Old and Young Actors Playing Novel Roles in the Drama of Multiple Myeloma Bone Marrow Microenvironment Dependent Drug Resistance.

Multiple myeloma (MM) is the second most frequent hematologic cancer. In addition to the deleterious effects of neoplastic plasma cell growth and spreading during the disease evolution, this tumor is characterized by the serious pathological consequences due to the massive secretion of monoclonal immunoglobulins and by the derangement of bone physiology with progressive weakening of the skeleton. Despite significant progresses having been made in the last two decades in the therapeutic management of this plasma cell tumor, MM remains invariably lethal, due to its extremely complex genetic architecture and to the constant protection it receives from the tumor niche, which is represented by the bone marrow microenvironment. While it is predictable that the discovery of novel therapies against the first of these two pathobiological features will take a longer time, the identification of the cellular and molecular mechanisms underlying the pro-growth effects of the myeloma milieu is a task that could lead to the development of novel treatments in a shorter timeframe. In this regard, aside from known "old" determinants of the cross-talk between bone marrow and MM cells, "young" cellular and molecular factors are now emerging, taking the scene of this complex neoplastic setting. In this review we aimed at giving insights on the latest evidence of potentially-targetable modes that MM cells exploit to increase fitness and gain a survival advantage. The benefits coming from the derangements of stress-managing pathways, autophagy, transcriptional rewiring, and non-coding RNAs are examples of such methods that MM cells utilize to escape cell death, but that hopefully will offer novel targets for the ever-increasing anti-MM therapeutic armamentarium.

Molecular Investigation of Recent Canine Parvovirus-2 (CPV-2) in Italy Revealed Distinct Clustering.

Canine parvovirus Type 2 (CPV-2) is a worldwide distributed virus considered the major cause of viral gastroenteritis in dogs. Studies on Italian CPV-2 are restricted to viruses circulating until 2017. Only one study provided more updated information on CPV-2 but was limited to the Sicily region. No information regarding the circulation and genetic characteristics of CPV-2 in Northeast Italy has been made available since 2015. The present study investigated the genetic characteristics of CPV-2 circulating in the dog population of Northeast Italy between 2013 and 2019. The VP2 gene of 67 CPV-2 was sequenced, and phylogenetic analysis was performed to identify patterns of distribution. Phylogenetic and molecular analysis highlighted unique characteristics of Northeast Italian CPV-2 and interestingly depicted typical genetic clustering of the Italian CPV-2 strains, showing the existence of distinct CPV-2 genetic groups. Such analysis provided insights into the origin of some Italian CPV-2 genetic clusters, revealing potential introductions from East European countries and the spread of CPV-2 from South/Central to North Italy. This is the first report that describes the genetic characteristics of recent Italian CPV-2. Tracking the genetic characteristics of CPV-2 nationally and globally may have impact on understanding the evolution and distribution of CPV-2, in particular in light of the current humanitarian emergency involving Ukraine, with the massive and uncontrolled movement of people and pet animals.

Unexpected Genetic Diversity of Two Novel Swine MRVs in Italy.

Mammalian Orthoreoviruses (MRV) are segmented dsRNA viruses in the family Reoviridae. MRVs infect mammals and cause asymptomatic respiratory, gastro-enteric and, rarely, encephalic infections. MRVs are divided into at least three serotypes: MRV1, MRV2 and MRV3. In Europe, swine MRV (swMRV) was first isolated in Austria in 1998 and subsequently reported more than fifteen years later in Italy. In the present study, we characterized two novel reassortant swMRVs identified in one same Italian farm over two years. The two viruses shared the same genetic backbone but showed evidence of reassortment in the S1, S4, M2 segments and were therefore classified into two serotypes: MRV3 in 2016 and MRV2 in 2018. A genetic relation to pig, bat and human MRVs and other unknown sources was identified. A considerable genetic diversity was observed in the Italian MRV3 and MRV2 compared to other available swMRVs. The S1 protein presented unique amino acid signatures in both swMRVs, with unexpected frequencies for MRV2. The remaining genes formed distinct and novel genetic groups that revealed a geographically related evolution of swMRVs in Italy. This is the first report of the complete molecular characterization of novel reassortant swMRVs in Italy and Europe, which suggests a greater genetic diversity of swMRVs never identified before.

Prosurvival autophagy is regulated by protein kinase CK1 alpha in multiple myeloma.

