Accelerating the speed of innovative anti-tumor drugs to first-in-human trials incorporating key de-risk strategies.

Pharmaceutical companies have recently focused on accelerating the timeline for initiating first-in-human (FIH) trials to allow quick assessment of biologic drugs. For example, a stable cell pool can be used to produce materials for the toxicology (Tox) study, reducing time to the clinic by 4-5 months. During the coronavirus disease 2019 (COVID-19) pandemic, the anti-COVID drugs timeline from DNA transfection to the clinical stage was decreased to 6 months using a stable pool to generate a clinical drug substrate (DS) with limited stability, virus clearance, and Tox study package. However, a lean chemistry, manufacturing, and controls (CMC) package raises safety and comparability risks and may leave extra work in the late-stage development and commercialization phase. In addition, whether these accelerated COVID-19 drug development strategies can be applied to non-COVID projects and established as a standard practice in biologics development is uncertain. Here, we present a case study of a novel anti-tumor drug in which application of "fast-to-FIH" approaches in combination with BeiGene's de-risk strategy achieved successful delivery of a complete CMC package within 10 months. A comprehensive comparability study demonstrated that the DS generated from a stable pool and a single-cell-derived master cell bank were highly comparable with regards to process performance, product quality, and potency. This accomplishment can be a blueprint for non-COVID drug programs that approach the pace of drug development during the pandemic, with no adverse impact on the safety, quality, and late-stage development of biologics.

Toxoplasma TgAtg8-TgAtg3 Interaction Primarily Contributes to Apicoplast Inheritance and Parasite Growth in Tachyzoite.

The apicoplast, which harbors key pathways involved in biosynthesis of vital metabolites, is a unique and essential nonphotosynthetic plastid organelle in apicomplexan parasites. Intriguingly, autophagy-related protein 8 (Atg8), a highly conserved eukaryotic protein, can localize to the outermost membrane of the apicoplast and modulate its inheritance in both Toxoplasma and Plasmodium parasites. The Atg8-Atg3 interaction plays a key role in Atg8 lipidation and localization, and our previously work in Toxoplasma has suggested that the core Atg8-family interacting motif (AIM) in TgAtg3, 239FADI242, and the R27 residue of TgAtg8 contribute to TgAtg8-TgAtg3 interaction in vitro. However, little is known about the function of this interaction or its importance in tachyzoite growth in Toxoplasma gondii. Here, we generated two complemented cell lines, TgAtg3F239A/I242A and TgAtg8R27E, based on the TgAtg3 and TgAtg8 conditional knockdown cell lines, respectively. We found that both mutant complemented cell lines were severely affected in terms of tachyzoite growth and displayed delayed death upon conditional knockdown of endogenous TgAtg3 or TgAtg8. Intriguingly, both complemented lines appeared to be defective in TgAtg8 lipidation and apicoplast inheritance. Moreover, we showed that the interaction of TgAtg8 and TgAtg3 is critical for TgAtg8 apicoplast localization. In addition, we found that the TgAtg3F239A/I242A complemented line exhibits an integral mitochondrial network upon ablation of endogenous TgAtg3, which is distinct from TgAtg3-depleted parasites with a fragmented mitochondrial network. Taken together, this work solidifies the contribution of the TgAtg8-TgAtg3 interaction to apicoplast inheritance and the growth of T. gondii tachyzoites. IMPORTANCEToxoplasma gondiiis a widespread intracellular parasite infecting a variety of warm-blooded animals, including humans. Current frontline treatment of toxoplasmosis suffers many drawbacks, including toxicity, drug resistance, and failure to eradicate tissue cysts, underscoring the need to identify novel drug targets for suppression or treatment of toxoplasmosis. TgAtg8 is thought to serve multiple functions in lipidation and is considered essential to the growth and development of both tachyzoites and bradyzoites. Here, we show that Toxoplasma gondii has adapted a conserved Atg8-Atg3 interaction, required for canonical autophagy in other eukaryotes, to function specifically in apicoplast inheritance. Our finding not only highlights the importance of TgAtg8-TgAtg3 interaction in tachyzoite growth but also suggests that this interaction is a promising drug target for the therapy of toxoplasmosis.

Conjugation of DM1 to anti-CD30 antibody has potential antitumor activity in CD30-positive hematological malignancies with lower systemic toxicity.

