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Chromosome; Cytogenetics; Myelodysplastic syndromes Cromosoma; Citogenética; Síndromes mielodisplásicos Cromosoma; Citogenètica; Síndromes mielodisplàsics Half of the myelodysplastic syndromes (MDS) have normal karyotype by conventional banding analysis. The percentage of true normal karyotype cases can be reduced by 20–30% with the complementary application of genomic microarrays. We here present a multicenter collaborative study of 163 MDS cases with a normal karyotype (≥10 metaphases) at diagnosis. All cases were analyzed with the ThermoFisher® microarray (either SNP 6.0 or CytoScan HD) for the identification of both copy number alteration(CNA) and regions of homozygosity (ROH). Our series supports that 25 Mb cut-off as having the most prognostic impact, even after adjustment by IPSS-R. This study highlights the importance of microarrays in MDS patients, to detect CNAs and especially to detect acquired ROH which has demonstrated a high prognostic impact. 2017 SGR288 (GRC) and CERCA Programme/Generalitat de Catalunya. Fundació Internacional Josep Carreras; Italy Ministry of Health.

In this study we want to evaluate the efficacy of yoga practice on dysfunctional stress, inflammation and QOL in breast cancer patients undergoing adjuvant radiotherapy.Patients with stage 0 to III breast cancer were recruited before starting radiotherapy (XRT) and were randomly assigned to yoga group (YG) two times a week during XRT or control group (CG). Self-report measures of QOL, fatigue and sleep quality, and blood samples were collected at day 1 of treatment, day 15, end of treatment and 1, 3 and 6 months later. Cortisol blood level, IL6, IL10, IL1RA, TNFα and lymphocyte-to-monocyte ratio were analyzed as measures of dysfunctional stress and inflammation.Patients started XRT and yoga classes in October 2019. Due to COVID-19 pandemic we closed the enrollment in March 2020. We analysed 24 patients, 12 YG and 12 CG. The analysis of blood cortisol levels revealed an interaction (p = 0.04) between yoga practice and time, in particular YG had lower cortisol levels at the end of XRT respect to CG (pTo our knowledge, this is the first study to evaluate the effects of yoga in a cancer population studying inflammation markers, cortisol trend and QOL during and until 6 months after XRT. This study suggests that yoga practice is able to reduce stress and inflammation levels over time. Besides including a larger number of patients to increase the power, future studies should consider other inflammatory or pro inflammatory factors and long-term yoga program to gain more evidence on yoga practice benefits.

Introduction: Three-dimensional (3D) multicellular spheroids are fundamental in vitro tools for studying in vivo tissues. Volume is the main feature used for evaluating the drug/treatment effects, but several other features can be estimated even from a simple 2D image. For high-content screening analysis, the bottleneck is the segmentation stage, which is essential for detecting the spheroids in the images and then proceeding to the feature extraction stage for performing morphotypic analysis. Problem: Today, several tools are available for extracting morphological features from spheroid images, but all of them have pros and cons and there is no general validated solution. Thanks to new deep learning models, it is possible to standardize the process and adapt the analysis to big data. Novelty: Starting from the first version of AnaSP, an open-source software suitable for estimating several morphological features of 3D spheroids, we implemented a new module for automatically segmenting 2D brightfield images of spheroids by exploiting convolutional neural networks. Results: Several deep learning segmentation models (i.e., VVG16, VGG19, ResNet18, ResNet50) have been trained and compared. All of them obtained very interesting results and ResNet18 ranked as the best-performing. Conclusions: A network based on an 18-layer deep residual architecture (ResNet-18) has been integrated into AnaSP, releasing AnaSP 2.0, a version of the tool optimized for high-content screening analysis. The source code, standalone versions, user manual, sample images, video tutorial, and further documentation are freely available at: https://sourceforge.net/p/anasp .