Multiple myeloma (MM) is a tumor of plasma cells (PCs). Due to the intense immunoglobulin secretion, PCs are prone to endoplasmic reticulum stress and activate several stress-managing pathways, including autophagy. Indeed, autophagy deregulation is maladaptive for MM cells, resulting in cell death. CK1α, a pro-survival kinase in MM, has recently been involved as a regulator of the autophagic flux and of the transcriptional competence of the autophagy-related transcription factor FOXO3a in several cancers. In this study, we investigated the role of CK1α in autophagy in MM. To study the autophagic flux we generated clones of MM cell lines expressing the mCherry-eGFP-LC3B fusion protein. We observed that CK1 inhibition with the chemical ATP-competitive CK1 α/δ inhibitor D4476 resulted in an impaired autophagic flux, likely due to an alteration of lysosomes acidification. However, D4476 caused the accumulation of the transcription factor FOXO3a in the nucleus, and this was paralleled by the upregulation of mRNA coding for autophagic genes. Surprisingly, silencing of CK1α by RNA interference triggered the autophagic flux. However, FOXO3a did not shuttle into the nucleus and the transcription of autophagy-related FOXO3a-dependent genes was not observed. Thus, while the chemical inhibition with the dual CK1α/δ inhibitor D4476 induced cell death as a consequence of an accumulation of ineffective autophagic vesicles, on the opposite, CK1α silencing, although it also determined apoptosis, triggered a full activation of the early autophagic flux, which was then not supported by the upregulation of autophagic genes. Taken together, our results indicate that the family of CK1 kinases may profoundly influence MM cells survival also through the modulation of the autophagic pathway.

Correction to: Role of protein kinases CK1α and CK2 in multiple myeloma: regulation of pivotal survival and stress-managing pathways.

The original article [1] contains an inadvertent error in the following sentence in the Abstract regarding the erroneous description of Ser/Thr kinases as 'phylogenetically related'.

The small GTPase RhoU lays downstream of JAK/STAT signaling and mediates cell migration in multiple myeloma.

Multiple myeloma is a post-germinal center B-cell neoplasm, characterized by the proliferation of malignant bone marrow plasma cells, whose survival and proliferation is sustained by growth factors and cytokines present in the bone marrow microenvironment. Among them, IL-6 triggers the signal downstream of its receptor, leading to the activation of the JAK/STAT pathway. The atypical GTPase RhoU lays downstream of STAT3 transcription factor and could be responsible for mediating its effects on cytoskeleton dynamics. Here we demonstrate that RHOU is heterogeneously expressed in primary multiple myeloma cells and significantly modulated with disease progression. At the mRNA level, RHOU expression in myeloma patients correlated with the expression of STAT3 and its targets MIR21 and SOCS3. Also, IL-6 stimulation of human myeloma cell lines up-regulated RHOU through STAT3 activation. On the other hand, RhoU silencing led to a decrease in cell migration with the accumulation of actin stress fibers, together with a decrease in cyclin D2 expression and in cell cycle progression. Furthermore, we found that even though lenalidomide positively regulated RhoU expression leading to higher cell migration rates, it actually led to cell cycle arrest probably through a p21 dependent mechanism. Lenalidomide treatment in combination with RhoU silencing determined a loss of cytoskeletal organization inhibiting cell migration, and a further increase in the percentage of cells in a resting phase. These results unravel a role for RhoU not only in regulating the migratory features of malignant plasma cells, but also in controlling cell cycle progression.

Role of protein kinases CK1α and CK2 in multiple myeloma: regulation of pivotal survival and stress-managing pathways.

Multiple myeloma (MM) is a malignant tumor of transformed plasma cells. MM pathogenesis is a multistep process. This cancer can occur de novo (rarely) or it can develop from monoclonal gammopathy of undetermined significance (most of the cases). MM can be asymptomatic (smoldering myeloma) or clinically active. Malignant plasma cells exploit intrinsic and extrinsic bone marrow microenvironment-derived growth signals. Upregulation of stress-coping pathways is also instrumental to maintain MM cell growth. The phylogenetically related Ser/Thr kinases CSNK1A1 (CK1α) and CSNK2 (CK2) have recently gained a growing importance in hematologic malignancies arising both from precursors and from mature blood cells. In multiple myeloma, CK1α or CK2 sustain oncogenic cascades, such as the PI3K/AKT, JAK/STAT, and NF-κB, as well as propel stress-related signaling that help in coping with different noxae. Data also suggest that these kinases modulate the delivery of growth factors and cytokines from the bone marrow stroma. The "non-oncogene addiction" phenotype generated by the increased activity of CK1α and CK2 in multiple myeloma contributes to malignant plasma cell proliferation and survival and represents an Achilles' heel for the activity of small ATP competitive CK1α or CK2 inhibitors.

Inactivation of CK1α in multiple myeloma empowers drug cytotoxicity by affecting AKT and ß-catenin survival signaling pathways.

Recent evidence indicates that protein kinase CK1α may support the growth of multiple myeloma (MM) plasma cells. Here, by analyzing a large cohort of MM cases, we found that high CK1α mRNA levels are virtually associated with all MM patients. Moreover, we provided functional evidence that CK1α activity is essential for malignant plasma cell survival even in the protective niche generated by co-cultures with bone marrow stromal cells. We demonstrated that CK1α inactivation, while toxic for myeloma cells, is dispensable for the survival of healthy B lymphocytes and stromal cells. Disruption of CK1α function in myeloma cells resulted in decreased Mdm2, increased p53 and p21 and reduced expression of ß-catenin and AKT. These effects were mediated partially by p53 and caspase activity. Finally, we discovered that CK1α inactivation enhanced the cytotoxic effect of both bortezomib and lenalidomide. Overall, our study supports a role for CK1α as a potential therapeutic target in MM in combination with proteasome inhibitors and/or immunomodulatory drugs.