Alok Ranjan Singh

-Leading one of the oncology program, LNPs (active/passive) characterization, in-situ CAR, CAR-NK, CAR-T, tumor microenvironment, cold tumor to hot tumor, assay development (in-vitro/in-vivo functional assays), building tumor models, bio-distribution, toxicity, efficacy, and writing SOPs/Protocol/Technical reports. -Hand on experienced in Identification, Evaluation, and Management of outsourced services -Managing CROs and cross-functional team in 3 different programs: Study plan & Design (both in-vitro and in-vivo), data analysis, data validation, and making PowerPoint slides -Managing scientist/senior scientist in my group Skills: Managing CRO, program/project, cross-functional team and research associates/scientists. Tumor/tissues specific drug delivery (LNP based), expression of LNP-Luc-mRNA (in-vitro and in-vivo (tumors/other tissues)), Bio-distribution of LNPs-mRNA, and LNPs-siRNA. Payload expression (in-vitro/in-vivo using Bio-Glo/IVIS imaging). Target gene knockdown analysis using western blot/qPCR.

Managed a team of 2-3 Research Scientists, day-to-day planning, troubleshooting, SOP writing, project management and timelines prioritizations, and coaching of junior staff. Technical Skills: managed project , in-vivo study design (efficacy, toxicity, bio-distribution, PK analysis), Ex-vivo analysis (TILs), flowcytometry, cytokines analysis using MSD, tumor/tissue/blood/plasma collection, and data analysis. Tumor models: syngeneic, xenograft, pseudo-humanized and humanized

-Evaluated potential activity of BiTes/T cell engager in in-vitro (T cell activation/ proliferation assays and cytokines quantification using MSD), -Binding assay, plate-based reporter (NFkB, Luc, NFAT), ELISA assay, Ligand blocking assays, -Target cells killing/cytotoxicity assays (Promega based detection reagent, LDH, ADCC, degranulation assay, Excelligence based killing assay) -Phagocytosis, ADCP-flow based detection. -Evaluated potential synergy/combination effects of check point antibodies with costimulatory agonists in both in-vitro and in-vivo systems -SEB assay, Mixed lymphocyte reaction (MLR) assay, suppression assay -Ex-vivo analysis: TILs analysis using flowcytometry -Effectively coordinate project with cross-functional teams and CROs. -Generated KO cell lines by using CRISPR-Cas-9 technology -Managed research associates Skills: Developed mouse model for characterizing BiTes and co-stimulatory mAbs in in-vivo, T cell/PBMSc engraftment (in-vivo), PK analysis, and bio-distribution, immune cell based assays (for both NK and T cells) Route of injections: subcutaneous, intraperitoneal (IP), intravenous (IV), retro-orbital, intramuscular (IM) and oral gauge.

-Developed cell lines for reporter assays -Screening, validation and characterization of lead ICIs/co-stimulatory mAbs -I-O based assays using mouse and human-PBMCs (flow-based assays, Immune cells characterization) -Designed and evaluate the efficacy studies of lead costimulatory agonists and check point antibodies in syngeneic, pseudo-humanized and humanized mouse model -Dose titration, PK analysis studies of costimulatory agonists and check point antibodies In-vitro Skills: Developed cell lines using lentivirus for cell binding/reporter assays, PBMCs isolation, T cells/NK cells isolation, T/NK cells activation/proliferation, Intracellular cytokines staining, MSD, blocking assays, MLR assay SEB assays In-vivo Skills: tumor injection and treatment routes (subcutaneous, IV, IM, IP, retro-orbital, oral gauges), mouse handling, tumor/tissues (organs) harvest, blood collection (from retroorbital bleeding/tail), Ex-vivo TILs analysis using flowcytometry

-Investigated the preclinical efficacy of PI3Kinase inhibitors, PI3Kinase/BRD4 (SF1126/SF2523) and PI3Kinase/CDK4/6 dual inhibitors in targeting tumor and stromal compartment of HCC tumors o This study opened new avenues for treatment of patients with advanced HCC. -Efficacy study of SF1126 in treatment of grade 3 and grade 4 neuroblastoma tumors. o This study opened phase I clinical trial for the treatment of refractory neuroblastoma in 14 Children hospital in US and Canada. -Evaluated the effect of SF1126 inhibitor on innate and adaptive immune response. o Conversion of immune-suppressive tumor microenvironment to immune-stimulatory o Role of macrophages (M1 or M2/TAM) and other immune cells -Investigated role of Rac2-Syk signaling in CCL4-induced liver fibrosis. -Hand on experienced on lung fibrosis using bleomycin -Experienced on various cancer models including medulloblastoma, neuroblastoma, Ewing sarcoma, lymphoma, renal cell carcinoma, hepatocellular carcinoma, Head and neck squamous carcinoma and glioma. Skills: Isolation of immune cells (macrophage/T cells from mouse), Differentiation of macrophage into M1/M1 phenotype, adoptive transfer assay, flowcytometry, TILs analysis, Molecular Biology, cell Biology, In-vitro Assays, In-vivo (mouse Models).

-Generated patient derived xenografts (PDX) from medulloblastoma patient tumors -Investigated the functional role of PTEN/PI3K inhibitors in CD15+ cancer stem cells isolated from medulloblastoma mouse model and human patient derived xenografts. -Investigated the effect combination therapy (of PI3K inhibitors plus other kinase inhibitors) to provided new opportunity to treat medulloblastoma patients -Investigated and identified the role of PTEN and PI-3 Kinase inhibitors in LPS induced lymphoproliferation in B cell lymphoma model. -Discovered the role of hematopoietic specific Rac2 GTPase in inflammation and M2 macrophage differentiation and implication of this signaling axis in promoting bleomycin induced Idiopathic pulmonary fibrosis, liver fibrosis model. -Mentored, taught, and trained high school and rotating graduate students in basic biology, experimental design, data analysis, and career development. Skills: orthotopic brain surgery, pancreatic surgery, lung surgery, subcutaneous tumor injection, Isolation macrophages from lung’s lavage (BAL), cell cycle analysis, apoptosis, cell signaling, western blotting, qRT-PCR, microscopy, cell biology, Immuno-Histochemistry, ELISA, killing/cell viability assay, molecular cloning, drug discovery, cell migration, cancer stem cells, transfection, transduction, RNA/DNA isolation, sample preparation for RNAseq.

Worked in collaboration with Signal Rx Pharmaceutical as my mentor “Dr. Donald Durden” was co-founder and CEO of Signal Rx Pharmaceutical. I was involved in cell based assays and preclinical studies using their inhibitors.

My doctoral thesis work is mainly focused on the molecular, biochemical and biophysical characterization of hexokinase from Brugia malayi.