Supplementary Figures S1-S4. Suppplementary S1: Western Blot analysis o study the effect of the combinatory therapy on DDR Supplementary S2: Evaluation of the percentage of hyper damaged cells among the leukemic compartment Supplementary S3: Cell cycle analysis on OCI-AML2 treated cells Supplementary S4: Model of the mechanism of action of the combinatory therapy PARPi and 5FU

The existing treatments to cure acute leukemias seem to be nonspecific and suboptimal for most patients, drawing attention to the need of new therapeutic strategies. In the last decade the anticancer potential of poly ADP-ribose polymerase (PARP) inhibitors became apparent and now several PARP inhibitors are being developed to treat various malignancies. So far, the usage of PARP inhibitors has been mainly focused on the treatment of solid tumors and not too much about their efficacy on leukemias is known. In this study we test, for the first time on leukemic cells, a combined therapy that associates the conventional chemotherapeutic agent fluorouracil (5FU), used as a source of DNA damage, and a PARP inhibitor, rucaparib. We demonstrate the efficacy and the specificity of this combined therapy in killing both acute myeloid leukemia and acute lymphoid leukemia cells in vitro and in vivo. We clearly show that the inhibition of DNA repair induced by rucaparib is synthetic lethal with the DNA damage caused by 5FU in leukemic cells. Therefore, we propose a new therapeutic strategy able to enhance the cytotoxic effect of DNA-damaging agents in leukemia cells via inhibiting the repair of damaged DNA. Mol Cancer Ther; 14(4); 889–98. ©2015 AACR.

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The following information describes additional study design criteria and safety considerations used in the phase 1 clinical trial of RG7112 in patients with hematological malignancies

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Figure S1. Hematologic recovery after NK cell infusion in all treated patients.

Purpose:The stromal and immune bone marrow (BM) landscape is emerging as a crucial determinant for acute myeloid leukemia (AML). Regulatory T cells (Treg) are enriched in the AML microenvironment, but the underlying mechanisms are poorly elucidated. Here, we addressed the effect of IFNγ released by AML cells in BM Treg induction and its impact on AML prognosis.Experimental Design:BM aspirates from patients with AML were subdivided according to IFNG expression. Gene expression profiles in INFγhigh and IFNγlow samples were compared by microarray and NanoString analysis and used to compute a prognostic index. The IFNγ release effect on the BM microenvironment was investigated in mesenchymal stromal cell (MSC)/AML cell cocultures. In mice, AML cells silenced for ifng expression were injected intrabone.Results:IFNγhigh AML samples showed an upregulation of inflammatory genes, usually correlated with a good prognosis in cancer. In contrast, in patients with AML, high IFNG expression was associated with poor overall survival. Notably, IFNγ release by AML cells positively correlated with a higher BM suppressive Treg frequency. In coculture experiments, IFNγhigh AML cells modified MSC transcriptome by upregulating IFNγ-dependent genes related to Treg induction, including indoleamine 2,3-dioxygenase 1 (IDO1). IDO1 inhibitor abrogated the effect of IFNγ release by AML cells on MSC-derived Treg induction. In vivo, the genetic ablation of IFNγ production by AML cells reduced MSC IDO1 expression and Treg infiltration, hindering AML engraftment.Conclusions:IFNγ release by AML cells induces an immune-regulatory program in MSCs and remodels BM immunologic landscape toward Treg induction, contributing to an immunotolerant microenvironment.See related commentary by Ferrell and Kordasti, p. 2986

