Bone marrow (BM) metastasis remains one of the main causes of death associated with solid tumors as well as with Multiple Myeloma (MM). Targeting the BM niche to prevent or modulate metastasis has not been successful to date. Here we show that stromal cell derived factor-1 (SDF-1/CXCL12) is highly expressed in active MM, as well as in BM sites of tumor metastasis, and report on the discovery of the high affinity anti-SDF-1 PEGylated mirror-image l-oligonucleotide (olaptesed-pegol). In vivo confocal imaging showed that SDF-1 levels are increased within MM cell-colonized BM areas. Using in vivo murine and xenograft mouse models, we document that in vivo SDF-1 neutralization within BM niches leads to a microenvironment that is less receptive for MM cells and reduces MM cell homing and growth, thereby inhibiting MM disease progression. Targeting of SDF-1 represents a valid strategy for preventing or disrupting colonization of the BM by MM cells.