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The therapy of neuroblastoma relies, amongst other things, on administering chemotherapeutics and radioactive compounds, e.g., the (meta-iodobenzyl)guanidine [131I]mIBG. For special applications (conditioning before stem cell transplantation), busulfan and melphalan (M) proved to be effective. However, both drugs are not used for normal chemotherapy in neuroblastoma because of their side effects. The alkylating drug melphalan contains a (Cl-CH2-CH2-)2N- group in the para-position of the phenyl moiety of the essential amino acid phenylalanine (Phe) and can, therefore, be taken up by virtually all kinds of cells by amino acid transporters. In contrast, mIBG isotopologs are taken up more selectively by neuroblastoma cells via the noradrenaline transporter (NAT). The present study aimed at synthesising and studying hybrid molecules of benzylguanidine (BG) and the alkylating motif of M. Such hybrids should combine the preferential uptake of BGs into neuroblastoma cells with the cytotoxicity of M. Besides the hybrid of BG with the dialkylating group (Cl-CH2-CH2-)2N- bound in the para-position as in M (pMBG), we also synthesised mMBG, which is BG meta-substituted by a (Cl-CH2-CH2-)2N- group. Furthermore, two monoalkylating hybrid molecules were synthesised: the BG para-substituted by a (Cl-CH2-CH2-)NH- group (pM*BG) and the BG meta-substituted by a (Cl-CH2-CH2-)NH- group (mM*BG). The effects of the four new compounds were studied with human neuroblastoma cell lines (SK-N-SH, Kelly, and LS) with regard to uptake, viability, and proliferation by standard test systems. The dialkylating hybrid molecules pMBG and mMBG were at least as effective as M, whereas the monoalkylating hybrid molecules pM*BG and mM*BG were more effective than M. Considering the preferred uptake via the noradrenaline transporter by neuroblastoma cells, we conclude that they might be well suited for therapy. [ABSTRACT FROM AUTHOR]

(1) Lysosomal storage diseases are rare inherited disorders with no standardized or commercially available tests for biochemical diagnosis. We present factors influencing the quality of enzyme assays for metachromatic leukodystrophy (MLD) and gangliosidoses (GM1; GM2 variants B and 0) and validate the reliability and stability of testing in a retrospective analysis of 725 samples. (2) Patient leukocytes were isolated from ethylene-diamine-tetra-acetic acid (EDTA) blood and separated for subpopulation experiments using density gradient centrifugation or magnetic cell separation. Enzyme activities in whole leukocyte lysate and leukocyte subpopulations were determined. (3) The enzyme activities in leukocyte subpopulations differed significantly. Compared to lymphocytes, the respective enzyme activities were 2.31–4.57-fold higher in monocytes and 1.64–2.81-fold higher in granulocytes. During sample preparation, a considerable amount of the lysosomal enzymes was released from granulocytes. Nevertheless, with the sample preparation method used here, total leukocyte count proved to be more accurate than total protein amount as a reference unit for enzyme activities. Subsequent analysis of 725 individuals showed clear discrimination of enzyme activities in patient samples (48 MLD; 21 gangliosidoses), with a sensitivity of 100% and specificity of 98–99%. [ABSTRACT FROM AUTHOR]