Somatic variants in TET2 and DNMT3A are founding mutations in hematological malignancies that affect the epigenetic regulation of DNA methylation. Mutations in both genes often co-occur with activating mutations in genes encoding oncogenic tyrosine kinases such as FLT3ITD, BCR-ABL1, JAK2V617F, and MPLW515L, or with mutations affecting related signaling pathways such as NRASG12D and CALRdel52. Here, we show that TET2 and DNMT3A mutations exert divergent roles in regulating DNA repair activities in leukemia cells expressing these oncogenes. Malignant TET2-deficient cells displayed downregulation of BRCA1 and LIG4, resulting in reduced activity of BRCA1/2-mediated homologous recombination (HR) and DNA-PK–mediated non-homologous end-joining (D-NHEJ), respectively. TET2-deficient cells relied on PARP1-mediated alternative NHEJ (Alt-NHEJ) for protection from the toxic effects of spontaneous and drug-induced DNA double-strand breaks. Conversely, DNMT3A-deficient cells favored HR/D-NHEJ owing to downregulation of PARP1 and reduction of Alt-NHEJ. Consequently, malignant TET2-deficient cells were sensitive to PARP inhibitor (PARPi) treatment in vitro and in vivo, whereas DNMT3A-deficient cells were resistant. Disruption of TET2 dioxygenase activity or TET2—Wilms' tumor 1 (WT1)–binding ability was responsible for DNA repair defects and sensitivity to PARPi associated with TET2 deficiency. Moreover, mutation or deletion of WT1 mimicked the effect of TET2 mutation on DSB repair activity and sensitivity to PARPi. Collectively, these findings reveal that TET2 and WT1 mutations may serve as biomarkers of synthetic lethality triggered by PARPi, which should be explored therapeutically.Significance:TET2 and DNMT3A mutations affect distinct DNA repair mechanisms and govern the differential sensitivities of oncogenic tyrosine kinase–positive malignant hematopoietic cells to PARP inhibitors.

Supplementary Figure S2 - PDF file 145K, Representation from Chromosome Analysis Suite (Chas, Affymetrix) of LOH alterations in analyzed MDS patients.The figure represents the LOH of the 5 patients with this abnormality and shows its distribution and its size within the chromosomes

Philadelphia chromosome–positive acute lymphoblastic leukemia (Ph+ ALL) is currently treated with BCR-ABL1 tyrosine kinase inhibitors (TKI) in combination with chemotherapy. However, most patients develop resistance to TKI through BCR-ABL1–dependent and –independent mechanisms. Newly developed TKI can target Ph+ ALL cells with BCR-ABL1–dependent resistance; however, overcoming BCR-ABL1–independent mechanisms of resistance remains challenging because transcription factors, which are difficult to inhibit, are often involved. We show here that (i) the growth of Ph+ ALL cell lines and primary cells is highly dependent on MYB-mediated transcriptional upregulation of CDK6, cyclin D3, and BCL2, and (ii) restoring their expression in MYB-silenced Ph+ ALL cells rescues their impaired proliferation and survival. Levels of MYB and CDK6 were highly correlated in adult Ph+ ALL (P = 0.00008). Moreover, Ph+ ALL cells exhibited a specific requirement for CDK6 but not CDK4 expression, most likely because, in these cells, CDK6 was predominantly localized in the nucleus, whereas CDK4 was almost exclusively cytoplasmic. Consistent with their essential role in Ph+ ALL, pharmacologic inhibition of CDK6 and BCL2 markedly suppressed proliferation, colony formation, and survival of Ph+ ALL cells ex vivo and in mice. In summary, these findings provide a proof-of-principle, rational strategy to target the MYB "addiction" of Ph+ ALL.Significance: MYB blockade can suppress Philadelphia chromosome-positive leukemia in mice, suggesting that this therapeutic strategy may be useful in patients who develop resistance to imatinib and other TKIs used to treat this disease. Cancer Res; 78(4); 1097–109. ©2017 AACR.

Table S1. Percentage and absolute number of alloreactive NK cells in responders (A) and non-responders (B) Table S2. Description of cell processing results in responders (A) and non-responders (B)