An anti-CD30 antibody-drug conjugate incorporating the antimitotic agent DM1 and a stable SMCC linker, anti-CD30-MCC-DM1, was generated as a new antitumor drug candidate for CD30-positive hematological malignancies. Here, the in vitro and in vivo pharmacologic activities of anti-CD30-MCC-DM1 (also known as F0002-ADC) were evaluated and compared with ADCETRIS (brentuximab vedotin). Pharmacokinetics (PK) and the safety profiles in cynomolgus monkeys were assessed. Anti-CD30-MCC-DM1 was effective in in vitro cell death assays using CD30-positive lymphoma cell lines. We studied the properties of anti-CD30-MCC-DM1, including binding, internalization, drug release and actions. Unlike ADCETRIS, anti-CD30-MCC-DM1 did not cause a bystander effect in this study. In vivo, anti-CD30-MCC-DM1 was found to be capable of inducing tumor regression in subcutaneous inoculation of Karpas 299 (anaplastic large cell lymphoma), HH (cutaneous T-cell lymphoma) and L428 (Hodgkin's disease) cell models. The half-lives of 4 mg/kg and 12 mg/kg anti-CD30-MCC-DM1 were about 5 days in cynomolgus monkeys, and the tolerated dose was 30 mg/kg in non-human primates, supporting the tolerance of anti-CD30-MCC-DM1 in humans. These results suggest that anti-CD30-MCC-DM1 presents efficacy, safety and PK profiles that support its use as a valuable treatment for CD30-positive hematological malignancies.

Structural diversity of anti-pancreatic cancer capsimycins identified in mangrove-derived Streptomyces xiamenensis 318 and post-modification via a novel cytochrome P450 monooxygenase.

Polycyclic tetramate macrolactams (PTMs) were identified as distinct secondary metabolites of the mangrove-derived Streptomyces xiamenensis 318. Together with three known compounds-ikarugamycin (1), capsimycin (2) and capsimycin B (3)-two new compounds, capsimycin C (4) with trans-diols and capsimycin D (5) with trans-configurations at C-13/C-14, have been identified. The absolute configurations of the tert/tert-diols moiety was determined in 4 by NMR spectroscopic analysis, CD spectral comparisons and semi-synthetic method. The post-modification mechanism of the carbocyclic ring at C-14/C-13 of compound 1 in the biosynthesis of an important intermediate 3 was investigated. A putative cytochrome P450 superfamily gene, SXIM_40690 (ikaD), which was proximally localized to the ikarugamycin biosynthetic pathway, was characterized. In vivo gene inactivation and complementation experiment confirmed that IkaD catalysed the epoxide-ring formation reaction and further hydroxylation of ethyl side chain to form capsimycin G (3'). Binding affinities and kinetic parameters for the interactions between ikarugamycin (1) and capsimycin B (3) with IkaD were measured with Surface Plasmon Resonance. The intermediate compound 3' was isolated and identified as 30-hydroxyl-capsimycin B. The caspimycins 2 and 3, were transferred to methoxyl derivatives, 6 and 7, under acidic and heating conditions. Compounds 1-3 exhibited anti-proliferative activities against pancreatic carcinoma with IC50 values of 1.30-3.37 µM.

Endogenous network states predict gain or loss of functions for genetic mutations in hepatocellular carcinoma.

Cancers have been typically characterized by genetic mutations. Patterns of such mutations have traditionally been analysed by posteriori statistical association approaches. One may ponder the possibility of a priori determination of any mutation regularity. Here by exploring biological processes implied in a mechanistic theory recently developed (the endogenous molecular-cellular network theory), we found that the features of genetic mutations in cancers may be predicted without any prior knowledge of mutation propensities. With hepatocellular carcinoma (HCC) as an example, we found that the normal hepatocyte and cancerous hepatocyte can be represented by robust stable states of one single endogenous network. These stable states, specified by distinct patterns of expressions or activities of proteins in the network, provide means to directly identify a set of most probable genetic mutations and their effects in HCC. As the key proteins and main interactions in the network are conserved through cell types in an organism, similar mutational features may also be found in other cancers. This analysis yielded straightforward and testable predictions on accumulated and preferred mutation spectra in normal tissue. The validation of predicted cancer state mutation patterns demonstrates the usefulness and potential of a causal dynamical framework to understand and predict genetic mutations in cancer.