay, trans isomers of the stilbenoids appear to be more active than cis isomers: trans-resveratrol [50% inhibitory concentration (IC50) = 14.9 microM, 96%] vs. cis-resveratrol (IC50 = 55.4 microM). In the COX-2 assay, among the compounds tested, only trans- and cis-resveratrol exhibited significant inhibitory activity (IC50 = 32.2 and 50.2 microM, respectively). This is the first report showing the potential cancer-chemopreventive activity of trans-astringin, a plant stilbenoid recently found in wine. trans-Astringin and its aglycone trans-piceatannol were active in the mouse mammary gland organ culture assay but did not exhibit activity in COX-1 and COX-2 assays. trans-Resveratrol was active in all three of the bioassays used in this investigation. These findings suggest that trans-astringin and trans-piceatannol may function as potential cancer-chemopreventive agents by a mechanism different from that of trans-resveratrol.

A stilbene and dihydrochalcones with radical scavenging activities from Loiseleuria procumbens.

A new dihydrochalcone, 6''-acetylphloridzosid, was isolated from the whole plant of Loiseleuria procumbens (L.) Desv. and identified as 2'-O-(6''-O-acetylglucopyranosyl)-4,4',6'-trihydroxydihydrochal cone by spectroscopic methods. In addition, one stilbene and three other dihydrochalcones were identified as (E)-piceid, phloretin (2',4,4',6'-tetrahydroxydihydrochalcone), phloridzosid (2'-O-glucopyranosyl-4,4',6'-trihydroxydihydrochalcone) and asebotin (2'-O-glucopyranosyl-4'-methoxy-4,6'-dihydroxydihydrochalcone), respectively. Some of these compounds showed scavenging properties towards the 2,2-diphenyl-1-picrylhydrazyl radical and antioxidant properties in a test with lysozyme.

The role of cyclooxygenase and lipoxygenase in cancer chemoprevention.

The involvement of prostaglandins (PGs) and other eicosanoids in the development of human cancer has been known for over two decades. Importantly, an increase in PG synthesis may influence tumor growth in human beings and experimental animals, and numerous studies have illustrated the effect of PG synthesis on carcinogen metabolism, tumor cell proliferation and metastatic potential. PGs produced by cyclooxygenases (COXs) are represented by a large series of compounds that mainly enhance cancer development and progression, acting as carcinogens or tumor promoters, with profound effects on carcinogenesis. Further investigations suggest that arachidonic acid (AA) metabolites derived from lipoxygenase (LOX) pathways play an important role in growth-related signal transduction, implying that intervention through these pathways should be useful for arresting cancer progression. We discuss here the implications of COX and LOX in colon, pancreatic, breast, prostate, lung, skin, urinary bladder and liver cancers. Select inhibitors of COX and LOX are described, including nonsteroidal antiinflammatory drugs (NSAIDs), selective COX-2 inhibitors, curcumin, tea, silymarin and resveratrol, as well as a method useful for evaluating inhibitors of COX. Although a substantial amount of additional work is required to yield a better understanding of the role of COX and LOX in cancer chemoprevention, it is clear that beneficial therapeutic effects can be realized through drug-mediated modulation of these metabolic pathways.