Purpose: The 9p21 locus, encoding three important tumor suppressors (p16/CDKN2A, p14/ARF, and p15/CDKN2B), is a major target of inactivation in the pathogenesis of many human tumors.Patients and Methods: To explore, at high resolution, the frequency and size of alterations affecting this locus in adult BCR-ABL1–positive acute lymphoblastic leukemia (ALL) and to investigate their prognostic value, 112 patients (101 de novo and 11 relapsed cases) were analyzed by genome-wide single-nucleotide polymorphism arrays and gene candidate deep exon sequencing. Paired diagnosis–relapse samples were further available and analyzed for 19 (19%) cases.Results:CDKN2A/ARF and CDKN2B genomic alterations were identified in 29% and 25% of newly diagnosed patients, respectively. Deletions were monoallelic in 72% of cases, and in 43% of them, the minimal overlapping region of the lost area spanned only the CDKN2A/B gene locus. An analysis conducted at relapse showed an increase in the detection rate of CDKN2A/ARF loss (47%) compared with the time of diagnosis (P = 0.06). Point mutations within the 9p21 locus were found at very low levels, with only a nonsynonymous substitution in the exon 2 of CDKN2A. Of note, deletions of CDKN2A/B were significantly associated with poor outcomes in terms of overall survival (P = 0.0206), disease free-survival (P = 0.0010), and cumulative incidence of relapse (P = 0.0014).Conclusions: Inactivation of the 9p21 locus by genomic deletion is a frequent event in BCR-ABL1–positive ALL. Deletions are frequently acquired during leukemia progression and are a poor prognostic marker of long-term outcomes. Clin Cancer Res; 17(23); 7413–23. ©2011 AACR.

Supplementary Fig. 1. Day 10 dose-mean PK profiles in patients with leukemia; Supplementary Figure 2. RG7112 induces p53-mediated apoptosis in circulating lymphoma cells from a patient with SLL/CLL (810 mg to 1500 mg BID)

Purpose: This phase II prospective study aimed to evaluate the efficacy and safety of 5-days azacytidine (5d-AZA) in patients with low-risk myelodysplastic syndromes (MDS). Second, single-nucleotide polymorphism (SNP) genetic profile and phosphoinositide-phospholipase C (PI-PLC) β1 levels were studied to evaluate possible biologic markers able to predict the hematologic response.Experimental Design: The study tested a lower intensity schedule of azacytidine. The treatment plan consisted of 75 mg/sqm/d subcutaneous administered for 5 days every 28 days, for a total of 8 cycles.Results: Thirty-two patients were enrolled in the study. The overall response rate was 47% (15 of 32) on intention-to-treat and 58% (15 of 26) for patients completing the treatment program. In this latter group, 5 (19%) achieved complete remission (CR) and 10 (38%) had hematologic improvement, according to the International Working Group (IWG) criteria. Three patients have maintained their hematologic improvement after 37, 34, and 33 months without other treatments. Moreover, 21 and 2 of 26 cases completing 8 cycles were transfusion-dependent for red blood cells and platelets at baseline, respectively. Of these, 7 (33%) and 2 (100%) became transfusion-independent at the end of the treatment program, respectively. Grade 3–4 neutropenia occurred in 28% of patients and 4 patients died early due to infections or hemorrhage. SNP results were not significantly correlated to the clinical outcome, whereas PI-PLCβ1 level anticipated either positive or negative clinical responses.Conclusions: 5d-AZA is safe and effective in a proportion of patients with low-risk MDS. PI-PLCβ1 gene expression is a reliable and dynamic marker of response that can be useful to optimize azacytidine therapy. Clin Cancer Res; 19(12); 3297–308. ©2013 AACR.

Supplementary Methods and Results

List of primers and antibodies used.

Figure S3. Correlation between relapse rate and percentage of alloreactive clones at day 3 after infusion.

Figure S4. Impact of the absolute number of infused donor alloreactive NK cells on the probability of response.

Supplementary Table S1 - PDF file 69K, Summary of macroscopic CNAs and LOH alterations identified in MDS patients and response to fourth cycle of 5d-Aza. The table summarizes the results of SNP karyotyping, LOH and CNAs in the 26 MDS patients for whom the an

Supplementary Table 1. Study stratification and dosing regimen; Supplementary Table 2. p53 target genes examined in this study; Supplementary Table 3. Characteristics of evaluable patients treated at the MTD; Supplementary Table 4. Patient demographic data by cohort; Supplementary Table 5. Adverse events by grade observed in > 10% of the patients/stratum; Supplementary Table 6. Stratum pharmacokinetic parameters based on RG7112 dose regimen and formulation; Supplementary Table 7. Summary of steady state AUC for AML patients treated at the MTD; Supplementary Table 8. Clinical activity for patients with p53 mutations

S1. MYB silencing reduces viability and proliferation of Ph+ ALL cell lines; S2. Expression of shMYB-resistant MYB cDNA rescues the effects of MYB silencing; S3. MYB silencing suppresses leukemia in NSG mice injected with BV173 or SUP-B15 cells; S4. MYB silencing alters the expression of cell cycle regulatory genes in SUP-B15 cells; S5. Restoring CDK4 nuclear localization rescues the impaired proliferation of MYB-silenced BV173 cells; S6. CDK6, Cyclin D3 and BCL2 but not p21 cooperate to rescue the cell growth of MYB silenced BV173 cells; S7. MYB regulation of CDK6 but not of BCL2 is dependent on p300/CBP recruitment; S8. Palbociclib reduces Ph+ ALL proliferation in vitro and leukemia formation in vivo; S9. Venetoclax or Sabutoclax in combination with the CDK4/6 inhibitor palbociclib synergistically reduce the viability of Ph+ ALL cells.

Supplementary Data from Valproic Acid at Therapeutic Plasma Levels May Increase 5-Azacytidine Efficacy in Higher Risk Myelodysplastic Syndromes

Purpose: In acute myeloid leukemia (AML), alloreactive natural killer (NK) cells are crucial mediators of immune responses after haploidentical stem cell transplantation. Allogeneic NK cell infusions have been adoptively transferred with promising clinical results. We aimed at determining whether the composition of NK graft in terms of frequency of alloreactive NK cells influence the clinical response in a group of elderly AML patients undergoing NK immunotherapy.Experimental Design: Seventeen AML patients, in first complete remission (CR; median age 64 years, range 53–73) received NK cells from haploidentical KIR-ligand–mismatched donors after fludarabine/cyclophosphamide chemotherapy, followed by IL2. To correlate donor NK cell activity with clinical response, donor NK cells were assessed before and after infusion.Results: Toxicity was moderate, although 1 patient died due to bacterial pneumonia and was censored for clinical follow-up. With a median follow-up of 22.5 months (range, 6–68 months), 9 of 16 evaluable patients (0.56) are alive disease-free, whereas 7 of 16 (0.44) relapsed with a median time to relapse of 9 months (range, 3–51 months). All patients treated with molecular disease achieved molecular CR. A significantly higher number of donor alloreactive NK cell clones was observed in responders over nonresponders. The infusion of higher number of alloreactive NK cells was associated with prolonged disease-free survival (0.81 vs. 0.14, respectively; P = 0.03).Conclusions: Infusion of purified NK cells is feasible in elderly AML patients as post-CR consolidation strategy. The clinical efficacy of adoptively transferred haploidentical NK cells may be improved by infusing high numbers of alloreactive NK cells. Clin Cancer Res; 22(8); 1914–21. ©2016 AACR.See related commentary by Muntasell and López-Botet, p. 1831

Purpose: RG7112 is a small-molecule MDM2 antagonist. MDM2 is a negative regulator of the tumor suppressor p53 and frequently overexpressed in leukemias. Thus, a phase I study of RG7112 in patients with hematologic malignancies was conducted.Experimental Design: Primary study objectives included determination of the dose and safety profile of RG7112. Secondary objectives included evaluation of pharmacokinetics; pharmacodynamics, such as TP53-mutation status and MDM2 expression; and preliminary clinical activity. Patients were divided into two cohorts: Stratum A [relapsed/refractory acute myeloid leukemia (AML; except acute promyelocytic leukemia), acute lymphoblastic leukemia, and chronic myelogenous leukemia] and Stratum B (relapsed/refractory chronic lymphocytic leukemia/small cell lymphocytic leukemia; CLL/sCLL). Some Stratum A patients were treated at the MTD to assess clinical activity.Results: RG7112 was administered to 116 patients (96 patients in Stratum A and 20 patients in Stratum B). All patients experienced at least 1 adverse event, and 3 dose-limiting toxicities were reported. Pharmacokinetic analysis indicated that twice-daily dosing enhanced daily exposure. Antileukemia activity was observed in the 30 patients with AML assessed at the MTD, including 5 patients who met International Working Group (IWG) criteria for response. Exploratory analysis revealed TP53 mutations in 14% of Stratum A patients and in 40% of Stratum B patients. Two patients with TP53 mutations exhibited clinical activity. p53 target genes were induced only in TP53 wild-type leukemic cells. Baseline expression levels of MDM2 correlated positively with clinical response.Conclusions: RG7112 demonstrated clinical activity against relapsed/refractory AML and CLL/sCLL. MDM2 inhibition resulted in p53 stabilization and transcriptional activation of p53-target genes. We provide proof-of-concept that MDM2 inhibition restores p53 function and generates clinical responses in hematologic malignancies. Clin Cancer Res; 22(4); 868–76. ©2015 AACR.

Figure S2. Immunofluorescence analyses of 2 representative donor alloreactive NK cell clones expressing the KIR for which there is no class I ligand in the recipient as their only inhibitory receptor for self.

Supplementary Data from Release of IFNγ by Acute Myeloid Leukemia Cells Remodels Bone Marrow Immune Microenvironment by Inducing Regulatory T Cells

supplemental figure legend

Primary human Ph+ ALL samples information.

Supplementary Figure 1 from Synergistic Proapoptotic Activity of Recombinant TRAIL Plus the Akt Inhibitor Perifosine in Acute Myelogenous Leukemia Cells

Supplementary Table 2 from Protein Tyrosine Phosphatase Receptor Type γ Is a Functional Tumor Suppressor Gene Specifically Downregulated in Chronic Myeloid Leukemia

Chronic myelogenous leukemia (CML) is the most common myeloproliferative disease. Protein tyrosine phosphatase receptor type γ (PTPRG) is a tumor suppressor gene and a myeloid cell marker expressed by CD34+ cells. Downregulation of PTPRG increases colony formation in the PTPRG-positive megakaryocytic cell lines MEG-01 and LAMA-84 but has no effect in the PTPRG-negative cell lines K562 and KYO-1. Its overexpression has an oncosuppressive effect in all these cell lines and is associated with myeloid differentiation and inhibition of BCR/ABL-dependent signaling. The intracellular domain of PTPRG directly interacts with BCR/ABL and CRKL, but not with signal transducers and activators of transcription 5. PTPRG is downregulated at the mRNA and protein levels in leukocytes of CML patients in both peripheral blood and bone marrow, including CD34+ cells, and is reexpressed following molecular remission of disease. Reexpression was associated with a loss of methylation of a CpG island of PTPRG promoter occurring in 55% of the patients analyzed. In K562 cell line, the DNA hypomethylating agent 5-aza-2′-deoxycytidine induced PTPRG expression and caused an inhibition of colony formation, partially reverted by downregulation of PTPRG expression. These findings establish, for the first time, PTPRG as a tumor suppressor gene involved in the pathogenesis of CML, suggesting its use as a potential diagnostic and therapeutic target. Cancer Res; 70(21); 8896–906. ©2010 AACR.

Supplementary Table 1 from Protein Tyrosine Phosphatase Receptor Type γ Is a Functional Tumor Suppressor Gene Specifically Downregulated in Chronic Myeloid Leukemia

Supplementary Figure 2 from Synergistic Proapoptotic Activity of Recombinant TRAIL Plus the Akt Inhibitor Perifosine in Acute Myelogenous Leukemia Cells

Background Trastuzumab is the cornerstone of adjuvant therapy for HER2-positive breast cancer (BC), but due to de novo or primary resistance, >20% of early-stage patients become resistant. Thus, predicting the mechanisms of trastuzumab resistance would facilitate the planning of specific therapeutic strategies. We performed a case-control study of 26 HER2-positive BC patients who relapsed within 5 years of starting adjuvant treatment and 26 controls, no relapsed, analyzing an extensive gene expression profile. Methods Total RNA was isolated from formalin-fixed paraffin-embedded primary tumors with AllPrep DNA/RNA FFPE Kit (Qiagen) and quantified by Nanodrop, before performing Nanostring gene expression profiling. nCounter Breast Cancer 360 Panel (Nanostring Technologies) was used according to the manufacturer’s instructions to analyze the expression of 770 genes and important signatures for BC (e.g. PAM50). For the statistical analyses, genes were normalized using a ratio of the expression value to the geometric mean of all housekeeping genes on the panel. Housekeeper-normalized data were then log(2) transformed. These data were evaluated by unsupervised selection of genes using consensus clustering with the Partitioning Around Medoids algorithm followed by the association of cluster “representative” genes (i.e. the medoids) with respect to the presence of relapse. The optimal number of clusters was determined by inspecting the consensus matrix cumulative distribution. Given the small sample size, the association between the "representative" genes, demographic and clinical covariates and the risk of relapse was evaluated using a logistic regression model with ELASTIC-NET regularization with α and λ tuned by 5-fold cross-validation. Penalized regression coefficients β are reported. The analyses were performed using R version 4.0.4 and package “ConsensusClusterPlus” and “glmnet”. Results Patient clinical characteristics are reported in Table 1. Relapsed patients showed a higher tumor stage, larger tumor size and greater lymph node involvement than controls (Table 1). Six samples failed the quality control check and so the analyses on gene expression data were performed on 46 subjects who had a distribution of clinical covariates similar to that reported in Table 1. The PAM50 intrinsic subtype, Risk Of Recurrence (ROR) score and ROR category were not associated with relapse status (P = 0.5, 0.5 and 0.1, respectively). The consensus clustering analysis revealed that the optimal number of clusters was 5, and the corresponding medoids were: HDAC6 for cluster 1 (n1=192 genes), WNT4 for cluster 2 (n2=102 genes), BMPR2 for cluster 3 (n3=162 genes), PALB2 for cluster 4 (n4=191 genes), and PARP1 for cluster 5 (n5=84 genes). HDAC6 and BMPR2 showed the highest correlation (r = 0.53, P Table 1.Patient characteristics in relation to Trastuzumab outcome.VariableAllCasesControlsP(n=52)(n=26)(n=26)n(%)n(%)n(%)Age at start adiuvant therapyMean ± sd53.4 ± 11.953.1 ± 12.653.7 ± 11.40.854Menopausal StatusPre-menopause21(40.4)10(38.5)11(42.3)0.777Post-menopause31(59.6)16(61.5)15(57.7)Stage of DiseaseI9(18.8)2(8.0)7(30.4)0.001II20(41.7)7(28.0)13(56.5)III19(39.6)16(64.0)3(13.0)Unknown413Lymph Node StatusNegative18(36.0)4(16.7)14(53.9)0.008Positive32(64.0)20(83.3)12(46.2)Unknown220Tumor SizeT128(53.9)7(26.9)21(80.8) Citation Format: Sara Ravaioli, Roberta Maltoni, Elisabetta Petracci, Michela Palleschi, William Balzi, Francesca Pirini, Maria Maddalena Tumedei, Michele Zanoni, Michela Cortesi, Giovanni Martinelli, Andrea Rocca, Sara Bravaccini. HDAC6 is an unfavorable prognostic factor in HER2-positive breast cancer patients treated with adjuvant trastuzumab [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P2-13-44.

IntroductionNeuroendocrine neoplasms (NENs) are a rare group of tumors exceptionally heterogeneous, with clinical presentation ranging from well differentiated more indolent tumors to poorly differentiated very aggressive forms. Both are often diagnosed after the metastatic spread and require appropriate medical treatment. A high priority need in the management of this disease is the identification of effective therapeutic strategies for advanced and metastatic patients. The recent TALENT trial demonstrated the efficacy of lenvatinib, a multi-tyrosine kinase inhibitor, in patients with gastroenteropancreatic neuroendocrine tumors (GEP-NETs) with no other treatment indication. Further development of this drug in advanced NETs is warranted.MethodsWe investigated potential clinical and molecular determinants of lenvatinib response in human primary cultures derived from patients with GEP-NET of different grades and sites of origin. We correlated response to treatment with patient clinical characteristics, with the mutational status of 161-cancer associated genes and with the expression levels of MKI-related genes.ResultsLenvatinib exerted a significant antitumor activity in primary GEP-NET cells, with median survival inhibitions similar or higher than those of standard frontline treatments. Of the 11 primary cultures analyzed in our case series, 6 were classified as responder showing a significant survival inhibition, and 5 as non-responder. We observed that the overexpression of HRAS in the original tumor tissue compared to the matched healthy tissue significantly correlated with responsiveness of primary cells to lenvatinib (p=.048). All 5 non-responder cultures showed normal HRAS expression, while of the 6 responder cultures, 4 had HRAS overexpression. Overexpression of HRAS was not associated with gene mutation. None of the other evaluated clinical variables (grade, Ki67, site of origin and syndromic disease) or molecular markers correlated with response.DiscussionLenvatinib appears to be a highly effective drug for the treatment of NETs. The evaluation of HRAS expression in the tumor tissue might improve patient selection and optimize therapeutic outcome.

BACKGROUND According to Europe’s Beating Cancer Plan, the number of cancer survivors is growing every year and is now estimated at over 12 million in Europe. A main objective of the European Commission is to ensure that cancer survivors can enjoy a high quality of life, underlining the role of digital technology and eHealth apps and tools to achieve this. OBJECTIVE The main objective of this study is the development of a user-centered artificial intelligence system to facilitate the input and integration of patient-related biopsychosocial data to improve posttreatment quality of life, well-being, and health outcomes and examine the feasibility of this digitally assisted workflow in a real-life setting in patients with colorectal cancer and acute myeloid leukemia. METHODS A total of 60 patients with colorectal cancer and 30 patients with acute myeloid leukemia will be recruited from 2 clinical centers: Universitätsmedizin der Johannes Gutenberg-Universität Mainz (Mainz, Germany) and IRCCS Istituto Romagnolo per lo Studio dei Tumori “Dino Amadori” (IRST, Italy). Psychosocial data (eg, emotional distress, fatigue, quality of life, subjective well-being, sleep problems, and appetite loss) will be collected by questionnaires via a smartphone app, and physiological data (eg, heart rate, skin temperature, and movement through step count) will be collected by a customizable smart wrist-worn sensor device. Each patient will be assessed every 2 weeks over their 3-month participation in the ONCORELIEF study. Inclusion criteria include patients with the diagnosis of acute myeloid leukemia or colorectal cancer, adult patients aged 18 years and older, life expectancy greater than 12 months, Eastern Cooperative Oncology Group performance status ≤2, and patients who have a smartphone and agree to use it for the purpose of the study. Exclusion criteria include patients with a reduced cognitive function (such as dementia) or technological illiteracy and other known active malignant neoplastic diseases (patients with a medical history of treated neoplastic disease are included). RESULTS The pilot study started on September 1, 2022. As of January 2023, we enrolled 33 patients with colorectal cancer and 7 patients with acute myeloid leukemia. As of January 2023, we have not yet started the data analysis. We expect to get all data in June 2023 and expect the results to be published in the second semester of 2023. CONCLUSIONS Web-based and mobile apps use methods from mathematical decision support and artificial intelligence through a closed-loop workflow that connects health professionals and patients. The ONCORELIEF system has the potential of continuously identifying, collecting, and processing data from diverse patient dimensions to offer health care recommendations, support patients with cancer to address their unmet needs, and optimize survivorship care. CLINICALTRIAL German Clinical Trials Register (DRKS) 00027808; https://drks.de/search/en/trial/DRKS00027808 INTERNATIONAL REGISTERED REPORT DERR1-10.2196/45475